ArticlePDF Available

Abstract and Figures

The gut microbiota of termites plays critical roles in the symbiotic digestion of lignocellulose. While phylogenetically “lower termites” are characterized by a unique association with cellulolytic flagellates, higher termites (family Termitidae) harbor exclusively prokaryotic communities in their dilated hindguts. Unlike the more primitive termite families, which primarily feed on wood, they have adapted to a variety of lignocellulosic food sources in different stages of humification, ranging from sound wood to soil organic matter. In this study, we comparatively analyzed representatives of different taxonomic lineages and feeding groups of higher termites to identify the major drivers of bacterial community structure in the termite gut, using amplicon libraries of 16S rRNA genes from 18 species of higher termites. In all analyses, the wood-feeding species were clearly separated from humus and soil feeders, irrespective of their taxonomic affiliation, offering compelling evidence that diet is the primary determinant of bacterial community structure. Within each diet group, however, gut communities of termites from the same subfamily were more similar than those of distantly related species. A highly resolved classification using a curated reference database revealed only few genus-level taxa whose distribution patterns indicated specificity for certain host lineages, limiting any possible cospeciation between the gut microbiota and host to short evolutionary time scales. Rather, the observed patterns in the host-specific distribution of the bacterial lineages in termite guts are best explained by diet-related differences in the availability of microhabitats and functional niches.
Content may be subject to copyright.
Diet is the primary determinant of bacterial community
structure in the guts of higher termites
ARAM MIKAELYAN,*CARSTEN DIETRICH,* TIM K
OHLER,* MICHAEL POULSEN,DAVID
SILLAM-DUSS
ES§¶ and ANDREAS BRUNE*
*Department of Biogeochemistry, Max Planck Institute for Terrestrial Microbiology, Karl-von-Frisch-Str. 10, 35043, Marburg,
Germany, LOEWE Center for Synthetic Microbiology, SYNMIKRO, Philipps-Universit
at Marburg, Marburg, Germany,
Section for Ecology and Evolution, Department of Biology, Centre for Social Evolution, University of Copenhagen, Copenhagen
East, Denmark, §Laboratory of Experimental and Comparative Ethology (LEEC), University of Paris 13, Sorbonne Paris Cit
e,
Villetaneuse, France, Institute of Ecology and Environmental Sciences Paris (iEES-Paris), Institute of Research for
Development, Sorbonne Universit
es, Bondy, France
Abstract
The gut microbiota of termites plays critical roles in the symbiotic digestion of ligno-
cellulose. While phylogenetically ‘lower termites’ are characterized by a unique associ-
ation with cellulolytic flagellates, higher termites (family Termitidae) harbour
exclusively prokaryotic communities in their dilated hindguts. Unlike the more primi-
tive termite families, which primarily feed on wood, they have adapted to a variety of
lignocellulosic food sources in different stages of humification, ranging from sound
wood to soil organic matter. In this study, we comparatively analysed representatives
of different taxonomic lineages and feeding groups of higher termites to identify the
major drivers of bacterial community structure in the termite gut, using amplicon
libraries of 16S rRNA genes from 18 species of higher termites. In all analyses, the
wood-feeding species were clearly separated from humus and soil feeders, irrespective
of their taxonomic affiliation, offering compelling evidence that diet is the primary
determinant of bacterial community structure. Within each diet group, however, gut
communities of termites from the same subfamily were more similar than those of dis-
tantly related species. A highly resolved classification using a curated reference data-
base revealed only few genus-level taxa whose distribution patterns indicated
specificity for certain host lineages, limiting any possible cospeciation between the gut
microbiota and host to short evolutionary timescales. Rather, the observed patterns in
the host-specific distribution of the bacterial lineages in termite guts are best explained
by diet-related differences in the availability of microhabitats and functional niches.
Keywords: gut microbiota, insects, pyrosequencing, termites
Received 25 June 2015; revision received 20 August 2015; accepted 1 September 2015
Introduction
Termites are eusocial cockroaches that have specialized
on a diet of lignocellulose in various stages of humifica-
tion (Brune 2014). The evolutionary transition from an
omnivorous to wood-feeding lifestyle, which occurred
more than 150 million years ago, was accompanied by
several digestive modifications, including a distinctive
enlargement of the hindgut and the acquisition of cellu-
lolytic flagellates (Brune & Dietrich 2015). The flagel-
lates play a critical role in symbiotic digestion in the
hindgut of all evolutionary ‘lower’ termite families, but
were lost in the most derived family, the Termitidae or
‘higher’ termites, which appeared about 50 million
years ago and whose gut microbiota is entirely prokary-
otic (Brune & Ohkuma 2011; Bourguignon et al. 2015).
Representing over 85% of all termite genera, the higher
termites are the most diverse of all termite families
(Krishna et al. 2013). While higher termites have
Correspondence: Andreas Brune, Fax: +49 6421 178999; E-mail:
brune@mpi-marburg.mpg.de
©2015 John Wiley & Sons Ltd
Molecular Ecology (2015) doi: 10.1111/mec.13376
diversified in many aspects, the most important special-
ization concerns their diet, which extends far beyond the
wood-feeding lifestyle of ‘lower termites’ (Eggleton &
Tayasu 2001). Some higher termites feed on sound ligno-
cellulose such as wood or dry grass, others on partially
degraded material such as leaf litter, herbivore dung or
humus, or in the case of the true soil feeders, on soil
organic matter in an advanced stage of humification
(Donovan et al. 2001; Eggleton & Tayasu 2001). A special
case are the fungus-cultivating termites, which grow
basidiomycete fungi (Termitomyces spp.) to predigest
their lignocellulosic diet and, in addition, also consume
the mycelia of the fungal garden (Poulsen 2015). While
fungus feeders consist exclusively of Macrotermitinae,
other feeding groups usually comprise representatives
from various subfamilies (Donovan et al. 2001).
Higher termites offer the unique opportunity to study
the co-evolution of gut microbiota and host. The
tremendous diversity in their dietary specializations
and corresponding digestive adaptations in gut anat-
omy and physiology are reflected in the composition of
their bacterial communities (Brune & Ohkuma 2011;
Brune 2014). Recent studies using deep-sequencing of
16S rRNA genes to examine the gut microbiota across a
wide range of higher termites have detected obvious
patterns in community structure, which were
interpreted to reflect both phylogeny and diet of the
respective host groups (Dietrich et al. 2014; Otani et al.
2014; Rahman et al. 2015). Nevertheless, limited taxon
sampling within the higher termites has so far pre-
vented an effective identification of the major drivers of
community structure of their gut microbiota.
Therefore, to investigate potential drivers of community
structure in higher termites, it is essential that the analysis
include a critical number of representatives from the same
feeding groups that belong to evolutionarily independent
host lineages. Building on our previous analysis of bacte-
rial community structure in a wide range of termites and
cockroaches, which already included 10 species of higher
termites (Dietrich et al. 2014), we investigated eight addi-
tional species from previously undersampled feeding
groups. We sequenced the amplified 16S rRNA (V3V4
region) genes using the Illumina MiSeq platform and
analysed the communities across all samples including
both similarity-based metrics and the identification of
individual lineages using a highly resolved taxonomic
framework (DictDb; Mikaelyan et al. 2015).
Materials and methods
Termites: collection, identification and dissection
Atlantitermes sp., Cornitermes sp., Neocapritermes taracua,
Termes fatalis and Velocitermes sp. were collected near
Petit-Saut dam in French Guiana (5°40N, 52°590W), Ter-
mes hospes and Microcerotermes parvus were collected
near Pointe-Noire in the Democratic Republic of the
Congo (4°410S, 11°510E), and Promirotermes sp. was col-
lected in ARC-PPRI Rietondale Research Station, Preto-
ria in South Africa, in the year 2013. The origin of the
remaining species has been previously described
(Dietrich et al. 2014). Species identity was confirmed by
sequence analysis of the gene encoding cytochrome oxi-
dase subunit 2 (COII; for accession numbers, see
Table 1). A maximum-likelihood tree was calculated for
the COII gene sequences using FASTTREE (Price et al.
2009) under the general time-reversible (GTR) model of
evolution to ascertain the phylogenetic position of the
sequences with respect to each other and to close rela-
tives in GenBank (Fig. S1, Supporting information). All
termites (worker caste only) were dissected with fine-
tipped forceps within a few days of arrival in the labo-
ratory in Marburg (for details, see Table 1). Hindguts
(1020 per sample) were pooled in 2-mL tubes contain-
ing 750 lL sodium phosphate buffer (120 mM; pH 8.0)
and homogenized. DNA was extracted and purified
using a bead-beating protocol as previously described
(Paul et al. 2012).
Library preparation and sequencing
The V3V4 region of the 16S rRNA gene was amplified
from each sample using the primers M13-343Fmod and
M13-784Rmod, which were based on the universal
bacterial primers 343Fmod and 784Rmod (K
ohler et al.
2012) respectively, but additionally included universal
M13-specific priming sites on their 50ends (Daigle et al.
2011). The cycling conditions for this PCR step were as
described previously (K
ohler et al. 2012). The resulting
amplicons, tagged with the M13 tails, were used as tem-
plate for a subsequent PCR step using the Herculase II
Fusion DNA Polymerase Kit (Agilent Technologies,
USA). The following PCR conditions were used: 94 °C for
3 min, followed by 28 cycles (94 °C for 20 s, 58 °Cfor
20 s and 72 °C for 50 s), and finally 72 °C for 2 min. Puri-
fied PCR products were mixed in equimolar amounts
and sequenced commercially (paired-end; 2 9350 nt;
Illumina MiSeq; GATC Biotech, Konstanz, Germany). The
quality of the final products was checked by gel elec-
trophoresis. The quality-checked MiSeq data sets were
submitted to the MG-RAST server (Meyer et al. 2008) and
can be accessed under the numbers 4639074.34639081.3.
Processing of sequence data
Both iTag (Degnan & Ochman 2012) libraries (this
study) and pyrotag libraries (Dietrich et al. 2014;
GenBank accession nos SAMN02228091101) were
©2015 John Wiley & Sons Ltd
2A. MIKAELYAN ET AL.
processed using the UPARSE pipeline (Edgar 2013).
Only reads with a minimum length of 250 bp and a
maximum expected error of 0.5 were selected and sepa-
rated by sample into different fastq files using the sam-
ple-specific barcodes included in the sequences. For the
iTag libraries, the UPARSE pipeline was used to merge
paired reads; pairs with mismatches in the overlapping
region were discarded. After removal of barcodes and
primers, the reads from each sample were clustered at
a threshold of 99% sequence similarity to form opera-
tional taxonomic units (OTUs) using UPARSE. A repre-
sentative sequence from each OTU was selected and
aligned with the MOTHUR aligner, using the Silva refer-
ence alignment (SSURef release 119) as a template.
Analysis of community structure
For the analysis of taxonomic composition of each sam-
ple, the OTUs in the de-replicated data sets were classi-
fied using the RDP classifier (Wang et al. 2007)
implemented in the MOTHUR software suite (Schloss et al.
2009), using a confidence cut-off of 80% and the Dicty-
optera taxonomic reference database (DICTDB) v. 3.0
(Mikaelyan et al. 2015). Genus-level lineages were
ranked by determining their cumulative contribution to
the principal component analysis (PCA) of the gut com-
munities as previously described (Dietrich et al. 2014;
Otani et al. 2014). BrayCurtis distances between all
communities at the genus level were calculated using
the VEGAN package (Oksanen et al. 2015) in the Rstatisti-
cal software suite (R Core Team 2015). Data sets were
subsampled to a thousand sequences, and distances
were visualized using principal coordinate analysis
(PCoA) implemented in vegan. The BrayCurtis dis-
tances were additionally subjected to hierarchical clus-
ter analysis and visualized as a dendrogram with the
PVCLUST package (Suzuki & Shimodaira 2006) of R.
Similarities in the phylogenetic structure of the com-
munities were determined using the taxonomy-indepen-
dent weighted UniFrac metric (Lozupone et al. 2007)
implemented in MOTHUR. A maximum-likelihood tree
was constructed using FASTTREE (Price et al. 2009) with
subsamples of 1100 sequences per sample (correspond-
ing to the number of sequences in the smallest library).
This tree served as input for the calculation of pairwise
distances between all 18 samples with UniFrac, which
were ordinated using PCoA.
The significance of clustering at the community level
was tested for both metrics using the adonis function
implemented in the VEGAN package (Oksanen et al.
2015), including taxonomy, diet and geography as pos-
sible variables. Additionally, the KruskalWallis non-
parametric test implemented in Rwas used to assess
whether differences in the relative abundance of indi-
vidual bacterial genera can be explained by differences
in diet or host taxonomy.
Phylogenetic analysis of short reads
Representatives of each OTU from each sample were
subjected to phylogenetic analysis using FASTTREE (Price
et al. 2009) under maximum-likelihood criteria and the
GTR model. The relative abundance of each OTU in the
tree was annotated as circles using the APE package
(Paradis et al. 2004) written for the Rsoftware suite
(R Core Team 2015).
Table 1 Higher termites used in the current study, their phylo-
genetic affiliations and dietary preferences
ID Host species COII gene
Feeding
group* Diet
Macrotermitinae
1Macrotermes sp. KT184483 F Lignocellulose,
fungus
2Macrotermes
subhyalinus
KT184482 F Lignocellulose,
fungus
3Odontotermes sp. KT184484 F Lignocellulose,
fungus
Apicotermitinae
4Alyscotermes
trestus
KT184481 S Soil organic
matter
Syntermitinae
5Cornitermes sp. AIZ68247
L Litter
§
Termitinae
6Microcerotermes
parvus
AIZ68273
W Wood
7Microcerotermes
sp.
KT184480 W Wood
8Neocapritermes
taracua
AIZ68299
H Humus
9Cubitermes
ugandensis
AIZ68260
S Soil organic
matter
10 Ophiotermes sp. KT184477 S Soil organic
matter
11 Termes hospes AIZ68312
H Humus
12 Termes fatalis KT184478 H Humus
13 Promirotermes sp. KT184479 H Humus
Nasutitermitinae
14 Atlantitermes sp. KT184476 H Humus
15 Velocitermes sp. KT184475 L Litter
16 Trinervitermes sp. KT184474 W Dry grass
17 Nasutitermes
corniger
AIZ68286
W Wood
18 Nasutitermes
takasagoensis
KT184473 W Wood
*Feeding groups: F, fungus feeders; W, wood or grass feeders;
L, litter feeders; H, humus feeders; S, ‘true’ soil feeders.
Based on dietary information in Bignell et al. (2011).
COII genes annotated in the mitochondrial genomes recon-
structed by Dietrich & Brune (2014).
§
Based on dietary information in Gontijo & Domingos (1991).
©2015 John Wiley & Sons Ltd
DIET SHAPES TERMITE GUT COMMUNITY STRUCTURE 3
Results
Quality processing of the amplicon libraries yielded
109721 828 sequence reads for each pyrotag and
57 028222 116 sequence reads for each iTag library.
Classification with the taxonomic framework of DictDb
successfully assigned 99% of the reads at the phylum
level. Classification success decreased with taxonomic
depth (4890% at the genus level; for details, see
Table S1, Supporting information). The lowest classifica-
tion success was observed with some humus-feeding
taxa, which is indicative of the bacterial diversity that
still remains to be explored in this diet group, and the
need to add more full-length 16S rRNA reference
sequences to DictDb. For a detailed overview of the
classification results, see Tables S2 and S3 (Supporting
information).
At the phylum level, termites within a subfamily dif-
fered considerably in community structure, with the
distribution of dominant phyla being strongly reflective
of the termite feeding group. Irrespective of their phylo-
genetic affiliation, termites belonging to the same
feeding group showed striking similarities in the distri-
bution of dominant bacterial phyla (Fig. 1). Members of
Spirochaetes, Fibrobacteres and/or the TG3 phylum
were more prevalent among termites feeding on sound
wood or grass than among termites of other feeding
groups, although the proportion of each phylum dif-
fered among host species. Large proportions of Spiro-
chaetes, Fibrobacteres and TG3 phylum were also
found among litter-feeding termites. However, with the
prevalence of Firmicutes and Bacteroidetes encountered
in these taxa, the overall pattern among the litter
feeders resembled that of the humus feeders. Humus
feeders, soil feeders and fungus feeders also shared
similarities in community structure, particularly in the
large proportions of Firmicutes, Bacteroidetes and Pro-
teobacteria, but differed in the abundance of Spiro-
chaetes, which was lower in soil feeders and almost
absent from fungus feeders (Fig. 1). Soil feeders also
harboured a larger proportion of Actinobacteria than
most other species. Fibrobacteres and TG3 phylum,
which were characteristic of wood-feeding and litter-
feeding termites, were virtually absent from most repre-
sentatives of the other feeding groups.
The diet-specific pattern in gut community structure
observed at the phylum level was even more evident at
higher taxonomic or phylogenetic resolution. PCoA of
community structure showed a clustering of samples by
host diet with both BrayCurtis (at the genus level) and
UniFrac metrics, with the first two axes explaining
approximately half of the total variability in the data
sets (Fig. 2). In addition, hierarchical cluster analysis
revealed a signal of host taxonomy within each diet
group; that is, phylogenetically related termites are
more similar in community structure than termites from
different subfamilies (Fig. 3). Both BrayCurtis and
UniFrac dissimilarities in community structure were
explained by host diet and taxonomy (considering a sig-
nificance threshold at P0.05), and not by geographi-
cal location (see Table S5, Supporting information for
details). Typically, differences in the relative abundance
of individual bacterial genera were better explained by
differences in diet in comparison with host taxonomy
(Tables S6 and S7, Supporting information).
To identify the genus-level taxa that contribute most
to the observed clustering pattern shown in Fig. 2, we
conducted a PCA and ranked the genus-level taxa in
descending order of their cumulative contribution to
the loading factors. The relative abundances of the 20
taxa contributing the most to the clustering observed in
PCA illustrate a preferential enrichment of different
Fig. 1 Abundance of dominant bacterial
phyla in the gut microbiota of higher ter-
mites. Numbers below the heatmap cor-
respond to hosts in Table 1. The
cladogram represents the phylogenetic
relationship among the hosts and is
based on the latest multilocus analysis of
concatenated mitochondrial genes (Bour-
guignon et al. 2015). The subfamily
assignment is based on the taxonomy
proposed by Inward et al. (2007). The
letters above the heatmap indicate the
feeding groups: F, fungus-feeding; S,
soil-feeding; W, wood or grass; H,
humus; L, litter (see Table 1 for more
information on the hosts’ dietary prefer-
ences).
©2015 John Wiley & Sons Ltd
4A. MIKAELYAN ET AL.
bacterial lineages among certain termite feeding groups
(Fig. 4). With only a few exceptions, the reads classified
as Spirochaetes (assigned to the genus-level lineages
Treponema Ia, Ic or If) decreased in abundance with
increasingly humified host diets. Treponema Ia was most
abundant in wood/grass feeders, and more abundant
in soil feeders than in humus/litter feeders (Fig. 4;
Table S2, Supporting information). The relative abun-
dance of Treponema Ic rose from 0.1% to 1.3% in the
fungus-feeding termites, through 0.252.1% in the soil
feeders, to as high as 16.0% in humus/litter feeders.
The highest abundance of Treponema Ic, however, was
observed among termites feeding on sound lignocellu-
lose, where the reads accounted for 25.148.7% of the
communities; an exception is the grass-feeding
Trinervitermes sp. (4.2% of the community). Also Tre-
ponema If was significantly more abundant among the
wood/grass feeders than the other diet groups, with
the exception of the wood-feeding Nasutitermes takasa-
goensis.
Our assessment of the gut communities at greater tax-
onomic depth revealed patterns of preferential associa-
tion of genus-level groups in Fibrobacteres and TG3
phylum with particular host lineages (Fig. 4). Subcluster
Ia of Fibrobacteres and Subcluster IIIb of TG3 phylum
were preferentially associated with Microcerotermes spp.
and Termes spp., both members of Termitinae. Contrast-
ingly, Subcluster Ib (of Fibrobacteres) and Subcluster
IIIa (of TG3 phylum) were found associated with Nasu-
titermes corniger and Trinervitermes sp., both members of
Nasutitermitinae. Reads from Cornitermes sp., a member
of Syntermitinae, were classified to Subcluster IIIb,
while those from Macrotermes sp. that were assigned to
Fibrobacteres could be binned to genus-level clusters
within ‘Cockroach cluster II’.
One of the most predominant genus-level taxa among
fungus-feeding termites was ‘Alistipes II’ (family:
Rikenellaceae), which represented 619% of the reads
obtained from the three Macrotermitinae but was absent
from most other host species (Fig. 4; Table S2,
Fig. 2 Principal coordinate analysis (PCoA) of pairwise distances among bacterial communities based on the BrayCurtis and Uni-
Frac metrics. Each data point in the plots represents the community harboured by a given termite species; colours indicate the differ-
ent host subfamilies. For more details on the samples, see Table 1.
Fig. 3 Hierarchical cluster analysis of pairwise distances
between bacterial communities based on the BrayCurtis met-
ric. Nodes in the dendrogram that are strongly supported by
high approximately uniform probability values are marked by
open (>70%) and closed (>90%) circles.
©2015 John Wiley & Sons Ltd
DIET SHAPES TERMITE GUT COMMUNITY STRUCTURE 5
Supporting information). In addition, the fungus feeders
were characterized by the abundant presence of several
genus-level groups within Lachnospiraceae and
Ruminococcaceae. The former include the genus-level
‘Gut cluster 13’ and ‘Ca. Arthromitus’, which were rep-
resented also in other feeding groups and most abun-
dant among the soil feeders (Fig. 4).
An analysis of the representative phylotypes from
selected genus-level lineages in the Treponema I clade
revealed that the highly divergent phylotypes clustered
according to the subfamilies of their respective host
species (e.g. Treponema Ia, Ic; Fig. 5A). A tendency of
the phylotypes to cluster by host subfamily was
observed also in the Fibrobacteres (Cluster I) and the
related TG3 phylum (Subcluster III) (Fig. 5B). However,
such host-specific clustering was observed mostly with
the dominant phylotypes, whereas many of the rarer
phylotypes clustered with sequences from other
subfamilies (Fig. S2, Supporting information).
Discussion
Previous studies have attempted to assess in how far
host phylogeny or dietary specialization of termites
determines the composition of their gut microbiota
(Colman et al. 2012; Dietrich et al. 2014; Rahman et al.
2015). However, because dietary diversification of
higher termites is part of their evolutionary history, the
influence of diet on gut community structure cannot be
easily disentangled from that of host phylogeny (Brune
& Dietrich 2015). Dietary diversification apparently
involved concerted adaptations in mandible morphol-
ogy (Donovan & Jones 2000), intestinal anatomy (Noirot
2001) and physicochemical gut conditions (Bignell &
Eggleton 1995; Brune et al. 1995; Brune & K
uhl 1996).
At the same time, there is considerable phylogenetic
conservatism in the dietary habits of closely related spe-
cies, for example wood-feeding within the genus Micro-
cerotermes, and soil-feeding within the ‘Cubitermes
group’ (Bourguignon et al. 2015).
Termite guts are small but complex ecosystems with
a wide range of physicochemically distinct microhabi-
tats, the distribution of which strongly reflects adapta-
tions to different diets (Brune & Dietrich 2015). In this
section, we discuss how differences in the availability
of these microhabitats and the dynamics of ecological
niches for the gut microbiota in the course of dietary
diversification can explain the differences in the distri-
bution and abundance of many bacterial lineages and
provide tenable ecological and evolutionary explanations
Fig. 4 Abundance of selected bacterial genus-level groups in higher termites. Numbers below the heatmap correspond to hosts in
Table 1; colours of circles represent the different subfamilies of higher termites (see Fig. 1 legend for colour key). The letters above
the heatmap indicate the different feeding groups: F, fungus-feeding; S, soil-feeding; W, wood or grass; H, humus; L, litter. See
Tables S2 and S3 (Supporting information) for a more detailed taxonomic overview of the communities.
©2015 John Wiley & Sons Ltd
6A. MIKAELYAN ET AL.
that reconcile the artificial dichotomy in the effects of
diet and host phylogeny on gut community structure in
higher termites.
Bacterial communities in wood feeders
The remarkable convergence of bacterial community
structure among wood-feeding Nasutitermitinae and
Termitinae serves to illustrate how diet can engineer
major changes in the gut microbiota through a redistri-
bution of niches and microhabitats in the gut. In the gut
of wood-feeding ‘lower termites’, microbial niches con-
nected to lignocellulose digestion are mostly occupied
by gut flagellates, which sequester wood particles in
their digestive vacuoles (Brugerolle & Radek 2006; Ni &
Tokuda 2013). In higher termites, the loss of flagellates
opened up these niches to fibre-digesting bacteria
(Brune & Dietrich 2015). This hypothesis is sparked by
the upshift in abundance in wood-feeding higher ter-
mites of bacterial lineages from the Fibrobacteres and
the TG3 phylum, which are extremely rare in the gut
microbiota of ‘lower termites’ and in higher termites of
other feeding groups (Hongoh et al. 2006; Dietrich et al.
2014; Rahman et al. 2015).
The deep taxon sampling of higher termites and the
highly resolved classification of the present study
revealed that bacterial lineages from Termite cluster I
(phylum: Fibrobacteres), Termite cluster III (TG3 phy-
lum) and the Treponema clusters Ic and If (phylum: Spir-
ochaetes) are more abundant in wood-feeding
representatives than in other feeding groups. Members
of these bacterial lineages dominate the cellulolytic
community that is specifically associated with the wood
particles in the hindgut of Nasutitermes spp. (Mikaelyan
et al. 2014), and are also abundantly represented in the
guts of other wood-feeding termites (Hongoh et al.
2006; Warnecke et al. 2007; He et al. 2013). The high
abundance of these cellulose-digesting, fibre-associated
bacterial lineages among wood and litter feeders identi-
fies the important structuring role played by both niche
(cellulose degradation) and microhabitat (wood fibres)
selection in the guts of higher termites.
Bacterial communities in litter, humus and soil feeders
One of the most notable adaptations to humus or soil
based diet is the increased alkalinity of the gut fluid,
particularly in the first-proctodeal compartment of the
(A) (B)
Fig. 5 Maximum-likelihood tree of 16S rRNA phylotypes from the different subfamilies of higher termites (see Fig. 1 legend for col-
our key and Fig. S2, Supporting information for tip labels). (A) Treponema Iac genus-level groups. (B) Phylum Fibrobacteres and
TG3. Size of the circles at the tips of the tree is proportional to the relative abundance of the phylotypes in the respective hosts. Col-
ours of the circles correspond to different subfamilies of higher termites.
©2015 John Wiley & Sons Ltd
DIET SHAPES TERMITE GUT COMMUNITY STRUCTURE 7
hindgut (Bignell & Eggleton 1995; Brune & K
uhl 1996).
We observed the association of several genus-level lin-
eages among Ruminococcaceae and Lachnospiraceae
(phylum Firmicutes) that are specifically enriched in the
alkaline first-proctodeal (P1) compartments of wood
feeders (Thongaram et al. 2005; K
ohler et al. 2012),
humus feeders (Thongaram et al. 2005) and soil feeders
(Schmitt-Wagner et al. 2003), clearly suggesting a prefer-
ence of these clostridial lineages for alkaline habitats in
the gut. Moreover, in comparison with the generally
tubular compartments of wood feeders, humus- and
soil-feeding termites are characterized by dilated P1
compartments that probably further magnify the rela-
tive abundance of these alkali-adapted clostridia in their
hindguts. Interestingly, guts of humivorous Pachnoda
scarab beetle larvae possess alkaline midguts (Lemke
et al. 2003), and many phylotypes in clone libraries from
Pachnoda spp. (Egert et al. 2003; Andert et al. 2010) have
been found to cluster in close phylogenetic neighbour-
hood of clones from P1 hindgut compartments of
humivorous higher termites (Schmitt-Wagner et al.
2003; Thongaram et al. 2005). The convergent enrich-
ment of related bacterial lineages in distantly related
orders of insects further underscores the importance of
habitat selection in the structuring of gut communities
in analogous gut environments, both characterized by
elevated intestinal pH.
Although the exact role of these alkali-adapted clos-
tridia remains unknown, their presence in humus- and
soil-feeding termite species may suggest an important
role in the digestion of peptides. This hypothesis is sup-
ported by the presence of alkaline proteolytic activity
(Ji & Brune 2005) and the accumulation of large
amounts of ammonia (Ngugi & Brune 2012) in the
anterior hindgut of soil feeders. Several genes encoding
proteases in the metagenome of the humivorous (dung-
feeding) Amitermes wheeleri have been binned to the
Clostridiales (He et al. 2013). Interestingly, the distribu-
tion patterns of genus-level groups within the Lach-
nospiraceae and Ruminococcaceae observed in
A. wheeleri are quite similar to those in termites feeding
on litter or humus (see Table S4, Supporting informa-
tion), suggesting important roles of these lineages in the
digestion of peptides in dung and humus.
In addition to the proteolytic activities (Ji & Brune
2005), the hindgut content of soil-feeding termites also
shows small amounts of xylanolytic and cellulolytic
activities (Rouland & Chararas 1986), which have been
suggested to be of bacterial origin (Brauman 2000).
Again, the abundance of clostridial genes encoding
xylanases in the hindgut metagenome and metatran-
scriptome of A. wheeleri (He et al. 2013) suggests that
the clostridial lineages in the soil-feeding and humus-
feeding termites investigated in the current study play
a role also in the digestion of residual polysaccharides
of lignocellulose.
Gut content analyses have also shown that members
of the humus-feeding termites consume a greater pro-
portion of plant material and/or wood fibres than the
true soil feeders (Sleaford et al. 1996; Donovan et al.
2001), and the composition of artificial lignocellulosic
diets strongly affects community structure in wood-
feeding N. takasagoensis, particularly the abundance of
the phyla Fibrobacteres, Spirochaetes and TG3 (Miyata
et al. 2007). Therefore, differences in the proportion of
plant material in the diets of higher termites could
explain the higher abundance of fibre-associated bacte-
rial lineages in humus feeders compared to soil feeders,
that is Treponema clusters Ic and If (Spirochaetes), Ter-
mite cluster I (Fibrobacteres) and Termite cluster III
(TG3 phylum). Considerable differences in the distribu-
tion of these lineages even among the humus feeders
agree with differences in the degree of humification in
their lignocellulosic diets (Bourguignon et al. 2011).
Bacterial communities in fungus feeders
The similarity in gut community structure we observed
among fungus-feeding termites from different genera
confirms the presence of a core set of bacterial lineages
that are either absent or rare in the communities of
other higher termites (Dietrich et al. 2014; Otani et al.
2014). This similarity in community structure among
the fungus feeders is driven by their highly specialized
diet, which has been shown to include fungal mycelia,
in addition to wood or litter (Donovan et al. 2001). The
shift to a relatively proteinaceous diet among the fun-
gus feeders could be also responsible for the conver-
gence of community structure with that of omnivorous
cockroaches (Dietrich et al. 2014), particularly in the
preferential enrichment of genus-level lineages such as
Alistipes clusters II, III and IV in both distantly related
insect groups (Mikaelyan et al. 2015). However, the role
of the Alistipes clusters in the guts of the termites is still
unknown (Poulsen et al. 2014). As another result of pos-
sible habitat selection, Macrotermes sp. harbours Cock-
roach cluster II of Fibrobacteres, a lineage so far
detected only among litter-feeding cockroaches (Mikael-
yan et al. 2015), which could again be a consequence of
dietary similarities between the two insect groups.
Evidence for co-evolution or cospeciation?
Our analyses clearly identify diet as the primary deter-
minant of community structure in higher termites. Nev-
ertheless, each diet group conceals a significant signal
of host taxonomy, which is reflected in the clustering of
bacterial phylotypes by termite subfamily (Mikaelyan
©2015 John Wiley & Sons Ltd
8A. MIKAELYAN ET AL.
et al. 2015; Fig. 5). This explains the previously reported
patterns of host phylogeny in the bacterial gut micro-
biota in a broader selection of termite and cockroach
hosts (Dietrich et al. 2014; Rahman et al. 2015), which
led to the suggestion of vertical transmission being the
primary force shaping the composition of the termite
gut microbiota (Rahman et al. 2015). However, even a
thorough assessment of phylogenetic similarity of gut
communities in different hosts (e.g. by the unweighted
UniFrac metric) may be misleading, because not all
members of the gut community are necessarily co-
evolving with the host (Brune & Dietrich 2015).
Using a wider selection of host taxa that comprises
members of the same feeding groups from different
subfamilies, we were able to demonstrate that the co-
evolutionary signal in community structure is masked
by an overwhelming signal of host diet. Although a
preferential association of certain bacterial lineages with
particular host groups is evident already in the classifi-
cation (Fig. 4) and phylogenetic analyses of short reads
(Fig. 5), the co-evolutionary signal is better resolved in
phylogenetic analyses of full-length 16S rRNA
sequences. For instance, detailed analyses of Fibrobac-
teres and the TG3 phylum revealed several mono-
phyletic, genus-level lineages that are specifically
associated with termites of the same subfamilies
(Hongoh et al. 2006; Mikaelyan et al. 2015).
Although co-evolution may lead to cospeciation, the
degree of cocladogenesis will depend on the intimacy
of interaction among the partners. In contrast to the
hereditary symbioses between insects and their intracel-
lular bacteria (e.g. Buchnera,Blattabacterium), which have
resulted in perfectly congruent phylogenies of symbiont
and host (Clark et al. 2000; Lo et al. 2003), the vertical
inheritance of gut microbiota is less reliable (Brune &
Dietrich 2015). The exchange of stomodeal or proc-
todeal fluids among nestmates (trophallaxis) and the
consumption of faeces (coprophagy) or nest material
(which is composed of faeces) increase the possibility of
an uptake of bacteria from the environment, including
gut microbiota of other termite species (Nalepa et al.
2001).
However, the contribution of horizontal transfer of
gut microbiota among different termite lineages is
apparently not very large. Already an early study of
several Reticulitermes and Microcerotermes species indi-
cated that the bacterial communities in the guts of con-
generic termites are very similar, irrespective of
geographical location (Hongoh et al. 2005). Also the
clustering patterns in our larger and more diverse data
set (Fig. 2) cannot be explained by the geographical ori-
gin of the samples (Table S5, Supporting information).
Nonetheless, it would be important to investigate
whether sympatric termite species with more frequent
opportunities to exchange gut microbiota, for example
inquilines such as Velocitermes spp. that cohabit the ter-
mitaria of other termites (Florencio et al. 2013), share
gut microbiota with their respective host. A horizontal
transmission of microbial symbionts, for example dur-
ing territory defence, has been used to explain high
similarities in the flagellate assemblages of Termopsidae
and certain Rhinotermitidae (Reticulitermes spp.), which
sometimes occur even within the same log of wood
(Kitade 2004).
Although the phylogenetic continuity in the identity
of other microbial lineages that occupy important niches
in the gut is in agreement with the vertical transmission
of symbionts, some of the co-evolutionary patterns
observed at higher taxonomic levels may merely be the
result of a diffuse selective pressure that the gut habitat
exerts on microbial lineages that are transferred via the
environment (Won et al. 2003; Fontanez & Cavanaugh
2014). Convincing evidence for cospeciation cannot be
obtained from the analysis of short reads but will
require the highly resolved phylogenetic analysis of
candidate symbionts from a sufficient number of closely
related termites.
Acknowledgements
We thank Katja Meuser for excellent laboratory assistance. This
study was funded by a grant of the Deutsche Forschungsge-
meinschaft (DFG) in the Collaborative Research Center SFB 987
and the LOEWE Programme of the State of Hessen (Synmikro).
AM was supported by the Synmikro Post-Doc Programme. CD
received a scholarship from the International Max Planck
Research School for Environmental, Cellular and Molecular
Microbiology (IMPRS-MIC). MP was supported by a STENO
grant from the Danish Agency for Science, Technology and
Innovation.
References
Andert J, Marten A, Brandl R, Brune A (2010) Inter- and
intraspecific comparison of the bacterial assemblages in the
hindgut of humivorous scarab beetle larvae (Pachnoda spp.).
FEMS Microbiology Ecology,74, 439449.
Bignell DE, Eggleton P (1995) On the elevated intestinal pH of
higher termites (Isoptera: Termitidae). Insectes Sociaux,42,
5769.
Bignell DE, Roisin Y, Lo N (2011) Global biogeography of ter-
mites. In: Biology of Termites: A Modern Synthesis(eds Bignell
DE, Roisin Y, Lo N), pp. 477498. Springer Netherlands, Do-
drecht.
Bourguignon T,
Sobotn
ık J, Lepoint G et al. (2011) Feeding ecol-
ogy and phylogenetic structure of a complex neotropical ter-
mite assemblage, revealed by nitrogen stable isotope ratios.
Ecological Entomology,36, 261269.
Bourguignon T, Lo N, Cameron SL et al. (2015) The evolution-
ary history of termites as inferred from 66 mitochondrial
genomes. Molecular Biology and Evolution,32, 406421.
©2015 John Wiley & Sons Ltd
DIET SHAPES TERMITE GUT COMMUNITY STRUCTURE 9
Brauman A (2000) Effect of gut transit and mound deposit
on soil organic matter transformations in the soil feeding
termite: a review. European Journal of Soil Biology,36, 117
125.
Brugerolle G, Radek R (2006) Symbiotic protozoa of termites.
In: Intestinal Microorganisms of Termites and Other Invertebrates
(eds K
onig H, Varma A), pp. 243269. Springer, Berlin, Hei-
delberg.
Brune A (2014) Symbiotic digestion of lignocellulose in termite
guts. Nature Reviews Microbiology,12, 168180.
Brune A, Dietrich C (2015) The gut microbiota of termites:
digesting the diversity in the light of ecology and evolution.
Annual Review of Microbiology,69, in press. doi: 10.1146/an-
nurev-micro-092412-155715.
Brune A, K
uhl M (1996) pH profiles of the extremely alkaline
hindguts of soil-feeding termites (Isoptera: Termitidae) deter-
mined with microelectrodes. Journal of Insect Physiology,42,
11211127.
Brune A, Ohkuma M (2011) Role of the termite gut microbiota
in symbiotic digestion. In: Biology of Termites(eds Bignell DE,
Roisin Y, Lo N), pp. 439475. Springer, Dodrecht.
Brune A, Emerson D, Breznak JA (1995) The termite gut micro-
flora as an oxygen sink: microelectrode determination of oxy-
gen and pH gradients in guts of lower and higher termites.
Applied and Environmental Microbiology,61, 26812687.
Clark MA, Moran NA, Baumann P et al. (2000) Cospeciation
between bacterial endosymbionts (Buchnera) and a recent
radiation of aphids (Uroleucon) and pitfalls of testing for
phylogenetic congruence. Evolution,54, 517525.
Colman DR, Toolson EC, Takacs-Vesbach CD (2012) Do diet
and taxonomy influence insect gut bacterial communities?
Molecular Ecology,21, 51245137.
Daigle D, Simen BB, Pochart P (2011) High-throughput
sequencing of PCR products tagged with universal primers
using 454 life sciences systems. Current Protocols in Molecular
Biology,96,114.
Degnan P, Ochman H (2012) Illumina-based analysis of micro-
bial community diversity. The ISME Journal,6, 183194.
Dietrich C, Brune A (2014) The complete mitogenomes of six
higher termite species reconstructed from metagenomic data-
sets (Cornitermes sp., Cubitermes ugandensis,Microcerotermes par-
vus,Nasutitermes corniger,Neocapritermes taracua, and Termes
hospes). Mitochondrial DNA. doi: 10.3109/19401736.2014.987257.
Dietrich C, K
ohler T, Brune A (2014) The cockroach origin of
the termite gut microbiota: patterns in bacterial community
structure reflect major evolutionary events. Applied and Envi-
ronmental Microbiology,80, 22612269.
Donovan SE, Jones DT (2000) Morphological phylogenetics of
termites (Isoptera). Biological Journal of the Linnean Society,70,
467513.
Donovan SE, Eggleton P, Bignell DE (2001) Gut content analy-
sis and a new feeding group classification of termites. Ecolog-
ical Entomology,26, 356366.
Edgar RC (2013) UPARSE: highly accurate OTU sequences
from microbial amplicon reads. Nature Methods,10, 996998.
Egert M, Wagner B, Lemke T et al. (2003) Microbial community
structure in midgut and hindgut of the humus-feeding larva
of Pachnoda ephippiata (Coleoptera: Scarabaeidae). Applied and
Environmental microbiology,69, 66596668.
Eggleton P, Tayasu I (2001) Feeding groups, lifetypes and the
global ecology of termites. Ecological Research,16, 941960.
Florencio DF, Marins A, Rosa CS et al. (2013) Diet segregation
between cohabiting builder and inquiline termite species.
PLoS ONE,8, e66535.
Fontanez KM, Cavanaugh CM (2014) Evidence for horizontal
transmission from multilocus phylogeny of deep-sea mussel
(Mytilidae) symbionts. Environmental Microbiology,16,
36083621.
Gontijo TA, Domingos DJ (1991) Guild distribution of some
termites from cerrado vegetation in south-east Brazil. Journal
of Tropical Ecology,7, 523529.
He S, Ivanova N, Kirton E et al. (2013) Comparative metage-
nomic and metatranscriptomic analysis of hindgut paunch
microbiota in wood- and dung-feeding higher termites. PLoS
ONE,8, e61126.
Hongoh Y, Deevong P, Inoue T et al. (2005) Intra- and inter-
specific comparisons of bacterial diversity and community
structure support coevolution of gut microbiota and termite
host. Applied and Environmental Microbiology,71, 65906599.
Hongoh Y, Deevong P, Hattori S et al. (2006) Phylogenetic
diversity, localization, and cell morphologies of members of
the candidate phylum TG3 and a subphylum in the phylum
Fibrobacteres, recently discovered bacterial groups dominant
in termite guts. Applied and Environmental Microbiology,72,
67806788.
Inward DJG, Vogler AP, Eggleton P (2007) A comprehensive
phylogenetic analysis of termites (Isoptera) illuminates key
aspects of their evolutionary biology. Molecular Phylogenetics
and Evolution,44, 953967.
Ji R, Brune A (2005) Digestion of peptidic residues in humic
substances by an alkali-stable and humic-acid-tolerant prote-
olytic activity in the gut of soil-feeding termites. Soil Biology
and Biochemistry,37, 16481655.
Kitade O (2004) Comparison of symbiotic flagellate faunae
between termites and a wood-feeding cockroach of the genus
Cryptocercus.Microbes and Environments,19, 215220.
K
ohler T, Dietrich C, Scheffrahn RH, Brune A (2012) High-res-
olution analysis of gut environment and bacterial microbiota
reveals functional compartmentation of the gut in wood-
feeding higher termites (Nasutitermes spp.). Applied and Envi-
ronmental Microbiology,78, 46914701.
Krishna K, Grimaldi DA, Krishna V, Engel MS (2013) Treatise
on the Isoptera of the world. Bulletin of the American Museum
of Natural History. American Museum of Natural History,
New York.
Lemke T, Stingl U, Egert M, Friedrich MW, Brune A (2003)
Physicochemical conditions and microbial activities in the
highly alkaline gut of the humus-feeding larva of Pachnoda
ephippiata (Coleoptera: Scarabaeidae). Applied and Environmen-
tal Microbiology,69, 66506658.
Lo N, Bandi C, Watanabe H, Nalepa C, Beninati T (2003) Evi-
dence for cocladogenesis between diverse dictyopteran lin-
eages and their intracellular endosymbionts. Molecular
Biology and Evolution,20, 907913.
Lozupone CA, Hamady M, Kelley ST, Knight R (2007) Quanti-
tative and qualitative bdiversity measures lead to different
insights into factors that structure microbial communities.
Applied and Environmental Microbiology,73, 15761585.
Meyer F, Paarmann D, D’Souza M et al. (2008) The metage-
nomics RAST servera public resource for the automatic
phylogenetic and functional analysis of metagenomes. BMC
Bioinformatics,9, 386.
©2015 John Wiley & Sons Ltd
10 A. MIKAELYAN ET AL.
Mikaelyan A, Strassert JFH, Tokuda G, Brune A (2014) The
fibre-associated cellulolytic bacterial community in the hind-
gut of wood-feeding higher termites (Nasutitermes spp.). En-
vironmental Microbiology,16, 27112722.
Mikaelyan A, K
ohler T, Lampert N et al. (2015) Classifying the
bacterial gut microbiota of termites and cockroaches: a
curated phylogenetic reference database (DictDb). Systematic
and Applied Microbiology, in press. doi: 10.1016/
j.syapm.2015.07.004.
Miyata R, Noda N, Tamaki H et al. (2007) Influence of feed
components on symbiotic bacterial community structure in
the gut of the wood-feeding higher termite Nasutitermes
takasagoensis.Bioscience, Biotechnology, and Biochemistry,71,
12441251.
Nalepa CA, Bignell DE, Bandi C (2001) Detritivory, copro-
phagy, and the evolution of digestive mutualisms in Dicty-
optera. Insectes Sociaux,48, 194201.
Ngugi DK, Brune A (2012) Nitrate reduction, nitrous oxide for-
mation, and anaerobic ammonia oxidation to nitrite in the
gut of soil-feeding termites (Cubitermes and Ophiotermes
spp.). Environmental Microbiology,14, 860871.
Ni J, Tokuda G (2013) Lignocellulose-degrading enzymes from
termites and their symbiotic microbiota. Biotechnology
Advances,31, 838850.
Noirot C (2001) The gut of termites (Isoptera). Annales de la
Soci
et
e Entomologique de France,37, 431471.
Oksanen JF, Blanchet G, Kindt R et al. (2015) vegan: Community
ecology package. R package version 2.3-0. Available from
http://CRAN.R-project.org/package=vegan.
Otani S, Mikaelyan A, Nobre T et al. (2014) Identifying the core
microbial community in the gut of fungus-growing termites.
Molecular Ecology,23, 46314644.
Paradis E, Claude J, Strimmer K (2004) APE: analyses of phylo-
genetics and evolution in R language. Bioinformatics,20,
289290.
Paul K, Nonoh JO, Mikulski L, Brune A (2012) “Methanoplas-
matales”, Thermoplasmatales-related archaea in termite guts
and other environments, are the seventh order of methano-
gens. Applied and Environmental Microbiology,78, 82458253.
Poulsen M (2015) Towards an integrated understanding of the
consequences of fungus domestication on the fungus-grow-
ing termite gut microbiota. Environmental Microbiology. doi:
10.1111/1462-2920.12765.
Poulsen M, Hu H, Li C et al. (2014) Complementary symbiont
contributions to plant decomposition in a fungus-farming
termite. Proceedings of the National Academy of Sciences of the
USA,111, 1450014505.
Price MN, Dehal PS, Arkin AP (2009) FastTree: computing
large minimum evolution trees with profiles instead of a dis-
tance matrix. Molecular Biology and Evolution,26, 16411650.
R Core Team (2015) R: A Language and Environment for Statisti-
cal Computing. R Foundation for Statistical Computing, Vi-
enna, Austria. Available from http://www.R-project.org/.
Rahman NA, Parks DH, Willner DL et al. (2015) A molecular
survey of Australian and North American termite genera
indicates that vertical inheritance is the primary force shap-
ing termite gut microbiomes. Microbiome,3,116.
Rouland C, Chararas C (1986)
Etude compar
ee des osidases de
trois esp
eces de termites africains
ar
egime alimentaire
diff
erent. Comptes rendus de l’Acad
emie des sciences,302, 341
345.
Schloss PD, Westcott SL, Ryabin T et al. (2009) Introducing
mothur: open-source, platform-independent, community-
supported software for describing and comparing microbial
communities. Applied and Environmental Microbiology,75,
75377541.
Schmitt-Wagner D, Friedrich MW, Wagner B, Brune A (2003)
Phylogenetic diversity, abundance, and axial distribution of
Bacteria in the intestinal tract of two soil-feeding termites
(Cubitermes spp.). Applied and Environmental Microbiology,69,
60076017.
Sleaford F, Bignell DE, Eggleton P (1996) A pilot analysis of
gut contents in termites from the Mbalmayo Forest Reserve,
Cameroon. Ecological Entomology,21, 279288.
Suzuki R, Shimodaira H (2006) Pvclust: an R package for
assessing the uncertainty in hierarchical clustering. Bioinfor-
matics,22, 15401542.
Thongaram T, Hongoh Y, Kosono S et al. (2005) Comparison
of bacterial communities in the alkaline gut segment
among various species of higher termites. Extremophiles,9,
229238.
Wang Q, Garrity GM, Tiedje JM, Cole JR (2007) Na
ıve Bayesian
classifier for rapid assignment of rRNA sequences into the
new bacterial taxonomy. Applied and Environmental Microbiol-
ogy,73, 52615267.
Warnecke F, Luginb
uhl P, Ivanova N et al. (2007) Metagenomic
and functional analysis of hindgut microbiota of a wood-
feeding higher termite. Nature,450, 560565.
Won YJ, Hallam SJ, O’Mullan GD et al. (2003) Environmental
acquisition of thiotrophic endosymbionts by deep-sea mus-
sels of the genus Bathymodiolus.Applied and Environmental
Microbiology,69, 67856792.
A.M., T.K. and A.B. conceived and designed the experi-
ments. M.P. and D.S.-D. collected and identified termites.
A.M. and C.D. performed the experiments and analysed
the data. C.D. and D.S.-D. contributed to the writing the
manuscript. A.M. and A.B. wrote the manuscript.
Data accessibility
The de-replicated, quality-checked MiSeq data sets have
been archived at MG-RAST under the accession num-
bers 4639074.34639081.3. The COII nucleotide align-
ment and tree, the phylogenetic tree of OTU
representatives (encoding abundance and sample infor-
mation in the labels), the OTU table for each sample,
and the BrayCurtis and UniFrac dissimilarity matrices
are archived at Dryad (http://dx.doi.org/10.5061/
dryad.v46f0).
Supporting information
Additional supporting information may be found in the online ver-
sion of this article.
©2015 John Wiley & Sons Ltd
DIET SHAPES TERMITE GUT COMMUNITY STRUCTURE 11
Fig. S1 Maximum-likelihood tree constructed using COII DNA
sequences from 18 termite hosts included in this study in the
context of close relatives in public databases.
Fig. S2 Maximum-likelihood tree of 16S rRNA phylotypes from
the different subfamilies of higher termites.
Table S1 Characteristics of the 16S rRNA amplicon libraries
and the classification success obtained for each library with
DictDb v 3.0 at different taxonomic levels.
Table S2 Relative abundance of bacterial taxa in iTag and
pyrotag libraries of 16S rRNA amplicons from different higher
termites (arranged by host phylogeny).
Table S3 Relative abundance of bacterial taxa in the iTag and
pyrotag amplicon libraries of 16S rRNA genes from the guts of
different higher termites (arranged by host diet).
Table S4 Comparison of the distribution of genus-level clusters
in Lachnospiraceae and Ruminococcaceae (order Clostridiales)
in a library from Amitermes wheeleri (He et al. 2013) and the ter-
mites investigated in the current study.
Table S5 The correlation of the diet, host taxonomy and geo-
graphical origin with BrayCurtis and UniFrac distances tested
for significance tested with adonis.
Table S6 Pairwise KruskalWallis tests for significant differ-
ences in the distribution of bacterial lineages for different diet
groups of termites (F, fungus feeders; S, soil feeders; H,
humus/litter feeders; W, wood/grass feeders).
Table S7 Pairwise KruskalWallis tests for significant differ-
ences in the distribution of bacterial lineages for different com-
binations of taxonomic groups of termites (At, Apicotermitinae;
Mt, Macrotermitinae; Nt, Nasutitermitinae; Tt, Termitinae; St,
Syntermitinae).
©2015 John Wiley & Sons Ltd
12 A. MIKAELYAN ET AL.
... Their vertical mode of inheritance is supported by the observations that differences among termite gut prokaryotic and protist communities tend to increase as phylogenetic distances among termite hosts increase [1,132]. In addition, the diet of the termite host, which broadly correlates with the termite phylogeny [12], also shapes the termite gut microbial communities [36,88]. Whether the termite phylogeny is recapitulated by gut microbial functions, as it is recapitulated by the taxonomic composition of microbial communities, remains unknown. ...
... We compared the bacterial community composition and abundance inferred from marker gene data and 16S ribosomal RNA (rRNA) gene amplicon sequences of 74 termite gut samples ( Figure S2). Our estimates of the bacterial community composition and abundance using the 16S rRNA gene were consistent with previously published estimates using the same marker [16,36,88]. These estimates also showed similarities at the phylum level to that inferred from marker gene data. ...
... The tree represents a simplified time-calibrated phylogenetic tree reconstructed using host termite mitochondrial genome sequences. Prokaryotic taxa presenting significant phylogenetic autocorrelation with the host phylogeny at a 5% false discovery rate (FDR) are indicated with an asterisk (*p < 0.05; **p < 0.01) a bacterial family of Acidobacteriota previously reported from the gut of three humus-feeding termite species [88] and two species of Nasutitermitinae [36], is widely distributed in Nasutitermitinae, Foraminitermitinae, the Cephalotermes-group, and the Pericapritermes-group (Fig. 1). Altogether, our results demonstrate that termite phylogeny is remarkably predictive of the gut bacterial community composition, at least at the family level, as shown for termite gut protists [132]. ...
Article
Full-text available
Background Termites primarily feed on lignocellulose or soil in association with specific gut microbes. The functioning of the termite gut microbiota is partly understood in a handful of wood-feeding pest species but remains largely unknown in other taxa. We intend to fill this gap and provide a global understanding of the functional evolution of termite gut microbiota. Results We sequenced the gut metagenomes of 145 samples representative of the termite diversity. We show that the prokaryotic fraction of the gut microbiota of all termites possesses similar genes for carbohydrate and nitrogen metabolisms, in proportions varying with termite phylogenetic position and diet. The presence of a conserved set of gut prokaryotic genes implies that essential nutritional functions were present in the ancestor of modern termites. Furthermore, the abundance of these genes largely correlated with the host phylogeny. Finally, we found that the adaptation to a diet of soil by some termite lineages was accompanied by a change in the stoichiometry of genes involved in important nutritional functions rather than by the acquisition of new genes and pathways. Conclusions Our results reveal that the composition and function of termite gut prokaryotic communities have been remarkably conserved since termites first appeared ~ 150 million years ago. Therefore, the “world’s smallest bioreactor” has been operating as a multipartite symbiosis composed of termites, archaea, bacteria, and cellulolytic flagellates since its inception.
... Social insects including ants, termites, and honeybees can exhibit phenotype-specific differences in gut microbial community diversity and composition (Shimada et al. 2013;Tarpy et al. 2015;Inagaki and Matsuura 2016;Jones et al. 2018;Otani et al. 2019;Sinotte et al. 2020). In social insects, differences in gut microbial communities among social phenotypes, including reproductive and non-reproductive subcastes, tend to be strongly associated with differences in diet and environmental exposure (Hongoh et al. 2006;Mikaelyan et al. 2015;Jones et al. 2018). However, given the bidirectional relationship between the animal brain and gut microbiota (i.e., the microbiota-gut-brain axis), it is plausible that different social phenotypes harbor unique microbiota independent of differences in diet or exposure (Sherwin et al. 2019). ...
... However, environmental conditions were otherwise very similar; consequently, our common environment design allows us to compare the gut bacteria of these two host species more directly than past studies that use wild animal or captive zoo collections (Debebe et al. 2017). Many of the taxa enriched in the gut microbiota of naked mole-rats (members of the Fibrobacteres, Spirochaetes, and Elusimicrobia; Dataset S1) are also prominent members of the gut bacteria found in higher termites (Mikaelyan et al. 2015), which feed on diverse lignocellulosic diets. Thus, it seems likely that some of the observed differences in the gut microbiota between mice and naked mole-rats are also driven by diet differences in captivity and/or naked mole-rat traits associated with adaptation to a diet high in cellulosic material in the wild (Lacey and Sherman 1991;Dyer 1998). ...
Article
Full-text available
Group living occurs across the animal kingdom and can shape fundamental aspects of individual biology, including the microbes inhabiting the animal gut. The naked mole-rat, Heterocephalus glaber, exhibits extreme cooperative breeding (eusociality) and presents an ideal opportunity to study the effects of social structure on the mammalian gut microbiota. Within colonies reproduction is limited to few animals, while the remaining members are non-reproductive, form linear dominance hierarchies, and show evidence of task specialization with stable yet plastic behavioral phenotypes. Here, we sequenced the gut bacteria of naked mole-rats across 6 replicate captive colonies and longitudinally during a colony removal experiment. Colony had much larger effects than social phenotype or status on the diversity and composition of gut bacteria. Our longitudinal experiment revealed that over the course of 2 months, the gut bacteria of previously unfamiliar, newly paired animals did not become more similar. In comparison to mice housed in the same facility, the naked mole-rat gut microbiome exhibited substantial compositional differences including the near absence of Lactobacillaceae and the enrichment of Prevotellaceae. Our study provides new insight into the factors shaping gut bacterial communities in social animals and indicates a prominent role of social group membership but not social phenotype. Significance statement The social groups to which animals belong can be strong determinants of the composition of their associated gut microbial communities (i.e., gut microbiome). However, less is known about the link between social status within animal groups and the gut microbiome. We studied this relationship in the naked mole-rat, Heterocephalus glaber, which exhibits extreme cooperative breeding (eusociality). Animals within naked mole-rat colonies show clear social hierarchies and evidence of social phenotypes. Across replicate colonies, we found that colony but not social phenotype or status had large effects on the diversity and composition of gut bacteria. During a colony removal experiment, the gut bacteria of newly paired but previously unfamiliar animals did not become more similar over the course of 2 months. Our results show that when decoupled from differences in diet or exposure to microbes, within colony social roles and their accompanying unique physiology and social experience may have little effect on gut microbiota. In contrast, gut bacterial diversity and composition was strongly shaped by colony membership despite being exposed to identical environmental conditions.
... Some arthropod groups form obligatory internal mutualistic relationships with microbes. For instance, higher termites (Termitidae) harbor exclusively prokaryotic communities in their dilated hindguts [79]. By contrast, their actual community composition is largely determined by their diet and microhabitat [79], suggesting low taxonomic affiliation between prokaryote and host. ...
... For instance, higher termites (Termitidae) harbor exclusively prokaryotic communities in their dilated hindguts [79]. By contrast, their actual community composition is largely determined by their diet and microhabitat [79], suggesting low taxonomic affiliation between prokaryote and host. ...
Article
Coevolution has driven speciation and evolutionary novelty in functional traits across the Tree of Life. Classic coevolutionary syndromes such as plant–pollinator, plant–herbivore, and host–parasite have focused strongly on the fitness consequences during the lifetime of the interacting partners. Less is known about the consequences of coevolved traits for ecosystem-level processes, in particular their 'afterlife' legacies for litter decomposition, nutrient cycling, and the functional ecology of decomposers. We review the mechanisms by which traits resulting from coevolution between plants and their consumers, microbial symbionts, or humans, and between microbial decomposers and invertebrates, drive plant litter decomposition pathways and rates. This supports the idea that much of current global variation in the decomposition of plant material is a legacy of coevolution.
... The V3-V4 regions of the 16S rDNA gene of the bacterial community were amplified by using a bacteria-specific primer set, 341F (5′-CCT ACG GGNGGC WGC AG-3′) and 785R (5′-GAC TAC HVGGG TAT CTA ATC C-3′) and then tagged with hexameric barcodes (Mikaelyan et al. 2015). The PCR products were amplified and subsequently normalized. ...
... The expected species richness (Horner 2010), diversity (Huang et al. 2018), and evenness (Ifediegwu et al. 2015) of the bacterial communities were determined. Each OTU was assigned to respective taxonomic groups using Ribosomal Database Project (RDP) naïve Bayesian classifier implemented in MOTHUR maintaining a confidence limit of 80% and manually curated reference DictDb database v. 3.0 (Mikaelyan et al. 2015). The examination of the core taxa of bacteria residing in the gut of chironomid larvae was determined by considering a threshold of ≥ 60%. ...
Article
Full-text available
Chironomids are the most abundant aquatic insects in freshwater habitats that can survive in extreme conditions. In this study, as the microbiome provides extended genotype to the host to perform various functions, we explored the microbiota of the Chironomus circumdatus larvae to find out the putative role played by the symbiotic bacteria for the host. The metabarcoding analyses of the larvae revealed that the insect harbors 1771 phylotypes. Out of the various microbial communities found, the majority corresponded to the phyla Proteobacteria (52.59%) and Actinobacteria (20.56%), respectively. The midges also harbored Klebsiella (2.57%), Enterobacter (1.32%), Bacillus (2.29%), and Acinetobacter (2.13%) genera that are involved in detoxification of xenobiotics present in the water. The presence of radiation-resistant genera like Deinococcus, including bacterial species like radiodurans, a highly radiation-resistant bacterium, indicates its potential to support the host's ability to sustain in adverse environments. The functional profiling of the bacteria showed the relative abundance of many enzyme groups, such as transferases (40.62%), oxidoreductases (23.49%), and hydrolases (3.77%). The results indicate that the larvae harbor a considerable variety of bacteria that help the host adapt and survive in the polluted waters. The present study provides thorough insights into the microbiome of the C. circumdatus larvae that can be exploited for the bioremediation of certain pollutants through biomimetic strategies. It also gives us a wake-up call to take a good look at the guts of these disease-carrying insects' inabilities to spread deadly human diseases.
... Most Actinobacteria are widely distributed in water and soil. These bacteria are easily transmitted to birds, suggesting that the bacterial community structure of birds is influenced by external bacterial transmission (Mikaelyan et al., 2015). Actinobacteria, which dominant in waders, are closely related to host fiber intake (Lee et al., 2015). ...
Article
Full-text available
Gut microbiota plays a vital role in maintaining the health and immunity of wild birds. However, less is known about the comparison of fecal microbiota between different ecological groups of wild birds, particularly in the Yellow River National Wetland in Baotou, China, an important transit point for birds migrating all over the East Asia-Australian and Central Asian flyways. In this study, we characterized the fecal microbiota and potential microbial function in nine bird species of raptors, waders, and waterfowl using 16S rRNA gene amplicon sequencing to reveal the microbiota differences and interaction patterns. The results indicated that there was no significant difference in α-diversity, but a significant difference in β-diversity between the three groups of birds. The fecal bacterial microbiota was dominated by Firmicutes, Proteobacteria, Actinobacteria, and Bacteroidetes in all groups of birds. Furthermore, we identified five bacterial genera that were significantly higher in raptors, five genera that were significantly higher in waders, and two genera that were more abundant in waterfowl. The bacterial co-occurrence network results revealed 15 and 26 key genera in raptors and waterfowls, respectively. The microbial network in waterfowl exhibited a stronger correlation pattern than that in raptors. PICRUSt2 predictions indicated that fecal bacterial function was significantly enriched in the antibiotic biosynthesis pathway in all three groups. Metabolic pathways related to cell motility (bacterial chemotaxis and flagellar assembly) were significantly more abundant in raptors than in waders, whereas waders were enriched in lipid metabolism (synthesis and degradation of ketone bodies and fatty acid biosynthesis). The fecal microbiota in waterfowl harbored more abundant vitamin B6 metabolism, RNA polymerase, and tyrosine and tryptophan biosynthesis. This comparative study revealed the microbial community structure, microbial co-occurrence patterns, and potential functions, providing a better understanding of the ecology and conservation of wild birds. Future studies may focus on unraveling metagenomic functions and dynamics along with the migration routine or different seasons by metagenomics or metatranscriptomics.
... Cellulose as potential biomass can be naturally degraded by the bacteria in termite gut. Almost low-level termites are diet of wood (Hongoh 2011), while high-level termites are diet of lignocellulosic biomass, like wood, plant litter, grass litter, and dirt (Hongoh 2011;Brune 2014;Ferbiyanto et al. 2015;Mikaelyan et al. 2015). Recently, research regarding termite has significantly increased, because of their ability to degrade lignocellulose by bacteria symbionts in their gut, becoming specialist agents to hydrolyze cellulose biomass. ...
Article
Full-text available
The termite Macrotermes gilvus Hagen from Tanjung Senai, Inderalaya peatland region, is classified as a high-level termite. Termites like eusocial insects break down cellulosic biomass into glucose by bacteria that produce cellulolytic enzymes in their gut. In the present study, new species of cellulolytic bacteria from Macrotermes gilvus Hagen gut were determined by biomolecular assay. Bacterial isolates were isolated and purified by using DNA kit and the optical density of DNA bacterial isolates was obtained by using spectrophotometer nanodrop. A total of 24 bacterial isolates were amplified using PCR to determine the fragment of 16S rRNA gene, performed using BLAST-N program and compared with NCBI database. Result from the sequences of 16S rRNA gene showed that the new species were identify as
... Microbial communities of insect guts are affected by their trophic associations and encounter with microbial sources in their natural habitats and subsequently by gut physiology that determines the survival and colonization outcomes of ingested microbes (Engel & Moran, 2013;Nayduch & Burrus, 2017;Ravenscraft et al., 2019). Food choice significantly influences gut microbial community composition and diversity across many insect taxa including termites, wasps, honeybees, fruit flies, mosquitos, and butterflies both in natural habitats and controlled environments (Chandler et al., 2011;Colman et al., 2012;Kešnerová et al., 2020;Mikaelyan et al., 2015;Ravenscraft et al., 2019). For example, gut microbial diversity in detritivorous insect species, irrespective of species relatedness, is more similar compared to the same order or genus of insect species that are xylophagous (Colman et al., 2012). ...
Article
Adult house flies feed and breed in a variety of microbe‐rich habitats and serve as vectors for human and animal pathogens. To better understand their role in harbouring and disseminating bacteria, we characterized the composition and diversity of bacterial communities in the gut of female house flies collected from three different habitats in Kansas: agricultural (dairy farm), urban (business area dumpsters) and mixed (business located between residential and animal agriculture areas). Bacterial community composition and diversity were influenced more by the house flies' habitat than by sampling time. The most abundant taxa were also highly prevalent in the house flies collected from all three habitats, potentially representing a ‘core microbiome’ attributable to the fly's trophic and reproductive associations with substrates and food sources comprised of decaying matter and/or animal waste. Bacterial taxa associated with vertebrate guts/faeces and potential pathogens were highly abundant in agricultural fly microbial communities. Interestingly, taxa of potential pathogens were highly abundant in flies from the mixed and urban sites. House flies harboured diverse bacterial communities influenced by the habitat in which they reside, including potential human and animal pathogens, further bolstering their role in the dissemination of pathogens, and indicating their utility for pathogen surveillance. Adult female house fly gut bacterial communities were diverse and were shaped mostly by their habitat (agriculture, mixed and urban), and to a lesser extent by sampling time. Although house flies from all sites harboured ‘core bacterial communities’ acquired through filth feeding, individual house fly guts harboured unique sets of bacterial communities. House flies across all habitats harboured several potential human and animal pathogenic bacterial taxa supporting their role as reservoirs and disseminators of disease agents.
... Food is an important source of gut microorganisms [25,26], and host plants feeding could affect the species and functions of gut flora in herbivorous insects. The structure and diversity of gut microorganisms vary with host plants [27]. ...
Article
Full-text available
The fall armyworm (FAW), Spodoptera frugiperda, is one of the most important invasive species and causes great damage to various host crops in China. In this study, the diversity and function of gut bacteria in the 5th instar larvae of FAW fed on maize, wheat, potato and tobacco leaves were analyzed through 16S rRNA sequencing. A total of 1324.25 ± 199.73, 1313.5 ± 74.87, 1873.00 ± 190.66 and 1435.25 ± 139.87 operational taxonomic units (OTUs) from the gut of FAW fed on these four different host plants were detected, respectively. Firmicutes, Proteobacteria and Bacteroidetes were the most abundant bacterial phyla. Beta diversity analysis showed that the gut bacterial community structure of larvae fed on different host plants was significantly differentiated. At the genus level, the abundance of Enterococcus in larvae fed on wheat was significantly lower than those fed on the other three host plants. Enterobacter and ZOR0006 were dominant in FAW fed on tobacco leaves, and in low abundance in larvae fed on wheat. Interestingly, when fed on Solanaceae (tobacco and potato) leaves which contained relative higher levels of toxic secondary metabolites than Gramineae (wheat and maize), the genera Enterococcus, Enterobacter and Acinetobacter were significantly enriched. The results indicated that gut bacteria were related to the detoxification and adaptation of toxic secondary metabolites of host plants in FAW. Further analysis showed that replication, repair and nucleotide metabolism functions were enriched in the gut bacteria of larvae fed on tobacco and potato. In conclusion, the gut bacterial diversity and community composition in FAW larvae fed on different host plants showed significant differences, and the insect is likely to regulate their gut bacteria for adaptation to different host plants.
Article
Feeding habits were the primary factor affecting the gut bacterial communities in Ensifera. However, the interaction mechanism between the gut microbiota and feeding characteristics is not precisely understood. Here, the gut microbiota of Ensifera with diverse feeding habits was analyzed by shotgun metagenomic sequencing to further clarify the composition and function of the gut microbiota and its relationship with feeding characteristics. Our results indicate that under the influence of feeding habits, the gut microbial communities of Ensifera showed specific characteristics. Firstly, the gut microbial communities of the Ensifera with different feeding habits differed significantly, among which the gut microbial diversity of the herbivorous Mecopoda niponensis was the highest. Secondly, the functional genes related to feeding habits were in high abundance. Thirdly, the specific function of the gut microbial species in the omnivorous Gryllotalpa orientalis showed that the more diverse the feeding behavior of Ensifera, the worse the functional specificity related to the feeding characteristics of its gut microbiota. However, feeding habits were not the only factors affecting the gut microbiota of Ensifera. Some microorganisms’ genes, whose functions were unrelated to feeding characteristics but were relevant to energy acquisition and nutrient absorption, were detected in high abundance. Our results were the first to report on the composition and function of the gut microbiota of Ensifera based on shotgun metagenomic sequencing and to explore the potential mechanism of the gut microbiota’s association with diverse feeding habits.
Article
Diaphorina citri Kuwayama (Hemiptera: Liviidae) can cause severe damage to citrus plants, as it transmits Candidatus Liberibacter spp., a causative agent of Huanglongbing disease. Symbiotic bacteria play vital roles in the ecology and biology of herbivore hosts, thereby affecting host growth and adaptation. In our research, the effects of Rutaceous plants (i.e., Citrus reticulata cv. Shatangju, Citrus poonensis cv. Ponkan, Murraya paniculata (orange jasmine), Citrus limon (lemon), and Citrus sinensis (navel orange)) on the gut microbiota (GM) and microbial diversity of D. citri adults were investigated by 16S rRNA high-throughput sequencing. It was found that Proteobacteria dominated the GM communities. The gut microbe diversity was the highest in the ponkan-feeding population, and the lowest in the Shatangju-feeding population. The NMDS analysis revealed that there were obvious differences in the GM communities among the different hosts. PICRUSt function prediction indicated significant differences in host function, and those pathways were crucial for maintaining population reproduction, growth, development, and adaptation to environmental stress in D. citri. Our study sheds new light on the interactions between symbionts, herbivores, and host plants and expands our knowledge on host adaptation related to GM in D. citri.
Article
Full-text available
Termites and their microbial gut symbionts are major recyclers of lignocellulosic biomass. This important symbiosis is obligate but relatively open and more complex in comparison to other well-known insect symbioses such as the strict vertical transmission of Buchnera in aphids. The relative roles of vertical inheritance and environmental factors such as diet in shaping the termite gut microbiome are not well understood. The gut microbiomes of 66 specimens representing seven higher and nine lower termite genera collected in Australia and North America were profiled by small subunit (SSU) rRNA amplicon pyrosequencing. These represent the first reported culture-independent gut microbiome data for three higher termite genera: Tenuirostritermes, Drepanotermes, and Gnathamitermes; and two lower termite genera: Marginitermes and Porotermes. Consistent with previous studies, bacteria comprise the largest fraction of termite gut symbionts, of which 11 phylotypes (6 Treponema, 1 Desulfarculus-like, 1 Desulfovibrio, 1 Anaerovorax-like, 1 Sporobacter-like, and 1 Pirellula-like) were widespread occurring in ≥50% of collected specimens. Archaea are generally considered to comprise only a minority of the termite gut microbiota (<3%); however, archaeal relative abundance was substantially higher and variable in a number of specimens including Macrognathotermes, Coptotermes, Schedorhinotermes, Porotermes, and Mastotermes (representing up to 54% of amplicon reads). A ciliate related to Clevelandella was detected in low abundance in Gnathamitermes indicating that protists were either reacquired after protists loss in higher termites or persisted in low numbers across this transition. Phylogenetic analyses of the bacterial communities indicate that vertical inheritance is the primary force shaping termite gut microbiota. The effect of diet is secondary and appears to influence the relative abundance, but not membership, of the gut communities. Vertical inheritance is the primary force shaping the termite gut microbiome indicating that species are successfully and faithfully passed from one generation to the next via trophallaxis or coprophagy. Changes in relative abundance can occur on shorter time scales and appear to be an adaptive mechanism for dietary fluctuations.
Article
Digestive osidases from three higher termite species with different nutritive regimes, the fungus growing termite Macrotermes mulleri, the xylophageous termite Sphaerotermes sphaerothorax and the soil eating termite Thoracotermes macrothorax were investigated. Very clear differences in the set of osidases were noticed between the three species.
Book
Biology of Termites, a Modern Synthesis brings together the major advances in termite biology, phylogenetics, social evolution and biogeography made in the decade since Abe et al Termites: Evolution, Sociality, Symbioses, Ecology became the standard modern reference work on termite science. Building on the success of the Kluwer book, David Bignell, Yves Roisin and Nathan Lo have brought together in the new volume most of the world's leading experts on termite taxonomy, behaviour, genetics, caste differentiation, physiology, microbiology, mound architecture, distribution and control. Very strong evolutionary and developmental themes run through the individual chapters, fed by new data streams from molecular sequencing, and for the first time it is possible to compare the social organisation of termites with that of the social Hymenoptera, focusing on caste determination, population genetics, cooperative behaviour, nest hygiene and symbioses with microorganisms. New chapters have been added on termite pheromones, termites as pests of agriculture and on destructive invasive species, and new molecular and cladistic frameworks are presented for clarifying taxonomy, especially in the higher termites which dominate many tropical ecosystems. Applied entomologists, developmental and evolutionary biologists, microbial ecologists, sociobiologists and tropical agriculture specialists will all benefit from the new insights provided by this work. © Springer Science+Business Media B.V. 2011. All rights reserved.
Article
Recent developments in sequencing technology have given rise to a large number of studies that assess bacterial diversity and community structure in termite and cockroach guts based on large amplicon libraries of 16S rRNA genes. Although these studies have revealed important ecological and evolutionary patterns in the gut microbiota, classification of the short sequence reads is limited by the taxonomic depth and resolution of the reference databases used in the respective studies. Here, we present a curated reference database for accurate taxonomic analysis of the bacterial gut microbiota of dictyopteran insects. The Dictyopteran gut microbiota reference Database (DictDb) is based on the Silva database but was significantly expanded by the addition of clones from 11 mostly unexplored termite and cockroach groups, which increased the inventory of bacterial sequences from dictyopteran guts by 26%. The taxonomic depth and resolution of DictDb was significantly improved by a general revision of the taxonomic guide tree for all important lineages, including a detailed phylogenetic analysis of the Treponema and Alistipes complexes, the Fibrobacteres, and the TG3 phylum. The performance of this first documented version of DictDb (v. 3.0) using the revised taxonomic guide tree in the classification of short-read libraries obtained from termites and cockroaches was highly superior to that of the current Silva and RDP databases. DictDb uses an informative nomenclature that is consistent with the literature also for clades of uncultured bacteria and provides an invaluable tool for anyone exploring the gut community structure of termites and cockroaches. Copyright © 2015 Elsevier GmbH. All rights reserved.
Article
Termite guts harbor a dense and diverse microbiota that is essential for symbiotic digestion. The major players in lower termites are unique lineages of cellulolytic flagellates, whereas higher termites harbor only bacteria and archaea. The functions of the mostly uncultivated lineages and their distribution in different diet groups are slowly emerging. Patterns in community structure match changes in the biology of different host groups and reflect the availability of microbial habitats provided by flagellates, wood fibers, and the increasing differentiation of the intestinal tract, which also creates new niches for microbial symbionts. While the intestinal communities in the closely related cockroaches seems to be shaped primarily by the selective forces of microhabitat and functional niche, the social behavior of termites reduces the stochastic element of community assembly, which facilitates coevolution and may ultimately result in cospeciation.
Article
Isoptera (termites) are an ecologically important order, with both a high abundance and biomass in tropical ecosystems. However, there have been few phylogenetic hypotheses for termites, and we present here the first comprehensive cladistic analysis for the group. We analysed relationships between all seven termite families, including representatives of all known feeding group, plus a number of systematically critical taxa. Termite species richness is biased towards the higher termites (Termitidae), and our taxon sampling reflects this. Our analysis was based essentially on morphological characters (96 workers, 93 soldiers) plus seven biological characters. The cladistic analysis gave four equally parsimonious trees, representing two islands of topologies. The strict consensus tree is fully resolved for the higher termites, but less so for the lower termites. Overall there is low statistical support for the suggested topology, and this can be explained by the high incongruence between the data sets (worker, soldier and biological). This study highlights the particular problems of coding morphological characters in social insects with multiple castes. Without the input of additional data sets, e.g. alates, biological, behavioural and molecular, it will not be possible to obtain a well-supported termite phylogeny.