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A centimeter-long bacterium with DNA contained in metabolically active, membrane-bound organelles



Cells of most bacterial species are around 2 micrometers in length, with some of the largest specimens reaching 750 micrometers. Using fluorescence, x-ray, and electron microscopy in conjunction with genome sequencing, we characterized Candidatus ( Ca. ) Thiomargarita magnifica, a bacterium that has an average cell length greater than 9000 micrometers and is visible to the naked eye. These cells grow orders of magnitude over theoretical limits for bacterial cell size, display unprecedented polyploidy of more than half a million copies of a very large genome, and undergo a dimorphic life cycle with asymmetric segregation of chromosomes into daughter cells. These features, along with compartmentalization of genomic material and ribosomes in translationally active organelles bound by bioenergetic membranes, indicate gain of complexity in the Thiomargarita lineage and challenge traditional concepts of bacterial cells.
A centimeter-long bacterium with DNA contained in
metabolically active, membrane-bound organelles
Jean-Marie Volland
*, Silvina Gonzalez-Rizzo
Natalia Ivanova
, Frederik Schulz
, Danielle Goudeau
, Nathalie H. Elisabeth
, Nandita Nath
Daniel Udwary
, Chantal Guidi-Rontani
, Susanne Bolte-Kluge
Karen M. Davies
, Maïtena R. Jean
, Jean-Louis Mansot
, Esther R. Angert
Tanja Woyke
*, Shailesh V. Date
Cells of most bacterial species are around 2 micrometers in length, with some of the largest specimens
reaching 750 micrometers. Using fluorescence, x-ray, and electron microscopy in conjunction with
genome sequencing, we characterized Candidatus (Ca.) Thiomargarita magnifica, a bacterium that has
an average cell length greater than 9000 micrometers and is visible to the naked eye. These cells
grow orders of magnitude over theoretical limits for bacterial cell size, display unprecedented polyploidy
of more than half a million copies of a very large genome, and undergo a dimorphic life cycle
with asymmetric segregation of chromosomes into daughter cells. These features, along with
compartmentalization of genomic material and ribosomes in translationally active organelles bound
by bioenergetic membranes, indicate gain of complexity in the Thiomargarita lineage and challenge
traditional concepts of bacterial cells.
Bacteria and archaea are taxonomically
and metabolically the most diverse and
abundant organisms on Earth, but with
only a small fraction of them isolated
in culture, we remain grossly ignorant
of their biology (1). Although most model
bacteria and archaea are small, some remark-
ably large cells, referred to as giant bacteria,
are evident in atleast four phyla (2), and have
cellular sizes in the range of tens or even hun-
dreds of micrometers (3,4). Some exceptional
members of the sulfur-oxidizing gammapro-
teobacteria Thiomargarita namibiensis,for
instance, are known to reach up to 750 mm
(average size: 180 mm) (46). Such bacterial
giants raise the question of whether other
lineages of previously unidentified macro-
bacteria might exist.
Thiomargarita species from a marine sulfidic
environment that is larger than all other known
giant bacteria by ~50-fold. Our multifaceted
imaging analyses revealed massive polyploidy
and a dimorphic developmental cycle in which
genome copies are asymmetrically segregated
into apparent dispersive daughter cells. We
show that centimeter-long Thiomargarita fila-
ments represent individual cells with genetic
material and ribosomes compartmentalized
into a metabolically active, membrane-bound
organelle. Sequencing and analysis of genomes
from five single cells revealed insights into dis-
tinct cell division and cell elongation mecha-
nisms. These cellular features likely allow the
organism to grow to an unusually large size
and circumvent some of the biophysical and
bioenergetic limitations on growth. In refer-
ence to its exceptional size, we propose to
name this species Thiomargarita magnifica,
which is hereafter referred to as Candidatus
(Ca.) Thiomargarita magnifica.
Ca. T. magnifica is a centimeter-long, single
bacterial cell
Some large sulfur bacteria (LSB) form very long
filaments that may reach several centimeters in
length but are composed of thousands of indi-
vidual cells that do not exceed 200 mm(710).
We observed seasonal bouquetsof centimeter-
long white filamentous Thiomargarita cells
attached to sunken leaves of Rhizophora
mangle (fig. S1) in shallow tropical marine
mangroves from Guadeloupe, Lesser Antilles.
Thiomargarita spp. are sulfur-oxidizing gam-
maproteobacteria known to be morpholog-
ically diverse and to display polyphenism (11).
The morphology of the filaments observed
in Guadeloupe resembled those of sessile
Thiomargarita-like cells reported from deep-
sea methane seeps (12). They had a stalk-like
shape for most of their length and constricted
gradually toward the apical end, forming buds
live buried in sediment, these filaments were
smooth in appearance and free of epibiotic
bacteria or any extracellular mucus matrix
(figs. S1 and S2) (11). Budding filaments had
an average length of 9.72 ± 4.25 mm, and
only the most apical constrictions closed
completely to form one to four separate, rod-
sha ped ce lls of 0.21 ± 0.05 mm. We also
noted that some filaments reached a length of
20.00 mm (Fig. 1A and figs. S1 and S3), much
larger than any previously described single-
celled prokaryote.
To further characterize Ca. T. magnifica cells,
we highlighted membranes using osmium te-
troxide or the fluorescent dye FM 1-43X, and
visualized entire filaments in three dimensions
(3D) with hard x-ray tomography (n=4;Fig.1C
and movies S1 and S2) and confocal laser scan-
ning microscopy (CLSM) (n= 6; Fig. 1, C and
D, and movie S3). Filament sections (up to
850.6 mm in length) were visualized with trans-
mission electron microscopy (TEM) (n=15;
Fig. 1, E to G, and fig. S5). All techniques
consistently showed that each filament was
one continuous cell for nearly its entire length,
with no division septa, including the partial
constrictions toward the apical pole. Only the
most apical few buds were separated from the
filament by a closed constriction and these
represented daughter cells (Fig. 1, fig. S5, and
movies S1 to S4).
Similar to other LSB, Ca. T. magnifica cells
has a large central vacuole that reduces the
cytoplasmatic space. This may minimize growth
limitation due to the reliance on chemical diffu-
sion because bacteria lack an active intracellu-
lar transport system (2,4). In Ca.T.magnifica,
the central vacuole was continuous along the
whole filament and accounted for 73.2 ± 7.5%
(n= 4) of the total volume (Fig. 1, D and E; fig.
S5; and table S2). The cytoplasm was 3.34 ±
1.48 mm thick and was constrained to the pe-
riphery of the cell, so it was preserved from
such chemical diffusion limitations (Fig. 1, E
and F, and fig. S5) (4).
Within the cytoplasm, TEM revealed numer-
ous lucent vesicles 2.40 ± 1.03 mmindiameter,
which corresponded to the refractile gran-
ules observed with bright-field microscopy
and represented sulfur granules, as shown
by energy dispersive x-ray spectroscopy (Fig.
1F, figs. S5 and S13, and supplementary text).
The cytoplasm of Ca. T. magnifica appeared
to contain many electron-dense membrane-
bound compartments 1.31 ± 0.70 mmindiam-
eter (Fig. 1, F and G). Similar structures have
Volland et al., Science 376, 14531458 (2022) 24 June 2022 1of6
Department of Energy Joint Genome Institute, Lawrence
Berkeley National Laboratory, Berkeley, CA, USA.
for Research in Complex Systems, Menlo Park, CA, USA.
Institut de Systématique, Evolution, Biodiversité, Université
des Antilles, Muséum National d'Histoire Naturelle, CNRS,
Sorbonne Université, EPHE, Campus de Fouillole, Pointe-à-
Pitre, France.
Centre Commun de Caractérisation des
Matériaux des Antilles et de la Guyane, Université des
Antilles, UFR des Sciences Exactes et Naturelles, Pointe-à-
Pitre, Guadeloupe, France.
Department of Energy Molecular
Biophysics and Integrated Bioimaging, Lawrence Berkeley
National Laboratory, Berkeley, CA, USA.
Institut de
Systématique, Evolution, Biodiversité CNRS UMR 7205,
Museum National dHistoire Naturelle, Paris, France.
Sorbonne Universités, UPMC Univ. Paris 06, CNRS FRE3631,
Institut de Biologie Paris Seine, Paris, France.
of Molecular and Cell Biology, University of California,
Berkeley, USA.
Cornell University, College of Agriculture and
Life Sciences, Department of Microbiology, Ithaca, NY, USA.
University of California Merced, School of Natural
Sciences, Merced, CA, USA.
University of California
San Francisco, San Francisco, CA, USA.
San Francisco
State University, San Francisco, CA, USA.
*Corresponding author. Email: (J.-M.V.); (O.G.); (T.W.); (S.V.D.)
These authors contributed equally to this work.
Present address: Electron Bio-Imaging Centre, Diamond
Light Source, Harwell Science and Innovation Campus,
Didcot, UK.
Downloaded from at Cornell University on June 23, 2022
occasionally been observed in other LSB and
have been referred to as blebs of cytoplasm,
putative endobionts,”“intracytoplasmic
structures which appear to contain nuclear
material,or membrane-enclosed cytoplas-
mic compartments with ribosomes and DNA
fibrils,but their nature and function re-
mained elusive in these primarily ultrastruc-
tural studies (7,10,13). We hypothesized that
some of these membrane-bound compart-
ments within the cytoplasm may contain
dispersed genomic material because poly-
ploidy is evident in many giant bacteria
DNA and ribosomes within a membrane-bound
bacterial organelle
Although bacteria were once presumed to be
uncompartmentalized bags of enzymes,re-
cent studies have revealed the presence of or-
ganelles with functions as diverse as anaerobic
ammonium oxidation, photosynthesis, and
magnetic orientation (16). No bacteria or
archaea are known to unambiguously segre-
gate their genetic material in the manner of
eukaryotes. There is, however, some evidence
of membrane-bound nucleoids in one mem-
ber of the Atribacteria, which has a compart-
ment containing DNA that occupies most of
the cellsvolume(17). Plancotomycetes such
as Gemmata obscuriglobus also have cytosolic
Volland et al., Science 376, 14531458 (2022) 24 June 2022 2of6
Fig. 1. Morphology and ultrastructure of
Ca. T. magnifica. (A) Size comparison of selected
bacterial (green) and eukaryotic (blue) model
systems on a log scale. (B) Light microscopy
montage of the upper half of a Ca. T. magnifica
cell, with a broken basal part revealing a tube-like
morphology due to the large central vacuole
and numerous spherical intracellular sulfur
granules (a tardigrade is shown for scale). (C)3D
rendering of segmented cells from hard x-ray
tomography (movies S1, S2, and S6) and CLSM
(movie S3) putatively at various stages of the
developmental cycle. From left to right, 3D
rendered cells D,B,F,G,andD(table S2). Note
that the smallest stage corresponds to the
cell Dterminal segment and was added to the left
for visualization purposes. (D)CLSMobservation
of cell K(table S2) after fluorescent labeling
of membranes with FM 1-43X showing the
continuity of the cell from the basal pole to the
first complete constriction at the apical end.
(E) TEM montage of the apical constriction
of a cell, with the cytoplasm constrained to
the periphery. (F) Higher magnification of the
area marked in (E), with sulfur granules and
pepins at various stages of development.
(G) Higher magnification of the area marked
in (F) showing two pepins (arrowheads).
S, sulfur granule; V, vacuole.
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compartments with DNA, and recent work
has shown that these compartments repre-
sent deep invaginations of a Gram-negative
cytoplasmic membrane rather than a closed,
membrane-bound organelle (16).
4,6-Diamidino-2-phenylindole (DAPI) stain-
ing revealed that the DNA in Ca.T.magnifica
cells was concentrated in the membrane-bound
though we did not observe any connection of
these compartments to the cell envelope, their
mechanism of formation, which may include
cell membrane invagination, is yet to be studied.
They also harbored electron-dense structures
10 to 20 nm in size, similar to signatures of
Volland et al., Science 376, 14531458 (2022) 24 June 2022 3of6
Fig. 2. Characterization of the pepin organelles
by FISH and correlative TEM, as well as
membrane and DNA staining, immunohisto-
chemistry, and BONCAT. (Ato D) FISH of pepins
(arrows) in the cytoplasm of Ca. T. magnifica
(class gammaproteobacteria). Pepins are labeled
with the general bacterial probe EUB labeled with
Alexa Fluor 488 [(A), green], the gammaproteo-
bacteria-specific probe Gam42a labeled with
Cy3 [(B), yellow], and the Thiomargarita-specific
probe Thm482 labeled with Cy5 [(C), red] and with
DAPI [(D), blue] (see the supplementary text
for details). (E) TEM of a serial thin section
consecutive to the semithin section used for FISH.
The FISH- and DAPI-positive pepins appear as
electron-dense organelles under TEM. (Fand
G) Pepins from (E) under higher magnification.
Pepins are delimited by a membrane (arrowheads)
and contain numerous ribosomes that appear as
small, electron-dense granules throughout the
sections of the pepins. (Hto J) Fluorescent
labeling of membranes using FM 1-43X (H) and of
DNA using DAPI (I) and overlay (J) on a cross
section of a cell. The pepins labeled with DAPI are
also labeled with the dye FM 1-43X, confirming
the presence of a membrane. (Kto M)Visualization
of ATP synthase localization using immunohisto-
chemistry. AntiATP synthase antibodies label (K)
reveals the presence of bioenergetic membranes
around pepins [labeled with DAPI in (L)] and
throughout the cytoplasm. (N) 3D visualization of
a central portion of a cell after DAPI staining
(blue) showing the multitude of DNA clusters
spread throughout the cytoplasm (cell M; table S2
and movie S5). (Oto Q) Translational activity
revealed by BONCAT showing active protein
biosynthesis within an entire cell, including hot-
spots at constriction sites and pepins [enlarged
in (P) and (Q), respectively].
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ribosomes (Fig. 2, F and G, and fig. S6, G to K).
Fluorescence in situ hybridization (FISH) with
probes specifically targeting ribosomal RNA
sequences of Thiomargarita confirmed that
ribosomes were indeed present and were con-
centrated in these membrane-bound structures
(Fig. 2, A to G, and figs. S7 to S9) and spread
throughout the entire cell, including the apical
buds (fig. S9). This compartmentalization of DNA
and ribosomes is reminiscent of the genomic
nuclear compartmentalization in eukaryotes. By
analogy with pips, the numerous small seeds in
fruits such as watermelon or kiwi, we propose to
name this bacterial organelle a pepin (singular
expressive creation used to express smallness).
Cytoplasmic localization of bioenergetic
membranes and translational activity
Adenosine triphosphate (ATP), the principal
and universal energy currency of cells, is pro-
duced by ATP synthase, a molecular machine
embedded in bioenergetic membranes and
powered by proton motive force. Bacterial and
archaeal ATP synthases are often observed
to be localized to the cell envelope, in contrast
to eukaryotes, in which ATP is generated by
mitochondria. This cell envelope localization
potentially constrains bacterial cell size be-
cause of the surface-to-volume ratio required
to satisfy energy needs; a theoretical maxi-
mum cell size of ~10
has been estimated
recently (18). Giant Thiomargarita cells are
often cited as being exceptions to such ener-
getic constraints and are thought to rely on
their cell surface for ATP production (1921).
We used immunohistochemistry to assess the
localization of ATP synthase in Ca. T. magnifica.
We observed the distribution of ATP synthases
around pepins and throughout the complex
membrane network of the entire cytoplasm,
but they were absent from the outer cell en-
velope (Fig. 2, K to M, and fig. S20).
To examine the localization of activity
throughout the whole cell, we used bioor-
thogonal noncanonical amino acid tagging
(BONCAT) to detect protein biosynthesis (22).
Live fil ament s in cubated with a clickable
amino acid analog were labeled throughout
(Fig. 2O and fig. S10). Consistent with the
detection of ribosomes by FISH and TEM,
BONCAT showed protein biosynthesis activ-
ity in small, round-shaped areas that were
similar to pepins in size and localization, but
not all pepins appeared to be labeled (Fig.
2Q and fig. S10D). Labeled hotspots were also
observed at the site of constriction in the
apical part of cells, suggesting higher trans-
lational activity or concentration of newly
synthesized proteins in these areas (Fig. 2P).
The establishment of stable continuous lab-
oratory cultures of Ca. T. magnifica will likely be
necessary to undertake detailed st ud ie s of the
formation, biochemistry, and functions of pepins.
Based on these data, the metabolically active
biovolume of Ca. T. magnifica (excluding the
central vacuole) is two orders of magnitude
above the predicted maximum mentioned
above (table S2). This does not appear to con-
tradict previous studies; indeed, such models
have excluded bacteria with structural adap-
tations such as endomembrane systems and
slower growth rates, which allow much larger
cell volumes (18). Whereas most bacteria have
doubling times ranging from minutes to hours,
Ca. T. magnifica may be similar to other
Thiomargarita species, which require up
to 2 weeks to produce daughter cells (5). The
predicted maximum volume of bacteria as-
sumes binary fission as a division mode, but
Ca. T. magnifica does not have to double its
volume to produce a daughter cell because
only a small portion of the apex constricts
and detaches from the mother cell.
A highly polyploid cell with a
large genome
All previously described giant bacteria are
polyploid (2,3,14),meaningthattheircells
contain large numbers of genome copies
ranging from tens to tens of thousands that
are dispersed throughout the cell, supporting
the local need for molecular machineries and
overall cellular growth (15,23,24). Polyploidy
has been shown to decrease the selective pres-
sure on genes, allowing intracellular gene dup-
lication, reassortment, and divergence, and to
lead to extreme intracellular genetic diversity
in some LSB (25). Conversely, polyploidy may
allow balancing of genome copies through
homologous recombination and support a
high level of genome conservation (26). Ca.
T. magnifica, like all bacterial giants, appeared
to be polyploid; counts of DAPI-stained DNA
clusters on three CLSM 3D datasets suggested
an average of 36,880 ± 7956 genome copies per
millimeter of filament (737,598 ± 159,115 for a
fully grown 2-cm cell; see table S2, Fig. 2N,
movie S5, and details in supplementary text).
With its number of genome copies being one
order of magnitude above that of other giant
bacteria (2,24), Ca. T. magnifica accounts for
the highest estimated number of genome
copies within a single cell. Understanding the
mechanisms underlying the regulation of such
a large number of genome copies will require
additional work.
To genomically characterize Ca. T. magnifica,
we amplified, sequenced, and assembled the
DNA of five individual cells collected from a
single sunken leaf (tables S3 and S4). All five
draft genomes were highly similar to each
other, with an average nucleotide identity
>99.5% (table S5). Although extreme intra-
cellular genetic diversity has been shown in
some LSB (25), our variant analysis of DNA se-
quences recovered from a single cell revealed a
largely homogenous genome population (1.22 ±
0.18 single-nucleotide polymorphisms/100 kbp;
table S6) (27), which is similar to other poly-
ploid bacteria (26,28). The Ca. T. magnifica
genome assemblies were estimated to be nearly
complete at 91.0 to 93.7%, with total sequence
lengths between 11.5 and 12.2 Mb. This value
is twice as large as the only other sequenced
Thiomargarita species, Ca. T. nelsonii (29,30),
and at the upper range of bacterial genome
sizes; bacterial genomes are on average 4.21 ±
1.77 Mb (fig. S11). The Ca.T.magnificagenomes
contained up to 11,788 genes (only half with a
functional annotation; table S4), more than
three times the median gene count of pro-
karyotes (3935 genes) (31). For comparison
with eukaryotic organisms, Ca.T.magnifica
has a genome as large as the bakers yeast
Saccharomyces cerevisiae (12.1 Mb) and contains
more genes than the model fungus Aspergillus
nidulans (9500 genes).
Analysis of the Ca.T.magnificagenome
revealed a large set of genes for sulfur oxida-
tion and carbon fixation, suggesting chemo-
autotrophy, which is consistent with other
evidence for thioautotrophy (figs. S12 to S14
and supplementary text). Like its sister lineage
Ca. T. nelsonii, Ca. T. magnifica encoded a wide
range of metabolic capabilities with one no-
table difference: It lacked nearly all genes in-
volved in dissimilatory and assimilatory nitrate
reduction and denitrification except for Nar and
Nap nitrate reductases (fig. S12). This absence
suggests that nitrate can solely be used as
an electron acceptor to be reduced to nitrite,
which is not further reduced (29,30) (see the
supplementary text for extended genome
analysis). The absence of epibiotic bacteria
encoding secondary metabolism. With 25.9%
of sequences dedicated to biosynthetic gene
clusters (Fig. 3A), the genome encoded dozens
of modular nonribosomal peptide synthetase
and polyketide synthase systems, hinting at
numerous secondary metabolism pathways
(similar to the Actinobacteria; see table S7)
that are indicative of antibiotic or bioactive
compound production (32).
Giant bacteria have been shown to repurpose
their cell division machinery as an adaptation
to extreme cell size (33). The Ca. T. magnifica
genome also holds clues for its unusual cell
morphology in the form of an atypical com-
plement of cell division and elongation genes.
Many genes that encode core cell division pro-
teins, including the core components of Z ring
assembly and regulation, FtsA, ZipA, and FtsE-
By contrast, genes that encode the cyto-
skeletal protein FtsZ, which is part of the
well-conserved dcw (division and cell wall)
operon and the core component of the Z
ring, were conserved. Proteins ZapA, ZapB,
and ZapD, which interact with FtsZ and reg-
ulate Z ring assembly, were likewise conse rv ed
Volland et al., Science 376, 14531458 (2022) 24 June 2022 4of6
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(34).The entire set of genes that encode late
divisome proteins, including peptidoglycan
polymerases FtsI and FtsW, as well as FtsQ,
FtsL, FtsB, and FtsK, was absent from all Ca.
Thiomargarita genomes (Fig. 3B, fig. S19,
and table S8). In contrast to the conspicuous
lack of cell division genes, a complete set of
genes encoding cell elongation proteins was
present, three of which, mreD,rodZ, and the
peptidoglycan transpeptidase mrdA,have
undergone recent dupl icati on s, wi th bo th
copies located next t o eac h oth er on the
chromosome (Fig. 3B, figs. S16 to S18, and
table S8) (34). It is possible that an increased
number of cell elongation genes, coupled
with the lack of key cell division genes, may be
responsible for producing the unusually
long filaments of Ca. T. magnifica (see the
supplementary text).
Dimorphic developmental cycle of
Ca. T. magnifica
Laboratory observations of live Ca. T. magnifica
revealed eventual apical bud detachment from
the filament and release into the environment,
likely representing a dispersive stage of the
developmental cycle (Fig. 1C; fig. S1, B to F;
and supplementary text). We observed dozens
of cells at all intermediary stages, from the
smallest attached cells resembling terminal
segments recently settled to the largest fila-
ments with apical constrictions (fig. S1 and
movies S1 to S3 and S6). Such a dimorphic life
cycle resembles the aquatic single-celled model
system Caulobacter crescentus, as well as the
multicellular segmented filamentous bacteria,
albeit at a different scale, in which stalked
parentcells produce free-living daughter
cells (35,36). Because of this asymmetrical
division mode, only a small fraction of the ge-
nome copies present within pepins in the most
apical bud (1%) were transmitted to the
daughter cell (fig. S9). Like the polyploid giant
bacterium Epulopiscium spp., Ca. T. magnifica
apparently transmits only a subset of its ge-
nomes, so-called germline genomes,to the
offspring (14,24). If terminal buds are indeed
daughter cells, then such a developmental
cycle may have evolved to enhance disper-
sion similar to the fruiting bodies of the so-
cial myxobacteria or to the aerial hyphae of
Streptomyces spp. (37). This apparent life cycle
is also somewhat analogous to the sulfur-
oxidizing giant ciliate symbiosis, Zoothamnium
niveum (38), possibly representing a case
of convergent evolution of developmental
cyc le a cross domains (see the supplementary
text). As with other aspects of Ca. T. mag-
nifica biology, detailed investigations of cell
division and its regulation will require the
establishment of stable laboratory cultures,
and considering the cell size, spatial omics
approaches within a single cell might be
Concluding remarks
Confirmation bias related to viral size pre-
vented the discovery of giant viruses for more
than a century, and their ubiquity is only now
being recognized (39,40). The discovery of
Ca. T. magnifica suggests that large and more
complex bacteria may be hiding in plain sight.
Volland et al., Science 376, 14531458 (2022) 24 June 2022 5of6
Fig. 3. Genome analysis and proposed model for the subcellular organization of Ca. T. magnifica.
(A) Genome phylogenetic tree with added information about genome quality [red: low quality, orange:
medium quality, and yellow: high quality (46)], estimated level of completeness, assembly size, coding
sequence (CDS) count, and percentage of sequence dedicated to biosynthetic gene clusters (BGCs).
Pattern 1 corresponds to complete gene cluster for cell division of model bacteria.Pattern 2
corresponds to mreD,mrdA,androdZ genes are duplicated.(B) Gene neighborhoods centered on
the ddl,mreB,androdZ genes showing the incomplete set of divisome genes (lack of ftsQ and ftsA)
in both Thiomargarita species, as well as the duplication of elongasome genes (mreD,mrdA,androdZ)in
Ca. T. magnifica. Note that the Beggiatoa sp. PS, Achromatium sp. WMS3, and Ca. Thiomargarita sp.
Thio36 draft genomes were too fragmented and thus are not included here. (C) Light microscopy
image and model proposed for the subcellular organization in Ca. T. magnifica showing how the pepin
organelles might develop into other cellular compartments, resulting in an increase of surface area of
the bioenergetic membranes.
Downloaded from at Cornell University on June 23, 2022
Investigating the biology, energy metabolism,
and the formation, nature, and role of pepins
will take us a step closer to understanding the
evolution of biological complexity.
Although cells of most bacteria and archaea
are ~2 mm, eukaryotic cells are usually be-
tween 10 and 20 mm, with some of the largest
single-cell eukaryotes reaching 3 to 4 cm
(41). Several theoretical frameworks explain
the restriction of bacteria and archaea to mi-
croscopic sizes, including: (i) the lack of active
intracellular transport and the reliance on
chemical diffusion, which is efficient only along
micrometer distances (4); (ii) a predicted maxi-
mum cell volume constraining the number of
needed ribosomes should the cell grow larger
(21); or even (iii) a decrease in energy effi-
ciency due to mismatched surface area to
volume ratio when considering placement of
membrane-bound ATP synthases (18,20).
These frameworks all suggest that with in-
creasing size, the physiological or metabolic
needs of a bacterial cell grow faster than the
cells capacity to sustain it and should reach a
limit. The next largest prokaryote known after
Ca. T. magnifica, Ca. T. nelsonii, has a meta-
bolically active biovolume of 1.05 × 10
(excluding the central vacuole), close to the
predicted maximum due to ribosome limita-
tions, 1.39 × 10
, and to the bioenergetic
membrane limitation of 10
Our precise 3D measurements on a 4.27-mm
Ca. T. magnifica cell revealed a cytoplasm
biovolume several orders of magnitude above
that limit (5.91 × 10
; table S2). It is
possible that changes in spatial organization
of cellular components, such as DNA and
ribosome compartmentalization and rear-
rangement of the bioenergetic membrane
system, may allow Ca. T. magnifica to overcome
many such limitations (Fig. 3C).
Distributed in at least 23 phyla are 19 known
types of bacterial organelles, of which only
seven are membrane bound (16,42). Cyano-
bacteria can form multicellular, centimeter-long
filaments and are capable of cell differentia-
tion (43). Planctomycetes have special energy
transduction organelles called anammoxosomes
and a compartmentalized cell, and some are
even capable of phagocytosis (16,44). The
social Myxobacteria have large genomes and
a complex developmental cycle, and are ca-
pable of moving and feeding cooperatively
in predatory groups (45). Through its gi-
gantic cell size, its large genome, and its
dimorphic life cycle, but most importantly
through its compartmentalization of genetic
material in membrane-bound pepins, Ca. T.
magnifica adds to the list of bacteria that
have evolved a high level of morphological
complexity. Because it segregates its genet-
ic material in membrane-bound organelles,
Ca. T. magnifica challenges our concept of a
bacterial cell.
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We are thankful to the following centers where the electron
microscopy analyses were performed: the Centre Commun de
Caractérisation des Matériaux des Antilles et de la Guyane in
Guadeloupe, F.W.I., which is supported by The European Regional
Development Fund, the Regional Council of Guadeloupe, and the
French Research Department; the Electron Microscope Lab
(EML) of theUniversity of CaliforniaBerkeley; the ElectronMicroscopy
Resource in Donner at LBNL, Berkeley; and the FEI Eindhoven
Center. We are particularly grateful to D. Jorgens at EML for advice
and assistance in electron microscopy sample preparation and
data collection. The x-ray tomography data were acquired at
the Stanford Nano Shared Facilities at Stanford University (Palo
Alto, CA), and we are particularly grateful to A. Vailionis for his
technical support during hard x-ray tomography scan acquisitions.
The confocal microscopy observations were performed at the
Advanced Microscopy Facility at LBNL (Berkeley, CA). Preliminary
confocal microscopy observations were acquired at the IBPS
Imaging Facility, which is supported by Conseil Regional dIle-de-
France.We thank S. Volland for his help with 3D rendering
animations and H. Maughan for copyediting this manuscript.
The work (proposal: 10.46936/10.25585/60001074) conducted
by the US Department of Energy Joint Genome Institute
(, a Department of Energy (DOE)
Office of Science User Facility, is supported by the Office of Science
of the DOE operated under contract no. DE-AC02-05CH11231.
The Nagoya permit TREL1820249A/50 (unique identifier ABSCH-
IRCC-FR-246822-1) can be consulted publicly online at: Funding: This work was
supported by the John Templeton Foundation (grant 60973 to
J.-M.V., S.V.D., and T.T.), the Gordon and Betty Moore
Foundation (grant GBMF7617 to J.-M.V., S.V.D., and T.T.),
DARPA (award no. HR001120036 to J.-M.V., S.V.D., and T.T.);
the DOE Office of Science (contract no. DE-AC02-05CH11231 to
T.W., F.S., T.T., J.-M.V., K.M.D., and N.H.E.), and Region
Guadeloupe (F.W.I. grant to M.R.J.). Author contributions:
Conceptualization: O.G., S.G.R., J.-M.V., T.W., S.V.D., F.S., R.R.M.,
N.I., T.T., N.H.E.; Data curation: N.I., F.S., J.-M.V., D.U., S.G.R.;
Formal analysis: J.-M.V., S.G.R., F.S., N.I., D.U., O.G.; Funding
acquisition: T.W., S.V.D., O.G., J.L.M.; Investigation: J.-M.V.,
S.G.R., O.G., T.T., F.S., D.U., D.G., N.N., N.I., C.G.R., S.B.K., N.H.E.,
M.R.J., J.L.M.; Methodology: O.G., T.T., J.-M.V., S.V.D., S.G.R.,
D.G., N.N., R.R.M., T.W., C.G.R., S.B.K.; Resources: K.M.D., T.W.,
R.R.M., O.G., J.L.M., N.H.E., N.J.M.; Supervision: O.G., T.W., S.V.D.;
Visualization: O.G., S.G.R., J.-M.V., F.S., N.I., D.U.; Writing
original draft: J.-M.V.; Writing review and editing: J.-M.V., S.G.R.,
O.G., T.W., S.V.D., E.A., N.I., R.R.M. Competing interests: S.V.D.
serves as the CEO of Sample Exchange. The remaining authors
declare no competing interests. Materials and data availability:
All data needed to evaluate the conclusions in this study are
present in the main manuscript or the supplementary materials.
magnifi ca draft ge nomes hav e been depo sited in IM G (https:// The raw reads have been deposited in SRA
( Accession numbers are
provided in tables S3 and S4. License information: Copyright ©
2022 the authors, some rights reserved; exclusive licensee
American Association for the Advancement of Science. No claim to
original US government works.
Materials and Methods
Supplementary Text
Figs. S1 to S20
Tables S1 to S11
References (4783)
Movies S1 to S6
MDAR Reproducibility Checklist
View/request a protocol for this paper from Bio-protocol.
Submitted 16 October 2021; resubmitted 26 March 2022
Accepted 9 May 2022
Volland et al., Science 376, 14531458 (2022) 24 June 2022 6of6
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to original U.S. Government Works
A centimeter-long bacterium with DNA contained in metabolically active,
membrane-bound organelles
Jean-Marie VollandSilvina Gonzalez-RizzoOlivier GrosTomáš TymlNatalia IvanovaFrederik SchulzDanielle
GoudeauNathalie H. ElisabethNandita NathDaniel UdwaryRex R. MalmstromChantal Guidi-RontaniSusanne Bolte-
KlugeKaren M. DaviesMaïtena R. JeanJean-Louis MansotNigel J. MounceyEsther R. AngertTanja WoykeShailesh V. Date
Science, 376 (6600), • DOI: 10.1126/science.abb3634
A magnificent megabacterium
We usually think of bacteria as microscopic isolated cells or colonies. Sampling a mangrove swamp, Volland et al.
found an unusually large, sulfur-oxidizing bacterium with a complex membrane organization and predicted life cycle
(see the Perspective by Levin). Using a range of microscopy techniques, the authors observed highly polyploid cells
with DNA and ribosomes compartmentalized within membranes. Single cells of the bacterium, dubbed Candidatus
Thiomargarita magnifica, although thin and tubular, stretched more than a centimeter in length. —MAF
View the article online
Downloaded from at Cornell University on June 23, 2022
... If the latter, they would indicate that by 3,200 Ma at least one lineage of cells had found a solution to the challenges of large size faced by typical modern prokaryotic organisms, including the limitations of diffusion in transporting nutrients in and waste out (119), and a decrease in energy efficiency associated with a relatively limited membrane area for respiration (80,117). Both eukaryotes and several different bacteria (119,129) have solved these challenges in a variety of ways, for example, through cell compartmentalization via the presence of a large, inert interior space that reduces cell volume to a layer a few micrometers thick; a system of intracellular transport; an interior membrane system where respiration can occur; the presence of multiple ATP-producing endosymbionts; and/or compartmentalization of DNA and ribosomes into membrane-bound organelles (117,119,129). It is reasonable to assume that these early organisms also possessed one or more of these features. ...
... If the latter, they would indicate that by 3,200 Ma at least one lineage of cells had found a solution to the challenges of large size faced by typical modern prokaryotic organisms, including the limitations of diffusion in transporting nutrients in and waste out (119), and a decrease in energy efficiency associated with a relatively limited membrane area for respiration (80,117). Both eukaryotes and several different bacteria (119,129) have solved these challenges in a variety of ways, for example, through cell compartmentalization via the presence of a large, inert interior space that reduces cell volume to a layer a few micrometers thick; a system of intracellular transport; an interior membrane system where respiration can occur; the presence of multiple ATP-producing endosymbionts; and/or compartmentalization of DNA and ribosomes into membrane-bound organelles (117,119,129). It is reasonable to assume that these early organisms also possessed one or more of these features. ...
... Grypania spiralis, a carbonaceous filament ∼1 mm in diameter and up to 90 mm in length, forms 5-to 30-mm-diameter coils (54) that could be aggregates of microscopic prokaryotes (though see 75,115,121) or unusually large, possibly coenocytic, single cells [cf. giant bacteria (129) or eukaryotes (54)]. ...
The origin of modern eukaryotes is one of the key transitions in life's history, and also one of the least understood. Although the fossil record provides the most direct view of this process, interpreting the fossils of early eukaryotes and eukaryote-grade organisms is not straightforward. We present two end-member models for the evolution of modern (i.e., crown) eukaryotes—one in which modern eukaryotes evolved early, and another in which they evolved late—and interpret key fossils within these frameworks, including where they might fit in eukaryote phylogeny and what they may tell us about the evolution of eukaryotic cell biology and ecology. Each model has different implications for understanding the rise of complex life on Earth, including different roles of Earth surface oxygenation, and makes different predictions that future paleontological studies can test.
... This is not only true for large and complex eukaryotic cells, but also for prokaryotes. In recent years, significant progress has been made to highlight that bacterial cells are highly organized entities often relying on proteinbased strategies to coordinate and compartmentalize complex metabolic functions 6,9,10,11,12,13 . One of these strategies are protein organelles and compartments which represent nano-sized functional analogues of eukaryotic membrane organelles and utilize semipermeable protein shells to sequester specific enzymes and processes. ...
... For example, bacterial microcompartments (BMCs) sequester combinations of enzymes in self-assembling protein shells and are involved in the anabolic fixation of carbon 14,15 and catabolic processes like carbon and nitrogen source utilization 7,16 . Besides serving as nanoscale reaction chambers, another important use of protein compartments is the storage of nutrients 1,6,10,13 . The most widely distributed protein-based storage system is ferritin, an 8-12 nm protein cage used by eukaryotic and prokaryotic cells to store iron 2 . ...
... Many cells contain further systems for storing nutrients such as polyphosphate-17 , polyhydroxyalkanoate- 18 , and sulfur-storage granules or globules whose detailed functions, compositions, and formation are still being debated 13,19 . In general, storage compartments enable organisms to accumulate and retain high-value compounds for later use when encountering changing, nutrient-limited, or stress conditions 17,18,20 . ...
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Intracellular compartmentalization is essential for all cells and enables the regulation and optimization of metabolism ¹ . One of the main functions of subcellular compartments is the storage of nutrients ²⁻⁴ . As bacteria do generally not possess membrane-bound organelles, they often have to rely on functionally analogous protein-based compartments 2,5-7 . Encapsulin nanocompartments are one of the most prevalent protein-based compartmentalization strategies found in prokaryotes 5,8 . Here we show that desulfurase encapsulins represent a novel sulfur storage compartment in bacteria able to sequester large amounts of crystalline elemental sulfur. We determined the 1.78 Å cryo-EM structure of a 24 nm desulfurase-loaded encapsulin highlighting the molecular details of the protein shell and desulfurase encapsulation. We found that elemental sulfur crystals can be formed inside encapsulin shells in a desulfurase-dependent manner with L-cysteine acting as the sulfur donor. Intracellular sulfur accumulation can be influenced by the concentration and type of sulfur source in growth media. The selectively permeable protein shell allows the long-term intracellular storage of redox-labile elemental sulfur by excluding cellular reducing agents from its interior. We found that encapsulation substantially improves desulfurase activity and stability while also preventing substrate inhibition. These findings represent the first example of a dedicated and widespread storage system for the essential element sulfur in bacteria and provide the basis for understanding how this novel protein-based storage compartment is integrated within bacterial metabolism.
... A mediados del 2022, y aun con la resaca de la pandemia COVID 19, fue emocionante leer en el teléfono, un par de notas en la que se hacia mención del descubrimiento de una bacteria gigante, "visible a simple vista" (Mishra, 2022), "del tamaño de una pestaña" (BBC, 2022). Fue tan grande mi asombro que inmediatamente busqué la publicación fuente y me encontré con un artículo publicado en la revista Science (Volland et al., 2022), resultado de una colaboración de 20 autores de 12 laboratorios de Estados Unidos y Francia, en el que se describe a Thiomargarita magnifica, la bacteria mas grande encontrada en la naturaleza (Fig. 1). ...
Full-text available
The discovery of a giant bacteria is reported and compared in size with other microorganisms used as reference in biology. The need to carry out more studies in extreme environments due to the great possibility of discovering new organisms with physiological and / or physicochemical new key characteristics (with potential importance in biotechnology) is discussed.
... Single-cell genomics is particularly useful for decoding individual genomes from highly diverse microbial samples or rare target microbes, which is difficult in the metagenomic binning approach. Examples of its application include the analysis of bacteria visible to the naked eye (Volland et al. 2022), a comprehensive survey of marine bacteria in surface seawater (Pachiadaki et al. 2019), the identification of secondary metabolite producers from marine sponges (Wilson et al. 2014;Kogawa et al. 2022), the assessment of subspecies and intraspecific recombination in environmental bacterial species (Zaremba-Niedzwiedzka et al. 2013;Kashtan et al. 2014), and the identification of gut bacteria that degrade soluble dietary fiber (Chijiiwa et al. 2020). Single-cell genomics provides previously inaccessible insights into microbial ecosystems and functions. ...
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The advent of next-generation sequencing technologies has facilitated the acquisition of large amounts of DNA sequence data at a relatively low cost, leading to numerous breakthroughs in decoding microbial genomes. Among the various genome sequencing activities, metagenomic analysis, which entails the direct analysis of uncultured microbial DNA, has had a profound impact on microbiome research and has emerged as an indispensable technology in this field. Despite its valuable contributions, metagenomic analysis is a “bulk analysis” technique that analyzes samples containing a wide diversity of microbes, such as bacteria, yielding information that is averaged across the entire microbial population. In order to gain a deeper understanding of the heterogeneous nature of the microbial world, there is a growing need for single-cell analysis, similar to its use in human cell biology. With this paradigm shift in mind, comprehensive single-cell genomics technology has become a much-anticipated innovation that is now poised to revolutionize microbiome research. It has the potential to enable the discovery of differences at the strain level and to facilitate a more comprehensive examination of microbial ecosystems. In this review, we summarize the current state-of-the-art in microbial single-cell genomics, highlighting the potential impact of this technology on our understanding of the microbial world. The successful implementation of this technology is expected to have a profound impact in the field, leading to new discoveries and insights into the diversity and evolution of microbes.
... Most of them were originally interpreted as macroalgae or multicellular eukaryotes, but few of them can be unambiguously accepted as eukaryotic, due to their simple morphology and lack of informative characters. In most cases, they cannot be differentiated from microbial mat fragments, bacterial macrocolonies (e.g., filamentous and sheet-like colonies built by Nostoc flagelliforme (Feng et al., 2012)), and/or even giant bacteria (e.g., centimeter long Thiomargarita magnifica (Volland et al., 2022)). The blade-like macrofossils from the Gaoyuzhuang Formation, in contrast, possess morphological and dimensional features which provide compelling evidence to identify them to be macroscopic eukaryotic organisms, despite their relatively simple morphology. ...
... Since then, advances in molecular techniques have allowed the further discovery of the diversity and functions of the mangrove sediment microbiome. New microorganisms (among others, Sefrji et al., 2021Sefrji et al., , 2022 and microbial roles in nutrient cycling have been recognized to determine profound effects (Volland et al., 2022), which will be reviewed here, on sediment microbial communities and overall ecosystem functioning. In this review, we first introduce the main groups of bioturbating macrofaunal species and their ecology and then, reviewing the more recent literature, explore the diversity, dynamics and function of the sediment microbiome considering the ...
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Globally, soils and sediments are affected by the bioturbation activities of benthic species. The consequences of these activities are particularly impactful in intertidal sediment, which is generally anoxic and nutrient-poor. Mangrove intertidal sediments are of particular interest because, as the most productive forests and one of the most important stores of blue carbon, they provide global-scale ecosystem services. The mangrove sediment microbiome is fundamental for ecosystem functioning, influencing the efficiency of nutrient cycling and the abundance and distribution of key biological elements. Redox reactions in bioturbated sediment can be extremely complex, with one reaction creating a cascade effect on the succession of respiration pathways. This facilitates the overlap of different respiratory metabolisms important in the element cycles of the mangrove sediment, including carbon, nitrogen, sulphur and iron cycles, among others. Considering that all ecological functions and services provided by mangrove environments involve microorganisms, this work reviews the microbial roles in nutrient cycling in relation to bioturbation by animals and plants, the main mangrove ecosystem engineers. We highlight the diversity of bioturbating organisms and explore the diversity, dynamics and functions of the sediment microbiome, considering both the impacts of bioturbation. Finally, we review the growing evidence that bioturbation, through altering the sediment microbiome and environment, determining a 'halo effect', can ameliorate conditions for plant growth, highlighting the potential of the mangrove microbiome as a nature-based solution to sustain mangrove development and support the role of this ecosystem to deliver essential ecological services.
Chapter 1 demonstrates the differences between science and philosophy, as well as why researchers should have a philosophical mindset. In addition, we will discuss what a PhD is all about and the reasons for enrolling in a doctoral program. Additionally, we discussed the differences between fixed-mindset and growth-mindset as well as the difference between intrinsic motivation and extrinsic motivation. Ultimately, philosophical mindset, growth-mindset, and intrinsic motivation are some of the few essential elements for high-achieving researchers.
The origin of eukaryotes is one of the most fundamental problems in the entire history of life. How did eukaryotes arise? Previous attempts to solve the problem are very far from an answer, at best they propose a solution to one of the various innovations that ended up culminating in eukaryotes. Based on a hypothetical-deductive methodology, as usual in evolutionary issues, I propose that eukaryotes emerged from the endosymbiotic association between a flagellate parasite and its host, of which the sperm is the main vestige. The hypothesis unifies the solution to the vast array of acquisitions shared by eukaryotes that differentiate them from other beings, remarkably cell nucleus, mitosis, meiosis and sexual reproduction. The solution has a deep impact on understanding the origin and functioning of all complex life forms.
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Discovery of Thiomargarita magnifica – an exceptionally large giant sulfur bacterium – urges us to pay additional attention to the giant sulfur bacteria and to revisit our recent bioinformatic finding of lipoxygenases in the representatives of the genus Beggiatoa. These close relatives of Thiomargarita magnifica meet the similar size requirements by forming multicellular structures. We hypothesize that their lipoxygenases are a part of the oxylipin signaling system that provides high level of cell-to-cell signaling complexity which, in turn, enables them to reach large sizes.
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The origin of eukaryotic cell size and complexity is thought by some to have required an energy excess provided by mitochondria, whereas others claim that mitochondria provide no energetic boost to eukaryotes. Recent observations show that energy demand scales continuously and linearly with cell volume across both prokaryotes and eukaryotes, and thus suggest that eukaryotes do not have an increased energetic capacity over prokaryotes. However, amounts of respiratory membranes and ATP synthases scale super-linearly with cell surface area. Furthermore, the energetic consequences of the contrasting genomic designs between prokaryotes and eukaryotes have yet to be precisely quantified. Here, we investigated (1) potential factors that affect the cell volumes at which prokaryotes become surface area-constrained, and (2) the amount of energy that is divested to increasing amounts of DNA due to the contrasting genomic designs of prokaryotes and eukaryotes. Our analyses suggest that prokaryotes are not necessarily constrained by their cell surfaces at cell volumes of 10 ⁰ ‒10 ³ μm ³ , and that the genomic design of eukaryotes is only slightly advantageous at genomes sizes of 10 ⁶ ‒10 ⁷ bp. This suggests that eukaryotes may have first evolved without the need for mitochondria as these ranges hypothetically encompass the Last Eukaryote Common Ancestor and its proto-eukaryotic ancestors. However, our analyses also show that increasingly larger and fast-dividing prokaryotes would have a shortage of surface area devoted to respiration and would disproportionally divest more energy to DNA synthesis at larger genome sizes. We thus argue that, even though mitochondria may not have been required by the first eukaryotes, the successful diversification of eukaryotes into larger and more active cells was ultimately contingent upon the origin of mitochondria. Significance There has been a lot of theorizing about the evolution of eukaryotes from prokaryotes, but no consensus seems to be on the horizon. Our quantitative analyses on the required amount of respiratory membrane, and the amount of energy diverted to DNA synthesis, by both prokaryotes and eukaryotes, suggest that mitochondria provided rather small advantages to the first eukaryotes, but were advantageous for the macro-evolutionary diversification of eukaryotes. This conclusion provides a middle road in the debate between those that claim that the origin of eukaryotes required a massive energy boost provided by mitochondria, and those that argue that the origin of mitochondria did not represent a quantum leap in energetic advantages to eukaryotes.
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After the first discovery in the 1980s in F-plasmids as a plasmid maintenance system, a myriad of toxin-antitoxin (TA) systems has been identified in bacterial chromosomes and mobile genetic elements (MGEs), including plasmids and bacteriophages. TA systems are small genetic modules that encode a toxin and its antidote and can be divided into seven types based on the nature of the antitoxin molecules and their mechanism of action to neutralise toxins. Among them, type II TA systems are widely distributed in chromosomes and plasmids and the best studied so far. Maintaining genetic material may be the major function of type II TA systems associated with MGEs, but the chromosomal TA systems contribute largely to functions associated with bacterial physiology, including the management of different stresses, virulence and pathogenesis. Due to growing interest in TA research, extensive work has been conducted in recent decades to better understand the physiological roles of these chromosomally encoded modules. However, there are still controversies about some of the functions associated with different TA systems. This review will discuss the most current findings and the bona fide functions of bacterial type II TA systems.
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Many microorganisms produce natural products that form the basis of antimicrobials, antivirals, and other drugs. Genome mining is routinely used to complement screening-based workflows to discover novel natural products. Since 2011, the "antibiotics and secondary metabolite analysis shell—antiSMASH" ( has supported researchers in their microbial genome mining tasks, both as a free-to-use web server and as a standalone tool under an OSI-approved open-source license. It is currently the most widely used tool for detecting and characterising biosynthetic gene clusters (BGCs) in bacteria and fungi. Here, we present the updated version 6 of antiSMASH. antiSMASH 6 increases the number of supported cluster types from 58 to 71, displays the modular structure of multi-modular BGCs, adds a new BGC comparison algorithm, allows for the integration of results from other prediction tools, and more effectively detects tailoring enzymes in RiPP clusters.
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Most studies of bacterial reproduction have centered on organisms that undergo binary fission. In these models, complete chromosome copies are segregated with great fidelity into two equivalent offspring cells. All genetic material is passed on to offspring, including new mutations and horizontally acquired sequences. However, some bacterial lineages employ diverse reproductive patterns that require management and segregation of more than two chromosome copies. Epulopiscium spp., and their close relatives within the Firmicutes phylum, are intestinal symbionts of surgeonfish (family Acanthuridae). Each of these giant (up to 0.6 mm long), cigar-shaped bacteria contains tens of thousands of chromosome copies. Epulopiscium spp. do not use binary fission but instead produce multiple intracellular offspring. Only ∼1% of the genetic material in an Epulopiscium sp. type B mother cell is directly inherited by its offspring cells. And yet, even in late stages of offspring development, mother-cell chromosome copies continue to replicate. Consequently, chromosomes take on a somatic or germline role. Epulopiscium sp. type B is a strict anaerobe and while it is an obligate symbiont, its host has a facultative association with this intestinal microorganism. Therefore, Epulopiscium sp. type B populations face several bottlenecks that could endanger their diversity and resilience. Multilocus sequence analyses revealed that recombination is important to diversification in populations of Epulopiscium sp. type B. By employing mechanisms common to others in the Firmicutes, the coordinated timing of mother-cell lysis, offspring development and congression may facilitate the substantial recombination observed in Epulopiscium sp. type B populations.
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A key feature that differentiates prokaryotic cells from eukaryotes is the absence of an intracellular membrane surrounding the chromosomal DNA. Here, we isolate a member of the ubiquitous, yet-to-be-cultivated phylum ‘Candidatus Atribacteria’ (also known as OP9) that has an intracytoplasmic membrane apparently surrounding the nucleoid. The isolate, RT761, is a subsurface-derived anaerobic bacterium that appears to have three lipid membrane-like layers, as shown by cryo-electron tomography. Our observations are consistent with a classical gram-negative structure with an additional intracytoplasmic membrane. However, further studies are needed to provide conclusive evidence for this unique intracellular structure. The RT761 genome encodes proteins with features that might be related to the complex cellular structure, including: N-terminal extensions in proteins involved in important processes (such as cell-division protein FtsZ); one of the highest percentages of transmembrane proteins among gram-negative bacteria; and predicted Sec-secreted proteins with unique signal peptides. Physiologically, RT761 primarily produces hydrogen for electron disposal during sugar degradation, and co-cultivation with a hydrogen-scavenging methanogen improves growth. We propose RT761 as a new species, Atribacter laminatus gen. nov. sp. nov. and a new phylum, Atribacterota phy. nov.
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The Pfam database is a widely used resource for classifying protein sequences into families and domains. Since Pfam was last described in this journal, over 350 new families have been added in Pfam 33.1 and numerous improvements have been made to existing entries. To facilitate research on COVID-19, we have revised the Pfam entries that cover the SARS-CoV-2 proteome, and built new entries for regions that were not covered by Pfam. We have reintroduced Pfam-B which provides an automatically generated supplement to Pfam and contains 136 730 novel clusters of sequences that are not yet matched by a Pfam family. The new Pfam-B is based on a clustering by the MMseqs2 software. We have compared all of the regions in the RepeatsDB to those in Pfam and have started to use the results to build and refine Pfam repeat families. Pfam is freely available for browsing and download at
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Bacteria surround their cell membrane with a net-like peptidoglycan layer, called sacculus, to protect the cell from bursting and maintain its cell shape. Sacculus growth during elongation and cell division is mediated by dynamic and transient multiprotein complexes, the elongasome and divisome, respectively. In this Review we present our current understanding of how peptidoglycan synthases are regulated by multiple and specific interactions with cell morphogenesis proteins that are linked to a dynamic cytoskeletal protein, either the actin-like MreB or the tubulin-like FtsZ. Several peptidoglycan synthases and hydrolases require activation by outer-membrane-anchored lipoproteins. We also discuss how bacteria achieve robust cell wall growth under different conditions and stresses by maintaining multiple peptidoglycan enzymes and regulators as well as different peptidoglycan growth mechanisms, and we present the emerging role of ld-transpeptidases in peptidoglycan remodelling.
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In the field of correlative microscopy, light and electron microscopy form a powerful combination for morphological analyses in zoology. Due to sample thickness limitations, these imaging techniques often require sectioning to investigate small animals and thereby suffer from various artefacts. A recently introduced nanoscopic X-ray computed tomography (NanoCT) setup has been used to image several biological objects, none that were, however, embedded into resin, which is prerequisite for a multitude of correlative applications. In this study, we assess the value of this NanoCT for correlative microscopy. For this purpose, we imaged a resin-embedded, meiofaunal sea cucumber with an approximate length of 1 mm, where microCT would yield only little information about the internal anatomy. The resulting NanoCT data exhibits isotropic 3D resolution, offers deeper insights into the 3D microstructure, and thereby allows for a complete morphological characterization. For comparative purposes, the specimen was sectioned subsequently to evaluate the NanoCT data versus serial sectioning light microscopy (ss-LM). To correct for mechanical instabilities and drift artefacts, we applied an alternative alignment procedure for CT reconstruction. We thereby achieve a level of detail on the subcellular scale comparable to ss-LM images in the sectioning plane.
Advances in imaging technologies have revealed that many bacteria possess organelles with a proteomically defined lumen and a macromolecular boundary. Some are bound by a lipid bilayer (such as thylakoids, magnetosomes and anammoxosomes), whereas others are defined by a lipid monolayer (such as lipid bodies), a proteinaceous coat (such as carboxysomes) or have a phase-defined boundary (such as nucleolus-like compartments). These diverse organelles have various metabolic and physiological functions, facilitating adaptation to different environments and driving the evolution of cellular complexity. This Review highlights that, despite the diversity of reported organelles, some unifying concepts underlie their formation, structure and function. Bacteria have fundamental mechanisms of organelle formation, through which conserved processes can form distinct organelles in different species depending on the proteins recruited to the luminal space and the boundary of the organelle. These complex subcellular compartments provide evolutionary advantages as well as enabling metabolic specialization, biogeochemical processes and biotechnological advances. Growing evidence suggests that the presence of organelles is the rule, rather than the exception, in bacterial cells. Advances in imaging techniques have revealed an unexpected abundance and diversity of organelles in bacteria. In this Review, Greening and Lithgow outline the different types of bacterial organelles and discuss common themes in their formation and function.
How mitochondria shaped the evolution of eukaryotic complexity has been controversial for decades. The discovery of the Asgard archaea, which harbor close phylogenetic ties to the eukaryotes, supports the idea that a critical endosymbiosis between an archaeal host and a bacterial endosymbiont transformed the selective constraints present at the origin of eukaryotes. Cultured Asgard archaea are typically prokaryotic in both size and internal morphology, albeit featuring extensive protrusions. The acquisition of the mitochondrial predecessor by an archaeal host cell fundamentally altered the topology of genes in relation to bioenergetic membranes. Mitochondria internalised not only the bioenergetic membranes but also the genetic machinery needed for local control of oxidative phosphorylation. Gene loss from mitochondria enabled expansion of the nuclear genome, giving rise to an extreme genomic asymmetry that is ancestral to all extant eukaryotes. This genomic restructuring gave eukaryotes thousands of fold more energy availability per gene. In principle, that difference can support more and larger genes, far more non-coding DNA, greater regulatory complexity, and thousands of fold more protein synthesis per gene. These changes released eukaryotes from the bioenergetic constraints on prokaryotes, facilitating the evolution of morphological complexity.