MICROBIOLOGY AND MOLECULAR BIOLOGY REVIEWS, June 2007, p. 295–347
Copyright © 2007, American Society for Microbiology. All Rights Reserved.
Vol. 71, No. 2
Sponge-Associated Microorganisms: Evolution, Ecology,
and Biotechnological Potential†
Michael W. Taylor,* Regina Radax, Doris Steger, and Michael Wagner
Department of Microbial Ecology, University of Vienna, Althanstr. 14, A-1090 Vienna, Austria
EVOLUTION AND DIVERSITY OF SPONGE-ASSOCIATED MICROORGANISMS...........................................297
Known Diversity of Microorganisms from Sponges ..........................................................................................298
Existing Evidence for Sponge-Specific Microorganisms ...................................................................................299
Census of Sponge-Associated Microorganisms...................................................................................................300
Sponge-Associated Microorganisms: Ancient Partners or Recent Visitors That Have Come To Stay?.....314
Scenario 1: Ancient symbioses maintained by vertical transmission..........................................................314
Scenario 2: Parental and environmental symbiont transmission................................................................316
Scenario 3: Environmental acquisition............................................................................................................316
ECOLOGICAL ASPECTS: FROM SINGLE CELLS TO THE GLOBAL SCALE............................................318
Establishment and Maintenance of Sponge-Microbe Associations.................................................................318
Physiology of Sponge-Associated Microorganisms.............................................................................................320
Other aspects of microbial metabolism in sponges.......................................................................................323
The Varied Nature of Sponge-Microbe Interactions..........................................................................................323
Microorganisms as a food source for sponges ...............................................................................................324
Harming the host: pathogenesis, parasitism, and fouling............................................................................325
The Big Picture: Temporal and Biogeographic Variability in Microbial Communities of Sponges...........326
BIOTECHNOLOGY OF SPONGE-MICROBE ASSOCIATIONS: POTENTIAL AND LIMITATIONS..............329
Biologically Active Chemicals from Marine Sponge-Microbe Consortia and Their Commercial-Scale
Methods for Accessing the Hidden Chemistry of Marine Sponges.................................................................331
Cultivation of metabolite-producing microorganisms ...................................................................................331
Cell separation and metabolite localization....................................................................................................335
Other Biotechnologically Relevant Aspects of Sponges.....................................................................................336
CONCLUSIONS AND FUTURE DIRECTIONS....................................................................................................337
Marine sponges represent a significant component of benthic
communities throughout the world, in terms of both biomass
and their potential to influence benthic or pelagic processes
(73, 74, 124, 220). Sponges (phylum Porifera) are among the
oldest of the multicellular animals (Metazoa) and possess rel-
atively little in the way of differentiation and coordination of
tissues (26, 371). They are sessile, filter-feeding organisms
which, despite a simple body plan, are remarkably efficient at
obtaining food from the surrounding water (290, 308, 443).
The more than 6,000 described species of sponges inhabit a
wide variety of marine and freshwater (somewhat more re-
stricted) systems and are found throughout tropical, temper-
ate, and polar regions (167). Sponges have been the focus of
much recent interest (Fig. 1) due to the following two main
(and often interrelated) factors: (i) they form close associa-
tions with a wide variety of microorganisms and (ii) they are a
rich source of biologically active secondary metabolites. This
increasing research interest has greatly improved our knowl-
edge of sponge-microbe interactions, and yet, as apparent
throughout this article, many gaps remain in our knowledge of
these enigmatic associations. For example, we still lack a clear
picture of microbial diversity—and the factors which influence
it—in these hosts. Similarly, the physiology of most sponge-
associated microorganisms remains unclear, as do many fun-
damental aspects of sponge symbiont ecology. (Throughout
this article, the terms “symbiont” and “symbiosis” are used in
their loosest possible definitions, to refer simply to two [or
more] different organisms that live together over a long period
* Corresponding author. Present address: School of Biological Sci-
ences, Faculty of Science, The University of Auckland, Private Bag
92019, Auckland, New Zealand. Phone: 64 9 373 7599, ext. 82280. Fax:
64 9 373 7416. E-mail: firstname.lastname@example.org.
† Supplemental material for this article may be found at http://mmbr
of time, similar to the original de Bary definition. No judgment
is made regarding benefit to either partner.) Here we aim to
provide a comprehensive review of the current knowledge of
the evolution, ecology, and biotechnological potential of sponge-
We begin with an introduction to the host organism. The
phylum Porifera is a paraphyletic grouping consisting of three
major sublineages (classes), namely, the Hexactinellida (glass
sponges), Calcarea (calcareous sponges), and Demospongiae
(demosponges), with the last group containing the majority of
extant species (38, 167). Sponge architecture is unlike that for
any other taxon, and sponge morphology greatly affects many
aspects of sponge biology, including interactions with micro-
organisms. The basic body plan comprises several different cell
layers (Fig. 2) (371). The outer surface, or pinacoderm, is
formed by epithelial cells known as pinacocytes. Through pores
FIG. 1. Increasing research interest in marine sponge-microorganism associations. (A) Number of publications retrieved from the ISI Web of
Science database by using the following search string: (sponge* or porifera* or demospong* or sclerospong* or hexactinellid*) and (bacteri* or
prokaryot* or microbe* or microbial or microorganism* or cyanobacteri* or archaeon or archaea* or crenarchaeo* or fung* or diatom* or
dinoflagellate* or zooxanthella*) not (surgery or surgical). (B) Number of sponge-derived 16S rRNA gene sequences deposited in GenBank per
year. The 2006 value includes the 184 sequences submitted to GenBank from this article. The search string used to recover sequences was as
follows: (sponge* or porifera*) and (16S* or ssu* or rRNA*) not (18S* or lsu* or large subunit or mitochondri* or 23S* or 5S* or 5.8S* or 28S*
or crab* or alga* or mussel* or bivalv* or crustacea*).
FIG. 2. Schematic representation of a sponge. Arrows indicate the direction of water flow through the sponge. (Adapted from reference 328
with permission of Brooks/Cole, a division of Thomson Learning.)
296 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
(ostia) on the sponge surface, these cells also extend along the
interior canals which permeate the sponge. Inside the sponge,
specialized flagellated cells (choanocytes) form a series of
chambers where feeding takes place. In these chambers, col-
lectively called the choanoderm, the flagellated choanocytes
beat to pump water in through the ostia and along the often
elaborate aquiferous systems within the sponge. Choanocytes
also filter out food particles (including bacteria and microal-
gae) from the water, and these are transferred to the mesohyl,
an extensive layer of connective tissue (Fig. 2). In the mesohyl,
food particles are digested via phagocytosis by another group
of sponge cells, the archaeocytes. These totipotent cells are
capable of differentiating into any of the other sponge cell
types. Also present in the mesohyl of many sponges are dense
communities of microorganisms (106, 430, 471–473). The ex-
istence of these putative symbionts alongside bacterium-digest-
ing archaeocytes is somewhat paradoxical and implies either
recognition of different microbial types by the sponge cells or
shielding of symbiont cells to prevent consumption (482). Once
filtered in the choanocyte chambers, water is eventually ex-
pelled from the sponge via the exhalant opening, or osculum.
It has been estimated that up to 24,000 liters of water can be
pumped through a 1-kg sponge in a single day (443).
Beyond the basic body plan described above, sponge mor-
phology is highly diverse. Inspection of any marine “sponge
garden” will reveal a colorful array of encrusting, branching,
cup-shaped, and massive (amorphous) types (Fig. 3), with in-
dividuals ranging in size from a few millimeters to more than a
meter in diameter (328). Sponge morphology can also reflect
ecological function, as seen in the many cyanobacterium-con-
taining species whose flattened shapes allow optimal light re-
ception for their photosynthetic symbionts (337, 474, 477).
Structural integrity is conferred upon most sponges by siliceous
or calcareous spicules (371), and these skeletal components
are the basis for much of sponge biology and taxonomy. A wide
range of spicule types are secreted, many of which are charac-
teristic of particular taxa (167). Collagenous tissues, such as
spongin, also play a role in providing structural support and,
together with spicules, allow the development of very large
individuals, such as those found among many tropical species.
Sessile organisms such as sponges and other marine inver-
tebrates (including corals and ascidians) rely heavily on the
production of chemicals as a form of defense against natural
enemies, such as predators and competitors. Marine sponges
have attracted particularly intense scrutiny in this regard, with
a wide variety of sponge natural products characterized to date
(see reference 32 and its preceding versions). More novel bio-
active metabolites are obtained from sponges each year than
from any other marine taxon, and a range of pharmacological
properties have been demonstrated (32, 250). Various ecolog-
ical roles have also been proposed for these compounds, in-
cluding defense against predators (20, 55, 275), competitors
(94, 395, 411), fouling organisms (363, 487), and microbes (19,
254, 398). Interestingly, in at least some cases, the compounds
appear to be produced by associated microorganisms rather
than by the sponge (27, 285, 351). Continued investigations of
sponge-derived compounds and their biotechnological and
ecological implications should guarantee vigorous interest in
sponge-microbe associations for some time to come.
Interactions between sponges and microorganisms occur in
many forms. To a sponge, different microbes can represent
food sources (290, 307, 308), pathogens/parasites (16, 171, 199,
455), or mutualistic symbionts (474, 477). Microbial associates
can comprise as much as 40% of sponge tissue volume (427),
with densities in excess of 109microbial cells per ml of sponge
tissue (159, 453), several orders of magnitude higher than
those typical for seawater. The diversity in types of interaction
is matched by the phylogenetic diversity of microbes that occur
within host sponges. It was already evident from early micros-
copy and culturing studies of sponge-associated microbes that
high levels of morphological and metabolic diversity were
present (62, 218, 336, 430, 471–473). The application of mo-
lecular tools over the past decade has greatly extended the
known diversity of microorganisms within these hosts (100,
106, 146, 214, 294, 390, 458). Each of the three domains of life,
i.e., Bacteria, Archaea, and Eukarya, are now known to reside
within sponges. We now consider in detail this enormous di-
versity together with the evolutionary mechanisms driving its
EVOLUTION AND DIVERSITY OF SPONGE-ASSOCIATED
Marine sponges are widely considered the most primitive of
the metazoans, arising at least as early as the Precambrian,
some 600 million years ago (206). According to molecular
FIG. 3. Sponges of diverse size, shape, and color. The encrusting sponge Tedania digitata (left), the branching sponge Axinella cannabina
(center), and the giant barrel sponge Xestospongia testudinaria (right) are shown. The last two images were kindly provided by Armin Svoboda
(Ruhr-Universita ¨t, Bochum, Germany).
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS297
clocks, the divergence of sponges from the ancestors of other
metazoans may have occurred even earlier, around 1.3 billion
years ago (144). During subsequent periods of the Paleozoic
era, sponges accounted for much of the biomass on marine
reefs (167, 491). Today, they remain important members of
both shallow- and deep-water communities, occupying as much
as 80% of available surfaces in some areas (74). Such sustained
evolutionary and ecological success is probably due, at least in
part, to their intimate associations with microbial symbionts.
However, unlike many other studied host-microbe associa-
tions, in which only a very small number of participants are
involved (e.g., squid-Vibrio fischeri , amoeba-Chlamydiae
, and Bugula-“Endobugula” symbioses [142, 210]), it is
apparent that sponge-associated microbial communities can be
highly diverse, with a range of different microorganisms con-
sistently associated with the same host species. In this section,
we describe the extent of this diversity, providing in-depth
phylogenetic analyses of all known sponge-associated microor-
ganisms. We summarize current evidence for the existence of
sponge-specific microorganisms and conclude by considering
whether sponge-microbe associations are evolutionarily an-
cient or are, instead, recently initiated relationships involving
microorganisms which are present in the surrounding sea-
Known Diversity of Microorganisms from Sponges
Prior to this review, the diversity of microorganisms
known from sponges was categorized in 14 recognized bacterial
phyla (and one candidate phylum), both major archaeal lin-
eages, and assorted microbial eukaryotes (145, 148, 477). Se-
quences representing the following bacterial phyla have been
recovered from 16S rRNA gene libraries and/or excised dena-
turing gradient gel electrophoresis (DGGE) bands: Acidobac-
teria, Actinobacteria, Bacteroidetes, Chloroflexi, Cyanobacteria,
Deinococcus-Thermus, Firmicutes, Gemmatimonadetes, Nitro-
spira, Planctomycetes, Proteobacteria (Alpha, Beta, Delta, and
Gammaproteobacteria), Spirochaetes, and Verrucomicrobia (7,
95, 123, 146, 148, 151, 154, 214, 317, 342, 383, 390, 391, 396,
404, 407, 421, 452, 454, 458; S. R. Longford, N. A. Tujula, G. R.
Crocetti, A. J. Holmes, C. Holmstro ¨m, S. Kjelleberg, P. D.
Steinberg, and M. W. Taylor, unpublished data). In addition, a
seemingly sponge-specific candidate phylum, “Poribacteria,”
has also been reported for several sponges (100). The most
frequently recovered sequences in general 16S rRNA gene
surveys of sponges include those from the Acidobacteria, Acti-
nobacteria, and Chloroflexi (148). Members of several bacterial
phyla, namely, the Actinobacteria, Bacteroidetes, Cyanobacteria,
Firmicutes, Planctomycetes, Proteobacteria, and Verrucomicro-
bia, have also been isolated in pure culture from marine
sponges (46, 47, 56, 81, 95, 147, 187, 188, 198, 202, 214, 235,
263, 264, 292, 334, 341, 365, 453, 458). Sequences from the
Chlorobi (green sulfur bacteria) have not been obtained from
sponges, although positive fluorescence in situ hybridization
(FISH) signals were obtained from Rhopaloeides odorabile with
a specific probe for this phylum (458). In contrast to the case
for marine sponges, the (limited) available evidence for fresh-
water species suggests that bacterial diversity and abundance
are both much lower. Only sequences from the Actinobacteria,
Chloroflexi, and Alpha- and Betaproteobacteria were recovered
in a recent 16S rRNA gene library constructed from the fresh-
water sponge Spongilla lacustris (123). Moreover, many of
these sequences were highly similar to those known previously
from freshwater habitats, suggesting that they may not represent
With a few exceptions in the Euryarchaeota (164, 456), ar-
chaea reported from marine sponges are members of the phy-
lum Crenarchaeota (164, 200, 226, 294, 454, 456). Lipid biomar-
kers also suggested the presence of both crenarchaeotes and
euryarchaeotes in a deep-water Arctic sponge, though no phy-
logenetic information was provided in that study (272). The
group I.1A Crenarchaeota are extremely prevalent in marine
environments (180), and almost all sponge-derived archaeal
sequences are affiliated with this group. The best-studied
sponge-associated archaeon is the psychrophilic crenarchaeote
“Candidatus Cenarchaeum symbiosum,” which comprises up
to 65% of prokaryotic cells within the Californian sponge
Axinella mexicana (135, 294, 343, 345).
Eukaryotic microbes also occur in sponges. Sponge-inhabit-
ing dinoflagellates (120, 152, 153, 338, 339, 355, 382, 454, 477)
and diatoms (16, 47, 51, 53, 65, 113, 305, 390, 409, 454) have
been reported, with the latter seemingly most prevalent in
polar regions (16, 51, 53, 113, 305, 409, 454). Freshwater
sponges often contain endosymbiotic microalgae, primarily
zoochlorellae (30, 108, 109, 331, 333, 475, 488). Two previous
reports of cryptomonads in sponges were noted by Wilkinson
(477), while marine sponge-derived fungi are receiving increas-
ing attention due to their biotechnological potential (44, 163,
191). Interestingly, of 681 fungal strains isolated worldwide
from 16 sponge species, most belonged to genera which are
ubiquitous in terrestrial habitats (e.g., Aspergillus and Penicil-
lium) (163). It thus remains unclear in most cases whether such
fungi are consistently associated with the source sponge, or
even whether they are obligate marine species. Compelling
evidence for symbiosis of a yeast with sponges of the genus
Chondrilla was obtained by extensive microscopy studies of
both adult sponge tissue and reproductive structures, with
strong indications of vertical transmission of the yeast symbi-
Little is known about viruses in sponges, although virus-like
particles were observed in cell nuclei in Aplysina (Verongia)
cavernicola (432). It was suggested that these particles could be
involved in sponge cell pathology. Infection of an Ircinia stro-
bilina-derived alphaproteobacterium by a bacteriophage iso-
lated from seawater has also been demonstrated (211), al-
though the propensity of this siphovirus to infect the bacterium
in nature is not known.
In addition to the realization of high microbial diversity per
se, we are now beginning to recognize more subtle patterns of
host-symbiont distribution. For example, it appears that a
given species of sponge contains a mixture of generalist and
specialist microorganisms (390) and that the associated micro-
bial communities are fairly stable in both space and time (105,
390, 391, 454). One particularly interesting pattern to emerge
is the apparent widespread existence of sponge-specific bacte-
rial clusters, i.e., closely related groups of bacteria which are
found only in sponges (146). In the following section, we ex-
amine the published evidence for such clusters.
298TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
Existing Evidence for Sponge-Specific Microorganisms
The notion that marine sponges might contain a specific
microbiota arose some 3 decades ago from the seminal work of
Vacelet et al. and Wilkinson et al. (427, 430, 469, 471–473,
483). Based on electron microscopy and bacterial cultivation
studies, these pioneers of sponge symbiont research proposed
the following three broad types of microbial associates in
sponges: (i) abundant populations of sponge-specific microbes
in the sponge mesohyl, (ii) small populations of specific bac-
teria occurring intracellularly, and (iii) populations of nonspe-
cific bacteria resembling those in the surrounding seawater
(427, 472). One type of bacterial isolate, regarded as a single
species, was recovered from 35 taxonomically diverse sponges
from several geographic regions, but never from seawater (469,
483). Immunological experiments in which these same isolates
cross-reacted with other “sponge-specific” bacteria but not
with seawater isolates were taken as further evidence of sponge
specificity (469). Another significant advance came in 2002,
when Hentschel and coworkers integrated these concepts into
the molecular age (146). They defined sponge-specific clusters
as sponge-derived groups of at least three 16S rRNA gene
sequences which (i) are more similar to each other than to
sequences from other, nonsponge sources; (ii) are found in at
least two host sponge species and/or in the same host species
but from different geographic locations; and (iii) cluster to-
gether irrespective of the phylogeny inference method used
The hypothesis of widespread, sponge-specific microbial
communities put forward by Hentschel and colleagues (146)
was compelling and was constrained only by the limited data
set available at that time. They performed phylogenetic anal-
yses with the 190 publicly available sponge-derived 16S rRNA
gene sequences, the majority of which were from Aplysina
aerophoba, Rhopaloeides odorabile, and Theonella swinhoei.
These three sponges are phylogenetically only distantly related
and were collected from the Mediterranean Sea, the Great
Barrier Reef, and Micronesia/Japan/Red Sea, respectively, yet
they contained largely overlapping microbial communities. To-
gether with the earlier work of Wilkinson and contemporaries
(e.g., see reference 483), these remarkable results suggested
that even unrelated sponges with nonoverlapping geographic
ranges might share a common core of bacterial associates.
Indeed, subsequent studies have lent further weight to this
notion, with numerous reports of similar (and in some cases
sponge-specific) bacteria found in different sponge species
(100, 151, 154, 198, 235, 342, 404, 407). Furthermore, both
cultivation-based and molecular methods have provided evi-
dence for distinct microbial communities between sponges and
the surrounding seawater (151, 265, 334, 391, 472). Taken
together, these results appear to indicate that sponge-associ-
ated microbial communities are indeed unique and at least
partially sponge specific, and the existence of sponge-specific
microorganisms has consequently become something of a par-
adigm in this field.
A total of 14 monophyletic, sponge-specific sequence clus-
ters were identified in the original study of Hentschel et al.
(146). These occurred in the Acidobacteria, Actinobacteria,
Bacteroidetes, Chloroflexi, Cyanobacteria, Nitrospira, and Pro-
teobacteria (Alpha, Delta, and Gammaproteobacteria) and, in
most cases, were strongly supported by bootstrap analyses (in
all cases, the clusters were found with three different tree
construction methods). Three further clusters—each sponge
specific, with the exception of a single nonsponge sequence—
were also identified in the Acidobacteria and in a lineage of
uncertain affiliation (later recognized as Gemmatimonadetes
(146, 499). Overall, 70% of the 190 sponge-derived sequences
available at the time fell into one of these monophyletic clus-
ters or the other. Interestingly, within-cluster 16S rRNA se-
quence similarities ranged down to as low as 77% (146), often
considered indicative of phylum-level differences (170). Sev-
eral subsequent, mostly cultivation-independent studies have
also led to the recovery of apparently sponge-specific se-
quences. Approximately 50% of 16S rRNA gene sequences in
a gene library obtained from the unidentified Indonesian
sponge 01IND 35 were most closely related to sequences de-
rived from other sponges (154). These included members of
the Acidobacteria, Nitrospira, Bacteroidetes, and Proteobacteria,
as well as several sequences in a group of uncertain affiliation
(our analyses indicate that these may be deltaproteobacterial
sequences [see Fig. 8]). A similar situation was reported for
Discodermia dissoluta, whereby three-quarters of 160 retrieved
16S rRNA sequences were most similar to other sponge-de-
rived sequences (342). Conversely, of 21 unique sequences
(each representing a unique restriction fragment length poly-
morphism [RFLP] type) obtained from the Caribbean sponge
Chondrilla nucula, only 5 retrieved other sponge-derived 16S
rRNA sequences during BLAST searches (although with the
advantage of our larger data set, we found indications that
several more of the C. nucula sequences are in fact from
members of sponge-specific clusters) (151). Perhaps the most
impressive sponge-specific cluster to be reported so far is the
candidate phylum “Poribacteria” (100). Fieseler and colleagues
found members of this lineage, which is moderately related to
the Planctomycetes, Verrucomicrobia, and Chlamydiae (446), in
several sponges from geographically diverse locations, but
never in adjacent seawater or sediment samples (100). It will
be especially interesting to see whether “Poribacteria” se-
quences are recovered from other environments in the future.
The sheer number of reports dealing with sponge-specific
microorganisms is compelling. However, one must be cautious
when proposing a sponge-specific cluster. Of crucial impor-
tance is the selection of nonsponge reference organisms for
phylogenetic analyses. In principle, any group of sequences can
appear sponge specific if the most appropriate reference or-
ganisms (i.e., those that are most closely related to the sponge-
derived sequences) are not also included. The length of analyzed
sequences is also of concern, with the level of phylogenetic
information obtainable increasing with sequence length. Every
effort should be made to obtain at least one near-full-length
sequence per sequence type (or operational taxonomic unit).
Decreasing sequence costs render this eminently achievable,
and in many cases, it would only involve performing a few
additional sequencing reactions. These are not new ideas
and we are certainly not the first to advocate the use of
full-length sequences (e.g., see reference 216), but during
our analyses of sponge-derived 16S rRNA sequences, it be-
came apparent that many of these sequences are rather
short and therefore phylogenetically not particularly infor-
mative. Indeed, we encountered many problems with inser-
VOL. 71, 2007SPONGE-ASSOCIATED MICROORGANISMS299
tion of short sponge-derived sequences into our phyloge-
netic trees, and in some cases, we were not even certain of
their phylum-level affiliation.
Census of Sponge-Associated Microorganisms
Increasing interest in sponge-microbe associations has re-
sulted in a concomitant increase in the amounts of 16S rRNA
sequence data obtained from sponges (Fig. 1B). There are
currently ?1,500 sponge-derived 16S rRNA gene sequences
available in GenBank (http://www.ncbi.nlm.nih.gov/), in con-
trast to only 190 such sequences available for the 2002 study by
Hentschel et al. (146). We carried out an extensive phyloge-
netic analysis of all currently available sponge-derived 16S
rRNA gene sequences, with two main objectives, as follows: (i)
to provide an overview of microbial diversity in sponges and
(ii) to critically assess the occurrence of monophyletic, sponge-
specific sequence clusters. As mentioned above, such clusters
are often discussed, yet their existence has not been reevalu-
ated rigorously in light of the rapidly expanding 16S rRNA
sequence databases. It is thus unclear whether these clusters
are truly sponge specific or merely reflect a greater sampling
effort for these communities than for others.
We began, using the ARB program package (217), by estab-
lishing an encompassing database that contains all sponge-
derived 16S rRNA sequences which were available in GenBank
on 28 February 2006. In addition to these 1,499 sequences
(plus 11 18S rRNA sequences amplified from eukaryotic mi-
crobes in sponges), we contributed a further 184 bacterial and
archaeal sequences from three hitherto unstudied sponges,
FIG. 4. 16S rRNA-based phylogeny showing representatives of all bacterial and archaeal phyla from which sponge-derived sequences have been
obtained. Sponge-derived sequences are shown in bold, with additional reference sequences also included. The displayed tree is based on a
maximum likelihood analysis. Bar, 10% sequence divergence.
300TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
namely, Agelas dilatata, Plakortis sp. (both from the Bahamas;
kindly provided by U. Hentschel), and Antho chartacea (from
southeastern Australia). Preliminary phylogenetic analyses
identified members of putative sponge-specific clusters, and for
each cluster, the most similar nonsponge sequences were re-
trieved by BLAST searches (from both regular NCBI and
environmental genome databases) and imported into ARB for
subsequent alignment (automatic and manual). The resulting
ARB database, containing an alignment of all sponge-derived
sequences and their nearest relatives (together with annotated
information [e.g., host species and collection location] for the
sponge sequences), is available upon request. Extensive phy-
logenetic analyses (see the supplemental material for details)
were conducted, taking all (n ? 1,694) sponge-derived se-
quences into account. In order to rigorously test the existence
of monophyletic, sponge-specific sequence clusters, we em-
ployed multiple tree construction methods (maximum likeli-
hood, neighbor joining, and maximum parsimony), together
with the use of various sequence conservation filters and cor-
In total, sequences representing 16 bacterial phyla and both
major archaeal lineages (Crenarchaeota and Euryarchaeota)
have been recovered from sponges (Fig. 4). In addition to
those known prior to this study (Acidobacteria, Actinobacteria,
Bacteroidetes, Chloroflexi, Cyanobacteria, Deinococcus-Ther-
mus, Firmicutes, Gemmatimonadetes, Nitrospira, Planctomycetes,
“Poribacteria,” Proteobacteria [Alpha-, Beta-, Delta-, and Gam-
maproteobacteria], Spirochaetes, Verrucomicrobia, and Chlorobi
FISH signals), we report for the first time the presence in
sponges of 16S rRNA sequences affiliated with the phylum
Lentisphaerae and the candidate phylum TM6. The number of
sequences representing each phylum varied widely, from single
sequences for the Lentisphaerae and TM6 to more than 250
sequences for each of the Actinobacteria, Alphaproteobacteria,
and Beta/Gammaproteobacteria (Table 1). The proportions of
sequences derived from cultivated versus noncultivated micro-
organisms also varied greatly among phyla.
The phylogenetic analyses presented here strongly support
the existence of monophyletic, sponge-specific 16S rRNA se-
quence clusters. These occurred in many of the bacterial and
archaeal phyla found in sponges, with approximately one-third
(32%) of all sponge-derived sequences falling into such clus-
ters (Table 1; Fig. 5 to 15; also see the supplemental material).
If sequences derived from cultured isolates are excluded, this
figure rises to 42%. This result was expected since tightly
linked symbionts—those presumed to occur in sponge-specific
clusters—are likely difficult to cultivate and therefore under-
represented in culture collections. Several additional clusters
each contained a single nonsponge sequence, with the extra
sequences often, but not always, obtained from marine envi-
TABLE 1. Summary of all publicly available sponge-derived 16S rRNA sequence data (as of 28 February 2006) plus 184 bacterial and
archaeal sequences contributed from this article
Total no. of
No. of sequences
of ?1,000 bp
No. of sequences% of sequences in clusters obtained from:
Sponges and one
Chloroflexi et al.
Total1,694 6081,259 435546 7443
aNumbers in parentheses are inclusive of clusters which are supported by only two of three tree construction methods.
bIncludes both 18S rRNA- and 16S rRNA (plastid)-derived sequences.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS301
FIG. 5. 16S rRNA-based phylogeny of sponge-associated cyanobacteria and chloroplasts. Sponge-derived sequences are shown in bold. The
displayed tree is a maximum likelihood tree constructed based on long (?1,000 nucleotides) sequences only. Shorter sequences were added using
the parsimony interactive tool in ARB and are indicated by dashed lines. Shaded boxes represent sponge-specific monophyletic clusters, as defined
302 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
ronments. It is also possible that sponge-specific microbes are
more prevalent in those sponges which contain very dense
microbial communities (Ute Hentschel, personal communica-
tion), i.e., the so-called bacteriosponges or high-microbial-
abundance sponges (148, 430). Due to a lack of microbial
abundance data for most host sponges, we did not attempt to
take this factor into account during our analyses. Overall, while
representation of sequences in sponge-specific clusters was
quite high, it should be noted that the proportions of se-
quences falling within such clusters differed greatly among the
More than three-quarters of the 119 available sponge-de-
rived Cyanobacteria sequences fell into monophyletic, sponge-
specific clusters (Table 1; Fig. 5). Most of these were in two
clusters, with one comprising 25 sequences from at least 7
sponge species and the other comprising 52 sequences from 21
sponges. The latter cluster represents the recently described
candidate species “Candidatus Synechococcus spongiarum”
(426) and was the sole Cyanobacteria cluster in the study of
Hentschel et al. (146), while the former corresponds to the
filamentous cyanobacterium Oscillatoria spongeliae (39, 157).
Sequences representing O. spongeliae were not available for
the 2002 study. Several other, smaller clusters are also evident
among the cyanobacteria (Fig. 5). Additionally, in a number of
cases, microalgal plastids have also been amplified using 16S
Another bacterial phylum containing many sponge-specific
sequence clusters is the Chloroflexi (Table 1; Fig. 6). Of the 109
sponge-derived sequences analyzed, 62% comprised such clus-
ters, while the occurrence of a further 13% of sequences in
clusters was weakly supported. In the new analyses, all but one
of the members of a sponge-specific cluster described by
Hentschel and coworkers (146) remained in a cluster, although
these sequences were now dispersed over four different clus-
ters. Such movement of sequences was frequently observed
and is not surprising given the much larger data set at our
disposal now (i.e., many new related sequences, both sponge-
and non-sponge-derived, were included in the phylogenetic
analyses described here). None of the sponge-derived se-
quences were closely related to the few described Chloroflexi
species, although many were similar to sequences from uncul-
tivated organisms, particularly from marine environments
Interestingly, many sponge-derived 16S rRNA sequences
formed exclusive monophyletic clusters with sequences ob-
tained from corals (Table 1). This was particularly apparent for
the Acidobacteria and Deltaproteobacteria (Fig. 7 and 8, respec-
tively) but was also evident for the Gemmatimonadetes (Fig. 9)
and Nitrospira (Fig. 10). No coral-derived sequences shared
monophyletic clusters with sponge sequences in the original
study of Hentschel et al. (146), no doubt reflecting the fact that
most of the relevant coral sequences were deposited in GenBank
since then. It is too early to speculate whether some sort of
marine invertebrate-specific sequence cluster exists, but fur-
ther sampling of taxa such as ascidians and bryozoans should
help to resolve this issue. A study of two marine macroalgae
and the cooccurring sponge Cymbastela concentrica gave no
indication of specific clusters spanning these taxonomically
disparate groups (Longford et al., unpublished data).
Sponge-specific sequence clusters were not prevalent for,
among others, the Bacteroidetes (see Fig. S1 in the supplemen-
tal material) and Firmicutes (see Fig. S2 in the supplemental
material), perhaps reflecting the relatively high proportions of
sequences derived from cultivated organisms in these phyla.
We report for the first time the recovery of Lentisphaerae
(Fig. 11) and candidate phylum TM6 (Fig. 12A) sequences
from sponges. Each phylum was represented by a single 16S
rRNA sequence, from the marine sponges Plakortis sp. and
Antho chartacea, respectively, and it cannot be ruled out that
these represent contaminating sequences from the sur-
rounding environment (although arguably this also applies
to many, more commonly recovered sequence types). The
Lentisphaerae phylum comprises part of the so-called Planc-
tomycetes-Verrucomicrobia-Chlamydiae (PVC) superphylum
(446), with sponge-derived sequences from the superphylum
additionally being found in the Verrucomicrobia, Planctomyce-
tes, and “Poribacteria” (Fig. 11). Members of the superphylum
are frequently associated with eukaryotes. There is also a
group of uncertain affiliation which falls near the PVC super-
phylum (but without strong bootstrap support) during phylo-
genetic analyses. This group includes sequences from many
sponges, such as Agelas dilatata, Aplysina aerophoba, Discoder-
mia dissoluta, and Theonella swinhoei. Those sequences most
closely related to the sponge sequences are also from marine
Several large sponge-specific clusters were found among the
Actinobacteria sequences, particularly in the family Acidimicro-
biaceae (Fig. 13). The largest comprised 54 sequences obtained
from sponges from the Caribbean (Agelas dilatata, Discodermia
dissoluta, Plakortis sp., and Xestospongia muta), Indonesia
(Xestospongia testudinaria), the Red Sea (Theonella swinhoei),
and the South China Sea (Dysidea avara). None of the se-
quences within this cluster were obtained from cultured bac-
teria, with the nearest (but still distantly related) cultured ac-
parvicella and the acidophilic Acidimicrobium spp. (Fig. 13).
Although not representing a sponge-specific cluster, the
group of sequences affiliated with the marine Pseudovibrio spp.
within the Alphaproteobacteria deserves special mention (Fig.
14). Members of this genus are frequently found in sponge-
by Hentschel et al. (146), i.e., a group of at least three sponge-derived 16S rRNA gene sequences which (i) are more similar to each other than
to sequences from other, nonsponge sources, (ii) are found in at least two host sponge species and/or in the same host species but from different
geographic locations, and (iii) cluster together irrespective of the phylogeny inference method used (all clusters shown here also occurred in
neighbor-joining and maximum parsimony analyses). Names outside wedges of grouped sequences represent the sponges from which the relevant
sequences were derived; the number in parentheses indicates the number of sequences in that wedge. Filled circles indicate bootstrap support
(maximum parsimony, with 100 resamplings) of ?90%, and open circles represent ?75% support. The outgroup (not shown) consisted of a range
of sequences representing several other bacterial phyla. Bar, 10% sequence divergence.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS303
derived cultivation-based and molecular studies (95, 96, 147,
187, 198, 263, 453), and there is strong evidence for its being a
true sponge symbiont (95, 453).
Only 28% of sponge-derived Archaea sequences fell into
well-supported sponge-specific clusters (Fig. 15), although
the fact that almost all of these were within the group I.1A
Crenarchaeota bears testimony to their high degree of phylo-
genetic relatedness. The recently isolated ammonia-oxidizing
archaeon “Candidatus Nitrosopumilus maritimus” (192) is the
only cultivated member of this group, with the well-studied
FIG. 6. 16S rRNA-based phylogeny of sponge-associated Chloroflexi organisms. Details are the same as those provided for Fig. 5, with the
following additions. Shaded boxes contained within dotted lines represent sponge-specific clusters supported by only two tree construction methods
(ML, maximum likelihood; MP, maximum parsimony; and NJ, neighbor joining), and new sequences from our laboratory have the prefix “AD”
(for the sponge Agelas dilatata), “AnCha” (Antho chartacea), or “PK” (Plakortis sp.).
304TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
FIG. 7. 16S rRNA-based phylogeny of sponge-associated Acidobacteria organisms. Details are the same as those provided for Fig. 5 and 6, with
the following two additions. Open boxes represent monophyletic clusters containing sponge-derived sequences and a single, nonsponge origin
sequence, and open boxes with asterisks outside them signify clusters containing only sponge- and coral-derived sequences (the number of asterisks
corresponds to the number of coral-derived sequences within the cluster).
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS305
(but still uncultivated) archaeon “Ca. Cenarchaeum symbio-
sum” being the best known sponge-associated member. A ge-
nome project for the latter has recently been completed (134).
At the time of sequence collection, 44 archaeal sequences had
been recovered from sponges, all of which were marine
sponges (Table 1; Fig. 15). All but one of these was from the
Crenarchaeota, with a single Euryarchaeota sequence from the
Great Barrier Reef sponge Rhopaloeides odorabile (456). An
FIG. 8. 16S rRNA-based phylogeny of sponge-associated Deltaproteobacteria organisms. Details are the same as those provided for Fig. 5 to 7.
306TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
article which appeared in mid-2006 (whose sequences were not
available on 28 February 2006 and were therefore not included
in our study) reported more euryarchaeotal sequences from
various sponges, although the majority of sequences in that
study were still affiliated with the Crenarchaeota (164).
All sponge-derived 16S rRNA sequences available on 28
February 2006 were analyzed phylogenetically, but for practi-
cal reasons the larger trees are available only in the supple-
mental material. Broadly speaking, the results of our analyses
are consistent with the earlier study by Hentschel et al. (146),
with, for example, the Actinobacteria, Nitrospira, and Acidobac-
teria still well represented by sponge-specific microorganisms.
As could be expected, some sponge-specific clusters from the
2002 study now form parts of several new clusters, while others
do not comprise clusters at all in the new data set. Conversely,
the addition of more sequences meant that many formerly
single sequences are now in specific clusters with other sponge-
FIG. 9. 16S rRNA-based phylogeny of sponge-associated Gemmatimonadetes organisms. Details are the same as those provided for Fig. 5 to 7.
FIG. 10. 16S rRNA-based phylogeny of sponge-associated Nitrospira organisms. Details are the same as those provided for Fig. 5 to 7.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS307
FIG. 11. 16S rRNA-based phylogeny of sponge-associated Verrucomicrobia, Planctomycetes, Lentisphaerae, and “Poribacteria” organisms and of
a lineage of uncertain affiliation. These and associated lineages comprising the PVC superphylum (446) are shown. Details are the same as those
provided for Fig. 5 to 7.
308 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
Very few sequences were available from sponge-associated
eukaryotic microbes at the time of database establishment
(since then, some 45 18S rRNA sequences derived from
sponge-associated fungi have been deposited in GenBank).
Those that are included in our database include 9 16S rRNA
sequences derived from diatom chloroplasts (Fig. 5) and 11
18S rRNA sequences obtained from diatoms and dinoflagel-
lates. All but one of the 18S rRNA sequences were obtained
from Antarctic sponges (454), with the remaining sequence
representing a zooxanthella (Symbiodinium sp.) from the Pa-
lauan sponge Haliclona koremella (49).
We endeavored to be as thorough and as careful as possible
throughout our analyses, yet there remain some caveats to our
results. Despite extensive BLAST searches using members of
all putative sponge-specific clusters, it is not inconceivable that
we failed to include some key sequences which would have
broken up otherwise specific sponge clusters. Another factor
relates to the short lengths of many sponge-derived 16S rRNA
sequences. We constructed our trees using only sequences
longer than 1,000 bp, but more than two-thirds of all sponge-
derived sequences are shorter than this (Table 1), and we
added these via the parsimony interactive tool in ARB. In
principle, this method allows the insertion of short sequences
without changing tree topology (217). However, when many
short sequences are added at once, they can influence each
other’s positioning (and potentially bias the analysis towards
the formation of sponge-specific clusters). We attempted to
gauge the severity of this problem by (for a selection of se-
quences) sequentially adding and removing individual short
sequences and comparing their placement to the outcome
FIG. 12. 16S rRNA-based phylogeny of sponge-associated members of the candidate phylum TM6 (A), Deinococcus-Thermus organisms (B),
and Spirochaetes organisms (C). Details are the same as those provided for Fig. 5 to 7. (B) In our analyses, the position of clone Dd-spU-11 (from
the sponge Discodermia dissoluta) was not stable, and we are not certain of its phylogenetic affiliation.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS309
when they were all added at once. The results were highly
consistent, but it should not be assumed that this will always be
the case. The alternative is to perform the entire phylogenetic
analysis with short sequences and to truncate longer sequences
to leave only the homologous region; this results in the loss of
much phylogenetic information and is not recommended un-
der any circumstances (216). Again, we reiterate the impor-
tance of obtaining at least one near-full-length sequence for
each operational taxonomic unit obtained. This is not possible
in some cases (e.g., excised DGGE bands) but is feasible in
It is prudent to consider whether the apparent occurrence of
sponge-specific sequence clusters could have a more dubious
origin, namely, laboratory contamination. Theoretically, a 16S
FIG. 13. 16S rRNA-based phylogeny of sponge-associated Actinobacteria organisms belonging to the family Acidimicrobiaceae. Other sponge-
derived actinobacteria are shown in Fig. S3 in the supplemental material. Details are the same as those provided for Fig. 5 to 7.
310 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
FIG. 14. 16SrRNA-basedphylogenyofsponge-associatedAlphaproteobacteriaorganismsaffiliatedwiththegenusPseudovibrioanditsrelatives.Other
sponge-derived alphaproteobacteria are shown in Fig. S4 and S5 in the supplemental material. Details are the same as those provided for Fig. 5 to 7.
FIG. 15. 16S rRNA-based phylogeny of sponge-associated archaeal organisms. Details are the same as those provided for Fig. 5 to 7.
312 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
rRNA gene-containing plasmid or PCR product could, if ac-
cidentally spread to DNAs from several sponges in the same
laboratory, appear to form its own sponge-specific cluster.
However, the available evidence strongly suggests that this is
not the case, since many or most clusters contain sequences
originating from several independent laboratories.
With almost 1,700 sponge-derived 16S rRNA sequences fall-
ing into some 16 or more bacterial and archaeal phyla, we
sought to address the following question: how well sampled are
marine sponge-associated microbial communities? If current
studies are recovering mainly sequences which were previously
obtained from sponges (as the presence of sponge-specific
clusters might imply), then we may have already uncovered
most of the microbial diversity in these hosts, suggesting
that our current descriptive phase might be nearing its log-
ical conclusion. Unfortunately, the available data are insuf-
ficient to satisfactorily address this issue for sponges. In a
recent article in this journal, Schloss and Handelsman (348)
used the program DOTUR to estimate richness at different
levels of phylogenetic relatedness for each bacterial phylum
represented in the Ribosomal Database Project (61). To per-
form an analogous study with the sponge symbiont data set, we
were restricted to sequences which met the following criteria:
(i) they were part of attempts at extensive microbial commu-
nity surveys using general 16S rRNA gene primers for the
construction of clone libraries; (ii) they overlapped a sufficient
distance to be useful (Escherichia coli positions 100 to 500
would have been appropriate for a reasonable portion of the
sponge data set); and (iii) they were not obtained from pre-
screened gene libraries (e.g., by RFLP analysis), as this would
heavily bias results—thus, all collected sequences must have
been deposited in GenBank. After applying these (in our eyes)
minimal criteria, only 317 sequences (of 1,694) were deemed
suitable for use with DOTUR or similar programs. For many
phyla, only a few sequences were retained (e.g., for Cyanobac-
teria, 8 of 119 sequences were kept, and for Alphaproteobacte-
ria, 21 of 311 sequences were kept), precluding meaningful
analyses. Furthermore, even if 50 or more sequences were
suitable (as in the case of the Beta/Gammaproteobacteria),
these were not necessarily representative of the known
(sponge-derived) diversity within that phylum, again prevent-
ing the drawing of meaningful conclusions. Although statisti-
cally robust analyses are therefore not possible at this stage,
data from two recent studies can add greatly to this discussion.
In the first, Lopez and colleagues at the Harbor Branch Ocean-
ographic Institution (213) obtained more than 700 sequences
from 20 different sponge-derived gene libraries by using gen-
eral 16S rRNA primers, with the vast majority of these belong-
ing to phyla already obtained from sponges, such as Chloroflexi,
Cyanobacteria, Nitrospira, Planctomycetes, and Spirochaetes. Of
the recovered sequences, Epsilonproteobacteria was the only
major taxonomic group not previously obtained from sponges.
In another study, examining the Adriatic sponges Chondrilla
nucula and Tethya aurantium, Thiel and coworkers recov-
ered representatives of only known sponge-associated phyla
(Acidobacteria, Actinobacteria, Bacteroidetes, Cyanobacteria, Gem-
matimonadetes, Planctomycetes, Proteobacteria [Alpha-, Beta-,
Delta-, and Gammaproteobacteria], Spirochaetes, and Verru-
comicrobia) (404; V. Thiel, T. Staufenberger, and J. F. Imhoff,
presented at the 11th International Symposium on Microbial
Ecology, Vienna, Austria, 20 to 25 August 2006). The lack of
new phyla in these data allow one to speculate that the major-
ity of sponge-associated microorganisms may have already
been encountered in gene libraries, at least at the phylum level.
However, two major caveats exist. Although we may arguably
be nearing the point of diminishing returns with respect to
using current techniques to recover novel lineages from
sponges (i.e., gene libraries constructed using general 16S
rRNA primers), it is highly likely that the use of phylum-
specific primers and/or metagenomic (i.e., PCR-independent)
approaches will reveal phyla previously unknown to exist in
these hosts or even unknown to science (e.g., “Poribacteria”)
(100). To our knowledge, there is no example of a sponge for
which the results of general versus specific 16S rRNA gene
libraries have been compared. A second point is that few gene
libraries, including those from sponges, are sequenced to full
coverage, and it is possible that the recurring sequences ob-
tained from different sponges are merely those that are most
abundant (or those that PCR is most biased toward) in each
sponge, with the unsequenced remainder of the library poten-
tially contributing new sequence types. The advent of high-
throughput sequencing technologies (e.g., see reference 227)
offers the potential to sequence gene libraries to much greater
depth, illuminating the rare biosphere within sponges (376).
Statistical comparisons of microbial community composi-
tions allow for the inclusion of more sequences (relative to
species richness estimates via DOTUR) due to less stringent
selection criteria. We thus used the so-called parsimony test,
implemented in the program TreeClimber (347), to compare
our three new gene libraries (from the sponges Agelas dilatata,
Antho chartacea, and Plakortis sp.) with selected sponge-
derived libraries from the literature and those deposited in
GenBank. The parsimony test compares phylogenetic trees
rather than sequence data per se (228, 347), and various tree
construction algorithms can be employed. Our criteria for se-
quence inclusion were that (i) general 16S rRNA gene primers
were used and (ii) at least 25 sequences were available from
each library. The main caveats are that prescreening of clones
(e.g., by RFLP analysis) with subsequent representation of
each operational taxonomic unit by a single sequence prevents
strict application of the parsimony test (347), while low se-
quencing coverage of some libraries may obscure true similar-
ities or differences among libraries by missing overlapping or
distinct sequences, respectively. With these considerations in
mind, we compared the three libraries obtained from this study
with those from the marine sponges Theonella swinhoei (146),
Aplysina aerophoba (146), Rhopaloeides odorabile (458), Cym-
bastela concentrica (Longford et al., unpublished data), Disco-
dermia dissoluta (342), and Chondrilla nucula (151) and the
freshwater sponge Spongilla lacustris (123). An initial analysis
comprising all 10 libraries yielded a highly significant (P ?
0.001) result (i.e., the differences in sequence composition
among the various libraries were not due to chance). Likewise,
comparisons of the marine versus freshwater (S. lacustris) li-
braries, as well as comparisons among the marine libraries and
among broad geographic locations, were all highly significant.
The usefulness of such analyses should increase as more 16S
rRNA gene libraries are sequenced from sponges (and with
greater sequencing coverage), including multiple species from
the same location and/or from the same genus or family.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS313
Sponge-Associated Microorganisms: Ancient Partners
or Recent Visitors That Have Come To Stay?
Based largely upon arguments centering on immunological
evidence dating back to the 1980s (469), it is often stated that
sponge-bacterium symbioses have existed for as long as 600
million years. This would date such associations back to the
Precambrian, prior to the bulk of taxonomic radiation in
sponges. Moreover, given the likely basal position of sponges
in the metazoan phylogenetic tree (38, 133), this would pre-
sumably make sponges and microorganisms the most ancient
of all metazoan-microorganism associations. So what is the
evidence for this oft-cited ancient symbiosis? In his 1984 study,
on which the majority of these arguments rest, Wilkinson used
a collection of 296 sponge isolates which, on the basis of mor-
phological and physiological characteristics, comprised one
bacterial species (469). In addition, 128 seawater and nonspe-
cific sponge isolates were included as control strains. It is
important to note that the sponge-specific isolates were ob-
tained from phylogenetically distant sponges in widely sepa-
rated geographical regions. From seven of the specific isolates
and five of the others, Wilkinson prepared antisera and per-
formed agglutination tests. Many of the “sponge-specific”
strains reacted positively in these tests to one or more of the
antisera derived from sponge-specific bacteria, but none of
them reacted with sera derived from non-sponge-specific
strains, nor did cross-reactions occur between the 128 non-
sponge-specific strains and the sera prepared from sponge-
specific bacteria. The implication of these results was that the
studied, widespread, sponge-specific bacterium did indeed
form a single species group distinct from isolates found in the
surrounding seawater (469). According to Wilkinson, the most
logical explanation for the occurrence of this specific bacterial
type in such diverse hosts and locations was that these bacteria
became associated with an ancestral sponge prior to the evo-
lution of current sponge classes (i.e., during the Precambrian).
One should bear in mind, however, that the enormous com-
plexity of microbial communities in seawater could have led to
this bacterium being missed in Wilkinson’s culture libraries.
In the 22 years since the Wilkinson study, a wealth of mo-
lecular data has become available for sponge-associated mi-
croorganisms, ranging from sequences of single genes to an
entire genome (for the archaeon “Candidatus Cenarchaeum
symbiosum”) (134). Here we ponder whether such data can be
exploited to address the issue of the ancientness (or otherwise)
of sponge-microbe associations. First, we consider some of the
many possible evolutionary scenarios (summarized in Fig. 16),
Scenario 1: Ancient symbioses maintained by vertical trans-
mission. A given sponge-specific cluster in the phylogenetic
tree of life may contain 16S rRNA gene sequences derived
from distantly related, geographically disparate sponge species.
If the microorganisms represented by these sequences do not
occur outside sponges today, then the ancestral strain (the
future symbiont) may have first inhabited a sponge during one
or several colonization events prior to sponge speciation (?600
million years ago) (the Precambrian acquisition hypothesis of
Wilkinson ). Such a symbiosis could have been main-
tained in the intervening years via vertical transmission (see
“Establishment and Maintenance of Sponge-Microbe Associ-
ations”), and the microbes evolved to become sponge (or even
species) specific. A related but subtly different hypothesis is
that an association could still be ancient but not predate the
bulk of sponge speciation. In this scenario, it is conceivable
that one sponge could have been colonized very early on,
resulting in the evolution of a sponge-adapted microorganism.
Millions (or hundreds of millions) of years later, this microbe
could have spread across the oceans and, upon encountering
other sponges, colonized them. Perhaps it is no longer present
in seawater, or perhaps it is still there but in very small num-
bers. Yet another scenario is that today’s sponge-specific mi-
crobes were once a generalist marine species, thriving in all
marine ecosystems, including sponges. Those strains that in-
habited sponges have since evolved to become genetically dis-
FIG. 16. Summary of various evolutionary scenarios for sponge-microorganism associations.
314TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
tinct from their free-living counterparts. Support for these
scenarios comes from another quarter, with various fatty acids
of likely microbial origin occurring in a wide range of sponges
irrespective of host phylogeny or geographic location (401,
403). The apparent absence of some of these biomarkers from
marine sediments and seawater led to the suggestion that the
compounds and their microbial producers have been present in
the sponges since ancient times (403).
It is likely that any ancient sponge-microbe symbiosis would
be obligate for one or both partners, potentially involving a
reduction in microbial genome size if the symbiont has devel-
oped a nutritional dependence on its host. This has been dem-
onstrated for many obligate insect endosymbionts (e.g., see
references 252, 437, and 501), but it is unknown whether such
tight host-symbiont coupling occurs in sponges. Integration of
host and symbiont genomes was discussed in the sponge con-
text by Sara and colleagues (337), while a recent paper offers
evidence for lateral gene transfer from a fungus to the mito-
chondrion of its host sponge (327). Such gene transfer would
not be without precedent among marine invertebrates, as it is
believed that the ascidian Ciona intestinalis laterally ac-
quired a cellulose synthase gene from a bacterium (253).
Future genome sequencing of sponges and their microbial
associates should offer valuable insights into the nature of
As noted earlier, not all sponge species harbor abundant
microbial communities, and it is worthwhile to take a moment
to consider these organisms. Freshwater sponges, for example,
typically contain a paucity of microbial associates, and it has
been suggested that this is due to an obligate requirement for
sodium ions by the symbiotic bacteria (469). When freshwater
sponges colonized their new habitat from the sea some 20 to 50
million years ago, it is presumed that existing symbionts were
lost. Many marine sponges also harbor only relatively small
numbers of microorganisms. These so-called low-microbial-
abundance sponges (148) often cooccur with the high-micro-
bial-abundance bacteriosponges, so habitat variation cannot be
invoked as an explanation for these differences. Whether these
sponges once contained, but later lost, large communities of
microbial symbionts is unknown. It is also unknown whether
the (comparatively few) microorganisms in low-microbial-
abundance sponges are phylogenetically similar to those in
their high-microbial-abundance counterparts.
Based on sequence information already at hand, the nearest
we can come to addressing these and the following hypotheses
is to consider estimated rates of 16S rRNA evolution for mem-
bers of given sponge-specific clusters and to attempt to infer
when the last common ancestor of sponge-specific microbes
from different sponges might have occurred. If one assumes
equal mutation rates in different bacterial lineages and asserts
that a 1 to 2% 16S rRNA sequence difference corresponds to
approximately 50 million years of evolution (259), then se-
quence differences of at least 10% would be required to place
a common ancestor of these organisms back in the late Pre-
cambrian (?600 million years ago). Here we consider two
examples, the cluster representing the cyanobacterium “Can-
didatus Synechococcus spongiarum” (426) and the “Poribacte-
ria” (100). The “Ca. Synechococcus spongiarum” cluster is one
of the largest of all sponge-specific sequence clusters, is well
supported by all tree construction methods, and contains 52
sequences from 21 sponges located around the world (Fig. 5).
We chose three of these sequences as an example, derived
from the sponges Theonella conica (sampled from east Africa;
GenBank accession number AY701309), Aplysina aerophoba
(from the Mediterranean Sea; GenBank accession number
AJ347056), and Antho chartacea (from southeastern Australia;
GenBank accession number EF076240). The minimum pair-
wise 16S rRNA similarity among these sequences (after cor-
recting for different sequence lengths) is 97.9%. This is a very
minor difference when one considers the phylogenetically dis-
parate hosts (the last two sponges are in different orders, while
T. conica is in a different subclass) and their geographically
distinct locations. Even if one assumes that cyanobacteria
evolve very slowly, we argue that greater sequence divergence
would be expected if these bacteria had indeed been living
(separately) within their host sponges for 600 million years.
This should be especially true for endosymbiotic microorgan-
isms, which are believed to evolve more rapidly due to in-
creased fixation of mutations within their small populations
(259). These members of the “Ca. Synechococcus spongiarum”
cluster may therefore have a much more recent common ori-
gin, reflecting a role of horizontal (i.e., environmental) trans-
mission consistent with scenario 2 or 3 in Fig. 16. However,
consideration of other sequences within the same cluster can
yield a quite different result. The two least similar sequences
within the “Ca. Synechococcus spongiarum” cluster are only
?93% similar, suggesting a much older separation of these
particular strains. A comparable degree of similarity is seen in
comparing sequences from the “Ca. Synechococcus spongia-
rum” cluster with those from free-living relatives. We suggest
that a combination of vertical and horizontal symbiont trans-
mission (scenario 2) could explain the observed data. Possible
vectors responsible for horizontal symbiont transmission could
include sponge-feeding animals, such as fishes and turtles (205,
274), analogous to the coral-feeding fireworm Hermodice
carunculata, which acts as a vector for the coral pathogen Vibrio
In our second example, we consider the “Poribacteria.” At
first glance, there appears to be a strong case for an evolution-
arily ancient relationship between these bacteria and their
sponge hosts. The members of this monophyletic, exclusively
sponge-specific bacterial lineage differ in their 16S rRNA se-
quences by up to 15% and are some 20% dissimilar to their
nearest nonsponge relative (derived from Antarctic sediment)
(Fig. 11). Such high divergence within the cluster, together
with the low similarity to the next most similar known organ-
ism, is suggestive of an ancient symbiosis with sponges. How-
ever, the two least similar “Poribacteria” sequences were taken
from closely related (same family) sponges collected at the
same Bahamas location, perhaps indicating horizontal symbi-
ont transfer between these hosts. If the associations were an-
cient and involved strict coevolution of host and symbiont, then
their respective phylogenies would be more congruent, with
the least similar microbes being hosted by the least similar
sponges. Furthermore, the long naked branch leading to the
“Poribacteria” in the 16S rRNA tree could potentially be ex-
plained by faster rates of evolution in these bacteria. “Pori-
bacteria” are a sister phylum to the Planctomycetes (446), which
are sometimes believed to exhibit higher rates of evolution
than other lineages (392). Like the case for “Ca. Synechococ-
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS 315
cus spongiarum,” a combination of vertical and horizontal sym-
biont transmission is thus the most likely scenario here, al-
though the acquisition of these bacteria exclusively from the
environment also cannot be ruled out.
Perhaps the most convincing evidence for a long-standing,
symbiotic relationship between sponges and at least some mi-
croorganisms comes from demonstrations of coevolution. De-
spite difficulties in addressing this issue due to the phylogenetic
complexity of sponge-associated microbial communities, sev-
eral authors have now shown coevolution between sponges and
microbes. In the first study, the mitochondrial cytochrome
oxidase subunit 1 (CO1) gene and its bacterial homolog were
amplified from several halichondrid sponges and their associ-
ated bacteria (98). A CO1-based phylogenetic tree of six pu-
tatively alphaproteobacterial symbionts was largely congruent
with a tree containing sequences from the corresponding host
sponges, suggesting that cospeciation had occurred (although
there also appeared to have been a host switch event at one
point). Subsequent studies of the filamentous cyanobacterium
Oscillatoria spongeliae indicated a high degree of host specific-
ity for various dictyoceratid sponges, with evidence of cospe-
ciation as well as indications of some host switching (316, 396).
Ongoing studies of this system by Thacker and coworkers
(R. W. Thacker, personal communication) should further elu-
cidate the complex evolutionary relationships among these
tropical sponges and their cyanobacterial associates. Coevolu-
tion requires that the host and symbiont maintain close asso-
ciations over evolutionary time, and as mentioned above, the
mechanism by which this presumably occurs in sponges is ver-
tical transmission of microorganisms in host eggs or larvae. An
additional point to consider at this stage is that the phylogeny
of sponges themselves is not fully resolved (40). Molecular
data are often incongruent with traditional sponge taxonomy,
which is based largely on morphological properties, such as
growth form and spicule characteristics (8, 37, 190). Accord-
ingly, our understanding of symbiont evolution in sponges
will continue to develop only in parallel with improvements
in our knowledge of host phylogeny. A recently initiated CO1
sequencing project for taxonomically diverse sponges (www
.spongebarcoding.org) is a step in the right direction for
achieving the latter goal.
The final type of evidence for ancient, close associations
between sponges and microorganisms comes from the fossil
record (43, 261, 377). Reef mounds constructed by siliceous
sponges and cyanobacterial mats, with the latter represented in
part by stromatolites still found today, flourished in (sub)trop-
ical marine waters as far back as the early Cambrian (43). The
fact that sponges and microbes closely coexisted hundreds of
millions of years ago is thus clear, but the nature of that
interaction (e.g., whether microbes lived within sponge tissues)
remains less certain.
Scenario 2: Parental and environmental symbiont transmis-
sion. Demonstrated vertical transmission is generally consid-
ered a strong indicator of symbiosis, yet this does not rule out
the possibility of horizontal (e.g., environmental) transmission
of the same microbe as an additional mechanism. Indeed, this
phenomenon has already been shown for insect-bacterium
symbioses (reviewed in reference 67), and here we borrow
from the well-developed literature on this topic. In aphids, the
primary (obligate) bacterial symbiont Buchnera aphidicola is
vertically transmitted, whereas secondary (facultative) symbi-
onts can be transferred either vertically or horizontally (329).
Given that facultative symbionts can confer fitness advantages
upon their hosts, maintenance of these populations—by what-
ever mechanism—is of clear benefit. Interestingly, it was re-
cently shown that secondary symbionts in aphids can also be
transmitted via the sperm, yielding a different infection pattern
from that which would be expected based on strictly maternal
transmission (237). As discussed in a subsequent section of this
article, both maternal and paternal transmissions of cyanobac-
terial symbionts have been documented for a marine sponge
(424). Another finding from the insect world which is relevant
to our discussion is that certain bacteria can invade novel host
species and form stable associations, perhaps using similar
mechanisms for invasion to those found in pathogenic bacteria
(67). Provided that host chemical and immune defenses can be
evaded, it therefore seems entirely plausible that marine mi-
crobes could invade, and establish themselves within, sponges
from which they were previously absent.
While phylogenetic trees of primary insect symbionts are
congruent with those of their hosts, episodes of horizontal
transfer in secondary symbionts obscure the coevolutionary
signal for these microorganisms. As mentioned in the preced-
ing section, in which the cases of the “Poribacteria” and “Can-
didatus Synechococcus spongiarum” were highlighted, the avail-
able molecular evidence for sponge-associated microorganisms
supports a combination of vertical and horizontal transmis-
sion (not just overall, but for specific individual lineages).
Another salient example is the alphaproteobacterial sponge
associate represented in Fig. 14; this bacterium occurs
widely in sponges (95, 453) and appears to be vertically
transmitted (95) yet is closely related to bacteria isolated
from seawater. Issues of 16S rRNA sequence resolution
notwithstanding (i.e., minor rRNA differences may hide ma-
jor ecological or even genomic differences) (178, 323), this
example underscores the complexities involved with consid-
erations of sponge-microbe evolution.
Scenario 3: Environmental acquisition. In the third sce-
nario, putatively sponge-specific microorganisms are, in fact,
also present in the surrounding seawater, but at such low abun-
dance that standard methods fail to detect them. Several mech-
anisms exist by which the same microbes may then be detected
upon contact with a sponge. Firstly, it is possible that sponges
absorb specific microbial types from seawater, a process which
would imply some degree of recognition of particular micro-
organisms (e.g., by the sponge’s innate immune system) (244).
Recognition of symbionts versus food bacteria has already
been proven experimentally (see the following section), and if
a given microbe encounters favorable conditions (e.g., high
nutrients), it may multiply to the extent to which it can then be
detected by the applied methods. Alternatively, a type of sub-
tractive enrichment may occur, whereby those microbes which
cannot resist phagocytosis by sponge cells are consumed and
hardier bacteria (e.g., those with protective capsules) (482)
survive and are physically enriched near the choanocyte cham-
bers due to the sponge’s filtering activities. If such resistant
bacteria are capable of out-competing other potential coloniz-
ers, they may establish themselves within the sponge tissue.
These possibilities can be placed under a banner of specific
enrichment. Unspecific enrichment is another, at least theo-
316 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
retical, alternative; in this case, microorganisms would simply
be concentrated by sponges during filtering to the extent to
which they can then be detected by the applied methods. Al-
though it is not easy to prove any of these hypotheses correct,
it is, in principle, even more difficult to prove them wrong.
Finding a sponge-specific microbe actively living in the
ocean—independent from a host sponge—would lend support
to the enrichment hypothesis. The converse is less convincing:
if such cells are not detected outside sponges, it may simply
reflect insufficient sampling.
Methodological considerations are of paramount impor-
tance in discussing the environmental acquisition scenario.
Even after many studies of marine microbiology, it still cannot
be discounted that many or most of the so-called sponge-
specific microbes are in fact also present in seawater, but only
at a low abundance which is not detected by standard methods.
But how likely is it that sponge-specific microbes are actually
present in other environments and that, due to methodological
limitations, we simply fail to detect them? Given recent find-
ings regarding the high level of diversity of uncommon micro-
organisms in the so-called rare biosphere (276, 376), the en-
richment scenarios seem entirely plausible. Deep sequencing
of seawater-derived 16S rRNA amplicons should provide fur-
ther insights into the diversity of marine microorganisms, per-
haps also yielding sequence types which are presently consid-
ered sponge specific. An interesting aside is that remarkably
few of the ?1,000 16S rRNA sequences obtained from the
Sargasso Sea metagenome (440) are closely related to those
sequences in sponge-specific clusters. That study employed a
direct cloning approach and was thus free of PCR biases which
might otherwise have resulted in the missing of certain se-
quence types. In this context, it will be very interesting to
examine the upcoming results of the Sorcerer II expedition
(www.venterinstitute.org), during which seawater samples are
being collected from around the world. Irrespective of what
such studies find, if the sponge specificity of any microbe is to
be disproven, then it will be necessary to demonstrate activity
of the said microbe outside a sponge host, as merely demon-
strating its presence is not sufficient.
Highly relevant to this discussion is an interesting point
made recently by R. Hill (154) regarding the abundance of
microorganisms in seawater and the potential consequences
for sponge microbiology. Central to this argument is the im-
mense filtering capacity of sponges, i.e., up to 24,000 liters of
seawater per day for a 1-kg sponge (443). Given that the typical
cell density of bacteria in seawater is about 106cells/ml, then
such a sponge could take in a staggering 2.4 ? 1013bacterial
cells per day. Thus, even if a specific bacterium is present
in seawater at only 1 cell/ml (i.e., one-millionth of all cells
present), then during a single day a sponge could still filter
some 24 million cells of this organism from the water column
(154). The implications are obvious: an organism which is
perennially extremely rare in seawater (and therefore never
detected by applied molecular methods) could conceivably be
concentrated within a pumping sponge (e.g., in the choanocyte
chambers, prior to phagocytotic ingestion) to an extent which
is readily detectable by PCR or even hybridization-based meth-
ods. If it occurs widely (but is undetected) in seawater, then
one can imagine a situation in which it is easily detected from
different sponges at different locations and erroneously con-
sidered a sponge symbiont. Hunting for sponge-specific mi-
crobes in other prolific filter feeders, such as ascidians (rather
understudied thus far, from a microbial perspective, but see
references 239 and 413), may lead to the identification of these
organisms from other, nonsponge sources. These arguments
might seem to paint a grim picture for proponents of the
sponge-specific microbe concept, but the following must also
be considered. If it were really the case that such nonspecific
enrichment of rare seawater microbes occurs in sponges, then
surely sponge-derived 16S rRNA gene libraries would be dom-
inated by other microbes which are known to be abundant in
seawater. This does not happen. To illustrate the point, we use
again the example of the cyanobacterium “Candidatus Syn-
echococcus spongiarum”: if “Ca. Synechococcus spongiarum”-
type organisms are very rare in the ocean but are concentrated
to sufficient levels to be detected in gene libraries from
sponges, then the exceedingly abundant (and closely related)
planktonic Prochlorococcus and Synechococcus strains should
be much better represented. However, a cursory look at Fig.
5 reveals only a few such sequences from sponges. We thus
feel there is a strong case for rejection of the unspecific
Regrettably, a lack of sufficient (appropriate) data prevents
us from ending this section with firm conclusions regarding the
origin of sponge-microbe associations. Even with almost 2,000
sponge-derived 16S rRNA sequences at our disposal, it is so-
bering how little can actually be inferred about the evolution of
sponge-microbe associations. For almost every argument in
favor of an ancient symbiosis, there exists a rational counter-
argument which invokes (recent) environmental acquisition as
the probable driver of the association. For example, a sponge-
specific cluster at the end of a long naked branch in a phylo-
genetic tree could reflect early divergence of this group from
its relatives, or it may simply reflect insufficient sampling of
closely related lineages and/or accelerated evolutionary rates
of the sponge-specific organisms. Additionally, while low levels
of sequence divergence within a sponge-specific cluster argue
against an ancient association, the corollary does not nec-
essarily hold: extensive intracluster sequence divergence could
equally indicate a long, separate evolutionary history (e.g.,
among different hosts) or selective enrichment of diverse (but
still monophyletic) organisms from seawater. So where do we
stand at the moment? At this stage, there is no clear indication
for scenario 1 (ancient symbioses), though this will be ad-
dressed more satisfactorily in the future once additional infor-
mation on sponge phylogeny is available. The existing data
point more towards scenario 2, whereby particular microbes
may be passed vertically between generations, but with some
horizontal exchange also occurring. Further consideration of
potential vectors for the latter process would be worthwhile.
Although we consider this scenario most likely, the supporting
data are also consistent with scenario 3, i.e., environmental
acquisition. This highlights a major current limitation from a
methodological perspective, namely, our inability to distin-
guish between facultative sponge symbionts and specifically
enriched microorganisms. Presumably, the former prefer to
inhabit sponges but can tolerate conditions in seawater, while
enriched microbes may simply tolerate sponges long enough to
be detected by applied methods. Such apparently complex
patterns of microbial transmission are not without precedent in
VOL. 71, 2007SPONGE-ASSOCIATED MICROORGANISMS317
the animal kingdom, with insect-bacterium symbioses provid-
ing a valuable framework for future considerations of sponge-
microbe symbioses. A well-studied marine system that has
contributed greatly to our understanding of symbiont trans-
mission—and, indeed, of host-microbe associations in gener-
al—is that of the squid Euprymna scolopes and its biolumines-
cent symbiont, Vibrio fischeri (195, 230, 258, 442). V. fischeri is
acquired from the surrounding seawater by the juvenile squid,
which, via a remarkable stepwise process coined “winnowing,”
prevents all other bacteria from being established, culminating
in a monoculture of V. fischeri in its light organ (258). The
phylogenetically less complex microbial communities encoun-
tered in hosts such as insects and the squid light organ (typi-
cally, in insects, there are one or two primary endosymbionts
and one or two secondary endosymbionts) facilitate a depth of
understanding of these processes thus far not achievable for
the highly diverse microbial communities in sponges.
ECOLOGICAL ASPECTS: FROM SINGLE CELLS
TO THE GLOBAL SCALE
Establishment and Maintenance of
The mechanisms by which associations between sponges and
microorganisms are established are not well understood. As
discussed at length in the previous section, the fundamental
question of symbiont origin (i.e., whether symbionts were
passed down from an ancestral sponge or obtained contempo-
raneously from seawater) remains unresolved, as do many of
the mechanisms of sponge-microbe interactions and the regu-
lation of microbial communities in these hosts. Extensive stud-
ies by Mu ¨ller and colleagues have attempted to address the
underlying bases of sponge-microbe interactions at the molec-
ular level (36, 241, 243, 249, 399, 400, 444, 465). Sponges are
often regarded as primitive animals, yet their morphological
simplicity belies the possession of a surprisingly complex im-
mune system (244). Indeed, the refinement of such a system
their long evolutionary history, especially when one consid-
ers the immense numbers of (potentially pathogenic) microor-
ganisms to which they are exposed due to their filter-feeding
activities. Detailed studies of the Adriatic sponge Suberites
domuncula have revealed immune responses against both
gram-negative (36, 243, 464) and gram-positive (400) bacteria,
illuminating one means by which sponges may select for and
against certain microbes from the surrounding environment. In
the case of the former, exposure of S. domuncula to the bac-
terial endotoxin lipopolysaccharide (LPS)—derived from
gram-negative cell walls—elicited an increase in synthesis of
two compounds with pronounced antibacterial activity (243).
Confirmation that the compounds were indeed synthesized by
the sponge was obtained by cloning of the gene encoding a key
enzyme in the relevant biosynthetic pathway. A receptor for
LPS on the sponge cell surface was later identified, as was a
signal transduction pathway which is upregulated upon expo-
sure to increased LPS levels (464). The immune response of S.
domuncula to gram-positive bacteria takes a quite different
form: upon exposure to peptidoglycan in the bacterial outer
cell wall, the sponge responds with a rapid activation of endo-
cytosis, followed by the release of lysozyme (400). Receptors
for fungi (277) and viruses (466) also occur in sponges. For
more detailed discussions of the various immune responses
and signal transduction pathways in sponges, the reader is
referred to recent reviews dealing with this topic (e.g., see
references 244 and 246).
In another recent study from the Mu ¨ller group, using S.
domuncula and its alphaproteobacterial symbiont SB2 as a
model system, the importance of oxygenation of sponge tissue
in mediating the relationship was demonstrated (241). Specif-
ically, it was shown that strain SB2 grew preferentially on
minimal media with the aromatic compound protocatechuate,
rather than glucose, as the carbon source. The bacterium can
obtain protocatechuate in situ from the sponge, which pro-
duces this and other diphenols via the activities of the enzyme
tyrosinase. Interestingly, tyrosinase activity and expression of
the tyrosinase-encoding gene in S. domuncula, as well as the
number of pcaDC genes in strain SB2 (responsible for bacterial
utilization of protocatechuate and used here as a proxy for SB2
abundance on the surface [exopinacoderm] of the sponge),
were all maximal under aerated conditions (241). Coupled with
the observed loss of SB2 cells from the sponge surface under
low-oxygen conditions, it was asserted that the oxygen level is
responsible for regulating the bacterial fauna in sponges.
Whether this type of mechanism is important in other sponge-
microbe systems remains to be determined.
The coexistence of microbial symbionts with bacterium-di-
gesting archaeocytes in the sponge mesohyl has long interested
sponge biologists. In a series of landmark experiments, Wilkin-
son and coworkers fed tritium-labeled sponge- and seawater-
derived bacteria back to host sponges and found that sponge
symbionts passed through uneaten, whereas seawater bacteria
were largely consumed (482). Two different mechanisms were
proposed to account for this: either (i) symbionts are specifi-
cally recognized by the sponge and deliberately not ingested or
(ii) bacteria use extracellular masking capsules to avoid detec-
tion by sponge cells (482, 483). While neither theory has been
tested explicitly (but see the preceding discussion on sponge
immune responses), the latter explanation is in favor today,
with several studies reporting the existence of slime layers and
sheaths on symbiotic bacteria (106, 427, 473). The results of
recent experiments by the Hentschel group were consistent
with earlier findings: seawater-derived bacteria were consumed
by Aplysina aerophoba some 2 orders of magnitude faster than
was a consortium of sponge-derived bacteria (459). In addi-
tion, when a green fluorescent protein-labeled food bacterium
was fed to the sponge, it was rapidly digested within the sponge
tissues. All of these findings carry interesting implications for
the evolution of sponge-microbe associations (also see the
previous section). If presumed symbionts are not taken from
the seawater (i.e., either as colonizers or as a food source),
then this suggests vertical transmission as the mechanism by
which these associations are maintained.
It is now established beyond doubt that many sponges (at
least among those in marine environments) harbor diverse and
abundant microbial communities. What is far from established
is how, if at all, the composition and density of these commu-
nities are regulated. The potential role of phages and protozoa
in regulating microbial communities within sponges is of inter-
est, but virtually nothing is known about this to date (but see
318 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
reference 211). Predatory bacteria, perhaps related to the
deltaproteobacterial genus Bdellovibrio, could also be involved
in structuring microbial communities in sponges (468). At least
for cyanobacteria, it has been suggested that their abundance is
directly proportional to the number of sponge cells, implying
some degree of influence by the sponge over symbiont growth
and reproduction (477). The high photosynthetic rates of cya-
nobacteria in sponges (see also the following section) should,
with all things being equal, result in cyanobacterial growth to
the extent that host tissues would be overwhelmed. It is thus
likely that the sponge exerts some control over its symbiont
populations, with several mechanisms being proposed, includ-
ing the following: sponges consume excess symbionts, sponges
eject symbionts when stressed, the host sponge steals photo-
synthate from the cyanobacteria, and sponges starve the sym-
bionts (477). With some debate over the extent of sponge
consumption of symbionts and no evidence for expulsion of
excess symbionts, Wilkinson argued that the last two scenarios
(steal and starve) are most likely. There is strong evidence for
stealing of photosynthate from symbionts in other systems
(e.g., coral-zooxanthella  and freshwater Hydra-Chlorella
 symbioses), and it seems plausible that sponges may also
produce some sort of host release factor to induce the release
of large quantities of fixed carbon from the cyanobacteria.
Along these lines, a host release factor was recently described
for the symbiosis between the sponge Haliclona cymaeformis
and its macroalgal symbiont Ceratodictyon spongiosum (129).
In contrast, the starve hypothesis holds that if a sponge can
somehow restrict the symbiont’s access to essential nutrients,
such as nitrogen and phosphorus, then symbiont protein syn-
thesis and cell division would be restricted. Consequently, an
excess supply of carbon-rich photosynthate would be excreted
from the symbiont (156, 477). To the best of our knowledge,
neither scenario has been proven unequivocally or disproven
for any sponge-microorganism association. More generally, re-
markably little is known about communication, or chemical
cross talk, between sponges and their microbial associates.
Marine sponges produce a wide variety of secondary metabo-
lites, some of which could potentially enable them to select for
or against particular types of microorganisms (185) (although
the nonspecific nature of many antimicrobial compounds sug-
gests that such selection may generally be limited to broader
classes of microbes, e.g., gram-positive versus gram-negative
bacteria). Conserved bacterial signaling systems, as exempli-
fied by the acyl homoserine lactone (AHL) regulatory systems
of many gram-negative bacteria (111, 369), often mediate col-
onization-related traits (e.g., biofilm formation, swarming, and
virulence) and offer one means by which sponges could interact
or interfere with bacteria. Bacteria capable of AHL production
have already been reported from marine sponges (361, 389), as
have other putative signaling molecules, such as diketopipera-
zines (1, 162, 182). It is highly likely that sponges produce
metabolites which allow them to disrupt AHL-regulated phe-
notypes, as shown for the macroalga Delisea pulchra (126, 127,
223, 224) and various terrestrial plants (302, 393). Indeed,
inhibition of bacterial swarming by chemical extracts from
sponges has recently been shown, though it has yet to be
clarified whether this is an AHL-specific effect (184).
Associations between sponges and microorganisms can be
maintained over different generations in either of the following
two ways: (i) microbes can be recruited from the surrounding
water by filter feeding (i.e., horizontal or environmental trans-
mission) or (ii) microbial symbionts can be passed on from the
parent sponge via reproductive stages (i.e., vertical transmis-
sion) (Fig. 17). Although horizontal transmission of symbionts
has been demonstrated convincingly for several marine symbi-
oses (e.g., squid-Vibrio fischeri  and hydrothermal vent
tubeworm-chemoautotrophic bacterium  symbioses), it is
the latter mechanism which has received most attention for
sponges. Bacteria have now been found in embryos or larvae
from all three classes of Porifera (see reference 97 and refer-
ences therein), including species with highly varied reproduc-
tive strategies. Sexual reproduction in sponges involves either
vivipary (where larvae are brooded within the animal) or
ovipary (whereby eggs, generally fertilized externally, develop
outside the sponge). Evidence for vertical transmission of bac-
teria has been reported for both types (97, 116, 118, 181, 362,
419, 422, 431), while asexual reproduction, i.e., budding, could
also contribute to symbiont transfer in some species (146).
Indeed, gemmules, the asexual buds of freshwater sponges,
contain symbiotic zoochlorellae in at least some species (372),
while a bud protruding from the surface of the marine sponge
Tethya orphei contained a symbiotic cyanobacterium (117).
The vast majority of reports dealing with vertical transmis-
sion in sponges have been based on transmission electron mi-
croscopy (TEM) observations. Such studies have contributed
greatly to our understanding of this phenomenon, with the
identification of several mechanisms by which symbiotic mi-
crobes can be incorporated from maternal mesohyl tissue into
eggs or embryos (reviewed in reference 97). These include
phagocytosis of microbial cells by the oocyte directly from the
adult mesohyl as well as transfer of microbes from parent
FIG. 17. Vertical transmission of microbial symbionts by a marine
sponge. A transmission electron micrograph of a Chondrilla australien-
sis larva is shown, indicating a diverse range of bacterial morphotypes.
Bar ? 1 ?m. (Modified from reference 420 with permission of the
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS319
sponge to embryo along an “umbilical cord.” Intriguingly, in
the Australian sponge Chondrilla australiensis, eggs containing
a cyanobacterial symbiont (of the “Candidatus Synechococcus
spongiarum” type ) were distributed throughout the
sponge mesohyl, whereas cyanobacteria are normally confined
to the better-illuminated periphery, or cortex, of the sponge
(422). Nurse cells, probably derived from choanocytes (425),
have been invoked as a possible mechanism by which cya-
nobacteria are transported to the eggs (422, 424). These cells,
which fuse with eggs and release their contents (including
cyanobacteria) into the egg cytoplasm prior to spawning, pre-
sumably phagocytose the symbionts in the cortex before mov-
ing to the developing eggs deeper within the sponge. Remark-
ably, Usher and coworkers were also able to demonstrate the
presence of cyanobacteria in sperm cells, indicating that both
parents are capable of transferring symbionts to offspring
(424). Sponges of the genus Chondrilla were also the subject of
another recent TEM study (which additionally employed im-
munocytochemical techniques), in which vertical transmission
of an endosymbiotic yeast was shown (221).
The drawback of the TEM approach is that, with some
exceptions (e.g., cyanobacteria [423, 424]), even phylum-level
identification of the relevant microorganisms is not possible
due to an insufficient number of distinguishing morphological
characters. Multiple bacterial morphotypes have been ob-
served in sponge larvae (suggesting transmittance of a complex
assemblage) (e.g., see reference 424), yet little or nothing is
known of their phylogenetic affiliations. The recent application
of molecular techniques in this area (95, 266) thus offers the
potential for exciting new insights into the phylogenetic (and,
in principle, metabolic) complexity of transmitted microbial
assemblages. A 16S rRNA gene library constructed using cya-
nobacterium-specific PCR primers confirmed the presence (as
indicated by TEM) of a single cyanobacterial type in both
larvae and adults of the Red Sea sponge Diacarnus erythraenus
(266). The transmitted cyanobacterium is highly similar to the
aforementioned “Ca. Synechococcus spongiarum”-type symbi-
ont of Chondrilla australiensis. A range of molecular tech-
niques were used to examine the bacterial community in larvae
of the Caribbean sponge Mycale laxissima, revealing a much
more diverse population than that recovered by cultivation
efforts (95). A single alphaproteobacterium, related to
Pseudovibrio denitrificans and previously reported from many
sponges (Fig. 14), was the only bacterium from the larvae that
could be grown on a standard marine medium. In contrast,
sequences representing a diverse assemblage comprising Acti-
nobacteria, Bacteroidetes, Cyanobacteria, Planctomycetes, and
Proteobacteria (including the isolated alphaproteobacterium)
were recovered from a 16S rRNA gene library based on DNAs
isolated from the larvae (95). Similarly diverse microbial com-
munities were found in larvae of the sponges Corticium sp.
(366) and Ircinia felix (352), using 16S rRNA-based ap-
proaches, such as gene libraries, FISH, and DGGE. Broad
congruence between larva- and adult-associated microbial
communities in these studies indicated that a significant subset
of the resident microbes is transferred in this way. Future
molecular studies could provide information on the metabolic
properties of the transferred symbionts (e.g., via analysis of
functional genes or FISH-microautoradiography), which should
improve our understanding of the mechanistic bases of these
Whatever the underlying mechanisms for the establishment
and maintenance of sponge-microbe associations, it is appar-
ent that in many cases such associations are highly stable and
resistant to external disturbance (reviewed in reference 148).
In at least some other instances (e.g., see reference 457), this is
not the case. Briefly, neither starvation, exposure to antibiotics,
nor transplantation to different depths could elicit major
changes in bacterial community composition in Aplysina aero-
phoba (105) and Aplysina cavernicola (407). Similarly, even
translocation of Aplysina fistularis from its natural depth of 4 m
to a new depth of 100 m was not enough to significantly affect
cyanobacterial abundance in this sponge (although the sponge
did die at depths of ?100 m) (222). In contrast, this change
resulted in a loss of cyanobacterial symbionts from the cooc-
curring sponge Ircinia felix. The importance of cyanobacterial
symbionts—at least to some sponges—was recently demon-
strated by Thacker in a series of elegant field experiments
(394). To test the hypothesis that a greater benefit to the host
is derived from more specialized symbionts, shading experi-
ments were conducted with the tropical sponges Lamellodys-
idea chlorea (containing the host-specific cyanobacterium Os-
cillatoria spongeliae) and Xestospongia exigua (which contains
the generalist symbiont “Ca. Synechococcus spongiarum”).
Shaded individuals of the encrusting sponge L. chlorea lost
?40% of their initial area, but the chlorophyll a concentration
in the remaining sponge tissue (a proxy of cyanobacterial abun-
dance) did not change, implying a close, putatively mutualistic
relationship between the sponge and O. spongeliae. In contrast,
X. exigua lost relatively little mass but did lose many of its “Ca.
Synechococcus spongiarum” symbionts, suggesting that this re-
lationship is less tight-knit than the other (394). While the
mechanism by which symbionts are lost from X. exigua is un-
clear, the existing data do suggest that the specialist O. spon-
geliae provides a greater benefit for its host sponge than does
the generalist “Ca. Synechococcus spongiarum” for its host.
Although the necessary data are currently lacking, one could
speculate that the degree of host specificity (of individual sym-
bionts or even entire communities) may explain some of the
different results obtained from previous perturbation experi-
ments (148). More generally, the extent of host specificity
among marine eukaryote-associated microbes may have sub-
stantial implications for microbial diversity on a wider scale
(390). If most symbionts are highly host specific, then their
overall diversity will be much higher than if the same hosts
harbor mostly generalist species. Finally, Roberts and cowork-
ers recently reported the results of experimental manipulations
with the photosynthetic symbiont-containing Australian sponge
Cymbastela concentrica (322). Shade and, to a lesser extent, silt
treatments (both designed to mimic the physical effects of
sewage effluent discharge) led to lowered chlorophyll a con-
centrations within the sponge, while increased salinity and nu-
trient loads had negligible effects.
Physiology of Sponge-Associated Microorganisms
A lack of pure cultures for most sponge-associated micro-
organisms has contributed to a paucity of knowledge about
their physiological characteristics. What we do know has arisen
320 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
from a combination of the existing pure-culture studies, pro-
cess data, and inferred metabolic properties from analysis of
16S rRNA, functional genes, and metagenome data. Collec-
tively, microbes in sponges are capable of, among other pro-
cesses, photosynthesis, methane oxidation, nitrification, nitro-
gen fixation, sulfate reduction, and dehalogenation. Here we
summarize the current knowledge by first examining, in turn,
the major nutrient cycles within sponges.
Carbon. Heterotrophy is a common form of carbon metab-
olism in sponges, either via consumption of microbes from
seawater or via microbial uptake of dissolved organic carbon
(495). However, for many sponges, particularly those in trop-
ical regions, carbon metabolism centers around the activities of
photosynthetic microorganisms, such as cyanobacteria (11, 58,
59, 381, 470, 474, 477, 486). Many tropical sponges contain
substantial populations of these oxygenic autotrophs, and no-
where is the contribution of microbial symbionts to the host
sponge more evident than in this case (see also the next sec-
tion). Translocation of photosynthates (mostly as glycerol)
from cyanobacteria to the host has been shown for marine
sponges (476), while glucose produced by a chlorella-like green
alga was passed to its freshwater sponge host, Ephydatia flu-
viatilis (475). Phototrophic sponges—those whose carbon nu-
trition depends heavily on cyanobacterial symbionts—receive
?50% of their energy requirements from cyanobacteria (474),
allowing these species to thrive in the low-nutrient, high-light
areas commonly found on tropical reefs. On the Great Barrier
Reef, phototrophic sponges comprise approximately half of
the total sponge biomass on outer reefs, where the water is
cleaner, but are much less common inshore, where terrestrial
runoff and turbidity are greater (470, 485). Similarly, phototro-
phic sponges are largely absent from Caribbean reefs, where
only small numbers of sponge-associated cyanobacteria are
present (470). Phototrophy has also been demonstrated for at
least one temperate sponge (57), while numerous others are
known to contain photosynthetic symbionts (321, 336, 390,
430). The sponge Cymbastela sp. from temperate South Aus-
tralia was capable of compensating photosynthetically (i.e., the
rate of photosynthesis equals its rate of respiration) at a 4- to
5-m depth in winter, while it was a net producer at the same
depth in summer (57). In contrast, Great Barrier Reef sponges
may derive much of their nutrition from photosynthetic sym-
bionts as deep as 15 to 30 m, due to the clearer water and,
therefore, decreased light attenuation (58). Some sponges are
apparently obligate phototrophs, with their lower depth limits
determined by the availability of light for photosynthesis (58).
Others function as mixotrophs, combining symbiont-derived
nutrition with filter feeding, while still others contain no pho-
toautotrophic symbionts and derive all of their carbon nutri-
tion from filter feeding (477).
The extent to which other photosynthetic associates (e.g.,
diatoms, dinoflagellates, and phototrophic sulfur bacteria)
contribute to carbon cycling within sponges is less clear. The
Mediterranean sponges Cliona viridis and Cliona nigricans both
contain symbiotic dinoflagellates (zooxanthellae), and for C.
viridis, at least, it appears that sponge metabolism depends on
the photosynthetic activity of these symbionts (353). Indeed,
the growth of C. viridis was greater in individuals maintained
under natural light conditions than in those maintained in
constant darkness, reflecting the contribution of photosyn-
thetic symbionts to host metabolism (324). Conversely, in at
least one case, it appears that diatoms in Antarctica may par-
asitize the sponge host, using its metabolic products as an
energy source (16). The sponge Cymbastela concentrica in
southeastern Australia contains high densities of diatom-like
cells in the illuminated periphery (M. W. Taylor, unpublished
data), but further work is required to elucidate whether this
association is phototrophic in nature. The occurrence in this
sponge of at least some cyanobacteria in addition to the dia-
toms (Longford et al., unpublished data) will complicate efforts
in this direction. Some freshwater sponges (e.g., Spongilla
lacustris) contain zoochlorellae, and although the symbiosis is
not obligate (aposymbiotic individuals occur in areas of deep
shade), it appears that algal photosynthesis can contribute to
host metabolism and growth (109, 333, 475).
An unusual form of nutritional symbiosis is that between
methanotrophic bacteria and deep-sea cladorhizid sponges
(428, 429, 431). These remarkable sponges, which possess no
aquiferous system but instead prey on tiny swimming organ-
isms, are believed to obtain a significant portion of their nu-
trition from the consumption of methanotrophs. Methane
serves as a carbon source and substrate for energy production
in methanotrophs, and in this particular system it is derived
from a deep-sea mud volcano (429). In other sponges, the
presence of methanogenic archaea may lead to methane pro-
duction within anoxic zones. A 16S rRNA gene sequence af-
filiated with the methanogens of the phylum Euryarchaeota
(456) is the sole piece of evidence for this at present, but the
documented existence of anoxic microhabitats within sponges
(159) suggests that these associations could be more wide-
spread. Other chemoautotrophic microbial processes that have
been observed in sponges and may also contribute to sponge
nutrition are nitrification and sulfur oxidation. These are dealt
with in the following sections.
Nitrogen. After carbon, nitrogen is the most important nu-
trient for life, as it is required for the synthesis of amino acids
and, subsequently, proteins. In oligotrophic waters where ni-
trogen levels are low (e.g., coral reefs), symbiotic microorgan-
isms may contribute to the nitrogen budget of sponges via
fixation of atmospheric nitrogen, N2(479, 484). The first evi-
dence for this came from measurements of nitrogenase activity
in three Red Sea sponges (479). The activity of this enzyme,
the catalyst for microbial N fixation, was estimated using an
acetylene reduction test (for caveats, see reference 484) and
could be measured only in Siphonochalina tabernacula and
Theonella swinhoei, both of which contained cyanobacteria. In
contrast, Inodes erecta, which contained only noncyanobacte-
rial microorganisms, showed no evidence of N fixation. Addi-
tionally, nitrogenase activity was higher in illuminated tissue
than in that maintained in the dark and did not correlate with
the abundance of the heterotrophic bacterial communities in S.
tabernacula and T. swinhoei. Taken together, these data sug-
gested that nitrogenase activity was due mainly to the presence
of cyanobacteria, many of which are capable of N fixation
(479). A subsequent study provided more concrete proof of N
fixation in sponges by demonstrating incorporation of the sta-
ble isotope15N2into various amino acids in Callyspongia muri-
cina (484). Whether microbial N fixation is of major ecological
significance for sponges remains uncertain, but it does appear
that its occurrence in sponges is not limited to cyanobacteria.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS321
Heterotrophic nitrogen-fixing bacteria were reported from a
Halichondria sp. (367), and the nifH gene (encoding a subunit
of the nitrogenase reductase enzyme) has been amplified from
both alpha- and gammaproteobacteria inhabiting several Ca-
ribbean sponges (N. M. Mohamed, Y. Tal, and R. T. Hill,
presented at the 11th International Symposium on Microbial
Ecology, Vienna, Austria, 20 to 25 August 2006).
Nitrification in sponges has also received attention. The two
steps of nitrification, i.e., the biological conversion of ammonia
to nitrite and then to nitrate, are catalyzed, in turn, by ammo-
nia-oxidizing and nitrite-oxidizing microorganisms (33, 194).
Ammonia, which can be toxic to eukaryotes, is a metabolic
waste product and could accumulate within sponge tissues,
particularly during periods of low pumping activity. The re-
lease of nitrate (and in some cases nitrite) from incubated
sponges provided the first indication of nitrification within
these organisms, with estimated rates often far exceeding those
for other benthic substrata (63, 80). These results suggested
the presence of nitrifying microorganisms, and indeed, 16S
rRNA sequences from both ammonia-oxidizing betaproteo-
bacteria (79; Taylor et al., unpublished data) and nitrite-
oxidizing bacteria of the genus Nitrospira (Fig. 10) (146; Long-
ford et al., unpublished data) have been recovered in molecular
surveys of sponges. The widespread presence of Nitrospira in
sponges may indicate low nitrite availability in these hosts, as
members of the Nitrospira typically favor low-nitrite habitats
(356, 448). Nitrifying microorganisms are among the few for
which metabolic capabilities can generally be inferred from
16S rRNA data, and sequences representing several types of
ammonia-oxidizing bacteria–in the genera Nitrosospira and Ni-
trosomonas–-were identified from the Australian sponge Cym-
bastela concentrica (Taylor et al., unpublished data). The find-
ing of only Nitrosomonas eutropha/europaea-affiliated ammonia
oxidizers in a previous study of six tropical sponges (79) may
have been due to the use of overly specific PCR primers (both
of the primers used have mismatches to all Nitrosospira-affili-
ated ammonia oxidizers and also to many Nitrosomonas spp.
[193, 300]). Nitrite oxidizers belonging to the Nitrospira genus
are frequently recovered in 16S rRNA gene surveys of sponges,
yet in at least one case (80), extensive release of nitrite indi-
cated that oxidation of ammonia and nitrite could be uncou-
pled. Interestingly, ammonia-oxidizing archaea, whose exis-
tence was just proven in 2005 (192), also exist in sponges.
Metagenomic reconstruction of “Candidatus Cenarchaeum
symbiosum,” the abundant and yet uncultivated crenarchaeote
in the Californian sponge Axinella mexicana (294), revealed the
existence of ammonia monooxygenase (amoA) genes (required
for ammonia oxidation) in this organism (135), while PCR-
based surveys for amoA indicated that archaeal ammonia ox-
idizers are widespread in marine sponges (D. Steger et al.,
unpublished data). Whether archaea or bacteria are the key
ammonia oxidizers in sponges remains to be determined, but in
at least one system (soils), ammonia-oxidizing archaea appear
to greatly outnumber their bacterial counterparts (204).
Numerous gaps remain in our knowledge of nitrogen cycling
in sponges (Fig. 18). For example, the existence of anoxic
zones in at least some sponges (159) suggests the potential for
mox). Neither process has been reported for sponges thus far,
and our own efforts to amplify 16S rRNA genes from known
anammox bacteria from several sponges have yielded no re-
sults. Denitrification is catalyzed by a phylogenetically diverse
range of microorganisms, and it is risky to infer the ability to
denitrify from 16S rRNA sequence data alone. Nonetheless, it
is worth noting that a common alphaproteobacterial associate
of marine sponges (95, 147, 453) (Fig. 14) is very closely related
to the marine denitrifier Pseudovibrio denitrificans (368), and at
least some of the sponge-derived strains have also tested pos-
itive for denitrification (95). The role of sponge filter feeding in
providing particulate organic nitrogen is also of interest (312),
with evidence that uptake of ultraplankton can yield sufficient
nitrogen to sustain both the tropical sponge Haliclona cymae-
formis and its macroalgal symbiont (72, 288).
FIG. 18. Current state of knowledge about the nitrogen cycle in sponges. Thick arrows signify those processes which have been demonstrated
in sponges; references (given in parentheses) pertain to either the process or the implicated microorganisms. PON, particulate organic nitrogen.
322 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
Sulfur. Several lines of evidence point to the widespread
occurrence of sulfur-metabolizing microorganisms in sponges.
For starters, two of the key microbial players in the sulfur cycle,
namely, sulfate reducers and sulfur oxidizers, have been found
in multiple sponges. Sulfur-oxidizing bacteria from the families
Rhodospirillaceae and Chromatiaceae (Alpha- and Gammapro-
teobacteria, respectively) were isolated from Ircinia sp. and
Euspongia officinalis in the 1970s (173). In that paper, the
bacteria were referred to as phototrophic sulfur bacteria, and
additional, earlier isolations of green sulfur bacteria (phylum
Chlorobi) were also discussed (see reference 173 and the Eim-
hjellen  citation within). FISH signals for Chlorobi were
later found in the Great Barrier Reef sponge Rhopaloeides
odorabile (458). The above-mentioned sulfur bacteria oxidize
reduced sulfur compounds such as hydrogen sulfide. This sub-
strate is presumably derived from the activities of sulfate-re-
ducing bacteria, which have also been obtained from sponges
(159, 160, 173, 225, 358). An endosymbiotic sulfur cycle com-
prising sulfate-reducing and sulfide-oxidizing bacteria has al-
ready been demonstrated for a marine oligochaete (83), and
the above data suggest that a similar process takes place in at
least some sponges.
The most extensive work on sulfur metabolism within
sponges has been conducted by Hoffmann, Reitner, and col-
leagues (159, 160, 225, 310, 358). They detected sulfate-reduc-
ing bacteria by FISH in the Mediterranean sponges Chondrosia
reniformis and Petrosia ficiformis (225, 358), as well as in the
cold-water sponge Geodia barretti (159, 160). In G. barretti,
FISH detection of sulfate reducers belonging to members of
the genera Desulfoarculus, Desulfomonile, and/or Syntrophus
(estimated abundance, 1.8% of the total bacterial community)
was complemented by isotopic measurements of sulfate reduc-
tion rates and analysis of oxygen profiles within the sponge
(159). Sulfate reduction is an anaerobic process, and through
the use of microelectrodes (354), these authors were able to
demonstrate the presence of anoxic zones within the sponge,
particularly during periods of pumping inactivity (159). The
estimated sulfate reduction rates in G. barretti, of up to 1,200
corded in natural systems. Intriguingly, analysis of lipid bio-
markers suggested that bacterially derived carboxylic acids
(perhaps from sulfate reducers) may be transferred to the host
for subsequent synthesis into other compounds (159). The
accumulation of toxic hydrogen sulfide was also addressed by
Hoffmann and coworkers, who calculated that the activities of
sulfide-oxidizing bacteria could, together with chemical reoxi-
dation processes and the use of oxidized iron from seawater as
an electron acceptor, be sufficient to balance microbial sulfide
production. Although it is pure speculation at this stage, it is
also possible that sulfur-oxidizing symbionts enable sponges to
occupy sulfide-rich environments. The base of the Micronesian
sponge Oceanapia sp., for example, can be buried up to 20 cm
deep in the sediment (360), where anoxic conditions with high
sulfide concentrations may prevail.
Interestingly, 16S rRNA gene sequences which are highly
similar to those from known sulfate reducers (e.g., Desulfobac-
terium or Desulfomicrobium spp.) (Fig. 8) have only rarely been
recovered from sponges. Ahn and colleagues did find Desulfo-
vibrio-related organisms in enrichment cultures grown on
Aplysina aerophoba-derived brominated phenolic compounds (3),
2?cm3sponge day?1, are among the highest re-
but these sequences do not appear to have been deposited in
GenBank and were therefore not included in our analyses.
Analysis of functional genes [e.g., dsrAB, encoding the dissim-
ilatory (bi)sulfite reductases] (447, 502) would be one way to
gain further insights into the composition of sulfur-metaboliz-
ing microbial guilds within sponges.
A final aspect of sulfur metabolism that has received much
attention in marine systems is that of dimethylsulfoniopropi-
onate (DMSP) and its cleavage product, dimethylsulfide (DMS).
These compounds are thought to play a role in global climate
regulation (497), while DMSP may also protect marine algae
and invertebrates from herbivores/predators and oxidative
damage (384, 436). In a recent survey of DMSP content in a
variety of coral reef invertebrates, high levels in corals were
attributed to symbiotic zooxanthellae, while the much lower
DMSP concentrations typical of sponges were presumed to be
diet derived (435). It is currently unknown whether the levels
in sponges are sufficient to play a role in predator deterrence
or whether those sponges with symbiotic zooxanthellae have
higher DMSP contents.
Other aspects of microbial metabolism in sponges. In con-
trast to the case for several major chemical elements (namely,
carbon, nitrogen, and sulfur), to our knowledge virtually noth-
ing is known about phosphorus cycling within marine sponges.
We assume that sufficient phosphorus is obtained from the
sponge’s diet of microorganisms.
The degradation of halogenated chemicals within the Med-
iterranean sponge Aplysina aerophoba was the subject of an
interesting recent study (3). This and other sponges are rich
sources of brominated compounds such as bromophenols and
bromoindoles, and it was predicted that microorganisms within
such sponges may be capable of dehalogenation. Indeed, by
establishing enrichment cultures from sponge tissue, in the
presence or absence of various electron acceptors, the authors
of that study were able to demonstrate reductive debromina-
tion under methanogenic and sulfidogenic, but not denitrify-
ing, conditions (3). Antibiotic inhibition of dehalogenation ac-
tivity indicated that it was the microbes, not the sponge, which
The production of a wide range of secondary metabolites by
sponge-associated microorganisms is well known. We provide
examples of some pharmacologically relevant metabolites in a
later section (see “Biologically Active Chemicals from Marine
Sponge-Microbe Consortia and Their Commercial-Scale Sup-
ply”) and, immediately below, outline the potential benefit(s)
of these metabolites to the sponge-microbe association.
The Varied Nature of Sponge-Microbe Interactions
Sponges and the microorganisms living within and around
them display the full gamut of interactions, from microbial
pathogenesis and parasitism (sometimes resulting in sponge
death) to microbes as the major food source for heterotrophic
sponges and to mutualistic (or at the very least commensalistic)
associations in which both partners appear to benefit. We first
consider the putative benefits of symbiotic microorganisms to
the host sponge.
Mutualism/commensalism. It is clear that sponges benefit
greatly from the diverse metabolic properties of their associ-
ated microorganisms (see the preceding section). The provi-
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS323
sion of photosynthates and (perhaps to a lesser extent) fixed
nitrogen from cyanobacteria (11, 474, 476, 477, 479, 486) is
presumably a key factor in the ecological success of many
sponges on nutrient-poor tropical reefs. Cyanobacterial sym-
bionts may be equally important to juvenile and adult sponges.
Sponge larvae are generally thought to be lecithotrophic (i.e.,
nourished from finite stored nutrients) (219), with no capacity
for filter feeding (though some may assimilate dissolved or-
ganic matter from seawater ), so the energy gained from
photosynthetic cyanobacteria should contribute to (i) gamete
and larval longevity in the water column (424) and (ii) (once
sponges are settled) the rapid growth required to outcompete
algae and other photosynthetic organisms for substratum in
illuminated areas (477, 478). Larval mortality may conse-
quently be lower for those harboring cyanobacterial symbionts.
The importance of photosynthetic symbionts to their hosts is
evident in the typically flattened morphologies of phototrophic
sponges, with the thinner species containing dense accumula-
tions of cyanobacteria throughout the tissue. In contrast, mixo-
trophic sponges—those which utilize both filter feeding and
photosynthetic symbionts for nutrition—may reduce their re-
liance on symbionts with age. Juveniles possess a high propor-
tion of symbiont-containing tissue, which reduces as the
sponge grows thicker and increases the amount of filter-feed-
ing tissue (474, 478). Interestingly, endosymbiont photosynthe-
sis can also bring with it certain costs for the sponge, such as
the following: (i) morphological adaptation for improved light
capture may occur at the expense of filter-feeding capacity
(477), and (ii) oxidative stress may result from the presence of
high levels of photosynthetically produced molecular oxygen,
necessitating an enhancement of antioxidant defenses com-
pared with those of asymbiotic specimens (303, 304). Another
role for cyanobacteria and their pigments has also been pro-
posed, namely, protection of sponges from excessive illumina-
tion (336). Although this role has not been proven experimen-
tally, one expects that this could be particularly important for
intertidal species, where radiation (UV and photosynthetically
active radiation) is especially high (381). The documented
occurrence of UV-absorbing mycosporine-like amino acids
in sponges harboring cyanobacteria (e.g., Dysidea herbacea
) also supports the hypothesis of shading by the symbionts.
Microbial metabolism may benefit the host sponge in other
ways. As mentioned earlier, Hoffmann and colleagues (159)
described the likely transfer of carboxylic acids from anaerobic
bacteria to the sponge Geodia barretti. Methanotrophic bacte-
ria may supplement the nutrition of non-filter-feeding, carniv-
orous sponges in methane-rich deep-sea habitats (429, 431),
while symbiotic zooxanthellae (dinoflagellates) enhance boring
and growth rates in clionid sponges (153). Elimination of toxic
metabolic by-products is another possible role played by sponge-
associated microbes. The sulfur-oxidizing bacteria mentioned
above oxidize reduced sulfur compounds, such as highly toxic
hydrogen sulfide, to less harmful forms. Sulfide may accumu-
late in anoxic zones due to the activities of sulfate reducers,
particularly during periods of low pumping activity (159). Sim-
ilarly, ammonia and nitrite, which can be toxic to eukaryotes,
are products of sponge and microbial metabolism but may be
oxidized to harmless forms via the activities of nitrifying mi-
croorganisms. While negative effects of nitrite on the develop-
ment of some juvenile freshwater sponges have been demon-
strated in laboratory experiments (179), it is less clear whether
ammonia or nitrite ever accumulates to sponge-harming levels
in nature. Microelectrode studies to address such questions
(354) would be of great interest and should further our under-
standing of the role of nitrifying microorganisms in sponges.
Also of interest will be the data derived in the future from
sponge genome projects. These should help to identify possible
absent metabolic pathways in the host, whose functions may
instead be filled by symbiont-derived factors.
Further putative benefits for sponges from their microbial
partners include increased structural rigidity (due to mucous
production by bacteria) (472), direct incorporation of dissolved
organic matter from seawater (480, 495), digestion and recy-
cling of insoluble sponge collagen (481), and microbial pro-
duction of secondary metabolites that are of use to the host. In
several cases, production of bioactive metabolites has tenta-
tively been ascribed to bacterial symbionts, and these may
serve to protect the sponge from pathogens, predators, and
foulers (e.g., see references 147, 351, and 417). In some other
cases, molecules produced by microbial symbionts could po-
tentially be used as precursors for the biosynthesis of defense
metabolites by sponges. Whatever the exact mechanism, it is
likely that the chemical defenses of many sponges include both
host- and symbiont-produced metabolites (see also “Harming
the host: pathogenesis, parasitism, and fouling” and Bio-
technology of Sponge-Microbe Associations: Potential and
The observant reader may have noticed that the preceding
discussion deals almost exclusively with putative benefits for
the host, with little mention of advantages for the symbiont(s).
Even when the benefits to the host sponge are obvious (e.g., in
phototrophic sponges), it is not necessarily clear what benefit
the microbial partner derives from the association. It is thus
difficult, and often impossible, to confidently assign a mutual-
istic rather than just a commensalistic label to a sponge-mi-
crobe association. Presumed benefits for microbial symbionts
of sponges include a generous supply of nutrients, as well as
shelter—from predators or high light levels—in the sponge
Microorganisms as a food source for sponges. With the
primary exception of phototrophic sponges (described above),
most sponges are thought to obtain the bulk of their carbon
nutrition via the consumption of microorganisms from the
water column (though uptake of dissolved organic carbon may
also be significant [e.g., see reference 495]). Bacteria (including
both cyanobacteria and presumed heterotrophs) as well as
eukaryotic microalgae can satisfy the entire food requirements
of sponges (307), with the potential for dense sponge commu-
nities to significantly deplete the surrounding water of micro-
bial cells (289, 309). Early studies of particle feeding in sponges
indicated that as much as 96% of bacterial cells were removed
from the inhalant seawater by the filtering activities of the
sponge (308). These results were supported by the later appli-
cation of more sophisticated techniques, in particular flow cy-
tometry (23, 289, 290). Pile and colleagues reported grazing of
the Atlantic sponge Mycale lingua on various types of plankton
(?10 ?m in size), with retention efficiencies ranging from 93%
for Prochlorococcus-type cyanobacteria down to 72% for the
smallest photosynthetic eukaryotes (290). Similar methodolo-
gies applied to the encrusting New Zealand sponge Polymastia
324 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
croceus showed the best retention of Synechococcus-type cya-
nobacteria (94%) and picoeukaryotes (88%), with somewhat
poorer retention of Prochlorococcus-type cyanobacteria and
other (noncyano-) bacteria (74 and 46%, respectively) (23).
The lower retention of some cell types suggested that P. cro-
ceus was selective in its feeding. Laboratory experiments in-
volving the feeding of symbiotic versus seawater bacteria to
other sponges lend strong support to the notion of selective
feeding (459, 482).
The generally highly efficient removal of particles from sea-
water is due largely to the extraordinarily large number of
choanocyte chambers (?1 ? 107per cm3) in sponge tissues
(306). With each chamber containing as many as 150 choano-
cytes (371), coupled with the ability of pinacocytes (epithelial
cells) to capture larger particles (308, 412), any food particle
passing through the intricate aquiferous system of a sponge is
subjected to intense grazing pressure. Interestingly, it now ap-
pears that even viruses can be retained by sponges, with some
23% of viral particles being removed from seawater by the Red
Sea sponge Negombata magnifica (131). Considering the enor-
mous abundance of viruses in seawater (1 million to 100 mil-
lion per ml) (387), this could represent a significant flux of
nutrients in ecosystems containing large sponge populations.
While most studies of sponge feeding have been conducted
with demosponges (e.g., see references 89, 196, 307, 313, and
379), microbial retention efficiencies of 90% or more have also
been reported for hexactinellids (494, 496). The deep-sea
hexactinellid Sericolophus hawaiicus was somewhat less effi-
cient, with microbial retention efficiencies ranging from 47%
for bacteria to 54% for photosynthetic eukaryotes of ?3 ?m
Consumption of symbiotic microorganisms has also been
raised as a possible food source for sponges (336, 337). The
first report of apparent widespread disintegration, both intra-
and extracellularly, of cyanobacterial symbionts was from Sara
in the early 1970s (336). His TEM observations suggested that
the Mediterranean sponge Ircinia variabilis actively degraded
Aphanocapsa-type cyanobacteria (now considered Synechococ-
cus spp.) (421) both in the sponge mesohyl and within certain
sponge cells, providing an important source of photosyntheti-
cally fixed carbon to the host sponge (336). These results have
since been questioned, with the suggestion that the observed
lysis of symbionts was in fact an artifact of the histology pro-
cedure (477). Wilkinson argued that while some cyanobacterial
cells may be digested intracellularly (e.g., see reference 473),
this is the exception rather than the rule. However, several
other reports of bacterial (including cyanobacterial) consump-
tion by sponges have since emerged (172, 222, 266, 422), lend-
ing weight to the notion that certain sponges may “farm”
bacteria as a food source (172). Indeed, phagocytosis and sub-
sequent intracellular digestion of bacteria are the presumed
mechanisms of nutrient transfer between a carnivorous Clado-
rhiza sponge and its methanotrophic symbionts (429, 431).
Harming the host: pathogenesis, parasitism, and fouling.
Deleterious effects of microbes on sponges may be direct (i.e.,
pathogenesis or parasitism) or indirect (e.g., microbial films
promoting surface fouling). The various reported instances of
sponge disease have generally been attributed to bacteria or
fungi (199), yet in most cases the responsible microbe(s) has
not been identified unequivocally. A notable exception is a
2002 study by Webster and colleagues (455) in which they
isolated a pathogenic alphaproteobacterium (designated strain
NW4327) from an infected individual of the Great Barrier
Reef sponge Rhopaloeides odorabile. Strain NW4327, which is
related to the tumor-forming symbionts of Prionitis sp. mac-
roalgae (12) and to the causative agent of juvenile oyster dis-
ease (34, 35), was shown to infect and kill healthy sponge tissue
(455). The mechanism by which this occurred was via degra-
dation of the collagenous spongin fibers, with almost the entire
sponge surface subject to tissue necrosis following experimen-
tal inoculation with strain NW4327 (Fig. 19). Similarly infected
tissue, with documented bacterial attack of spongin fibers, was
evident during a devastating outbreak of disease in commer-
cially important Mediterranean sponges during the late 1980s
(433). Mass mortalities of commercially important sponges
(e.g., Spongia spp.) have occurred several times in both the
Mediterranean (199, 298) and Caribbean (119, 199, 375), vir-
tually eliminating commercial sponge fisheries in some areas.
Not only sponges, but also corals and other epibenthic organ-
isms, experienced extensive mortality during a 1999 episode in
the northwestern Mediterranean (52). This outbreak coincided
with a sudden increase in seawater temperature, with subse-
quent laboratory studies suggesting the additional involvement
of both protozoans and fungi. Increased microbial virulence
FIG. 19. Effect of a bacterial pathogen on a marine sponge. Transmission electron micrographs of Rhopaloeides odorabile tissue are shown,
displaying (A) the diversity of bacterial morphotypes in healthy tissue, (B) a sponge experimentally infected with the alphaproteobacterial pathogen
strain NW4327, and (C) consequent necrosis of the sponge tissue. Bar ? 500 nm. (Reprinted from reference 455 with permission of the publisher.)
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS325
and/or compromised host resistance linked to global warming
has already been postulated as a cause of many mass mortal-
ities of marine organisms (139, 140), and it will be of great
interest (and concern) to see how marine sponges are affected
by predicted rises in seawater temperature in the future. Other
reports of diseases in sponges include the so-called Aplysina
red band syndrome, afflicting aplysinid sponges on Bahaman
reefs (262), cyanobacterial overgrowth of Geodia papyracea
(330), and repeated observations of diseased sponges on a
Panamanian coral reef over a 14-year period (492). Bleaching
of Xestospongia muta and other Caribbean sponges has also
been reported (64, 91, 251, 441), but it remains to be estab-
lished whether, as is the case for some corals (325, 326) and an
Australian macroalga (R. J. Case, A. Low, W. C. Chen, S.
Longford, G. R. Crocetti, N. A. Tujula, P. Steinberg, and S.
Kjelleberg, presented at the 11th International Symposium on
Microbial Ecology, Vienna, Austria, 20 to 25 August 2006),
bacterial pathogens are to blame. Bacteria of two genera (Ba-
cillus and Pseudomonas) were identified as possible agents of
disease in the Papua New Guinean sponge Ianthella basta (54).
This fan-shaped sponge has undergone significant mortality at
a number of inshore sites, leading to speculation that the
putative pathogen(s) may be of terrestrial origin. A simple
model was recently developed to describe the role of sponge
morphology in recovery from disease, with branching sponges
being the most likely to recover (493). To our knowledge,
nothing is known about diseases of freshwater sponges.
Parasitism of sponges by diatoms has been reported for
several Antarctic species (16, 51). Bavestrello and coworkers
found a negative correlation between chlorophyll a (used as a
proxy for diatom abundance) and sponge carbohydrate levels
(16), while in a parallel study of the hexactinellid sponge Sco-
lymastra joubini, they described a degradation of sponge inter-
nal tissue in areas of dense diatom aggregations (51). The
diatoms in S. joubini were of the genus Melosira and appeared
to enter the host either through the ostia (inhalant openings)
(Fig. 2) or via active incorporation by the sponge pinacoderm
(dermal membrane). Why sponges should actively incorporate
potentially harmful diatoms is not clear, although consumption
of diatoms as a food source is one possibility (113, 114). Al-
ternatively, the silica-encased diatoms may “trick” the sponge
cells into taking them up (51), a plausible explanation given the
tendency of some sponges to incorporate siliceous particles
from the surrounding environment (15, 17, 18, 107). Currently,
nothing is known about the nature of the interactions between
sponges and diatoms in tropical and temperate systems (65,
Microbes may also harm sponges in a less direct manner, for
example, by promoting the fouling of sponge surfaces. Any
surface in the marine environment, biotic or abiotic, is subject
to intense fouling pressure. During the colonization process, a
new surface will first develop a biochemical conditioning film,
followed by microbial fouling (e.g., colonization by bacteria
and diatoms). This biofilm then acts as a precursor to attach-
ment by macrofouling organisms, such as invertebrates and
macroalgae (71, 450), which in the worst cases can negatively
affect sponge nutrition by blocking feeding channels or can
increase hydrodynamic drag, resulting in sponge dislodgment
from the substratum.
It is important that sponges not be considered mere helpless
targets for potentially harmful microorganisms. Compounds
with antibacterial, antifungal, or antifouling properties are pro-
duced by many sponges (19, 32, 112), and those chemicals with
more specific effects may allow the host sponge to select for
harmless or even beneficial microbes while deterring deleteri-
ous types. Interestingly, the resident microbial community may
also participate in host defense, and there are numerous ex-
amples of the antimicrobial potential of apparently indigenous
microbes (56, 147, 203, 397, 402). In addition, at least one
sponge, Halichondria panicea, prevents fouling and sedimen-
tary clogging of its ostia by sloughing off its outer tissue layer
every few weeks (14), while the innate immune systems of
sponges (see “Establishment and Maintenance of Sponge-Mi-
crobe Associations”) are also believed to play a role in the
prevention of microbial invasion.
The Big Picture: Temporal and Biogeographic Variability in
Microbial Communities of Sponges
Variability in sponge-associated microbial communities has
been examined at a number of levels, such as over time (days
to months) (105, 201, 390, 462), within and among individuals
of the same sponge species (mm to thousands of km) (7, 146,
201, 365, 390, 391, 404, 454, 462), and among different host
species (146, 151, 208, 390). Other studies have investigated
the spatial and/or temporal distributions of specific microbial
taxa within sponges (100, 226, 294, 421, 453). If an emergent
theme is to be identified from these studies, it is that, with
some exceptions, sponge-associated microbial communities ap-
pear to be relatively stable, with little variation in time and
space (148). The main caveat to this statement is that the
methods employed may not always detect the variability which
is present. We review the main published findings on these
topics and, where appropriate, use our own phylogenetic anal-
yses (see Evolution and Diversity of Sponge-Associated Mi-
croorganisms) to aid our discussion of symbiont biogeography.
Considering first those studies in which the whole microbial
community was targeted, only a few papers deal with temporal
variability. The first examined such variation in aquarium-
maintained Aplysina aerophoba (105). The authors used several
methods to characterize the resident microbial community,
including cell counts (both 4?,6?-diamidino-2-phenylindole
[DAPI]- and cultivation-based), TEM, FISH, and DGGE.
What they revealed was an extremely abundant bacterial com-
munity (6.4 ? 108cells per g sponge tissue) which varied little
during an 11-day incubation period, even under starvation con-
ditions or upon exposure to antibiotics. Although DGGE
banding patterns changed slightly during the antibiotic treat-
ment, relative levels of abundance of the major bacterial
groups, as assessed by FISH, stayed fairly constant irrespective
of treatment (105). The lack of observed temporal variability as
well as the apparent resistance of the community to distur-
bance suggests that A. aerophoba harbors a highly stable mi-
crobial community. Similarly, cultivation for up to 8 months
did not seem to greatly alter the bacterial community in Geodia
barretti, at least at the broad phylogenetic levels targeted by the
applied FISH probes (160). Temporal variability has also been
examined among sponges in the field. A 16S rRNA gene-
DGGE study found bacterial communities in the temperate
Australian sponges Callyspongia sp., Stylinos sp., and Cymbas-
326TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
tela concentrica to be highly stable over the course of a year,
while additional sampling of the last species revealed a similar
lack of variation on a shorter (days to weeks) time scale (390).
Another DNA fingerprinting method, terminal RFLP analysis,
also identified only relatively minor temporal changes in bac-
terial community composition on the surface of the sponge
Mycale adhaerens from Hong Kong (201). In contrast to these
studies, the bacterial community profile of the North Sea
sponge Halichondria panicea, as assessed by 16S rRNA gene-
DGGE, varied considerably over a 10-month period (462). The
archaeal community, also assessed by DGGE, varied little.
Another study by the same group, on the North Sea sponge
Pachymatisma johnstonia, demonstrated stable bacterial com-
munities in specimens sampled at different times (2 years
apart) from two Orkney Isles collection sites (A. Wichels, S.
Kuppardt, and G. Gerdts, presented at the 10th International
Symposium on Microbial Ecology, Cancun, Mexico, 2004).
Spatial variability in sponge-associated microbial communi-
ties has been studied from the millimeter to the interocean
scale. Taylor and coworkers examined spatial variability within
and among individuals of three cooccurring Australian sponges
(390). In all cases, the variation in 16S rRNA gene-DGGE
banding patterns (and inferred community compositions) was
minor, with even the least similar samples for a species sharing
?70% of bands. For the 30% or less of bands which did vary,
most of the variation could be ascribed to differences among
rather than within individual sponges. Considerable differences
were seen among different host species, with one sponge in
particular harboring a distinct bacterial community compared
to those in the other two species (390). In another study,
Wichels et al. found differences between mesohyl-inhabiting
microorganisms and transient microbes present in the sponge
aquiferous system (462). The latter fraction of microbes was
targeted by gently compressing Halichondria panicea tissue
within a syringe and collecting the outflowing water. Although
incomplete separation of aquiferous system and tissue frac-
tions may have sometimes disguised differences (462), in gen-
eral there did seem to be distinct communities between the two
sample types. Marked differences were also evident between
outer (cortex) and inner (endosome) tissues in the Mediterra-
nean sponge Tethya aurantium (404). Cell separation tech-
niques used in natural product research on sponges have also
identified patterns of microbial distribution within sponge tis-
sues. For example, cyanobacterial symbionts in the ectosome
(outer tissues) of Theonella swinhoei were readily separated
from a filamentous bacterium (later identified as the delta-
proteobacterium “Candidatus Entotheonella palauensis” )
which occurs exclusively in the inner endosome (27, 28). It is
typical for phototrophic symbionts, such as cyanobacteria, to
be prevalent in the outer, well-illuminated surfaces of host
sponges, while other microorganisms may dominate the inner
Moving up to the next spatial scale, we consider geographic
patterns of variability. A 16S rRNA gene-DGGE study of the
Antarctic sponges Homaxinella balfourensis, Kirkpatrickia vari-
alosa, Latrunculia apicalis, Mycale acerata, and Sphaerotylus
antarcticus revealed that associated bacterial communities
were highly consistent, both among individual sponges at the
same sampling site and also among three different sampling
sites separated by some 10 km (454). The first molecular study
of large-scale biogeographic variability in sponges was the 2002
study by Hentschel and colleagues (146), which we discussed at
length in previous sections. The sponges Rhopaloeides odor-
abile (458), Aplysina aerophoba, and Theonella swinhoei con-
tained substantially overlapping microbial communities whose
sequences often fell in monophyletic, sponge-specific clusters,
despite wide (host) phylogenetic and geographic separation
(146). A 2005 study employed 16S rRNA gene-DGGE to in-
vestigate the bacterial community in the sponge Cymbastela
concentrica along the eastern Australian coast (391). At eight
sampling sites spanning 500 km of coastline within the tem-
perate range of the sponge, bacterial community composition
varied little. However, C. concentrica sponges from a tropical
location ?1,000 km away had a seemingly very different resi-
dent bacterial community. Seawater collected during sponge
sampling varied comparatively little between the tropical and
temperate locations. Allopatric speciation (resulting from ad-
aptation to geographically separated hosts) is one possible
explanation for the different communities, although latitudinal
changes in environmental factors (e.g., temperature and light)
could also be responsible. A more prosaic explanation is that
the C. concentrica individuals from the two locations could
simply be distinct (sub)species, although molecular taxonomy
studies would be needed to confirm this (391). Cultivation
efforts by Sfanos and colleagues in which 17,000 bacterial iso-
lates were obtained and more than 2,000 were screened by 16S
rRNA-based RFLP fingerprinting and/or sequencing often
yielded the same bacterium from many sponge hosts from
multiple locations (365). The most extreme case was that of an
alphaproteobacterium (GenBank accession no. AY362009),
which was recovered from 18 sponge species (plus several
nonsponge sources, such as a coral and a sea cucumber) spread
among various Caribbean and eastern Atlantic locations (365).
Several papers have examined the temporal and spatial vari-
ability of particular sponge-associated microorganisms, often
providing valuable clues to the nature of the sponge-microbe
association (i.e., true symbionts would be expected to maintain
a long and consistent relationship). For example, “Candidatus
Cenarchaeum symbiosum,” the sole archaeon present in the
marine sponge Axinella mexicana, was recorded from all 23
individuals of this sponge collected from the Californian coast
over a 9-month period (294). Furthermore, archaeal rRNA
levels stayed relatively constant (and high) for more than a
year in aquarium-maintained A. mexicana, indicating a highly
stable relationship between the sponge and its archaeal inhab-
itant. A subsequent study demonstrated temporally and spa-
tially stable associations between three Mediterranean ax-
inellid sponges (Axinella damicornis, Axinella verrucosa, and
Axinella sp.) and their single, host-specific crenarchaeal asso-
ciates (which, in all cases, were related to “Ca. Cenarchaeum
symbiosum”) (226). The cultivable fraction of the microbial
community in Rhopaloeides odorabile was always dominated,
irrespective of sampling time or location, by a specific alpha-
proteobacterium (453). This bacterium, which is closely related
to Pseudovibrio denitrificans (Fig. 14), comprised ?80% of the
total heterotrophic bacterial colony count in samples collected
over a 460-km portion of the Great Barrier Reef as well as
in those collected during four consecutive seasons at one
reef. Another alphaproteobacterium, affiliated with the genus
Rhodobacter, was found in all samples of the sponge Halichon-
VOL. 71, 2007SPONGE-ASSOCIATED MICROORGANISMS327
dria panicea from the Adriatic, Baltic, and North seas (7). In a
much earlier study, Wilkinson et al. isolated a particular bac-
terium (or at least a highly similar one, as molecular data were
not feasible at that time) from several sponges in the Mediter-
ranean and the Great Barrier Reef (483).
The biogeography of sponge-associated cyanobacteria has
recently come under close scrutiny. Usher and colleagues per-
formed an extensive survey of cyanobacterial symbionts, sam-
pling nine sponge species (from six genera) in the Mediterra-
nean Sea and the Pacific, Southern, and Indian Oceans (421).
In addition, one of these sponges, Chondrilla australiensis, was
sampled from eight Australian locations spanning several
thousand kilometers and a wide temperature range. 16S rRNA
gene sequences representing at least four closely related lin-
eages of Synechococcus spp. were recovered from the various
host sponges and included, most notably, “Candidatus Syne-
chococcus spongiarum” (426), which was present in four of the
sampled sponges, including all sampled individuals of C. aus-
traliensis (421). Interestingly, the “Ca. Synechococcus spongia-
rum” sequences from the Usher et al. study comprise, together
with other sequences obtained independently from the Carib-
bean (78, 151, 342, 383), Red Sea (383), east Africa (383),
Micronesia (146, 394), Mediterranean (146), and southeastern
Australia (this study), one of the largest documented mono-
phyletic, sponge-specific clusters (Fig. 5). In total, “Ca. Syn-
echococcus spongiarum”-like sequences have been recovered
from 21 sponge species from around the world, making this
organism similarly widely distributed as its free-living Syne-
chococcus/Prochlorococcus counterparts (273, 340).
Many of the sponge-specific clusters from other phyla are
also widely distributed. The “Poribacteria,” for example, have
so far been identified from sponges in the Mediterranean,
Caribbean, and eastern Pacific (100; this study), while a large
sponge-specific cluster within the Actinobacteria (Fig. 13) con-
tains sequences from the Red Sea (146), South China Sea
(data not shown), Indonesia (235), and various Caribbean lo-
cations (235, 342; this study). Numerous other such examples
exist, indicating an apparently global distribution of many
sponge-specific microbes (Fig. 20). Hill and colleagues (151)
attempted to relate the occurrence of major (sponge-associ-
ated) bacterial taxa to the geographic location where the host
sponge was collected. They suggested that some patterns could
be discerned whereby specific taxa might be better represented
in, e.g., tropical but not cold-temperate sponges. However, we
argue that such statements are premature and that there are
insufficient data to draw firm conclusions at present. For ex-
ample, many (or most) of the existing 16S rRNA gene libraries
from sponges have not been sampled exhaustively, and missing
taxa may well be represented in a library of clones but not yet
identified due to low sequence coverage. Furthermore, differ-
ent geographic areas (and different habitat types within an
area) have not been studied equally well, so there could be
biases towards apparently more diversity in some areas which
have received more attention. The construction of 16S rRNA
gene libraries from sponges in underrepresented locations
(e.g., Africa, South America, and western North America)
would go a long way towards improving our understanding of
sponge symbiont biogeography.
Throughout this review, we have attempted to carefully eval-
uate published data in the context of the methods and ap-
FIG. 20. Global distributions of selected monophyletic, sponge-specific clusters. Symbols refer to collection locations for representatives of the
“Ca. Synechococcus spongiarum” (Cyanobacteria) (circles), Actinobacteria (triangles), and Acidobacteria (stars) clusters. In the last cluster,
coral-derived sequences from the Mediterranean are also present.
328 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
proaches used for a particular study. This is never more critical
than when considering variability in complex microbial com-
munities. Appropriate sampling designs are important for in-
vestigations of any ecological system, yet in the past this has
often been overlooked in microbial ecology circles (238, 390).
The fact that one should analyze sufficient replicate samples to
encompass biological variability is beyond question, yet many
of the available methods for characterizing microbial commu-
nities are seemingly incompatible with this goal. DNA finger-
printing techniques such as DGGE, terminal RFLP analysis,
and automated ribosomal intergenic spacer analysis offer
high-throughput analyses of large numbers of samples, but
together they suffer from one major drawback: without con-
siderable efforts (e.g., sequencing of excised DGGE bands),
the bands or peaks representing particular microorganisms
have no identity. If banding patterns alone are compared as
proxies of community composition, one must acknowledge that
observed changes in a community could equally likely be due
to changes among two strains of the same microbial species or
to a much more significant shift, such as from a member of one
bacterial phylum to another. Conversely, FISH and 16S rRNA
gene library analyses can provide detailed quantitative and
phylogenetic information, respectively, yet neither approach is
well suited to analyzing large numbers of samples. Based on
our own experience as well as reports in the literature (e.g., see
reference 458), autofluorescence in sponges can also create
difficulties for FISH analyses. A need therefore exists for a
phylogenetically informative yet rapid means of assessing mi-
crobial community structure. Microarrays offer particular
promise in this regard, with a range of 16S rRNA- and func-
tional gene-based microarrays already available (reviewed in
references 122 and 215). The highly parallel nature of microar-
rays provides the potential, for example, to survey the presence
of multiple sponge-specific clusters in a single assay, something
which is not possible with other existing techniques. Impor-
tantly, symbiont function could also be addressed via the
so-called isotope array approach (2).
BIOTECHNOLOGY OF SPONGE-MICROBE
ASSOCIATIONS: POTENTIAL AND LIMITATIONS
Biologically Active Chemicals from Marine Sponge-Microbe
Consortia and Their Commercial-Scale Supply
An enormous number of biologically active compounds have
been isolated from marine sponges and their associated micro-
organisms. Indeed, sponges are the most prolific marine pro-
ducers of novel compounds, with more than 200 new metabo-
lites reported each year (see reference 32 and preceding
reviews in that series). Furthermore, more sponge-derived
compounds are in clinical and preclinical trials (e.g., as anti-
cancer or anti-inflammatory agents) than compounds from
any other marine phylum (31). The occurrence in unrelated
sponges of structurally similar compounds, particularly those
which were otherwise known exclusively from microorganisms,
led to speculation that such compounds (including some al-
ready in drug trials) were of microbial origin (27, 143, 280, 427)
(Fig. 21). Since chemical synthesis of natural products can be
problematic and expensive due to their structural complexity
(4, 48, 373), the realization that at least some compounds may
be produced by microbes raised hopes of obtaining a sustain-
able, essentially unlimited supply of compounds for testing and
subsequent drug production (e.g., via cultivation of the rele-
vant bacteria) (280, 297). Today, the true origin of most sponge
compounds has still not been proven unambiguously and re-
mains a key issue among marine natural product chemists. The
possibility of convergent evolution of biosynthetic pathways
among different sponges has also been raised (332). It is not
our intention to comprehensively review sponge-derived natu-
ral products; such reviews are the subject of chemistry rather
than microbiology per se, and many excellent reviews dedi-
cated to this topic already exist (31, 32, 191, 236, 279, 280).
Rather, we focus our attention on selected important examples
and highlight some of the difficulties involved with obtaining a
consumer-ready end product.
Sponge (or microbe)-derived compounds span a wide range
of chemical classes (e.g., terpenoids, alkaloids, peptides, and
FIG. 21. Chemical structures of jaspamide (left), from Jaspis sp. sponges, and chondramide D (right), from the deltaproteobacterium
Chondromyces crocatus. Note the remarkable structural similarities between the compounds.
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS329
polyketides) with an equally wide range of biotechnologically
relevant properties (e.g., anticancer, antibacterial, antifungal,
antiviral, anti-inflammatory, and antifouling) (31, 32, 112, 186,
229, 236, 279, 280). The attention of natural product chemists
and pharmaceutical companies, at present, is focused firmly on
anticancer drugs, with several promising sponge-derived com-
pounds in clinical and preclinical cancer trials (31, 255, 370).
The large number of novel, active metabolites being reported
from sponges every year begs the question of why such chem-
icals have not yet made it to pharmacy shelves. To date, and to
the best of our knowledge, not a single compound obtained
from a sponge has been approved as a drug, with a major brake
on progress being the so-called supply problem (138, 297, 408).
(The nucleoside analogs Ara-A and Ara-C, commercialized as
antiviral and anticancer agents, respectively, could arguably be
considered the sole exceptions. They were not isolated directly
from sponges but are synthetic derivatives based on com-
pounds from the Caribbean sponge Cryptotethia crypta [24,
25].) Biologically active natural products are often produced in
relatively small amounts, and often by rare animals whose
natural populations cannot sustain the extensive collections
required for clinical trials. Alternative means for producing
large amounts of metabolites are therefore required. We illus-
trate this issue by using two examples, the anticancer com-
pounds halichondrin B and peloruside A.
The halichondrins are a group of polyether macrolides that
exhibit potent antitumor activities (158, 415). First isolated
from the Japanese sponge Halichondria okadai in the mid-
1980s (158), they were subsequently found in several other
sponges from diverse geographic locations, including Axinella
spp., Phakiella carteri, Raspailia agminata, and Lissodendoryx
sp. (138). Halichondrin B (Fig. 22) was particularly sought
after due to its high cytotoxicity, and its total synthesis was
reported as early as 1992 (4). However, due to the structural
complexity of the compound, many steps were required for its
synthesis, rendering total synthesis impractical for industrial-
scale production. While the occurrence of halichondrins in
many unrelated sponges suggested a microbial origin, little was
known about the microbiology of the relevant sponges, and
thus alternative avenues were investigated (to our knowledge,
the precise [i.e., sponge versus microbial] origin of the hali-
chondrins has never been determined unambiguously). Lisso-
dendoryx sp., collected from the coast of southern New Zeal-
and, yielded the largest amounts of halichondrins and
therefore became a focus of drug supply efforts (138, 250).
Based on the potency of halichondrin B and its projected
demand if approved for human use, the requirement for clin-
ical trials was estimated to be ?10 g, with annual requirements
as a commercial drug of 1 to 5 kg (138). Given that 1 metric ton
of Lissodendoryx sp. sponges yielded only 300 mg of halichon-
drin B and that the entire natural biomass of the sponge was
estimated to be only 289 metric tons, collection from the wild
was quickly ruled out. Aquaculture of Lissodendoryx sp. was
then investigated, with promising initial results (250). How-
ever, scale-up of the operation to the levels necessary for
commercial production was not achieved due to a lack of
funding (M. J. Page, personal communication), and the small
amounts of compound present in the sponge tissue may render
the aquaculture option economically untenable in this case
anyway (373). Nevertheless, halichondrin B may yet prove to
be a success story, with a synthetic analog, E7389, in phase I
clinical trials as an anticancer compound (370). This simplified
version of halichondrin B is more amenable to chemical syn-
thesis but retains the biological activities of the original com-
Our second example concerns the macrocyclic lactone
peloruside A (460) (Fig. 23). Isolated from the New Zealand
demosponge Mycale hentscheli, peloruside A shows promising
FIG. 22. Chemical structure of halichondrin B.
FIG. 23. Chemical structure of peloruside A.
330 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
anticancer properties, acting in a similar manner and potency
to the widely used cancer drug paclitaxel (Taxol) (166). With
the compound currently in preclinical trials, two avenues are
being pursued in parallel to ensure a sufficient supply of the
compound for subsequent clinical trials. Chemical synthesis is
one approach, with several groups recently reporting partial or
total synthesis (e.g., see reference 177). A New Zealand con-
sortium, working together with a U.S. pharmaceutical com-
pany, is currently investigating whether cost-effective, industri-
al-scale synthesis is achievable (137). An alternative supply
option for peloruside A is being explored by the same group,
with aquaculture of M. hentscheli looking highly encouraging
(137, 271). With 200 kg of sponge yielding a mere 2 g of pure
peloruside A, scaling-up is a priority, with the goal of growing
?500 kg of sponge over the coming year (137). Other com-
pounds of pharmaceutical interest are also produced by M.
hentscheli, namely, the cytotoxic polyketide mycalamide A and
the macrolide pateamine (165, 256, 270, 278). Concentrations
of these metabolites in natural sponge populations vary signif-
icantly in time and/or space (270), suggesting that complex
ecological and physical factors may be involved in their pro-
duction. An improved understanding of the ecological roles of
these and other compounds could greatly benefit metabolite
harvesting programs, and indeed, ecological observations are
often used to guide the initial stages of drug discovery in
marine environments (295, 359). Ongoing microbiological in-
vestigations with M. hentscheli (452; S. A. Anderson, unpublished
data) should also benefit future drug development efforts with
Supply issues notwithstanding, the pharmacological po-
tential of marine sponges and other sessile invertebrates
(e.g., corals, bryozoans, and ascidians) is enormous. Although
progress toward the commercial product stage has been slow,
it is highly likely that at least one of the several compounds
now in clinical trials (or a synthetic analog) will be commer-
cialized within the next few years. A combination of improved
chemical synthesis methods with the various approaches out-
lined in the following section should ensure a bright future for
this field, with sponge-derived natural products being utilized
either in their natural form or as inspiration for new, labora-
tory-generated compounds (e.g., via chemical proteomics)
(287). As a footnote to this discussion, the freshwater sponges
should also be mentioned. Their chemistry has received much
less attention than that of their marine counterparts, and while
various lipids and a compound with antipredator activity have
been reported (77, 311), it is unclear whether these sponges
produce many, if any, compounds of pharmaceutical interest.
Methods for Accessing the Hidden Chemistry
of Marine Sponges
A number of (nonsynthesis) approaches are available for
accessing biologically active natural products from sponges and
the microorganisms within them (Fig. 24). For convenience, we
split these into the following three main themes: cultivation of
metabolite-producing microbes, sponge culture, and molecular
biological methods, such as metagenomics. In addition, we
highlight the importance of metabolite localization studies for
improving our knowledge of which partner (sponge or micro-
organism) is responsible for metabolite production.
Cultivation of metabolite-producing microorganisms. Culti-
vation of sponge-associated microorganisms that produce bio-
active compounds is the most direct method for large-scale
production of these chemicals (154), and cultivation ap-
proaches are widely practiced among those targeting bioactive
compounds (46, 47, 81, 130, 147, 154, 163, 176, 189, 235, 364,
365, 378, 453). The potential payoffs from the cultivation ap-
proach are obvious and substantial: if metabolite producers
can be isolated on artificial media and grown to significant cell
numbers (while continuing to produce the relevant metabo-
lite), then this obviates the need for large-scale harvesting of
natural sponge populations, with its environmentally and finan-
cially negative implications.
Two broad strategies for isolating microbial producers of
bioactive compounds were outlined by Hill in a recent review
(154). The first is to use a wide range of media in an effort to
grow as many different sponge-associated microbes as possible.
Since growth under different culture conditions may influence
which metabolites are produced, the use of many different
media and conditions should help to maximize the chemical
diversity from a given microorganism (154). Bacteria associ-
ated with deep-sea benthic invertebrates have been the subject
of extensive cultivation efforts by the Harbor Branch Oceano-
graphic Institution, with a range of nutrient-poor to nutrient-
rich media being utilized (130, 264, 365). Approximately 17,000
isolated microbes, most from deep water and mostly from
sponges, are present in the Harbor Branch Oceanographic
Marine Microbial Culture Collection (365). These include rep-
resentatives of the Proteobacteria, Bacteroidetes, Firmicutes,
and Actinobacteria and are the subject of natural product
screening. An alternative, more targeted approach is to go
after specific microbial groups with proven track records in the
production of bioactive compounds. Many such groups, includ-
ing cyanobacteria, fungi, and actinomycetes, are well known
from sponges (148, 163), with actinomycetes being the subject
of a particularly interesting success story. Sponge-derived
actinomycetes of the genus Micromonospora produce man-
zamines, alkaloids with, among other things, potent antimalar-
ial properties (10, 93, 155, 301). The first hint that manzamines
were of microbial origin came from the finding of these com-
pounds in many distantly related, geographically disparate
sponge species. Subsequent cultivation-dependent and -inde-
pendent characterization of the microbial communities in
two Indonesian manzamine-producing sponges, 01IND 35 and
01IND 52, revealed highly diverse assemblages, with the re-
covery of actinomycetes provoking intensive culturing efforts
in their direction (154). Growth of the sponge-derived Mi-
cromonospora sp. has since been achieved on a large scale in
20-liter fermentations, with maintenance of manzamine pro-
duction (R. T. Hill, personal communication). Improvements
in this process should greatly facilitate passage of these com-
pounds through the various stages of the drug-testing process.
Actinomycete-selective media were also used successfully with
the sponges Pseudoceratina clavata (188), Xestospongia spp.
(235), Hymeniacidon perlevis (498), and Craniella australiensis
(209). In the last study, many of the cultivated actinomycetes
displayed broad-spectrum antimicrobial activities.
There are numerous other examples of the production of
biologically active compounds by sponge-derived microbial iso-
lates. An antibacterial peptide was isolated from both the
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS331
sponge Hyatella sp. and an associated Vibrio sp. (260), while a
glycoglycerolipid produced by a Halichondria panicea-derived
Microbacterium sp. had antitumor properties (463). In another
study, several quinolones, including one with both antimicro-
bial and cytotoxic activities, were isolated from a pseudomonad
from the Pacific sponge Homophymia sp. (45). Although the
mechanistic basis was not identified, 27 bacteria isolated from
the Mediterranean sponges Aplysina aerophoba and A. caver-
nicola exhibited antimicrobial activities in a series of assays
(147). Given the activity of some of these isolates against
clinically important multiresistant Staphylococcus aureus and
Staphylococcus epidermidis strains, assay-guided fractionation
and subsequent chemical characterization of any active com-
ponents could prove particularly profitable. Terrestrial fungi
have a long-standing reputation as prolific producers of bioac-
tive natural products (44), and it is hardly surprising that
sponge-associated fungi also show promise in this regard.
Many examples exist of sponge-derived fungi that produce
bioactive compounds (29, 42, 44, 163, 174, 176, 295, 296, 451).
Although many of the isolated fungi are of suspected terres-
trial origin (i.e., they are closely related to typical terrestrial
species), to some extent this does not matter for drug discovery
purposes; even if sponges act only as mere accumulators of
contaminant fungi, these microorganisms can still be targeted,
and once they are isolated, there may be no need to attempt
reisolation from the original sponge hosts. Interestingly, it ap-
pears that unlike bacteria, fungi are not the source of any
natural products previously ascribed to marine sponges (191).
The success of efforts to isolate sponge-associated microor-
ganisms that produce bioactive compounds is dependent upon
a number of factors. Most significantly, the majority of envi-
ronmental microorganisms, including those in sponges, have
proven resistant to cultivation by standard techniques (105,
380). Although various authors have reported improved cul-
turability of sponge-associated bacteria via, for example, sup-
plementing the media with sponge tissue extracts (458) or
catalase and sodium pyruvate (264), the proportion of total
sponge bacteria that can be isolated has remained low. Only
0.06, 0.1, 0.15, and 0.7% of total bacteria could be cultured
from the sponges Candidaspongia flabellata (47), Rhopaloeides
odorabile (47, 453), Aplysina aerophoba (105), and 01IND 35/
01IND 52 (154), respectively. Santavy and colleagues were able
to achieve somewhat better recovery (3 to 11%) from the
Caribbean sclerosponge Ceratoporella nicholsoni, although ob-
viously, even in this case, some 90% of the resident bacteria
were not captured by cultivation attempts (335). While culti-
vation difficulties are hardly confined to sponges, it is never-
theless likely that many of the sponge associates are obligate
symbionts which may have evolved with the sponge over hun-
dreds of millions of years (see Evolution and Diversity of
FIG. 24. Approaches for obtaining bioactive metabolites from marine sponges.
332TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
Sponge-Associated Microorganisms) and will, due to nutri-
tional or other dependencies, be extremely difficult to obtain in
pure culture. Furthermore, those that can be isolated may not
necessarily produce the compound anymore, as they may re-
quire some as yet unknown cue or metabolic intermediates
from the host sponge. Additionally, and for unknown reasons,
some bacteria simply stop producing the compound of interest
after a certain time on artificial media (145). This could be due
to any of a number of genetic reasons relating to a lack of
selective pressure in pure culture, e.g., point mutation in a key
gene or loss of a mobile genetic element carrying the biosyn-
Sponge culture. Culturing sponges is another way to address
the supply problem, irrespective of whether compound pro-
duction is due to the sponge or the symbiont. Methods em-
ployed for the cultivation of sponges for drug production vary
widely in scale and sophistication, from sea-based aquaculture
to in vitro cultivation (in closed or semiclosed systems) to cell
and tissue cultures (22). The technical and economic potential
of each of these was reviewed recently (373).
In-sea sponge aquaculture has received considerable atten-
tion for its potential to cost-effectively address the metabolite
supply issue (Fig. 25) (84–88, 132, 137, 231, 248, 271, 299, 439).
Of all approaches, it has the advantage of most closely simu-
lating the conditions encountered by sponges in nature, and
practitioners have been able to draw upon more than a century
of experience in farming bath sponges. On the negative side,
the inherent unpredictability of marine environments can cre-
ate problems (e.g., due to atypical climatic conditions or storm
damage) (439) which are easily avoided in controlled in vitro
systems. The outcomes of sponge aquaculture trials have var-
ied widely, with success dependent upon a number of factors,
including the type of farming structure (84, 85, 132), sponge
growth form (86), farming location (271), and season of trans-
plantation (86, 87). If sponge survival can be ensured, then
growth increases of up to 5000% per year (relative to the
starting size) are achievable, depending on the sponge species
examined (271). Crucially, bioactive metabolites are typically
retained in farmed sponges (84, 87, 132, 248, 250, 271).
The consequences of environmental variability can be side-
stepped by cultivating sponges under semienclosed or even
fully closed conditions (22, 90, 141, 161, 267–269). Although
this is generally more expensive than sea-based aquaculture, an
obvious advantage is the ability to control environmental pa-
rameters, such as growth temperature, water movement, and
food supply, as well as to eliminate biomass loss due to storms
or disease outbreaks (see “The Varied Nature of Sponge-
Microbe Interactions”). Potential problems in recirculating
systems include a buildup of toxic secondary metabolites and
metabolic wastes, such as ammonia (22). Considerable growth
(?200% in 1 to 2 months) of Pseudosuberites andrewsi explants
was achieved in a bioreactor (268), with even more growth
(?1,000% in 45 days) observed for the sponge Crambe crambe
in a closed system (21). Like the case for sea-grown sponges,
metabolite production has been observed for sponges sub-
jected to in vitro cultivation (e.g., see references 75 and 90).
However, one must remember that despite its promise, this
technology remains in its infancy, and to our knowledge, there
are no examples of industrial-scale in vitro cultivation of
Sponge cell culture for the production of biologically active
metabolites represents the other extreme of the scale contin-
uum from sea-based aquaculture. Although sponge cells can be
dissociated readily and even induced to divide in suspension
for several cycles, it has so far proven impossible to establish
continuous cell lines (reviewed in references 293, 319, and
320). Primmorphs, which are three-dimensional aggregates
comprising proliferating and differentiating sponge cells, have
therefore generated much recent interest since they can be
maintained for long periods (66, 240, 242, 247, 293). Particu-
larly exciting was the finding by Mu ¨ller and coworkers that
primmorphs from Dysidea avara grown in a bioreactor pro-
duced the secondary metabolite avarol, which is both charac-
teristic of this sponge and of great pharmacological interest
due to its strong biological activities (e.g., antitumor, antibac-
terial, and antiviral activities) (242). In contrast, single D. avara
cells did not produce avarol. Another interesting aspect of
primmorphs, especially within the context of this review, is that
symbiotic microorganisms can be retained within them (247,
398), potentially allowing for primmorph production of both
sponge- and microbe-derived compounds. Since their initial
demonstration in Suberites domuncula (66, 247) and then D.
avara, primmorphs have been generated from a wide range of
sponges, including Axinella polypoides, Cliona celata, Halichon-
dria panicea, Petrosia ficiformis, and Stylotella agminata (374,
434, 500). In the coming years, it should become clear whether
primmorphs can be scaled up sufficiently to overcome the
supply problem for many promising drug leads.
Surprisingly little information exists on the microbiology of
cultured sponges (in any system), yet this could be of vital
importance if metabolites are produced by microbial associ-
ates. For example, if sponges are cultured away from their
natural environment, then metabolite-producing symbionts
may conceivably be lost, or if metabolites are diet derived (as
suspected for okadaic acid) (297), then a change in diet would
presumably result in a loss of compounds. Moreover, even if
the desired metabolite is produced by the sponge itself, micro-
bial symbionts may still be of direct (e.g., by providing meta-
bolic precursors) or indirect (e.g., by affecting general sponge
health) significance (154).
FIG. 25. In-sea aquaculture of the Great Barrier Reef sponge Rho-
paloeides odorabile. (Image courtesy of Rocky de Nys [James Cook
University, Australia], reproduced with permission.)
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS333
Metagenomics. One of the most exciting developments in
molecular biology from a drug discovery perspective has been
the advent of environmental genomics, or metagenomics (92,
104, 136, 207, 346, 438). Metagenomics refers to the analysis of
genome fragments from a complex microbial community and
offers the potential for large-scale, sustainable production of
bioactive metabolites, including those produced by unculti-
vated microorganisms. If biosynthesis genes can successfully be
cloned and expressed in another (cultivated) microorganism,
such as E. coli, then this could ensure an unlimited supply of a
specific metabolite (48, 143). Different approaches for the
cloning and heterologous expression of biosynthesis genes
from marine invertebrate symbionts were recently the subject
of a comprehensive review by Hildebrand and colleagues
(149). The successful application of many of these methods is
exemplified by studies by Haygood and coworkers on the sym-
biosis between the bryozoan Bugula neritina and its bryostatin-
producing gammaproteobacterial symbiont, “Candidatus En-
dobugula seritina” (69, 70, 142, 143, 149, 150). Here we focus
our attention on the results of recent metagenomic studies
Two main types of analysis have been used to extract bio-
technologically relevant information from metagenome librar-
ies: one is based on function, whereby libraries are screened for
the expression of specific traits, and the other is based on
screening for sequences themselves (346). While screening for
functional traits (e.g., antibiotic production or quorum-sensing
inhibitors) has been successful (to various degrees) in other
environments (68, 318, 489), studies of sponge metagenomics,
to our knowledge, have been exclusively sequence based. To
date, the major foci of such studies have been the polyketide
synthase (PKS) and nonribosomal peptide synthetase genes
(187, 284, 285, 342). The PKSs are responsible for the synthesis
of bacterial polyketides, a diverse group of pharmacologically
important natural products which include the antibiotics eryth-
romycin and tetracycline as well as antitumor, immunosuppres-
sive, and cholesterol-lowering agents (197). Type I PKSs are
organized in a modular fashion, lending themselves to the
combinatorial biosynthesis of novel polyketides with poten-
tially useful pharmaceutical properties (197). A number of
marine drug candidates, including the bryostatins, disco-
dermolide, and the aforementioned peloruside A, belong to
this class of compounds (104, 150). The modular nature of the
polyketides suggests that environmentally retrieved PKS frag-
ments, which may not produce intact bioactive compounds,
could still be useful by providing modules for combinatorial
polyketide synthesis (187, 197). A large part of the sponge-PKS
story comes from work by Piel and colleagues (279–286). Their
study of the polyketide onnamide in sponges was greatly facil-
itated by prior metagenomic investigations into the production
of a structurally highly similar antitumor compound, pederin,
in beetles of the genus Paederus (281, 283, 286). The microbial
community within these beetles is much less complex than that
of sponges, allowing easier access to the genome of the bacte-
rial pederin producer (349). Ultimately, pederin production
was linked to a beetle symbiont closely related to Pseudomonas
aeruginosa, although evidence for lateral gene transfer of the
pederin-type genes suggests that compound production may
not necessarily correlate with rRNA-based bacterial phylogeny
(283, 285). Armed with a sound understanding of the genetic
bases of pederin biosynthesis, Piel et al. investigated the pro-
duction of onnamide in the sponge Theonella swinhoei from
Japan (285). Ketosynthase (KS) fragments were PCR ampli-
fied directly from the sponge metagenome, revealing a di-
verse range of sequence types. More importantly, PCR-based
screening of a 60,000-clone cosmid library with the same prim-
ers yielded a single KS-positive clone, which was fully se-
quenced. Strong indications existed for a bacterial origin of this
genome fragment (e.g., a lack of introns and small intergenic
distances), which should correspond to almost the entire re-
gion of the polyketide structure needed to obtain an antitumor
compound (285). In addition to the obvious biotechnological
importance of this study, as heterologous expression of such
gene clusters could lead to an inexhaustible supply of target
metabolites for clinical trials, interesting ecological and evolu-
tionary questions were also raised. For example, how did such
similar biosynthetic pathways come to be present in symbionts
of such dissimilar hosts, i.e., marine sponges and terrestrial
PKSs have also been studied in other sponges by using meta-
genomic approaches. In an attempt to isolate genes encoding
the promising antitumor compound discodermolide from the
Caribbean sponge Discodermia dissoluta, Schirmer and col-
leagues (342) employed a two-step approach. First, degenerate
KS primers were used to amplify 256 sequences (85 different
KS sequences), including several from trans-AT-type PKS do-
mains of the pederin and onnamide types. A selection of the
derived sequences was then used to create a probe pool for the
screening of 155,000 macroarrayed fosmid and cosmid clones;
given the proportion of bacterial inserts in the studied libraries
(?90%) and the average insert size (35 kb), ?4 Gb (some
1,000 bacterial genome equivalents) were screened (342). In
total, 1,025 PKS-positive clones (0.7% of all analyzed clones)
were identified. Interestingly, sequencing of selected fosmid
and cosmid clones revealed suprisingly little overlap between
these KS domains and those derived from the same sample by
the direct PCR approach. A PKS consistent with the biosyn-
thesis of discodermolide was also not found (342). Direct am-
plification of KS domains was also combined with the construc-
tion of fosmid libraries to study PKSs in another sponge, the
Great Barrier Reef species Pseudoceratina clavata (187, 188).
Each approach led to the retrieval of five KS domains, all of
which fell into an apparently sponge-specific KS cluster (to-
gether with sequences obtained from Discodermia dissoluta,
Theonella swinhoei, and an unidentified sponge) following phy-
logenetic analysis (187). Cultivated bacteria from P. clavata
were also screened by PCR for KS domains, with 10 such
domains detected in representatives of the Actinobacteria,
Alphaproteobacteria, and Firmicutes. None of the KS do-
mains from isolates clustered with the metagenome-derived
sequences, highlighting the importance of a polyphasic ap-
proach to encompass as much of the PKS diversity as possible
Although not part of a drug isolation strategy per se, no
discussion of sponge metagenomics would be complete without
considering the seminal work of DeLong and colleagues (134,
135, 294, 343–345). Over the past decade, the symbiosis be-
tween the Californian sponge Axinella mexicana and the psy-
chrophilic crenarchaeote “Candidatus Cenarchaeum symbio-
sum” (294) has been a model system for environmental
334TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
genomics. “Ca. Cenarchaeum symbiosum” was the first known
symbiotic crenarchaeote, and despite its as yet uncultivated
status, several factors have made it a suitable target for
genomic studies: it is the only archaeon in the sponge and
dominates the microbiota, consistently comprising some 65%
of all prokaryotic cells; it is always associated with the sponge;
and the symbiosis is maintained for a long time in aquaria
(294). Relatively large amounts of biomass and DNA have
therefore been available, with physical enrichment for the ar-
chaeal cells greatly facilitating the construction of large-insert
genomic libraries for this organism. The first published results
from the genomic analyses outlined the characterization of a
DNA polymerase which was homologous to those of cultivated
archaeal hyperthermophiles and yet, as revealed by heterolo-
gous expression in E. coli, was inactivated at temperatures
above 40°C, reflecting the symbiont’s low-temperature lifestyle
(345). Initial studies based on the 16S rRNA gene indicated
the presence of a single archaeal phylotype in A. mexicana
(294), so subsequent genome-derived indications of the pres-
ence of two closely related variants were unexpected (343).
Although the two variants differed ?0.8% in their 16S and 23S
rRNA genes and had an identical gene order for a studied
28-kb region, variations in DNA identity of up to 20% were
observed for protein coding regions, with up to 30% variation
for intergenic regions (343). These findings thus highlighted
the difficulties created by genomic microheterogeneity in as-
sembling environmentally retrieved genome fragments, and
these difficulties remain today (76, 414, 440). Despite such
obstacles, the DeLong group was able to assemble a closed
genome for “Ca. Cenarchaeum symbiosum” (134), which has
already yielded important insights into the metabolism of the
sponge symbiont in particular and of marine Crenarchaeota in
general (135). For example, genome reconstruction revealed
the potential of “Ca. Cenarchaeum symbiosum” to function
either autotrophically (as an ammonia oxidizer) or mixotrophi-
cally. In an extension of the environmental genomics approach,
homologues of genes involved in carbon and nitrogen metab-
olism were also found in metagenome libraries from ocean
waters worldwide, demonstrating the ubiquity of these meta-
bolic pathways among marine crenarchaeotes (135). Con-
versely, certain genes encoding cell surface, regulatory, or de-
fense mechanisms were not recovered from the free-living
relatives of “Ca. Cenarchaerum symbiosum,” suggesting that
these could be involved specifically with the establishment and
maintenance of the symbiosis (134).
The organism-oriented approach taken for “Ca. Cenar-
chaeum symbiosum” was also employed in a recent study of
the sponge-specific candidate phylum “Poribacteria” (100,
101). Virtually nothing is known about the physiology and
genetic makeup of these bacteria, and since there are no
cultured representatives, metagenomics offered a promising
approach. The sole entry point into the “Poribacteria” ge-
nome was the 16S rRNA gene sequence, and a single fosmid
clone among 29,000 clones (corresponding to 1.1 Gb of
DNA in total) was positive in an initial 16S rRNA PCR-
based screening (101). Analysis of this 39-kb insert revealed
27 open reading frames, including one encoding a new kind
of molybdenum-containing oxidoreductase and several en-
coding unusual transmembrane proteins (101).
The potential of metagenomics and other cloning ap-
proaches to revolutionize natural product research with
sponges is undeniable, yet there remain considerable technical
challenges. For example, if the microbial communities under
study are highly complex, then the genomes of target organ-
isms (e.g., “Poribacteria”) may remain largely hidden against a
background of genomes from other symbionts (but see Con-
clusions and Future Directions for possible means of enriching
specific genomes). Other potential problems include the use of
inappropriate host organisms for expression studies (some
hosts may not express the genes of interest) (104, 136) and the
large sizes of many gene clusters, which can prohibit their
successful cloning (although there exists the theoretical possi-
bility of reconstructing biosynthetic pathways via the assembly
of many smaller, overlapping sequence reads). In still other
cases, nonclustering of biosynthesis genes may be a problem,
with the genes of interest spread across different parts of the
symbiont’s genome (281). On the bright side, the relevant
technology is developing quickly. Massive sequencing efforts,
such as the recent Sargasso Sea study (440), may provide one
means of accessing rare genomes and the biotechnologically
relevant information within them. Indeed, an ambitious small-
and large-insert sequencing study along these lines was re-
cently initiated for a temperate Australian sponge (S. Kjelle-
berg, P. Steinberg, and T. Thomas, personal communication).
Similarly, the new pyrosequencing technology from 454 Life
Sciences (227) has already been applied successfully to com-
plex environmental samples (204) and may prove valuable in
future sponge metagenomics projects. Furthermore, high-
throughput screening techniques have developed to the extent
that a single worker can now screen, from start to finish, a
library of 400,000 clones for a particular gene in a mere 4 days
Cell separation and metabolite localization. The aforemen-
tioned techniques each offer the potential to generate large
quantities of biologically active metabolites from sponge-mi-
crobe associations. All three approaches (microbial cultivation,
sponge culture, and metagenomics) can, in principle, be un-
dertaken without prior knowledge of which partner (sponge or
microorganism) produces a given compound. However, the
most rational approach is to first establish which cell type(s) is
responsible for metabolite production, as this can determine a
logical strategy for future efforts (Fig. 24). Many researchers
have employed cell separation techniques for this purpose (27,
28, 99, 103, 120, 314, 315, 332, 416, 417). A major breakthrough
in this area came with the realization that chemically fixed
sponge and microbial cells retain their natural products in a
form amenable to chemical analyses, such as high-performance
liquid chromatography and nuclear magnetic resonance spec-
troscopy (416). Coupled with the relative ease with which one
can dissociate sponge tissue, this opened up a number of pos-
sibilities for natural product research. In the 1990s, researchers
in Faulkner’s laboratory thus took advantage of cyanobacterial
autofluorescence to separate, via flow cytometry, the filamen-
tous cyanobacterium Oscillatoria spongeliae from sponge and
other microbial cells in dissociated tissue of Dysidea herbacea
(416, 417). Of three major types of metabolites known previ-
ously from this sponge, two—polychlorinated peptides (416)
and polybrominated biphenyl ethers (417)—were found exclu-
sively inside O. spongeliae cells. In contrast, sesquiterpenoids
were confined to the sponge cells. While autofluorescing cells
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS 335
(e.g., cyanobacteria) have been separated from other cell types
(e.g., sponge and noncyanobacterial microbes) by flow cytom-
etry, other characteristics, such as cell size and, provided the
natural products are retained, FISH probe-conferred fluores-
cence, could also be used to differentiate sponge and symbiont
cells by this technique.
A more common approach has been to separate different
cell types in dissociated sponge tissue via centrifugation (27,
28, 314). Again, the Faulkner group was instrumental in estab-
lishing these approaches for sponge natural product study. In
the chemically rich lithistid sponge Theonella swinhoei, Bewley
and colleagues (28) identified four distinct cell types by elec-
tron microscopy (sponge cells, unicellular cyanobacteria, uni-
cellular heterotrophic bacteria, and filamentous heterotrophic
bacteria [the term “heterotrophic bacteria” is used here in
reference to noncyanobacterial microorganisms; these cells
could, in principle, be from chemolithoautotrophic bacteria]).
By following gross dissection of the sponge (into the endosome
[inner tissues] and ectosome [outer tissues]) with dissociation
and centrifugation, they were able to obtain relatively clean
fractions comprising each cell type. The macrocyclic polyketide
cytotoxin swinholide A was found only in the fraction contain-
ing the unicellular heterotrophic bacteria, while the potent
antifungal glycopeptide theopalauamide (350) was associated
with a filamentous bacterium (later identified as a deltapro-
teobacterium, “Candidatus Entotheonella palauensis” )
(28). Density gradient centrifugation provides yet another op-
portunity for separating sponge and microbial cells. In this
case, dissociated sponge tissue is placed above a density gra-
dient (consisting of, e.g., Ficoll or Percoll), and following cen-
trifugation, various cell fractions are formed based on differing
densities (103, 120, 121, 315). Individual fractions can then be
examined chemically and microscopically to correlate the oc-
currence of compounds with specific cell types. For Haliclona
sp., Garson and coworkers used Percoll gradients to demon-
strate localization of the cytotoxic alkaloids haliclonacyclamine
A and B within the sponge cells rather than in associated
Depending on the properties of and prior knowledge about
the compounds under study, localization may be achieved with-
out the need for prior cell separation. For example, bromi-
nated compounds, which are often biologically active and can
be present in large amounts in sponges, may be visualized in
situ by X-ray energy-dispersive microanalysis (406, 410). Spe-
cific antibodies for the toxin latrunculin B were used to prove
its localization in sponge but not symbiont cells within the Red
Sea demosponge Negombata magnifica (125). In a novel ap-
proach for sponges, catalyzed reporter deposition-FISH was
recently used to demonstrate mRNA expression from the
dysB1 genes (responsible for the biosynthesis of polychlori-
nated peptides) in cells of the cyanobacterium Oscillatoria
spongeliae in Dysidea (Lamellodysidea) herbacea (102). The
biological result was consistent with earlier work by others
(103, 416), but more significantly, this study proved the utility
of the method for the investigation of target chemicals in
sponges. The main limitation of the method is that a certain
level of genetic information must first be available for the
studied, or a related, biosynthetic pathway.
Although strongly suggestive, even the localization of a me-
tabolite to a particular cell type is not unequivocal proof of its
production in that cell or, indeed, even in that organism at all.
A compound may diffuse or be exported away from its site of
synthesis, especially if it is toxic to the producer (149). The
recent demonstration of swinholide A production by free-living
marine cyanobacteria (9) is a case in point: while Bewley and
coworkers (28) found this compound only in heterotrophic
bacteria (outlined above), it now seems plausible that cya-
nobacterial symbionts in Theonella swinhoei may produce the
compound but excrete it to be stored elsewhere. Alternatively,
both the heterotrophic bacteria and the cyanobacteria may
share the capacity for swinholide A biosynthesis, perhaps due
to lateral gene transfer (9). Interphylum and even interdomain
transfer of natural product pathways has been reported a num-
ber of times (e.g., the ?-lactam biosynthesis genes, responsible
for the production of antibiotics, including penicillins and
cephalosporins, are found in both fungi and bacteria ). Yet
another potential explanation is that a given metabolite could
be derived from the sponge’s diet. There is evidence suggesting
that this may be the case for okadaic acid, which has been
isolated from various sponges but is known to be produced by
free-living marine dinoflagellates (297, 467). Interestingly,
while localization studies have very often implicated the host
sponge as the source of bioactive metabolites, in at least some
cases it appears likely that intracellular or cell surface-associ-
ated bacteria may have been present but overlooked (149). In
such cases, microbial production of metabolites cannot be
ruled out completely.
It is important to note that while any of the aforementioned
localization techniques may give an indication of which is the
metabolite-producing organism, none of them directly allows
the harvesting of large amounts of the desired compounds (in
contrast to cultivation of the relevant microorganism, sponge
culture, or heterologous expression of biosynthesis genes). The
chief benefit of such studies is therefore to improve our un-
derstanding of metabolite production within the sponge-mi-
crobe association, identifying the putative producer(s) and thus
providing the basis for targeted cultivation and/or molecular
Other Biotechnologically Relevant Aspects of Sponges
In addition to the by now well-known pharmacological po-
tential of marine sponges and their associated microbiota, sev-
eral other aspects are also worth highlighting. For example, the
farming of certain sponges, particularly those in the genera
Spongia and Hippospongia, for their use as bath sponges has
been going on for more than a century. The industry, based
mainly in the Caribbean and Mediterranean, has been hit
numerous times by mass outbreaks of sponge disease (see
“Harming the host: pathogenesis, parasitism, and fouling”). A
better understanding of sponge microbiology in general, and
disease ecology in particular, should contribute to the future
success of these endeavors. The enormous filtering capacity of
sponges has led to the suggestion that they be farmed in a
bioremediation context, e.g., to reduce the high bacterial loads
resulting from sewage discharges and marine fish farms (110,
233, 379). Such an approach could be optimized financially by
farming sponges that, in addition to their role as a biofilter, are
useful as bath sponges or produce bioactive metabolites which
can subsequently be harvested (233).
336 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
Another aspect relates to the remarkable structural proper-
ties of the silica skeletons of demosponges and hexactinellid
(glass) sponges. A series of recent studies examined in detail
the siliceous spicules of Euplectella aspergillum, a deep-sea
hexactinellid from the western Pacific (5, 6, 385). Individual
spicules exhibit, in addition to their role in providing structural
support, fiber-optic properties similar to those of fibers utilized
in the telecommunications industry (5, 385). Importantly from
a technical point of view, spicules are formed at ambient tem-
peratures, and the study of this process may allow some of the
inherent problems associated with the commercial (high-tem-
perature) manufacture of optical fibers to be overcome (385).
Further insights into how organisms synthesize such sophisti-
cated biological materials were gained from electron micros-
copy observations of spicule organization in the same sponge
(6). The spicules of E. aspergillum provide great structural
stability due to their intricate arrangement at several spatial
levels, ranging from nanometers to centimeters, perhaps in-
spiring the future development of new materials by humans
(6). As mentioned earlier, sponges (e.g., the hexactinellid
sponge Scolymastra joubini) (51) may derive at least some of
their silica from diatoms, while in turn, spicules may extend the
range of photosynthetic symbionts within sponge tissue via
their conduction of light (115). Ongoing work on sponge skel-
etogenesis (e.g., see references 245, 357, 418, and 490) should
provide fascinating insights into its biological and, potentially,
CONCLUSIONS AND FUTURE DIRECTIONS
As indicated at the beginning of this article, even the signif-
icant recent advances in our understanding of sponge-micro-
organism associations have not closed numerous gaping holes
in our knowledge of these systems. It is startling how little is
known about many fundamental aspects of sponge symbiont
biology, particularly in the areas of symbiont metabolism and
evolution. On the other hand, the ever-increasing research
interest in this topic (Fig. 1) promises a bright future for the
field. To close, we offer our thoughts on where some of this
research attention could best be directed.
Detailed studies of symbiont transmission and sponge-mi-
crobe coevolution will greatly facilitate our understanding of
the evolution of these systems. Improved host phylogenies,
from species to class levels, will be critical in this regard, and
recent efforts in this direction are highly encouraging. “Hunt-
ing” for apparently sponge-specific microorganisms in other,
nonsponge habitats (e.g., seawater and other filter-feeding in-
vertebrates) will also be important. However, merely proving
the presence of a sponge-specific microbe outside a sponge
host is not sufficient to disprove its sponge-specific nature. For
example, a sponge damaged by a predator or storm may dis-
integrate and spread its microbial inhabitants into the water
column. It is thus necessary to prove the activity of such mi-
croorganisms outside the sponge host, using methods which
link microbial identity with function (e.g., FISH-microautora-
diography or stable isotope probing). This will be no small feat
if the organism is extremely rare in the ocean, as will almost
certainly be the case.
The function and physiology of sponge-associated microbes
are increasingly important research topics, reflecting our cur-
rent paucity of knowledge about many of the microbial asso-
ciates of sponges. For many, or even most, symbionts in
sponges, all that is known so far is their 16S rRNA gene
sequence: their metabolism remains a black box. Despite var-
ious caveats, the 16S rRNA data assembled here, together with
analyses of so-called functional genes, will provide a solid
framework for the application of recently developed molecular
tools for ecophysiological analyses of uncultivated microbes (2,
445, 449). Moreover, future combinations of (hypothesis-gen-
erating) metagenomic data with such tools should be particu-
larly profitable, especially for enigmatic microbes such as the
“Poribacteria.” Another area warranting further attention is
that of the molecular and biochemical bases of sponge-microbe
interactions. Recent work on the innate immune responses of
sponges has already provided many important insights, but
much remains unknown about host-symbiont signaling and
Although many aspects of sponge-microbe associations are
interesting and important from a basic research perspective,
we acknowledge that it is largely biotechnological interest that
will sustain this field into the future. It is thus fitting to end
our review with this topic. The biotechnological potential of
sponge-microbe associations has been widely (and justifiably)
lauded, yet the transition from initial compound discovery to
large-scale commercial production remains difficult. Chemical
synthesis of sponge-derived compounds or their simpler deriv-
atives offers the most reliable option for sustainable, long-term
drug supply if the said compounds can be produced cost-effec-
tively. Emerging technologies such as metagenomics and high-
throughput microbial cultivation approaches offer exciting po-
tential for accessing those compounds which are produced by
microorganisms. Current problems for metagenomics due to
the complexity of microbial communities in sponges may be
overcome in the future by single-cell genomic approaches
(such as multiple displacement amplification) applied to spe-
cific cell types which have been separated by, e.g., FISH and
flow cytometry (e.g., see reference 183). The use of sponge
larvae as starting material for metagenomic studies of drug-
producing microbes has also been suggested, as these are most
likely to represent the true symbionts and, perhaps, may be
simpler communities (279). The demonstrated complexity of
vertically transmitted communities (reviewed in this article)
suggests, however, that even this will be challenging. Combi-
natorial biosynthesis, as exemplified by an elegant recent study
of Prochloron sp. symbionts of ascidians (82), provides fur-
ther potential for exploiting the chemical novelties present in
sponge-microbe associations. Looking to the future, it is clear
that even greater integration among microbiologists, chemists,
geneticists, zoologists, and aquaculture experts will be crucial
in order to wring the most (data) out of sponges and their
We gratefully acknowledge the constructive comments on various
sections of the manuscript provided by Rocky de Nys, Friederike
Hoffmann, Peter Schupp, Bob Thacker, and Torsten Thomas. The
ideas expressed in this article also benefitted from discussions and/or
collaborations with Rocky de Nys, Piers Ettinger-Epstein, Ute
Hentschel, Matthias Horn, Staffan Kjelleberg, Peter Schupp, Peter
Steinberg, and Steve Whalan. We thank Ute Hentschel for providing
samples of Plakortis sp. and Agelas dilatata and Megan Huggett and
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS337
Sharon Longford for collection of Antho chartacea. Ivica Letunic and
Peer Bork are thanked for their help with the phylogenetic tree dis-
played in Fig. 4, while we are indebted to Rocky de Nys for his
compilation of the chemical structures comprising Fig. 21 to 23. Jo ¨rg
Ott kindly facilitated access to some of the sponge photographs in
Fig. 3. We thank Kayley Usher, Russell Hill, and Nicole Webster for
their permission to reproduce published figures.
The research of M.W.T. was funded in part by a grant from the
German Federal Ministry for Education and Research (Biolog II) to
M.W. D.S. received a DOC-FORTE stipend from the Austrian Acad-
emy of Sciences. M.W. thanks the University of Vienna for financial
support in the framework of the faculty focus “Symbiosis” and the
university research focus “Symbiosis Research and Molecular Princi-
ples of Recognition.”
1. Adamczeski, M., A. R. Reed, and P. Crews. 1995. New and known dike-
topiperazines from the Caribbean sponge, Calyx cf. podatypa. J. Nat. Prod.
2. Adamczyk, J., M. Hesselsoe, N. Iversen, M. Horn, A. Lehner, P. H. Nielsen,
M. Schloter, P. Roslev, and M. Wagner. 2003. The isotope array, a new tool
that employs substrate-mediated labeling of rRNA for determination of
microbial community structure and function. Appl. Environ. Microbiol.
3. Ahn, Y. B., S. K. Rhee, D. E. Fennell, L. J. Kerkhof, U. Hentschel, and
M. M. Haggblom. 2003. Reductive dehalogenation of brominated phenolic
compounds by microorganisms associated with the marine sponge Aplysina
aerophoba. Appl. Environ. Microbiol. 69:4159–4166.
4. Aicher, T. D., K. R. Buszek, F. G. Fang, C. J. Forsyth, S. H. Jung, Y. Kishi,
M. C. Matelich, P. M. Scola, D. M. Spero, and S. K. Yoon. 1992. Total
synthesis of halichondrin B and norhalichondrin B. J. Am. Chem. Soc.
5. Aizenberg, J., V. C. Sundar, A. D. Yablon, J. C. Weaver, and G. Chen. 2004.
Biological glass fibers: correlation between optical and structural proper-
ties. Proc. Natl. Acad. Sci. USA 101:3358–3363.
6. Aizenberg, J., J. C. Weaver, M. S. Thanawala, V. C. Sundar, D. E. Morse,
and P. Fratzl. 2005. Skeleton of Euplectella sp.: structural hierarchy from
the nanoscale to the macroscale. Science 309:275–278.
7. Althoff, K., C. Schu ¨tt, R. Steffen, R. Batel, and W. E. G. Mu ¨ller. 1998.
Evidence for a symbiosis between bacteria of the genus Rhodobacter and
the marine sponge Halichondria panicea: harbor also for putatively toxic
bacteria? Mar. Biol. 130:529–536.
8. Alvarez, B., M. D. Crisp, F. Driver, J. N. A. Hooper, and R. W. M. van Soest.
2000. Phylogenetic relationships of the family Axinellidae (Porifera: Demo-
spongiae) using morphological and molecular data. Zool. Screen. 29:169–
9. Andrianasolo, E. H., H. Gross, D. Goeger, M. Musafija-Girt, K. McPhail,
R. M. Leal, S. L. Mooberry, and W. H. Gerwick. 2005. Isolation of swin-
holide A and related glycosylated derivatives from two field collections of
marine cyanobacteria. Org. Lett. 7:1375–1378.
10. Ang, K. K., M. J. Holmes, T. Higa, M. T. Hamann, and U. A. Kara. 2000.
In vivo antimalarial activity of the beta-carboline alkaloid manzamine A.
Antimicrob. Agents Chemother. 44:1645–1649.
11. Arillo, A., G. Bavestrello, B. Burlando, and M. Sara. 1993. Metabolic
integration between symbiotic cyanobacteria and sponges—a possible
mechanism. Mar. Biol. 117:159–162.
12. Ashen, J. B., and L. J. Goff. 2000. Molecular and ecological evidence for
species specificity and coevolution in a group of marine algal-bacterial
symbioses. Appl. Environ. Microbiol. 66:3024–3030.
13. Bandaranayake, W. M., J. E. Bemis, and D. J. Bourne. 1996. Ultraviolet
absorbing pigments from the marine sponge Dysidea herbacea: isolation and
structure of a new mycosporine. Comp. Biochem. Phys. C 115:281–286.
14. Barthel, D., and B. Wolfrath. 1989. Tissue sloughing in the sponge Hali-
chondria panicea: a fouling organism prevents being fouled. Oecologia
15. Bavestrello, G., A. Arillo, U. Benatti, C. Cerrano, R. Cattaneo-Vietti, L.
Cortesognoi, L. Gaggero, M. Giovine, M. Tonetti, and M. Sara. 1995.
Quartz dissolution by the sponge Chondrosia reniformis (Porifera, Demo-
spongiae). Nature 378:374–376.
16. Bavestrello, G., A. Arillo, B. Calcinai, R. Cattaneo-Vietti, C. Cerrano, E.
Gaino, A. Penna, and M. Sara. 2000. Parasitic diatoms inside Antarctic
sponges. Biol. Bull. 198:29–33.
17. Bavestrello, G., U. Benatti, B. Calcinai, R. Cattaneo-Vietti, C. Cerrano, A.
Favre, M. Giovine, S. Lanza, R. Pronzato, and M. Sara. 1998. Body polarity
and mineral selectivity in the demosponge Chondrosia reniformis. Biol. Bull.
18. Bavestrello, G., U. Benatti, R. Cattaneo-Vietti, C. Cerrano, and M. Giovine.
2003. Sponge cell reactivity to various forms of silica. Microsc. Res. Tech.
19. Becerro, M. A., N. I. Lopez, X. Turon, and M. J. Uriz. 1994. Antimicrobial
activity and surface bacterial film in marine sponges. J. Exp. Mar. Biol.
20. Becerro, M. A., R. W. Thacker, X. Turon, M. J. Uriz, and V. J. Paul. 2003.
Biogeography of sponge chemical ecology: comparisons of tropical and
temperate defenses. Oecologia 135:91–101.
21. Belarbi, E., M. R. Dominguez, M. C. C. Garcia, A. C. Gomez, F. G. Camacho,
and E. M. Grima. 2003. Cultivation of explants of the marine sponge
Crambe crambe in closed systems. Biomol. Eng. 20:333–337.
22. Belarbi, E. H., A. C. Gomez, Y. Chisti, F. G. Camacho, and E. M. Grima.
2003. Producing drugs from marine sponges. Biotechnol. Adv. 21:585–598.
23. Bell, A. H., P. R. Bergquist, and C. N. Battershill. 1999. Feeding biology of
Polymastia croceus. Mem. Qld. Mus. 44:51–56.
24. Bergmann, W., and R. J. Feeney. 1951. Contributions to the study of marine
products. 32. The nucleosides of sponges. J. Org. Chem. 16:981–987.
25. Bergmann, W., and R. J. Feeney. 1950. The isolation of a new thymine
pentoside from sponges. J. Am. Chem. Soc. 72:2809–2810.
26. Bergquist, P. R. 1978. Sponges. Hutchinson and Co. Ltd., London, United
27. Bewley, C. A., and D. J. Faulkner. 1998. Lithistid sponges: star performers
or hosts to the stars. Angew. Chem. Int. Ed. 37:2162–2178.
28. Bewley, C. A., N. D. Holland, and D. J. Faulkner. 1996. Two classes of
metabolites from Theonella swinhoei are localized in distinct populations of
bacterial symbionts. Experientia 52:716–722.
29. Bhadury, P., B. T. Mohammad, and P. C. Wright. 2006. The current status
of natural products from marine fungi and their potential as anti-infective
agents. J. Ind. Microbiol. Biotechnol. 33:325–337.
30. Bil, K., E. Titlyanov, T. Berner, I. Fomina, and L. Muscatine. 1999. Some
aspects of the physiology and biochemistry of Lubomirska baikalensis, a
sponge from Lake Baikal containing symbiotic algae. Symbiosis 26:179–191.
31. Blunt, J. W., B. R. Copp, M. H. Munro, P. T. Northcote, and M. R. Prinsep.
2005. Marine natural products. Nat. Prod. Rep. 22:15–61.
32. Blunt, J. W., B. R. Copp, M. H. Munro, P. T. Northcote, and M. R. Prinsep.
2006. Marine natural products. Nat. Prod. Rep. 23:26–78.
33. Bock, E., and M. Wagner. 3 October 2006, posting date. Oxidation of
inorganic nitrogen compounds as an energy source. In M. Dworkin, S.
Falkow, E. Rosenberg, K.-H. Schleifer, and E. Stackebrandt (ed.), The
prokaryotes, vol. 2. Ecophysiology and biochemistry. Springer Verlag, New
York, NY. http://www.springerlink.com/content/p17748465102002j.
34. Boettcher, K. J., B. J. Barber, and J. T. Singer. 2000. Additional evidence
that juvenile oyster disease is caused by a member of the Roseobacter group
and colonization of nonaffected animals by Stappia stellulata-like strains.
Appl. Environ. Microbiol. 66:3924–3930.
35. Boettcher, K. J., K. K. Geaghan, A. P. Maloy, and B. J. Barber. 2005.
Roseovarius crassostreae sp. nov., a member of the Roseobacter clade and the
apparent cause of juvenile oyster disease (JOD) in cultured Eastern oysters.
Int. J. Syst. Evol. Microbiol. 55:1531–1537.
36. Bo ¨hm, M., U. Hentschel, A. B. Friedrich, L. Fieseler, R. Steffen, V. Gamulin,
I. M. Mu ¨ller, and W. E. G. Mu ¨ller. 2001. Molecular response of the
sponge Suberites domuncula to bacterial infection. Mar. Biol. 139:1037–
37. Borchiellini, C., C. Chombard, B. Lafay, and N. Boury-Esnault. 2000.
Molecular systematics of sponges (Porifera). Hydrobiologia 420:15–27.
38. Borchiellini, C., M. Manuel, E. Alivon, N. Boury-Esnault, J. Vacelet, and Y.
Le Parco. 2001. Sponge paraphyly and the origin of metazoa. J. Evol. Biol.
39. Borowitzka, M. A., R. Hinde, and F. Pironet. 1988. Carbon fixation by the
sponge Dysidea herbacea and its endosymbiont Oscillatoria spongeliae. Proc.
6th Int. Coral Reef Symp. 3:151–156.
40. Boury-Esnault, N. 2006. Systematics and evolution of Demospongiae. Can.
J. Zool. 84:205–224.
41. Brakhage, A. A., Q. Al-Abdallah, A. Tuncher, and P. Sprote. 2005. Evolu-
tion of beta-lactam biosynthesis genes and recruitment of trans-acting fac-
tors. Phytochemistry 66:1200–1210.
42. Bringmann, G., G. Lang, T. A. M. Gulder, H. Tsuruta, J. Muhlbacher, K.
Maksimenka, S. Steffens, K. Schaumann, R. Stohr, J. Wiese, J. F. Imhoff,
S. Perovic-Ottstadt, O. Boreiko, and W. E. G. Mu ¨ller. 2005. The first
sorbicillinoid alkaloids, the antileukemic sorbicillactones A and B, from a
sponge-derived Penicillium chrysogenum strain. Tetrahedron 61:7252–7265.
43. Brunton, F. R., and O. A. Dixon. 1994. Siliceous sponge-microbe biotic
associations and their recurrence through the Phanerozoic as reef mound
constructors. Palaios 9:370–387.
44. Bugni, T. S., and C. M. Ireland. 2004. Marine-derived fungi: a chemically
and biologically diverse group of microorganisms. Nat. Prod. Rep. 21:143–
45. Bultel-Ponce, V., J.-P. Berge, C. Debitus, J.-L. Nicolas, and M. Guyot. 1999.
Metabolites from the sponge-associated bacterium Pseudomonas species.
Mar. Biotechnol. 1:384–390.
46. Burja, A. M., and R. T. Hill. 2001. Microbial symbionts of the Australian
Great Barrier Reef sponge, Candidaspongia flabellata. Hydrobiologia 461:
47. Burja, A. M., N. Webster, P. Murphy, and R. T. Hill. 1999. Microbial
symbionts of Great Barrier Reef sponges. Mem. Qld. Mus. 44:63–75.
338TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
48. Butzke, D., and J. Piel. 2006. Genomic and metagenomic strategies to
identify biosynthetic gene clusters in uncultivated symbionts of marine
invertebrates, p. 327–355. In P. Proksch and W. E. G. Mueller (ed.), Fron-
tiers in marine biotechnology. Horizon Bioscience, Norfolk, VA.
49. Carlos, A. A., B. K. Baillie, M. Kawachi, and T. Maruyama. 1999. Phylo-
genetic position of Symbiodinium isolates from tridacnids (Bivalvia), cardi-
ids (Bivalvia), a sponge (Porifera), a soft coral (Anthozoa) and a free-living
strain. J. Phycol. 35:1054–1062.
50. Reference deleted.
51. Cerrano, C., A. Arillo, G. Bavestrello, B. Calcinai, R. Cattaneo-Vietti, A.
Penna, M. Sara, and C. Totti. 2000. Diatom invasion in the Antarctic
hexactinellid sponge Scolymastra joubini. Polar Biol. 23:441–444.
52. Cerrano, C., G. Bavestrello, C. N. Bianchi, R. Cattaneo-Vietti, S. Bava, C.
Morganti, C. Morri, P. Picco, G. Sara, S. Schiaparelli, A. Siccardi, and F.
Sponga. 2000. A catastrophic mass-mortality episode of gorgonians and
other organisms in the Ligurian Sea (north-western Mediterranean), sum-
mer 1999. Ecol. Lett. 3:284–293.
53. Cerrano, C., B. Calcinai, E. Cucchiari, C. Di Camillo, C. Totti, and G.
Bavestrello. 2004. The diversity of relationships between Antarctic sponges
and diatoms: the case of Mycale acerata Kirkpatrick, 1907 (Porifera, Demo-
spongiae). Polar Biol. 27:231–237.
54. Cervino, J. M., K. Winiarski-Cervino, S. W. Polson, T. Goreau, and G. W.
Smith. 2006. Identification of bacteria associated with a disease affecting
the marine sponge Ianthella basta in New Britain, Papua New Guinea. Mar.
Ecol. Prog. Ser. 324:139–150.
55. Chanas, B., J. R. Pawlik, T. Lindel, and W. Fenical. 1997. Chemical defense
of the Caribbean sponge Agelas clathrodes (Schmidt). J. Exp. Mar. Biol.
56. Chelossi, E., M. Milanese, A. Milano, R. Pronzato, and G. Riccardi. 2004.
Characterisation and antimicrobial activity of epibiotic bacteria from Petro-
sia ficiformis (Porifera, Demospongiae). J. Exp. Mar. Biol. Ecol. 309:21–33.
57. Cheshire, A. C., A. J. Butler, G. Westphalen, B. C. Rowland, J. Stevenson,
and C. R. Wilkinson. 1995. Preliminary study of the distribution and phys-
iology of the temperate phototrophic sponge Cymbastela sp. from South
Australia. Mar. Freshw. Res. 46:1211–1216.
58. Cheshire, A. C., and C. R. Wilkinson. 1991. Modelling the photosynthetic
production by sponges on Davies Reef, Great Barrier Reef. Mar. Biol.
59. Cheshire, A. C., C. R. Wilkinson, S. Seddon, and G. Westphalen. 1997.
Bathymetric and seasonal changes in photosynthesis and respiration of the
phototrophic sponge Phyllospongia lamellosa in comparison with respiration
by the heterotrophic sponge Ianthella basta on Davies Reef, Great Barrier
Reef. Mar. Freshw. Res. 48:589–599.
60. Choi, H. W., D. Demeke, F. A. Kang, Y. Kishi, K. Nakajima, P. Nowak, Z. K.
Wan, and C. Y. Xie. 2003. Synthetic studies on the marine natural product
halichondrins. Pure Appl. Chem. 75:1–17.
61. Cole, J. R., B. Chai, R. J. Farris, Q. Wang, S. A. Kulam, D. M. McGarrell,
G. M. Garrity, and J. M. Tiedje. 2005. The Ribosomal Database Project
(RDP-II): sequences and tools for high-throughput rRNA analysis. Nucleic
Acids Res. 33:294–296.
62. Colwell, R. R., and J. Liston. 1962. The natural bacterial flora of certain
marine invertebrates. J. Insect Pathol. 4:23–33.
63. Corredor, J. E., C. R. Wilkinson, V. P. Vicente, J. M. Morell, and E. Otero.
1988. Nitrate release by Caribbean reef sponges. Limnol. Oceanogr. 33:
64. Cowart, J. D., T. P. Henkel, S. E. McMurray, and J. R. Pawlik. 2006.
Sponge orange band (SOB): a pathogenic-like condition of the giant barrel
sponge, Xestospongia muta. Coral Reefs 25:513.
65. Cox, G., and A. W. D. Larkum. 1983. A diatom apparently living in symbi-
osis with a sponge. Mar. Sci. B 33:943–945.
66. Custodio, M. R., I. Prokic, R. Steffen, C. Koziol, R. Borojevic, F. Bru ¨mmer,
M. Nickel, and W. E. G. Mu ¨ller. 1998. Primmorphs generated from disso-
ciated cells of the sponge Suberites domuncula: a model system for studies
of cell proliferation and cell death. Mech. Ageing Dev. 105:45–59.
67. Dale, C., and N. A. Moran. 2006. Molecular interactions between bacterial
symbionts and their hosts. Cell 126:453–465.
68. Daniel, R. 2004. The soil metagenome—a rich resource for the discovery of
novel natural products. Curr. Opin. Biotechnol. 15:199–204.
69. Davidson, S. K., S. W. Allen, G. E. Lim, C. M. Anderson, and M. G.
Haygood. 2001. Evidence for the biosynthesis of bryostatins by the bacterial
symbiont “Candidatus Endobugula sertula” of the bryozoan Bugula neritina.
Appl. Environ. Microbiol. 67:4531–4537.
70. Davidson, S. K., and M. G. Haygood. 1999. Identification of sibling species
of the bryozoan Bugula neritina that produce different anticancer bryost-
atins and harbor distinct strains of the bacterial symbiont “Candidatus
Endobugula sertula.” Biol. Bull. 196:273–280.
71. Davis, A. R., N. M. Targett, O. J. McConnel, and C. M. Young. 1989.
Epibiosis of marine algae and benthic invertebrates: natural products chem-
istry and other mechanisms inhibiting settlement and overgrowth. Bioorg.
Mar. Chem. 3:85–114.
72. Davy, S. K., D. A. Trautman, M. A. Borowitzka, and R. Hinde. 2002.
Ammonium excretion by a symbiotic sponge supplies the nitrogen require-
ments of its rhodophyte partner. J. Exp. Biol. 205:3505–3511.
73. Dayton, P. K. 1974. Biological accommodation in the benthic community at
McMurdo Sound, Antarctica. Ecol. Monogr. 44:105–128.
74. Dayton, P. K. 1989. Interdecadal variation in an Antarctic sponge and its
predators from oceanographic climate shifts. Science 245:1484–1486.
75. de Caralt, S., G. Agell, and M. J. Uriz. 2003. Long-term culture of sponge
explants: conditions enhancing survival and growth, and assessment of
bioactivity. Biomol. Eng. 20:339–347.
76. DeLong, E. F. 2005. Microbial community genomics in the ocean. Nat. Rev.
77. Dembitsky, V. M., T. Rezanka, and M. Srebnik. 2003. Lipid compounds of
freshwater sponges: family Spongillidae, class Demospongiae. Chem. Phys.
78. Diaz, M. C. 1997. Molecular detection and characterization of specific
bacterial groups associated with tropical sponges. Proc. 8th Int. Coral Reef
79. Diaz, M. C., D. Akob, and S. C. Cary. 2004. Denaturing gradient gel
electrophoresis of nitrifying microbes associated with tropical sponges. Boll.
Mus. Ist. Biol. Univ. Genova 68:279–289.
80. Diaz, M. C., and B. B. Ward. 1997. Sponge-mediated nitrification in tropical
benthic communities. Mar. Ecol. Prog. Ser. 156:97–107.
81. Dieckmann, R., I. Graeber, I. Kaesler, U. Szewzyk, and H. von Dohren.
2005. Rapid screening and dereplication of bacterial isolates from marine
sponges of the Sula Ridge by intact-cell-MALDI-TOF mass spectrometry
(ICM-MS). Appl. Microbiol. Biotechnol. 67:539–548.
82. Donia, M. S., B. J. Hathaway, S. Sudek, M. G. Haygood, M. J. Rosovitz, J.
Ravel, and E. W. Schmidt. 2006. Natural combinatorial peptide libraries in
cyanobacterial symbionts of marine ascidians. Nat. Chem. Biol. 2:729–735.
83. Dubilier, N., C. Mulders, T. Ferdelman, D. de Beer, A. Pernthaler, M.
Klein, M. Wagner, C. Erseus, F. Thiermann, J. Krieger, O. Giere, and R.
Amann. 2001. Endosymbiotic sulphate-reducing and sulphide-oxidizing
bacteria in an oligochaete worm. Nature 411:298–302.
84. Duckworth, A., and C. Battershill. 2003. Sponge aquaculture for the pro-
duction of biologically active metabolites: the influence of farming proto-
cols and environment. Aquaculture 221:311–329.
85. Duckworth, A. R., and C. N. Battershill. 2003. Developing farming struc-
tures for production of biologically active sponge metabolites. Aquaculture
86. Duckworth, A. R., C. N. Battershill, and P. R. Bergquist. 1997. Influence of
explant procedures and environmental factors on culture success of three
sponges. Aquaculture 156:251–267.
87. Duckworth, A. R., C. N. Battershill, and D. R. Schiel. 2004. Effects of depth
and water flow on growth, survival and bioactivity of two temperate sponges
cultured in different seasons. Aquaculture 242:237–250.
88. Duckworth, A. R., C. N. Battershill, D. R. Schiel, and P. R. Bergquist. 1999.
Farming sponges for the production of bioactive metabolites. Mem. Qld.
89. Duckworth, A. R., W. M. Bru ¨ck, K. E. Janda, T. P. Pitts, and P. J.
McCarthy. 2006. Retention efficiencies of the coral reef sponges Aplysina
lacunosa, Callyspongia vaginalis and Niphates digitalis determined by Coulter
counter and plate culture analysis. Mar. Biol. Res. 2:243–248.
90. Duckworth, A. R., G. A. Samples, A. E. Wright, and S. A. Pomponi. 2003. In
vitro culture of the tropical sponge Axinella corrugata (Demospongiae):
effect of food cell concentration on growth, clearance rate, and biosynthesis
of stevensine. Mar. Biotechnol. 5:519–527.
91. Dunlap, M., and J. R. Pawlik. 1998. Spongivory by parrotfish in Florida
mangrove and reef habitats. Mar. Ecol. 19:325–337.
92. Dunlap, W. C., M. Jaspars, D. Hranueli, C. N. Battershill, N. Peric-Concha,
J. Zucko, S. H. Wright, and P. F. Long. 2006. New methods for medicinal
chemistry: universal gene cloning and expression systems for production of
marine bioactive metabolites. Curr. Med. Chem. 13:697–710.
93. El Sayed, K. A., M. Kelly, U. A. Kara, K. K. Ang, I. Katsuyama, D. C.
Dunbar, A. A. Khan, and M. T. Hamann. 2001. New manzamine alkaloids
with potent activity against infectious diseases. J. Am. Chem. Soc. 123:
94. Engel, S., and J. R. Pawlik. 2000. Allelopathic activities of sponge extracts.
Mar. Ecol. Prog. Ser. 207:273–281.
95. Enticknap, J. J., M. Kelly, O. Peraud, and R. T. Hill. 2006. Characterization
of a culturable alphaproteobacterial symbiont common to many marine
sponges and evidence for vertical transmission via sponge larvae. Appl.
Environ. Microbiol. 72:3724–3732.
96. Enticknap, J. J., R. Thompson, O. Peraud, J. E. Lohr, M. T. Hamann, and
R. T. Hill. 2004. Molecular analysis of a Florida Keys sponge: implications
for natural products discovery. Mar. Biotechnol. 6:S288–S293.
97. Ereskovsky, A. V., E. Gonobobleva, and A. Vishnyakov. 2005. Morpholog-
ical evidence for vertical transmission of symbiotic bacteria in the viviparous
sponge Halisarca dujardini Johnston (Porifera, Demospongiae, Halisar-
cida). Mar. Biol. 146:869–875.
98. Erpenbeck, D., J. A. J. Breeuwer, H. C. van der Velde, and R. W. M. van
Soest. 2002. Unravelling host and symbiont phylogenies of halichondrid
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS 339
sponges (Demospongiae, Porifera) using a mitochondrial marker. Mar.
99. Faulkner, D. J., M. K. Harper, C. E. Salomon, and E. W. Schmidt. 1999.
Localisation of bioactive metabolites in marine sponges. Mem. Qld. Mus.
100. Fieseler, L., M. Horn, M. Wagner, and U. Hentschel. 2004. Discovery of the
novel candidate phylum “Poribacteria” in marine sponges. Appl. Environ.
101. Fieseler, L., A. Quaiser, C. Schleper, and U. Hentschel. 2006. Analysis of
the first genome fragment from the marine sponge-associated, novel can-
didate phylum Poribacteria by environmental genomics. Environ. Microbiol.
102. Flatt, P. M., J. T. Gautschi, R. W. Thacker, M. Musafija-Girt, P. Crews,
and W. H. Gerwick. 2005. Identification of the cellular site of polychlori-
nated peptide biosynthesis in the marine sponge Dysidea (Lamellodysidea)
herbacea and symbiotic cyanobacterium Oscillatoria spongeliae by CARD-
FISH analysis. Mar. Biol. 147:761–774.
103. Flowers, A. E., M. J. Garson, R. I. Webb, E. J. Dumdei, and R. D. Charan.
1998. Cellular origin of chlorinated diketopiperazines in the dictyoceratid
sponge Dysidea herbacea (Keller). Cell Tissue Res. 292:597–607.
104. Fortman, J. L., and D. H. Sherman. 2005. Utilizing the power of microbial
genetics to bridge the gap between the promise and the application of
marine natural products. Chembiochem 6:960–978.
105. Friedrich, A. B., I. Fischer, P. Proksch, J. Hacker, and U. Hentschel. 2001.
Temporal variation of the microbial community associated with the Med-
iterranean sponge Aplysina aerophoba. FEMS Microbiol. Ecol. 38:105–113.
106. Friedrich, A. B., H. Merkert, T. Fendert, J. Hacker, P. Proksch, and U.
Hentschel. 1999. Microbial diversity in the marine sponge Aplysina caver-
nicola (formerly Verongia cavernicola) analyzed by fluorescence in situ hy-
bridization (FISH). Mar. Biol. 134:461–470.
107. Frohlich, H., and D. Barthel. 1997. Silica uptake of the marine sponge
Halichondria panicea in Kiel Bight. Mar. Biol. 128:115–125.
108. Frost, T. M., L. E. Graham, J. E. Elias, M. J. Haase, D. W. Kretchmer, and
J. A. Kranzfelder. 1997. A yellow-green algal symbiont in the freshwater
sponge, Corvomeyenia everetti: convergent evolution of symbiotic associa-
tions. Freshw. Biol. 38:395–399.
109. Frost, T. M., and C. E. Williamson. 1980. In situ determination of the effect
of symbiotic algae on the growth of the freshwater sponge Spongilla
lacustris. Ecology 61:1361–1370.
110. Fu, W. T., L. M. Sun, X. C. Zhang, and W. Zhang. 2006. Potential of the
marine sponge Hymeniacidon perleve as a bioremediator of pathogenic bacteria
in integrated aquaculture ecosystems. Biotechnol. Bioeng. 93:1112–1122.
111. Fuqua, C., M. R. Parsek, and E. P. Greenberg. 2001. Regulation of gene
expression by cell-to-cell communication: acyl-homoserine lactone quorum
sensing. Annu. Rev. Genet. 35:439–468.
112. Fusetani, N. 2004. Biofouling and antifouling. Nat. Prod. Rep. 21:94–104.
113. Gaino, E., G. Bavestrello, R. Cattaneo-Vietti, and M. Sara. 1994. Scanning
electron microscope evidence for diatom uptake by two Antarctic sponges.
Polar Biol. 14:55–58.
114. Gaino, E., and M. Rebora. 2003. Ability of mobile cells of the freshwater
sponge Ephydatia fluviatilis (Porifera, Demospongiae) to digest diatoms.
Ital. J. Zool. 70:17–22.
115. Gaino, E., and M. Sara. 1994. Siliceous spicules of Tethya seychellensis
(Porifera) support the growth of a green alga: a possible light conducting
system. Mar. Ecol. Prog. Ser. 108:147–151.
116. Gaino, E., and M. Sara. 1994. An ultrastructural comparative study of the
eggs of 2 species of Tethya (Porifera, Demospongiae). Invertebr. Reprod.
117. Gaino, E., M. Sciscioli, E. Lepore, M. Rebora, and G. Corriero. 2006.
Association of the sponge Tethya orphei (Porifera, Demospongiae) with
filamentous cyanobacteria. Invertebr. Biol. 125:281–287.
118. Gallissian, M. F., and J. Vacelet. 1976. Ultrastructure de quelques stades de
l’ovogenese des spongiaires du genre Verongia (Dictyoceratida). Ann. Sci.
Nat. Zool. Biol. Anim. 18:381–404.
119. Galtsoff, P. S., H. H. Brown, C. L. Smith, and F. G. W. Smith. 1939. Sponge
mortality in the Bahamas. Nature 143:807–808.
120. Garson, M. J., A. E. Flowers, R. I. Webb, R. D. Charan, and E. J. McCaffrey.
1998. A sponge/dinoflagellate association in the haplosclerid sponge
Haliclona sp.: cellular origin of cytotoxic alkaloids by Percoll density gra-
dient centrifugation. Cell Tissue Res. 293:365–373.
121. Garson, M. J., M. P. Zimmermann, C. N. Battershill, J. L. Holden, and
P. T. Murphy. 1994. The distribution of brominated long-chain fatty acids
in sponge and symbiont cell types from the tropical marine sponge Am-
phimedon terpenensis. Lipids 29:509–516.
122. Gentry, T. J., G. S. Wickham, C. W. Schadt, Z. He, and J. Zhou. 2006.
Microarray applications in microbial ecology research. Microb. Ecol. 52:
123. Gernert, C., F. O. Glockner, G. Krohne, and U. Hentschel. 2005. Microbial
diversity of the freshwater sponge Spongilla lacustris. Microb. Ecol. 50:206–
124. Gili, J.-M., and R. Coma. 1998. Benthic suspension feeders: their para-
mount role in littoral marine food webs. Trends Ecol. Evol. 13:316–321.
125. Gillor, O., S. Carmeli, Y. Rahamim, Z. Fishelson, and M. Ilan. 2000.
Immunolocalization of the toxin latrunculin B within the Red Sea sponge
Negombata magnifica (Demospongiae, Latrunculiidae). Mar. Biotechnol.
126. Givskov, M., R. de Nys, M. Manefield, L. Gram, R. Maximilien, L. Eberl,
S. Molin, P. D. Steinberg, and S. Kjelleberg. 1996. Eukaryotic interference
with homoserine lactone-mediated prokaryotic signalling. J. Bacteriol. 178:
127. Gram, L., R. de Nys, R. Maximilien, M. Givskov, P. Steinberg, and S.
Kjelleberg. 1996. Inhibitory effects of secondary metabolites from the red
alga Delisea pulchra on swarming motility of Proteus mirabilis. Appl. Envi-
ron. Microbiol. 62:4284–4287.
128. Grant, A. J., M. Remond, and R. Hinde. 1998. Low molecular-weight factor
from Plesiastrea versipora (Scleractinia) that modifies release and glycerol
metabolism of isolated symbiotic algae. Mar. Biol. 130:553–557.
129. Grant, A. J., D. A. Trautman, I. Menz, and R. Hinde. 2006. Separation of
two cell signalling molecules from a symbiotic sponge that modify algal
carbon metabolism. Biochem. Biophys. Res. Commun. 348:92–98.
130. Gunasekera, A. S., K. S. Sfanos, D. K. Harmody, S. A. Pomponi, P. J.
McCarthy, and J. V. Lopez. 2005. HBMMD: an enhanced database of the
microorganisms associated with deeper water marine invertebrates. Appl.
Microbiol. Biotechnol. 66:373–376.
131. Hadas, E., D. Marie, M. Shpigel, and M. Ilan. 2006. Virus predation by
sponges is a new nutrient-flow pathway in coral reef food webs. Limnol.
132. Hadas, E., M. Shpigel, and M. Ilan. 2005. Sea ranching of the marine
sponge Negombata magnifica (Demospongiae, Latrunculiidae) as a first
step for latrunculin B mass production. Aquaculture 244:159–169.
133. Halanych, K. M. 2004. The new view of animal phylogeny. Annu. Rev. Ecol.
Evol. Syst. 35:229–256.
134. Hallam, S. J., K. T. Konstantinidis, N. Putnam, C. Schleper, Y. Watanabe,
J. Sugahara, C. Preston, J. de la Torre, P. M. Richardson, and E. F.
DeLong. 2006. Genomic analysis of the uncultivated marine crenarchaeote
Cenarchaeum symbiosum. Proc. Natl. Acad. Sci. USA 103:18296–18301.
135. Hallam, S. J., T. J. Mincer, C. Schleper, C. M. Preston, K. Roberts, P. M.
Richardson, and E. F. DeLong. 2006. Pathways of carbon assimilation and
ammonia oxidation suggested by environmental genomic analyses of ma-
rine Crenarchaeota. PLoS Biol. 4:e95.
136. Handelsman, J. 2004. Metagenomics: application of genomics to uncul-
tured microorganisms. Microbiol. Mol. Biol. Rev. 68:669–685.
137. Handley, S., M. Page, and P. Northcote. 2006. Anti-cancer sponge: the race
is on for aquaculture supply. Water Atmos. 14:14–15.
138. Hart, J. B., R. E. Lill, S. J. H. Hickford, J. W. Blunt, and M. H. G. Munro.
2000. The halichondrins: chemistry, biology, supply and delivery, p. 134–
153. In N. Fusetani (ed.), Drugs from the sea. Karger, Basel, Switzerland.
139. Harvell, C. D., K. Kim, J. M. Burkholder, R. R. Colwell, P. R. Epstein, D. J.
Grimes, E. E. Hofmann, E. K. Lipp, A. D. M. E. Osterhaus, R. M. Over-
street, J. W. Porter, G. W. Smith, and G. R. Vasta. 1999. Emerging marine
diseases: climate links and anthropogenic factors. Science 285:1505–1510.
140. Harvell, C. D., C. E. Mitchell, J. R. Ward, S. Altizer, A. P. Dobson, R. S.
Ostfeld, and M. D. Samuel. 2002. Climate warming and disease risks for
terrestrial and marine biota. Science 296:2158–2162.
141. Hausmann, R., M. P. Vitello, F. Leitermann, and C. Syldatk. 2006. Ad-
vances in the production of sponge biomass: Aplysina aerophoba—a model
sponge for ex situ sponge biomass production. J. Biotechnol. 124:117–127.
142. Haygood, M. G., and S. K. Davidson. 1997. Small-subunit rRNA genes and
in situ hybridization with oligonucleotides specific for the bacterial symbi-
onts in the larvae of the bryozoan Bugula neritina and proposal of “Candi-
datus Endobugula sertula”. Appl. Environ. Microbiol. 63:4612–4616.
143. Haygood, M. G., E. W. Schmidt, S. K. Davidson, and D. J. Faulkner. 1999.
Microbial symbionts of marine invertebrates: opportunities for microbial
biotechnology. J. Mol. Microbiol. Biotechnol. 1:33–43.
144. Hedges, S. B., J. E. Blair, M. L. Venturi, and J. L. Shoe. 2004. A molecular
timescale of eukaryote evolution and the rise of complex multicellular life.
BMC Evol. Biol. 4:2.
145. Hentschel, U., L. Fieseler, M. Wehrl, C. Gernert, M. Steinert, J. Hacker,
and M. Horn. 2003. Microbial diversity of marine sponges. Prog. Mol.
Subcell. Biol. 37:59–88.
146. Hentschel, U., J. Hopke, M. Horn, A. B. Friedrich, M. Wagner, J. Hacker,
and B. S. Moore. 2002. Molecular evidence for a uniform microbial com-
munity in sponges from different oceans. Appl. Environ. Microbiol. 68:
147. Hentschel, U., M. Schmid, M. Wagner, L. Fieseler, C. Gernert, and J.
Hacker. 2001. Isolation and phylogenetic analysis of bacteria with antimi-
crobial activities from the Mediterranean sponges Aplysina aerophoba and
Aplysina cavernicola. FEMS Microbiol. Ecol. 35:305–312.
148. Hentschel, U., K. M. Usher, and M. W. Taylor. 2006. Marine sponges as
microbial fermenters. FEMS Microbiol. Ecol. 55:167–177.
149. Hildebrand, M., L. E. Waggoner, G. E. Lim, K. H. Sharp, C. P. Ridley, and
M. G. Haygood. 2004. Approaches to identify, clone, and express symbiont
bioactive metabolite genes. Nat. Prod. Rep. 21:122–142.
150. Hildebrand, M., L. E. Waggoner, H. Liu, S. Sudek, S. Allen, C. Anderson,
340 TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
D. H. Sherman, and M. Haygood. 2004. bryA: an unusual modular
polyketide synthase gene from the uncultivated bacterial symbiont of the
marine bryozoan Bugula neritina. Chem. Biol. 11:1543–1552.
151. Hill, M., A. Hill, N. Lopez, and O. Harriott. 2006. Sponge-specific bacterial
symbionts in the Caribbean sponge, Chondrilla nucula (Demospongiae,
Chondrosida). Mar. Biol. 148:1221–1230.
152. Hill, M., and T. Wilcox. 1998. Unusual mode of symbiont repopulation after
bleaching in Anthosigmella varians: acquisition of different zooxanthellae
strains. Symbiosis 25:279–289.
153. Hill, M. S. 1996. Symbiotic zooxanthellae enhance boring and growth rates
of the tropical sponge Anthosigmella varians forma varians. Mar. Biol.
154. Hill, R. T. 2004. Microbes from marine sponges: a treasure trove of biodi-
versity for natural products discovery, p. 177–190. In A. T. Bull (ed.),
Microbial diversity and bioprospecting. ASM Press, Washington, DC.
155. Hill, R. T., O. Peraud, M. T. Hamann, and N. Kasanah. November 2005.
Manzamine-producing actinomycetes. U.S. patent 20050244938.
156. Hinde, R. 1988. Symbiotic nutrition and nutrient limitation. Proc. 6th Int.
Coral Reef Symp. 1:199–204.
157. Hinde, R., F. Pironet, and M. A. Borowitzka. 1994. Isolation of Oscillatoria
spongeliae, the filamentous cyanobacterial symbiont of the marine sponge
Dysidea herbacea. Mar. Biol. 119:99–104.
158. Hirata, Y., and D. Uemura. 1986. Halichondrins—antitumor polyether
macrolides from a marine sponge. Pure Appl. Chem. 58:701–710.
159. Hoffmann, F., O. Larsen, V. Thiel, H. T. Rapp, T. Pape, W. Michaelis, and
J. Reitner. 2005. An anaerobic world in sponges. Geomicrobiol. J. 22:1–10.
160. Hoffmann, F., H. T. Rapp, and J. Reitner. 2006. Monitoring microbial
community composition by fluorescence in situ hybridization during culti-
vation of the marine cold-water sponge Geodia barretti. Mar. Biotechnol.
161. Hoffmann, F., H. T. Rapp, T. Zoller, and J. Reitner. 2003. Growth and
regeneration in cultivated fragments of the boreal deep water sponge Geo-
dia barretti Bowerbank, 1858 (Geodiidae, Tetractinellida, Demospongiae).
J. Biotechnol. 100:109–118.
162. Holden, M. T., S. Ram Chhabra, R. de Nys, P. Stead, N. J. Bainton, P. J.
Hill, M. Manefield, N. Kumar, M. Labatte, D. England, S. Rice, M. Givskov,
G. P. Salmond, G. S. Stewart, B. W. Bycroft, S. Kjelleberg, and P. Williams.
1999. Quorum-sensing cross talk: isolation and chemical characterization of
cyclic dipeptides from Pseudomonas aeruginosa and other gram-negative
bacteria. Mol. Microbiol. 33:1254–1266.
163. Ho ¨ller, U., A. D. Wright, G. F. Matthee, G. M. Ko ¨nig, S. Draeger, H.-J.
Aust, and B. Schulz. 2000. Fungi from marine sponges: diversity, biological
activity and secondary metabolites. Mycol. Res. 104:1354–1365.
164. Holmes, B., and H. Blanch. 2006. Genus-specific associations of marine
sponges with group I crenarchaeotes. Mar. Biol. 150:759–772.
165. Hood, K. A., L. M. West, P. T. Northcote, M. V. Berridge, and J. H. Miller.
2001. Induction of apoptosis by the marine sponge (Mycale) metabolites,
mycalamide A and pateamine. Apoptosis 6:207–219.
166. Hood, K. A., L. M. West, B. Rouwe, P. T. Northcote, M. V. Berridge, S. J.
Wakefield, and J. H. Miller. 2002. Peloruside A, a novel antimitotic agent
with paclitaxel-like microtubule-stabilizing activity. Cancer Res. 62:3356–
167. Hooper, J. N. A., and R. W. M. van Soest. 2002. Systema Porifera: a guide
to the classification of sponges. Kluwer Academic/Plenum Publishers, New
168. Horn, M., and M. Wagner. 2004. Bacterial endosymbionts of free-living
amoebae. J. Eukaryot. Microbiol. 51:509–515.
169. Hrvatin, S., and J. Piel. 2007. Rapid isolation of rare clones from highly
complex DNA libraries by PCR analysis of liquid gel pools. J. Microbiol.
170. Hugenholtz, P., B. M. Goebel, and N. R. Pace. 1998. Impact of culture-
independent studies on the emerging phylogenetic view of bacterial diver-
sity. J. Bacteriol. 180:4765–4774.
171. Hummel, H., A. B. J. Sepers, L. de Wolf, and F. W. Melissen. 1988. Bacterial
growth on the marine sponge Halichondria panicea induced by reduced
waterflow rate. Mar. Ecol. Prog. Ser. 42:195–198.
172. Ilan, M., and A. Abelson. 1995. The life of a sponge in a sandy lagoon. Biol.
173. Imhoff, J. F., and H. G. Tru ¨per. 1976. Marine sponges as habitats of
anaerobic phototrophic bacteria. Microb. Ecol. 3:1–9.
174. Jadulco, R., P. Proksch, V. Wray, Sudarsono, A. Berg, and U. Grafe. 2001.
New macrolides and furan carboxylic acid derivative from the sponge-
derived fungus Cladosporium herbarum. J. Nat. Prod. 64:527–530.
175. Jaeckle, W. B. 1995. Transport and metabolism of alanine and palmitic acid
by field-collected larvae of Tedania ignis (Porifera, Demospongiae): esti-
mated consequences of limited label translocation. Biol. Bull. 189:159–167.
176. Jensen, P. R., and W. Fenical. 2000. Marine microorganisms and drug
discovery: current status and future potential, p. 6–19. In N. Fusetani (ed.),
Drugs from the sea. Karger, Basel, Switzerland.
177. Jin, M., and R. E. Taylor. 2005. Total synthesis of (?)-peloruside A. Org.
178. Johnson, Z. I., E. R. Zinser, A. Coe, N. P. McNulty, E. M. Woodward, and
S. W. Chisholm. 2006. Niche partitioning among Prochlorococcus ecotypes
along ocean-scale environmental gradients. Science 311:1737–1740.
179. Kahlert, M., and D. Neumann. 1997. Early development of freshwater
sponges under the influence of nitrite and pH. Arch. Hydrobiol. 139:69–81.
180. Karner, M. B., E. F. DeLong, and D. M. Karl. 2001. Archaeal dominance in
the mesopelagic zone of the Pacific Ocean. Nature 409:507–510.
181. Kaye, H. R. 1991. Sexual reproduction in four Caribbean commercial
sponges. II. Oogenesis and transfer of bacterial symbionts. Invertebr. Re-
prod. Dev. 19:13–24.
182. Kazlauskas, R., P. T. Murphy, and R. J. Wells. 1978. A diketopiperazine
derived from trichloroleucine from the sponge Dysidea herbacea. Tetrahe-
dron Lett. 49:4945–4948.
183. Keller, M., M. Walcher, K. Zengler, C. A. Abulencia, H. C. Chang, J. A.
Garcia, T. Holland, C. R. Kuske, and F. J. Brockman. 2006. On the way to
single cell microbiology, p. A160. Abstr. 11th Int. Symp. Microb. Ecol.,
Vienna, Austria, 20 to 25 August 2006.
184. Kelly, S. R., E. Garo, P. R. Jensen, W. Fenical, and J. R. Pawlik. 2005.
Effects of Caribbean sponge secondary metabolites on bacterial surface
colonization. Aquat. Microb. Ecol. 40:191–203.
185. Kelman, D., Y. Kashman, E. Rosenberg, M. Ilan, I. Ifrach, and Y. Loya.
2001. Antimicrobial activity of the reef sponge Amphimedon viridis from the
Red Sea: evidence for selective toxicity. Aquat. Microb. Ecol. 24:9–16.
186. Keyzers, R. A., and M. T. Davies-Coleman. 2005. Anti-inflammatory me-
tabolites from marine sponges. Chem. Soc. Rev. 34:355–365.
187. Kim, T. K., and J. A. Fuerst. 2006. Diversity of polyketide synthase genes
from bacteria associated with the marine sponge Pseudoceratina clavata:
culture-dependent and culture-independent approaches. Environ. Micro-
188. Kim, T. K., M. J. Garson, and J. A. Fuerst. 2005. Marine actinomycetes
related to the “Salinospora” group from the Great Barrier Reef sponge
Pseudoceratina clavata. Environ. Microbiol. 7:509–518.
189. Kim, T. K., A. K. Hewavitharana, P. N. Shaw, and J. A. Fuerst. 2006.
Discovery of a new source of rifamycin antibiotics in marine sponge acti-
nobacteria by phylogenetic prediction. Appl. Environ. Microbiol. 72:2118–
190. Klautau, M., C. A. M. Russo, C. Lazoski, N. Boury-Esnault, J. P. Thorpe,
and A. M. Sole-Cava. 1999. Does cosmopolitan result from overconserva-
tive systematics? A case study using the marine sponge Chondrilla nucula.
191. Ko ¨nig, G. M., S. Kehraus, S. F. Seibert, A. Abdel-Lateff, and D. Muller.
2006. Natural products from marine organisms and their associated mi-
crobes. Chembiochem 7:229–238.
192. Konneke, M., A. E. Bernhard, J. R. de la Torre, C. B. Walker, J. B.
Waterbury, and D. A. Stahl. 2005. Isolation of an autotrophic ammonia-
oxidizing marine archaeon. Nature 437:543–546.
193. Koops, H.-P., U. Purkhold, A. Pommerening-Ro ¨ser, G. Timmermann, and
M. Wagner. March 2003, posting date. The lithoautotrophic ammonia-
oxidizing bacteria. In M. Dworkin, S. Falkow, E. Rosenberg, K.-H. Schle-
ifer, and E. Stackebrandt (ed.), The prokaryotes: an evolving electronic
resource for the microbiological community, 3rd ed., release 3.13. Springer-
Verlag, New York, NY.
194. Koops, H. P., and A. Pommerening-Ro ¨ser. 2001. Distribution and ecophysi-
ology of the nitrifying bacteria emphasizing cultured species. FEMS Micro-
biol. Ecol. 37:1–9.
195. Koropatnick, T. A., J. T. Engle, M. A. Apicella, E. V. Stabb, W. E. Goldman,
and M. J. McFall-Ngai. 2004. Microbial factor-mediated development in a
host-bacterial mutualism. Science 306:1186–1188.
196. Kowalke, J. 2000. Ecology and energetics of two Antarctic sponges. J. Exp.
Mar. Biol. Ecol. 247:85–97.
197. Kumar, P., C. Khosla, and Y. Tang. 2004. Manipulation and analysis of
polyketide synthases. Methods Enzymol. 388:269–293.
198. Lafi, F. F., M. J. Garson, and J. A. Fuerst. 2005. Culturable bacterial
symbionts isolated from two distinct sponge species (Pseudoceratina clavata
and Rhabdastrella globostellata) from the Great Barrier Reef display similar
phylogenetic diversity. Microb. Ecol. 50:213–220.
199. Lauckner, G. 1980. Diseases of Porifera, p. 139–165. In O. Kinne (ed.),
Diseases of marine animals, vol. 1. John Wiley and Sons, Chichester,
200. Lee, E. Y., H. K. Lee, Y. K. Lee, C. J. Sim, and J. H. Lee. 2003. Diversity of
symbiotic archaeal communities in marine sponges from Korea. Biomol.
201. Lee, O. O., S. C. Lau, and P. Y. Qian. 2006. Consistent bacterial community
structure associated with the surface of the sponge Mycale adhaerens Bower-
bank. Microb. Ecol. 52:693–707.
202. Lee, O. O., S. C. Lau, M. M. Tsoi, X. Li, I. Plakhotnikova, S. Dobretsov,
M. C. Wu, P. K. Wong, and P. Y. Qian. 2006. Gillisia myxillae sp. nov., a
novel member of the family Flavobacteriaceae, isolated from the marine
sponge Myxilla incrustans. Int. J. Syst. Evol. Microbiol. 56:1795–1799.
203. Lee, O. O., and P. Y. Qian. 2004. Potential control of bacterial epibiosis on
the surface of the sponge Mycale adhaerens. Aquat. Microb. Ecol. 34:11–21.
204. Leininger, S., T. Urich, M. Schloter, L. Schwark, J. Qi, G. W. Nicol, J. I.
VOL. 71, 2007SPONGE-ASSOCIATED MICROORGANISMS341
Prosser, S. C. Schuster, and C. Schleper. 2006. Archaea predominate
among ammonia-oxidizing prokaryotes in soils. Nature 442:806–809.
205. Leon, Y. M., and K. A. Bjorndal. 2002. Selective feeding in the hawksbill
turtle, an important predator in coral reef ecosystems. Mar. Ecol. Prog. Ser.
206. Li, C. W., J. Y. Chen, and T. E. Hua. 1998. Precambrian sponges with
cellular structures. Science 279:879–882.
207. Li, X., and L. Qin. 2005. Metagenomics-based drug discovery and marine
microbial diversity. Trends Biotechnol. 23:539–543.
208. Li, Z.-Y., L.-M. He, J. Wu, and Q. Jiang. 2006. Bacterial community diver-
sity associated with four marine sponges from the South China Sea based
on 16S rDNA-DGGE fingerprinting. J. Exp. Mar. Biol. Ecol. 329:75–85.
209. Li, Z.-Y., and Y. Liu. 2006. Marine sponge Craniella austrialiensis-associated
bacterial diversity revelation based on 16S rDNA library and biologically
active actinomycetes screening, phylogenetic analysis. Lett. Appl. Micro-
210. Lim, G. E., and M. G. Haygood. 2004. “Candidatus Endobugula glebosa,” a
specific bacterial symbiont of the marine bryozoan Bugula simplex. Appl.
Environ. Microbiol. 70:4921–4929.
211. Lohr, J. E., F. Chen, and R. T. Hill. 2005. Genomic analysis of bacterio-
phage PhiJL001: insights into its interaction with a sponge-associated al-
pha-proteobacterium. Appl. Environ. Microbiol. 71:1598–1609.
212. Reference deleted.
213. Lopez, J., A. Ledger, C. Peterson, K. Sfanos, D. Harmody, S. Pomponi, and
P. McCarthy. 2006. Molecular census and comparison of cultured and
uncultured microbial symbiont diversity from an ancient metazoan host,
phylum Porifera, abstr. 6.15. Abstr. 5th Int. Symbiosis Soc. Congr., Vienna,
Austria, 4 to 10 August 2006.
214. Lopez, J. V., P. J. McCarthy, K. E. Janda, R. Willoughby, and S. A.
Pomponi. 1999. Molecular techniques reveal wide phyletic diversity of het-
erotrophic microbes associated with Discodermia spp. (Porifera: Demo-
spongiae). Mem. Qld. Mus. 44:329–341.
215. Loy, A., and L. Bodrossy. 2006. Highly parallel microbial diagnostics using
oligonucleotide microarrays. Clin. Chim. Acta 363:106–119.
216. Ludwig, W., O. Strunk, S. Klugbauer, N. Klugbauer, M. Weizenegger,
J. Neumaier, M. Bachleitner, and K. H. Schleifer. 1998. Bacterial phylog-
eny based on comparative sequence analysis. Electrophoresis 19:554–568.
217. Ludwig, W., O. Strunk, R. Westram, L. Richter, H. Meier, Yadhukumar, A.
Buchner, T. Lai, S. Steppi, G. Jobb, W. Forster, I. Brettske, S. Gerber, A. W.
Ginhart, O. Gross, S. Grumann, S. Hermann, R. Jost, A. Konig, T. Liss,
R. Lussmann, M. May, B. Nonhoff, B. Reichel, R. Strehlow, A. Stamatakis,
N. Stuckmann, A. Vilbig, M. Lenke, T. Ludwig, A. Bode, and K. H. Schle-
ifer. 2004. ARB: a software environment for sequence data. Nucleic Acids
218. Madri, P. P., M. Hermel, and G. Claus. 1971. The microbial flora of the
sponge Microciona prolifera Verrill and its ecological implications. Bot.
219. Maldonado, M. 2006. The ecology of the sponge larva. Can. J. Zool. 84:
220. Maldonado, M., C. Carmona, Z. Velasquez, A. Puig, A. Cruzado, A. Lopez,
and C. M. Young. 2005. Siliceous sponges as a silicon sink: an overlooked
aspect of benthopelagic coupling in the marine silicon cycle. Limnol.
221. Maldonado, M., N. Cortadellas, M. I. Trillas, and K. Rutzler. 2005. Endo-
symbiotic yeast maternally transmitted in a marine sponge. Biol. Bull.
222. Maldonado, M., and C. M. Young. 1998. Limits on the bathymetric distri-
bution of keratose sponges: a field test in deep water. Mar. Ecol. Prog. Ser.
223. Manefield, M., R. de Nys, N. Kumar, R. Read, M. Givskov, P. Steinberg,
and S. Kjelleberg. 1999. Evidence that halogenated furanones from Delisea
pulchra inhibit acylated homoserine lactone (AHL)-mediated gene expres-
sion by displacing the AHL signal from its receptor protein. Microbiology
224. Manefield, M., T. B. Rasmussen, M. Henzter, J. B. Andersen, P. Steinberg,
S. Kjelleberg, and M. Givskov. 2002. Halogenated furanones inhibit quo-
rum sensing through accelerated LuxR turnover. Microbiology 148:1119–
225. Manz, W., G. Arp, G. Schumann-Kindel, U. Szewzyk, and J. Reitner. 2000.
Widefield deconvolution epifluorescence microscopy combined with fluo-
rescence in situ hybridization reveals the spatial arrangement of bacteria in
sponge tissue. J. Microbiol. Methods 40:125–134.
226. Margot, H., C. Acebal, E. Toril, R. Amils, and J. L. Fernandez Puentes.
2002. Consistent association of crenarchaeal archaea with sponges of the
genus Axinella. Mar. Biol. 140:739–745.
227. Margulies, M., M. Egholm, W. E. Altman, S. Attiya, J. S. Bader, L. A.
Bemben, J. Berka, M. S. Braverman, Y. J. Chen, Z. Chen, S. B. Dewell, L.
Du, J. M. Fierro, X. V. Gomes, B. C. Godwin, W. He, S. Helgesen, C. H. Ho,
G. P. Irzyk, S. C. Jando, M. L. Alenquer, T. P. Jarvie, K. B. Jirage, J. B.
Kim, J. R. Knight, J. R. Lanza, J. H. Leamon, S. M. Lefkowitz, M. Lei, J.
Li, K. L. Lohman, H. Lu, V. B. Makhijani, K. E. McDade, M. P. McKenna,
E. W. Myers, E. Nickerson, J. R. Nobile, R. Plant, B. P. Puc, M. T. Ronan,
G. T. Roth, G. J. Sarkis, J. F. Simons, J. W. Simpson, M. Srinivasan, K. R.
Tartaro, A. Tomasz, K. A. Vogt, G. A. Volkmer, S. H. Wang, Y. Wang, M. P.
Weiner, P. Yu, R. F. Begley, and J. M. Rothberg. 2005. Genome sequencing
in microfabricated high-density picolitre reactors. Nature 437:376–380.
228. Martin, A. P. 2002. Phylogenetic approaches for describing and comparing
the diversity of microbial communities. Appl. Environ. Microbiol. 68:3673–
229. Matsunaga, S., and N. Fusetani. 2003. Nonribosomal peptides from marine
sponges. Curr. Org. Chem. 7:945–966.
230. McCann, J., E. V. Stabb, D. S. Millikan, and E. G. Ruby. 2003. Population
dynamics of Vibrio fischeri during infection of Euprymna scolopes. Appl.
Environ. Microbiol. 69:5928–5934.
231. Mendola, D. 2003. Aquaculture of three phyla of marine invertebrates to
yield bioactive metabolites: process developments and economics. Biomol.
232. Mews, L. K. 1980. The green hydra symbiosis. III. The biotrophic transport
of carbohydrate from alga to animal. Proc. R. Soc. Lond. B 209:377–401.
233. Milanese, M., E. Chelossi, R. Manconi, A. Sara, M. Sidri, and R. Pronzato.
2003. The marine sponge Chondrilla nucula Schmidt, 1862 as an elective
candidate for bioremediation in integrated aquaculture. Biomol. Eng. 20:
234. Reference deleted.
235. Montalvo, N. F., N. M. Mohamed, J. J. Enticknap, and R. T. Hill. 2005.
Novel actinobacteria from marine sponges. Antonie Leeuwenhoek 87:29–
236. Moore, B. S. 2006. Biosynthesis of marine natural products: macroorgan-
isms, part B. Nat. Prod. Rep. 23:615–629.
237. Moran, N. A., and H. E. Dunbar. 2006. Sexual acquisition of beneficial
symbionts in aphids. Proc. Natl. Acad. Sci. USA 103:12803–12806.
238. Morris, C. E., M. Bardin, O. Berge, P. Frey-Klett, N. Fromin, H. Girardin,
M. H. Guinebretiere, P. Lebaron, J. M. Thiery, and M. Troussellier. 2002.
Microbial biodiversity: approaches to experimental design and hypothesis
testing in primary scientific literature from 1975 to 1999. Microbiol. Mol.
Biol. Rev. 66:592–616.
239. Moss, C., D. H. Green, B. Perez, A. Velasco, R. Henriquez, and J. D.
McKenzie. 2003. Intracellular bacteria associated with the ascidian Ectein-
ascidia turbinata: phylogenetic and in situ hybridisation analysis. Mar. Biol.
240. Mu ¨ller, W. E., V. A. Grebenjuk, G. Le Pennec, H. Schroder, F. Brummer,
U. Hentschel, I. M. Mu ¨ller, and H. Breter. 2004. Sustainable production of
bioactive compounds by sponges—cell culture and gene cluster approach: a
review. Mar. Biotechnol. 6:105–117.
241. Mu ¨ller, W. E., V. A. Grebenjuk, N. L. Thakur, A. N. Thakur, R. Batel, A.
Krasko, I. M. Muller, and H. J. Breter. 2004. Oxygen-controlled bacterial
growth in the sponge Suberites domuncula: toward a molecular understand-
ing of the symbiotic relationships between sponge and bacteria. Appl.
Environ. Microbiol. 70:2332–2341.
242. Mu ¨ller, W. E. G., M. Bo ¨hm, R. Batel, S. De Rosa, G. Tommonaro, I. M.
Mu ¨ller, and H. C. Schro ¨der. 2000. Application of cell culture for the
production of bioactive compounds from sponges: synthesis of avarol by
primmorphs from Dysidea avara. J. Nat. Prod. 63:1077–1081.
243. Mu ¨ller, W. E. G., M. Klemt, N. L. Thakur, H. C. Schro ¨der, A. Aiello, M.
D’Esposito, M. Menna, and E. Fattorusso. 2004. Molecular/chemical ecol-
ogy in sponges: evidence for an adaptive antibacterial response in Suberites
domuncula. Mar. Biol. 144:19–29.
244. Mu ¨ller, W. E. G., and I. M. Mu ¨ller. 2003. Origin of the metazoan immune
system: identification of the molecules and their functions in sponges.
Integr. Comp. Biol. 43:281–292.
245. Mu ¨ller, W. E. G., K. Wendt, C. Geppert, M. Wiens, A. Reiber, and H. C.
Schroder. 2006. Novel photoreception system in sponges? Unique trans-
mission properties of the stalk spicules from the hexactinellid Hyalonema
sieboldi. Biosens. Bioelectron. 21:1149–1155.
246. Mu ¨ller, W. E. G., M. Wiens, T. Adell, V. Gamulin, H. C. Schroder, and I. M.
Mu ¨ller. 2004. Bauplan of urmetazoa: basis for genetic complexity of meta-
zoa. Int. Rev. Cytol. 235:53–92.
247. Mu ¨ller, W. E. G., M. Wiens, R. Batel, R. Steffen, H. C. Schroder, R.
Borojevic, and M. R. Custodio. 1999. Establishment of a primary cell cul-
ture from a sponge: primmorphs from Suberites domuncula. Mar. Ecol.
Prog. Ser. 178:205–219.
248. Mu ¨ller, W. E. G., W. Wimmer, W. Schatton, M. Bo ¨hm, R. Batel, and Z.
Filic. 1999. Initiation of an aquaculture of sponges for the sustainable
production of bioactive metabolites in open systems: example, Geodia
cydonium. Mar. Biotechnol. 1:569–579.
249. Mu ¨ller, W. E. G., R. K. Zahn, B. Kurelec, C. Lucu, I. Mu ¨ller, and G.
Uhlenbruck. 1981. Lectin, a possible basis for symbiosis between bacteria
and sponges. J. Bacteriol. 145:548–558.
250. Munro, M. H. G., J. W. Blunt, E. J. Dumdei, S. J. H. Hickford, R. E. Lill,
S. Li, C. N. Battershill, and A. R. Duckworth. 1999. The discovery and
development of marine compounds with pharmaceutical potential. J. Bio-
251. Nagelkerken, I. 2000. Barrel sponge bows out. Reef Encounter 28:14–15.
252. Nakabachi, A., A. Yamashita, H. Toh, H. Ishikawa, H. E. Dunbar, N. A.
342TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.
Moran, and M. Hattori. 2006. The 160-kilobase genome of the bacterial
endosymbiont Carsonella. Science 314:267.
253. Nakashima, K., L. Yamada, Y. Satou, J. Azuma, and N. Satoh. 2004. The
evolutionary origin of animal cellulose synthase. Dev. Genes Evol. 214:
254. Newbold, R. W., P. R. Jensen, W. Fenical, and J. R. Pawlik. 1999. Antimi-
crobial activity of Caribbean sponge extracts. Aquat. Microb. Ecol. 19:279–
255. Newman, D. J., and G. M. Cragg. 2004. Marine natural products and
related compounds in clinical and advanced preclinical trials. J. Nat. Prod.
256. Northcote, P. T., J. W. Blunt, and M. H. G. Munro. 1991. Pateamine: a
potent cytotoxin from the New Zealand marine sponge, Mycale sp. Tetra-
hedron Lett. 32:6411–6414.
257. Nussbaumer, A. D., C. R. Fisher, and M. Bright. 2006. Horizontal endo-
symbiont transmission in hydrothermal vent tubeworms. Nature 441:345–
258. Nyholm, S. V., and M. J. McFall-Ngai. 2004. The winnowing: establishing
the squid-vibrio symbiosis. Nat. Rev. Microbiol. 2:632–642.
259. Ochman, H., S. Elwyn, and N. A. Moran. 1999. Calibrating bacterial evo-
lution. Proc. Natl. Acad. Sci. USA 96:12638–12643.
260. Oclarit, J. M., H. Okada, S. Ohta, K. Kaminura, Y. Yamaoka, T. Iizuka, S.
Miyashiro, and S. Ikegami. 1994. Anti-bacillus substance in the marine
sponge, Hyatella species, produced by an associated Vibrio species bacte-
rium. Microbios 78:7–16.
261. Oloriz, F., M. Reolid, and F. J. Rodriguez-Tovar. 2003. A late Jurassic
carbonate ramp colonized by sponges and benthic microbial communities
(external Prebetic, southern Spain). Palaios 18:528–545.
262. Olson, J. B., D. J. Gochfeld, and M. Slattery. 2006. Aplysina red band
syndrome: a new threat to Caribbean sponges. Dis. Aquat. Organ. 71:163–
263. Olson, J. B., D. K. Harmody, and P. J. McCarthy. 2002. Alpha-proteobac-
teria cultivated from marine sponges display branching rod morphology.
FEMS Microbiol. Lett. 211:169–173.
264. Olson, J. B., C. C. Lord, and P. J. McCarthy. 2000. Improved recoverability
of microbial colonies from marine sponge samples. Microb. Ecol. 40:139–
265. Olson, J. B., and P. J. McCarthy. 2005. Associated bacterial communities of
two deep-water sponges. Aquat. Microb. Ecol. 39:47–55.
266. Oren, M., L. Steindler, and M. Ilan. 2005. Transmission, plasticity and the
molecular identification of cyanobacterial symbionts in the Red Sea sponge
Diacarnus erythraenus. Mar. Biol. 148:35–41.
267. Osinga, R. 2003. Biotechnological aspects of marine sponges. J. Biotechnol.
268. Osinga, R., H. el Belarbi, E. M. Grima, J. Tramper, and R. H. Wijffels. 2003.
Progress towards a controlled culture of the marine sponge Pseudosuberites
andrewsi in a bioreactor. J. Biotechnol. 100:141–146.
269. Osinga, R., P. B. de Beukelaer, E. M. Meijer, J. Tramper, and R. H.
Wijffels. 1999. Growth of the sponge Pseudosuberites (aff.) andrewsi in a
closed system. J. Biotechnol. 70:155–161.
270. Page, M., L. West, P. Northcote, C. Battershill, and M. Kelly. 2005. Spatial
and temporal variability of cytotoxic metabolites in populations of the New
Zealand sponge Mycale hentscheli. J. Chem. Ecol. 31:1161–1174.
271. Page, M. J., P. T. Northcote, V. L. Webb, S. Mackey, and S. J. Handley.
2005. Aquaculture trials for the production of biologically active metabo-
lites in the New Zealand sponge Mycale hentscheli (Demospongiae: Poecilo-
sclerida). Aquaculture 250:256–269.
272. Pape, T., F. Hoffmann, N.-V. Queric, K. von Juterzenka, J. Reitner, and W.
Michaelis. 2006. Dense populations of archaea associated with the demo-
sponge Tentorium semisuberites Schmidt, 1870 from Arctic deep-waters.
Polar Biol. 29:662–667.
273. Partensky, F., W. R. Hess, and D. Vaulot. 1999. Prochlorococcus, a marine
photosynthetic prokaryote of global significance. Microbiol. Mol. Biol. Rev.
274. Pawlik, J. R. 1998. Coral reef sponges: do predatory fishes affect their
distribution? Limnol. Oceanogr. 43:1396–1399.
275. Pawlik, J. R., B. Chanas, R. J. Toonen, and W. Fenical. 1995. Defenses of
Caribbean sponges against predatory reef fish. I. Chemical deterrency. Mar.
Ecol. Prog. Ser. 127:183–194.
276. Pedros-Alio, C. 2006. Marine microbial diversity: can it be determined?
Trends Microbiol. 14:257–263.
277. Perovic-Ottstadt, S., T. Adell, P. Proksch, M. Wiens, M. Korzhev, V. Gamulin,
I. M. Mu ¨ller, and W. E. G. Mu ¨ller. 2004. A (133)-beta-D-glucan recognition
protein from the sponge Suberites domuncula. Mediated activation of fi-
brinogen-like protein and epidermal growth factor gene expression. Eur.
J. Biochem. 271:1924–1937.
278. Perry, N. B., J. W. Blunt, M. H. G. Munro, and L. K. Pannell. 1988.
Mycalamide A, an antiviral compound from a New Zealand sponge of
genus Mycale. J. Am. Chem. Soc. 110:4850–4851.
279. Piel, J. 2006. Bacterial symbionts: prospects for the sustainable production
of invertebrate-derived pharmaceuticals. Curr. Med. Chem. 13:39–50.
280. Piel, J. 2004. Metabolites from symbiotic bacteria. Nat. Prod. Rep. 21:519–
281. Piel, J. 2002. A polyketide synthase-peptide synthetase gene cluster from an
uncultured bacterial symbiont of Paederus beetles. Proc. Natl. Acad. Sci.
282. Piel, J., D. Butzke, N. Fusetani, D. Hui, M. Platzer, G. Wen, and S.
Matsunaga. 2005. Exploring the chemistry of uncultivated bacterial symbi-
onts: antitumor polyketides of the pederin family. J. Nat. Prod. 68:472–479.
283. Piel, J., I. Hofer, and D. Hui. 2004. Evidence for a symbiosis island involved
in horizontal acquisition of pederin biosynthetic capabilities by the bacterial
symbiont of Paederus fuscipes beetles. J. Bacteriol. 186:1280–1286.
284. Piel, J., D. Hui, N. Fusetani, and S. Matsunaga. 2004. Targeting modular
polyketide synthases with iteratively acting acyltransferases from metage-
nomes of uncultured bacterial consortia. Environ. Microbiol. 6:921–927.
285. Piel, J., D. Hui, G. Wen, D. Butzke, M. Platzer, N. Fusetani, and S.
Matsunaga. 2004. Antitumor polyketide biosynthesis by an uncultivated
bacterial symbiont of the marine sponge Theonella swinhoei. Proc. Natl.
Acad. Sci. USA 101:16222–16227.
286. Piel, J., G. Wen, M. Platzer, and D. Hui. 2004. Unprecedented diversity of
catalytic domains in the first four modules of the putative pederin
polyketide synthase. Chembiochem 5:93–98.
287. Piggott, A. M., and P. Karuso. 2004. Quality, not quantity: the role of
natural products and chemical proteomics in modern drug discovery.
Comb. Chem. High Throughput Scr. 7:607–630.
288. Pile, A. J., A. Grant, R. Hinde, and M. A. Borowitzka. 2003. Heterotrophy
on ultraplankton communities is an important source of nitrogen for a
sponge-rhodophyte symbiosis. J. Exp. Biol. 206:4533–4538.
289. Pile, A. J., M. R. Patterson, M. Savarese, V. I. Chernykh, and V. A. Fialkov.
1997. Trophic effects of sponge feeding within Lake Baikal’s littoral zone. 2.
Sponge abundance, diet, feeding efficiency, and carbon flux. Limnol.
290. Pile, A. J., M. R. Patterson, and J. D. Witman. 1996. In situ grazing on
plankton ?10?m by the boreal sponge Mycale lingua. Mar. Ecol. Prog. Ser.
291. Pile, A. J., and C. M. Young. 2006. The natural diet of a hexactinellid
sponge: benthic-pelagic coupling in a deep-sea microbial food web. Deep-
Sea Res. 53:1148–1156.
292. Pimentel-Elardo, S., M. Wehrl, A. B. Friedrich, P. R. Jensen, and U.
Hentschel. 2003. Isolation of planctomycetes from Aplysina sponges. Aquat.
Microb. Ecol. 33:239–245.
293. Pomponi, S. A. 2006. Biology of the Porifera: cell culture. Can. J. Zool.
294. Preston, C. M., K. Y. Wu, T. F. Molinski, and E. F. DeLong. 1996. A
psychrophilic crenarchaeon inhabits a marine sponge: Cenarchaeum sym-
biosum gen. nov., sp. nov. Proc. Natl. Acad. Sci. USA 93:6241–6246.
295. Proksch, P., R. Ebel, R. A. Edrada, P. Schupp, W. H. Lin, Sudarsono, V.
Wray, and K. Steube. 2003. Detection of pharmacologically active natural
products using ecology: selected examples from Indopacific marine inver-
tebrates and sponge-derived fungi. Pure Appl. Chem. 75:343–352.
296. Proksch, P., R. Ebel, R. A. Edrada, V. Wray, and K. Steube. 2003. Bioactive
natural products from marine invertebrates and associated fungi. Prog.
Mol. Subcell. Biol. 37:117–142.
297. Proksch, P., R. A. Edrada, and R. Ebel. 2002. Drugs from the seas—current
status and microbiological implications. Appl. Microbiol. Biotechnol. 59:
298. Pronzato, R. 1999. Sponge-fishing, disease and farming in the Mediterra-
nean Sea. Aquat. Conserv. Mar. Freshw. Ecosyst. 9:485–493.
299. Pronzato, R., G. Bavestrello, C. Cerrano, G. Magnino, R. Manconi, J.
Pantelis, A. Sara, and M. Sidri. 1999. Sponge farming in the Mediterranean
Sea: new perspectives. Mem. Qld. Mus. 44:485–491.
300. Purkhold, U., A. Pommering-Ro ¨ser, S. Juretschko, M. C. Schmid, H.-P.
Koops, and M. Wagner. 2000. Phylogeny of all recognized species of am-
monia oxidizers based on comparative 16S rRNA and amoA sequence
analysis: implications for molecular diversity surveys. Appl. Environ. Mi-
301. Rao, K. V., M. S. Donia, J. Peng, E. Garcia-Palomero, D. Alonso, A.
Martinez, M. Medina, S. G. Franzblau, B. L. Tekwani, S. I. Khan, S.
Wahyuono, K. L. Willett, and M. T. Hamann. 2006. Manzamine B and E
and ircinal A related alkaloids from an Indonesian Acanthostrongylophora
sponge and their activity against infectious, tropical parasitic, and Alzhei-
mer’s diseases. J. Nat. Prod. 69:1034–1040.
302. Rasmussen, T. B., T. Bjarnsholt, M. E. Skindersoe, M. Hentzer, P. Kristoffersen,
M. Kote, J. Nielsen, L. Eberl, and M. Givskov. 2005. Screening for quorum-
sensing inhibitors (QSI) by use of a novel genetic system, the QSI selector. J.
303. Regoli, F., C. Cerrano, E. Chierici, S. Bompadre, and G. Bavestrello. 2000.
Susceptibility to oxidative stress of the Mediterranean demosponge Petrosia
ficiformis: role of endosymbionts and solar irradiance. Mar. Biol. 137:453–
304. Regoli, F., C. Cerrano, E. Chierici, M. C. Chiantore, and G. Bavestrello.
2004. Seasonal variability of prooxidant pressure and antioxidant adapta-
VOL. 71, 2007 SPONGE-ASSOCIATED MICROORGANISMS343
tion to symbiosis in the Mediterranean demosponge Petrosia ficiformis. Download full-text
Mar. Ecol. Prog. Ser. 275:129–137.
305. Regoli, F., M. Nigro, E. Chierici, C. Cerrano, S. Schiapparelli, C. Totti, and
G. Bavestrello. 2004. Variations of antioxidant efficiency and presence of
endosymbiotic diatoms in the Antarctic porifera Haliclona dancoi. Mar.
Environ. Res. 58:637–640.
306. Reiswig, H. M. 1975. The aquiferous systems of three marine Demo-
spongiae. J. Morphol. 145:493–502.
307. Reiswig, H. M. 1975. Bacteria as food for temperate-water marine sponges.
Can. J. Zool. 53:582–589.
308. Reiswig, H. M. 1971. Particle feeding in natural populations of three marine
demosponges. Biol. Bull. 141:568–591.
309. Reiswig, H. M. 1974. Water transport, respiration and energetics of three
tropical marine sponges. J. Exp. Mar. Biol. Ecol. 14:231–249.
310. Reitner, J., and G. Schumann-Kindel. 1997. Pyrite in mineralized sponge
tissue: product of sulfate reducing sponge related bacteria? Facies 36:272–
311. Rezanka, T., K. Sigler, and V. M. Dembitsky. 2006. Syriacin, a novel
unusual sulfated ceramide glycoside from the freshwater sponge Ephy-
datia syriaca (Porifera, Demospongiae, Spongillidae). Tetrahedron 62:
312. Ribes, M., R. Coma, M. J. Atkinson, and R. A. I. Kinzie. 2005. Sponges and
ascidians control removal of particulate organic nitrogen from coral reef
water. Limnol. Oceanogr. 50:1480–1489.
313. Ribes, M., R. Coma, and J.-M. Gili. 1999. Natural diet and grazing rate of
the temperate marine sponge Dysidea avara (Demospongiae, Dendrocer-
atida) throughout an annual cycle. Mar. Ecol. Prog. Ser. 176:179–190.
314. Richelle-Maurer, E., J.-C. Braekman, M. J. De Kluijver, R. Gomez, G. Van
de Vyver, R. W. M. van Soest, and C. Devijver. 2001. Cellular location of
(2R, 3R, 7Z)-2-aminotetradec-7-ene-1,3-diol, a potent antimicrobial metab-
olite produced by the Caribbean sponge Haliclona vansoesti. Cell Tissue
315. Richelle-Maurer, E., M. J. De Kluijver, S. Feio, S. Gaudencio, H. Gaspar,
R. Gomez, R. Tavares, G. van de Vyver, and R. W. M. van Soest. 2003.
Localization and ecological significance of oroidin and sceptrin in the Ca-
ribbean sponge Agelas conifera. Biochem. Syst. Ecol. 31:1073–1091.
316. Ridley, C. P., P. R. Bergquist, M. K. Harper, D. J. Faulkner, J. N. Hooper,
and M. G. Haygood. 2005. Speciation and biosynthetic variation in four
dictyoceratid sponges and their cyanobacterial symbiont, Oscillatoria spon-
geliae. Chem. Biol. 12:397–406.
317. Ridley, C. P., D. J. Faulkner, and M. G. Haygood. 2005. Investigation of
Oscillatoria spongeliae-dominated bacterial communities in four dictyocer-
atid sponges. Appl. Environ. Microbiol. 71:7366–7375.
318. Riesenfeld, C. S., P. D. Schloss, and J. Handelsman. 2004. Metagenomics:
genomic analysis of microbial communities. Annu. Rev. Genet. 38:525–552.
319. Rinkevich, B. 1999. Cell cultures from marine invertebrates: obstacles, new
approaches and recent improvements. J. Biotechnol. 70:133–153.
320. Rinkevich, B. 2005. Marine invertebrate cell cultures: new millenium
trends. Mar. Biotechnol. 7:429–439.
321. Roberts, D. E., S. P. Cummins, A. R. Davis, and C. Pangway. 1999. Evi-
dence for symbiotic algae in sponges from temperate coastal reefs in New
South Wales, Australia. Mem. Qld. Mus. 44:493–497.
322. Roberts, D. E., A. R. Davis, and S. P. Cummins. 2006. Experimental ma-
nipulation of shade, silt, nutrients and salinity on the temperate reef sponge
Cymbastela concentrica. Mar. Ecol. Prog. Ser. 307:143–154.
323. Rocap, G., F. W. Larimer, J. Lamerdin, S. Malfatti, P. Chain, N. A. Ahlgren,
A. Arellano, M. Coleman, L. Hauser, W. R. Hess, Z. I. Johnson, M. Land,
D. Lindell, A. F. Post, W. Regala, M. Shah, S. L. Shaw, C. Steglich, M. B.
Sullivan, C. S. Ting, A. Tolonen, E. A. Webb, E. R. Zinser, and S. W.
Chisholm. 2003. Genome divergence in two Prochlorococcus ecotypes re-
flects oceanic niche differentiation. Nature 424:1042–1047.
324. Rosell, D., and M. J. Uriz. 1992. Do associated zooxanthellae and the
nature of the substratum affect survival, attachment and growth of Cliona
viridis (Porifera: Hadromerida)? An experimental approach. Mar. Biol.
325. Rosenberg, E., and Y. Ben-Haim. 2002. Microbial diseases of corals and
global warming. Environ. Microbiol. 4:318–326.
326. Rosenberg, E., and L. Falkovitz. 2004. The Vibrio shiloi/Oculina patagonica
model system of coral bleaching. Annu. Rev. Microbiol. 58:143–159.
327. Rot, C., I. Goldfarb, M. Ilan, and D. Huchon. 2006. Putative cross-kingdom
horizontal gene transfer in sponge (Porifera) mitochondria. BMC Evol.
328. Ruppert, E. E., and R. D. Barnes. 1994. Invertebrate zoology, 6th ed.
Saunders College Publishing, Fort Worth, TX.
329. Russell, J. A., A. Latorre, B. Sabater-Munoz, A. Moya, and N. A. Moran.
2003. Side-stepping secondary symbionts: widespread horizontal transfer
across and beyond the Aphidoidea. Mol. Ecol. 12:1061–1075.
330. Ru ¨tzler, K. 1988. Mangrove sponge disease induced by cyanobacterial sym-
bionts: failure of a primitive immune system? Dis. Aquat. Org. 5:143–149.
331. Saller, U. 1989. Microscopical aspects on symbiosis of Spongilla lacustris
(Porifera, Spongillidae) and green algae. Zoomorphology 108:291–296.
332. Salomon, C. E., T. Deerinck, R. H. Ellisman, and D. J. Faulkner. 2001.
The cellular localization of dercitamide in the Palauan sponge Oceanapia
sagittaria. Mar. Biol. 139:313–319.
333. Sand-Jensen, K., and M. F. Pedersen. 1994. Photosynthesis by symbiotic
algae in the freshwater sponge, Spongilla lacustris. Limnol. Oceanogr. 39:
334. Santavy, D. L., and R. R. Colwell. 1990. Comparison of bacterial commu-
nities associated with the Caribbean sclerosponge Ceratoporella nicholsoni
and ambient seawater. Mar. Ecol. Prog. Ser. 67:73–82.
335. Santavy, D. L., P. Willenz, and R. R. Colwell. 1990. Phenotypic study of
bacteria associated with the Caribbean sclerosponge Ceratoporella nichol-
soni. Appl. Environ. Microbiol. 56:1750–1762.
336. Sara, M. 1971. Ultrastructural aspects of the symbiosis between two species
of the genus Aphanocapsa (Cyanophyceae) and Ircinia variabilis (Demo-
spongiae). Mar. Biol. 11:214–221.
337. Sara, M., G. Bavestrello, R. Cattaneo-Vietti, and C. Cerrano. 1998. Endo-
symbiosis in sponges: relevance for epigenesis and evolution. Symbiosis
338. Sara, M., and L. Liaci. Symbiotic associations between zooxanthellae and
Cliona. Nature 203:321–323.
339. Scalera-Liaci, L., M. Sciscioli, E. Lepore, and E. Gaino. 1999. Symbiotic
zooxanthellae in Cinachyra tarentina, a non-boring demosponge. Endocy-
tobiosis Cell Res. 13:105–114.
340. Scanlan, D. J., and N. J. West. 2002. Molecular ecology of the marine
cyanobacterial genera Prochlorococcus and Synechococcus. FEMS Micro-
biol. Ecol. 40:1–12.
341. Scheuermayer, M., T. A. Gulder, G. Bringmann, and U. Hentschel. 2006.
Rubritalea marina gen. nov., sp. nov., a marine representative of the phylum
“Verrucomicrobia,” isolated from a sponge (Porifera). Int. J. Syst. Evol.
342. Schirmer, A., R. Gadkari, C. D. Reeves, F. Ibrahim, E. F. DeLong, and C. R.
Hutchinson. 2005. Metagenomic analysis reveals diverse polyketide syn-
thase gene clusters in microorganisms associated with the marine sponge
Discodermia dissoluta. Appl. Environ. Microbiol. 71:4840–4849.
343. Schleper, C., E. F. DeLong, C. M. Preston, R. A. Feldman, K.-Y. Wu, and
R. V. Swanson. 1998. Genomic analysis reveals chromosomal variation in
natural populations of the uncultured psychrophilic archaeon Cenarchaeum
symbiosum. J. Bacteriol. 180:5003–5009.
344. Schleper, C., G. Jurgens, and M. Jonuscheit. 2005. Genomic studies of
uncultivated archaea. Nat. Rev. Microbiol. 3:479–488.
345. Schleper, C., R. V. Swanson, E. J. Mathur, and E. F. DeLong. 1997. Char-
acterization of a DNA polymerase from the uncultivated psychrophilic
archaeon Cenarchaeum symbiosum. J. Bacteriol. 179:7803–7811.
346. Schloss, P. D., and J. Handelsman. 2003. Biotechnological prospects from
metagenomics. Curr. Opin. Biotechnol. 14:303–310.
347. Schloss, P. D., and J. Handelsman. 2006. Introducing TreeClimber, a test
to compare microbial community structures. Appl. Environ. Microbiol.
348. Schloss, P. D., and J. Handelsman. 2004. Status of the microbial census.
Microbiol. Mol. Biol. Rev. 68:686–691.
349. Schmidt, E. W. 2005. From chemical structure to environmental biosynthetic
pathways: navigating marine invertebrate-bacteria associations. Trends Bio-
350. Schmidt, E. W., C. A. Bewley, and D. J. Faulkner. 1998. Theopalauamide,
a bicyclic glycopeptide from filamentous bacterial symbionts of the lithistid
sponge Theonella swinhoei from Palau and Mozambique. J. Org. Chem.
351. Schmidt, E. W., A. Y. Obraztsova, S. K. Davidson, D. J. Faulkner, and
M. G. Haygood. 2000. Identification of the antifungal peptide-containing
symbiont of the marine sponge Theonella swinhoei as a novel ?-proteobac-
terium, “Candidatus Entotheonella palauensis.” Mar. Biol. 136:969–977.
352. Schmitt, S., J. Weisz, N. Lindquist, and U. Hentschel. 2007. Vertical trans-
mission of a phylogenetically complex microbial consortium in the vivipa-
rous sponge Ircinia felix. Appl. Environ. Microbiol. 73:2067–2078.
353. Scho ¨nberg, C. H. L., D. de Beer, and A. Lawton. 2005. Oxygen microsensor
studies on zooxanthellate clionaid sponges from the Costa Brava, Mediter-
ranean Sea. J. Phycol. 41:774–779.
354. Scho ¨nberg, C. H. L., F. Hoffmann, and S. Gatti. 2004. Using microsensors
to measure sponge physiology. Boll. Mus. Ist. Biol. Univ. Genova 68:593–
355. Scho ¨nberg, C. H. L., and W. K. W. Loh. 2005. Molecular identity of the
unique symbiotic dinoflagellates found in the bioeroding demosponge
Cliona orientalis. Mar. Ecol. Prog. Ser. 299:157–166.
356. Schramm, A., D. de Beer, J. C. van den Heuvel, S. Ottengraf, and R.
Amann. 1999. Microscale distribution of populations and activities of Ni-
trosospira and Nitrospira spp. along a macroscale gradient in a nitrifying
bioreactor: quantification by in situ hybridization and the use of microsen-
sors. Appl. Environ. Microbiol. 65:3690–3696.
357. Schro ¨der, H. C., D. Brandt, U. Schlossmacher, X. Wang, M. N. Tahir, W.
Tremel, S. I. Belikov, and W. E. G. Mu ¨ller. 11 January 2007, posting date.
Enzymatic production of biosilica glass using enzymes from sponges: basic
aspects and application in nanobiotechnology (material sciences and med-
icine). Naturwissenschaften doi:10.1007/s00114-006-0192-0.
344TAYLOR ET AL.MICROBIOL. MOL. BIOL. REV.