Phylogenetic Analysis of the Gut Bacterial Microflora of the Fungus-Growing Termite Odontotermes formosanus

We constructed a bacterial 16S rRNA gene clone library from the gut microbial community of O. formosanus and phylogenetically analyzed it in order to contribute to the evolutional study of digestive symbiosis and method development for termite control. After screening by restriction fragment length polymorphism (RFLP) analysis, 56 out of 280 clones with unique RFLP patterns were sequenced and phylogenetically analyzed. The representative phylotypes were affiliated to four phylogenetic groups, Firmicutes, the Bacteroidetes/Chlorobi group, Proteobacteria, and Actinobacteria of the domain Bacteira. No one clone affiliated with the phylum Spirochaetes was identified, in contrast to the case of wood-feeding termites. The phylogenetic analysis revealed that nearly half of the representative clones (25 phylotypes) formed monophyletic clusters with clones obtained from other termite species, especially with the sequences retrieved from fungus-growing termites. These results indicate that the presence of termite-specific bacterial lineages implies a coevolutional relationship of gut microbes and host termites.     

Molecular phylogenetic diversity of the bacterial community in the gut of the termite Coptotermes formosanus

The phylogenetic diversity of the bacterial community in the gut of the termite Coptotermes formosanus was investigated using a 16S rRNA gene clone library constructed by PCR. After screening by restriction fragment length polymorphism (RFLP) analysis, 49 out of 261 clones with unique RFLP patterns were sequenced and phylogenetically analyzed. Many of the clones (94%) were derived from Bacteroidales, Spirochaetes, and low G + C content gram-positive bacteria consisting of Clostridiales, Mycoplasmatales, Bacillales, and Lactobacillales. In addition, a few clones derived from Actinobacteria, Proteobacteria, Planctomycetes, Verrucomicrobia, and the candidate phylum "Synergistes" were also found. The most frequently identified RFLP type, BCf1-03, was assigned to the order Bacteroideales, and it constituted about 70% of the analyzed clones. The phylogenetic analysis revealed that the representative clones found in this study tended to form some clusters with the sequences cloned from the termite gut in several other studies, suggesting the existence of termite-specific bacterial lineages.     

Intra- and interspecific comparisons of bacterial diversity and community structure support coevolution of gut microbiota and termite host

We investigated the bacterial gut microbiota from 32 colonies of wood-feeding termites, comprising four Microcerotermes species (Termitidae) and four Reticulitermes species (Rhinotermitidae), using terminal restriction fragment length polymorphism analysis and clonal analysis of 16S rRNA. The obtained molecular community profiles were compared statistically between individuals, colonies, locations, and species of termites. Both analyses revealed that the bacterial community structure was remarkably similar within each termite genus, with small but significant differences between sampling sites and/or termite species. In contrast, considerable differences were found between the two termite genera. Only one bacterial phylotype (defined with 97% sequence identity) was shared between the two termite genera, while 18% and 50% of the phylotypes were shared between two congeneric species in the genera Microcerotermes and Reticulitermes, respectively. Nevertheless, a phylogenetic analysis of 228 phylotypes from Microcerotermes spp. and 367 phylotypes from Reticulitermes spp. with other termite gut clones available in public databases demonstrated the monophyly of many phylotypes from distantly related termites. The monophyletic "termite clusters" comprised of phylotypes from more than one termite species were distributed among 15 bacterial phyla, including the novel candidate phyla TG2 and TG3. These termite clusters accounted for 95% of the 960 clones analyzed in this study. Moreover, the clusters in 12 phyla comprised phylotypes from more than one termite (sub)family, accounting for 75% of the analyzed clones. Our results suggest that the majority of gut bacteria are not allochthonous but are specific symbionts that have coevolved with termites and that their community structure is basically consistent within a genus of termites.     

Intra- and Interspecific Comparisons of Bacterial Diversity and Community Structure Support Coevolution of Gut Microbiota and Termite Host

We investigated the bacterial gut microbiota from 32 colonies of wood-feeding termites, comprising four Microcerotermes species (Termitidae) and four Reticulitermes species (Rhinotermitidae), using terminal restriction fragment length polymorphism analysis and clonal analysis of 16S rRNA. The obtained molecular community profiles were compared statistically between individuals, colonies, locations, and species of termites. Both analyses revealed that the bacterial community structure was remarkably similar within each termite genus, with small but significant differences between sampling sites and/or termite species. In contrast, considerable differences were found between the two termite genera. Only one bacterial phylotype (defined with 97% sequence identity) was shared between the two termite genera, while 18% and 50% of the phylotypes were shared between two congeneric species in the genera Microcerotermes and Reticulitermes, respectively. Nevertheless, a phylogenetic analysis of 228 phylotypes from Microcerotermes spp. and 367 phylotypes from Reticulitermes spp. with other termite gut clones available in public databases demonstrated the monophyly of many phylotypes from distantly related termites. The monophyletic “termite clusters” comprised of phylotypes from more than one termite species were distributed among 15 bacterial phyla, including the novel candidate phyla TG2 and TG3. These termite clusters accounted for 95% of the 960 clones analyzed in this study. Moreover, the clusters in 12 phyla comprised phylotypes from more than one termite (sub)family, accounting for 75% of the analyzed clones. Our results suggest that the majority of gut bacteria are not allochthonous but are specific symbionts that have coevolved with termites and that their community structure is basically consistent within a genus of termites.     

Phylogenetic and morphological diversity of Bacteroidales members associated with the gut wall of termites

The microbial community adherent directly or indirectly to the gut wall of termites is distinct from that of the other habitats in the gut. The bacterial 16S rRNA genes were identified from the fractionated gut walls of two termite species, Hodotermopsis sjoestedti and Neotermes koshunensis, and compared with those previously identified from Reticulitermes speratus. Surprisingly, the bacterial constituents were almost entirely different among the termites at the phylotype level (the criterion of the phylotype was >97% nucleotide identity). Bacteria in the order Bacteroidales, which were commonly abundant symbionts on gut walls, were phylogenetically analyzed. They were dispersed in a number of clusters formed by phylotypes from the guts of various termites. In situ hybridization with probes specific for some phylotypes and a phylogenetic cluster detected the cells of several Bacteroidales members with a significant variety of cell morphology in the gut wall fractions, which reflects the phylogenetic diversity of this order.     

Comparison of the gut microbiota from soldier and worker castes of the termite Reticulitermes grassei

The bacterial microbiota from the whole gut of soldier and worker castes of the termite Reticulitermes grassei was isolated and studied. In addition, the 16S rDNA bacterial genes from gut DNA were PCR-amplified using Bacteria-selective primers, and the 16S rDNA amplicons subsequently cloned into Escherichia coli. Sequences of the cloned inserts were then used to determine closest relatives by comparison with published sequences and with sequences from our previous work. The clones were found to be affiliated with the phyla Spirochaetes, Proteobacteria, Firmicutes, Bacteroidetes, Actinobacteria, Synergistetes, Verrucomicrobia, and candidate phyla Termite Group 1 (TG1) and Termite Group 2 (TG2). No significant differences were observed with respect to the relative bacterial abundances between soldier and worker phylotypes. The phylotypes obtained in this study were compared with reported sequences from other termites, especially those of phylotypes related to Spirochaetes, Wolbachia (an Alphaproteobacteria), Actinobacteria, and TG1. Many of the clone phylotypes detected in soldiers grouped with those of workers. Moreover, clones CRgS91 (soldiers) and CRgW68 (workers), both affiliated with 'Endomicrobia', were the same phylotype. Soldiers and workers also seemed to have similar relative protist abundances. Heterotrophic, poly-β-hydroxyalkanoate-accumulating bacteria were isolated from the gut of soldiers and shown to be affiliated with Actinobacteria and Gammaproteobacteria. We noted that Wolbachia was detected in soldiers but not in workers. Overall, the maintenance by soldiers and workers of comparable axial and radial redox gradients in the gut is consistent with the similarities in the prokaryotes and protists comprising their microbiota.     

The bacterial community in the gut of the Cockroach Shelfordella lateralis reflects the close evolutionary relatedness of cockroaches and termites

Termites and cockroaches are closely related, with molecular phylogenetic analyses even placing termites within the radiation of cockroaches. The intestinal tract of wood-feeding termites harbors a remarkably diverse microbial community that is essential for the digestion of lignocellulose. However, surprisingly little is known about the gut microbiota of their closest relatives, the omnivorous cockroaches. Here, we present a combined characterization of physiological parameters, metabolic activities, and bacterial microbiota in the gut of Shelfordella lateralis, a representative of the cockroach family Blattidae, the sister group of termites. We compared the bacterial communities within each gut compartment using terminal-restriction fragment length polymorphism (T-RFLP) analysis and made a 16S rRNA gene clone library of the microbiota in the colon-the dilated part of the hindgut with the highest density and diversity of bacteria. The colonic community was dominated by members of the Bacteroidetes, Firmicutes (mainly Clostridia), and some Deltaproteobacteria. Spirochaetes and Fibrobacteres, which are abundant members of termite gut communities, were conspicuously absent. Nevertheless, detailed phylogenetic analysis revealed that many of the clones from the cockroach colon clustered with sequences previously obtained from the termite gut, which indicated that the composition of the bacterial community reflects at least in part the phylogeny of the host.     

Niche heterogeneity determines bacterial community structure in the termite gut (Reticulitermes santonensis)

Differences in microenvironment and interactions of microorganisms within and across habitat boundaries should influence structure and diversity of the microbial communities within an ecosystem. We tested this hypothesis using the well characterized gut tract of the European subterranean termite Reticulitermes santonensis as a model. By cloning and sequencing analysis and molecular fingerprinting (terminal restriction fragment length polymorphism), we characterized the bacterial microbiota in the major intestinal habitats - the midgut, the wall of the hindgut paunch, the hindgut fluid and the intestinal protozoa. The bacterial community was very diverse (> 200 ribotypes) and comprised representatives of several phyla, including Firmicutes (mainly clostridia, streptococci and Mycoplasmatales-related clones), Bacteroidetes, Spirochaetes and a number of Proteobacteria, all of which were unevenly distributed among the four habitats. The largest group of clones fell into the so-called Termite group 1 (TG-1) phylum, which has no cultivated representatives. The majority of the TG-1 clones were associated with the protozoa and formed two phylogenetically distinct clusters, which consisted exclusively of clones previously retrieved from the gut of this and other Reticulitermes species. Also the other clones represented lineages of microorganisms that were exclusively recovered from the intestinal tract of termites. The termite specificity of these lineages was underscored by the finding that the closest relatives of the bacterial clones obtained from R. santonensis were usually derived also from the most closely related termites. Overall, differences in diversity between the different gut habitats and the uneven distribution of individual phylotypes support conclusively that niche heterogeneity is a strong determinant of the structure and spatial organization of the microbial community in the termite gut.     

Phylogenetic relationships of symbiotic spirochetes in the gut of diverse termites

Phylogenetic relationships of symbiotic spirochetes in the gut of diverse termites were analyzed without cultivation of these microorganisms. A portion of the 16S rDNA (ca. 850 bp) was amplified directly from DNA of the mixed population in the gut by PCR and cloned. A total of 30 spirochetal phylotypes affiliated with the treponemes were identified from four termite species and they were compared with those already reported from other termites. They represented separate lines of descent from any known species of Treponema, and they were divided into two discrete clusters; one was related to Spirochaeta stenostrepta and S. caldaria, and the other was grouped together with members of the Treponema bryantii subgroup. Although some sequences from evolutionarily related termites showed close similarity, most of the sequences of spirochetes were dissimilar among different termite species, and spirochetal sequences from a single termite species occurred in several distinct phylogenetic positions. These findings suggest that termites constitute a rich reservoir of novel spirochetal diversity and that evolution of the symbiosis is not simple.     

低等白蚁肠道共生微生物的多样性及其功能

低等白蚁肠道里存在着复杂的微生物区系,包括真核微生物鞭毛虫和原核生物,细菌及古细菌。低等白蚁的后肠以特别膨大的囊形胃及其氢氧浓度的明显梯度分布和丰富的微生物区系为特征,是白蚁进行木质纤维素消化的主要器官。后肠内的鞭毛虫能将纤维素水解并发酵为乙酸,二氧化碳和氢,为白蚁提供营养和能源。系统发育研究表明,低等白蚁肠道共生细菌的主要类群为白蚁菌群1、螺旋体、拟杆菌,低G C mol%含量的革兰氏阳性菌和紫细菌等。而古细菌主要为甲烷短杆菌属的产甲烷菌。共生原核生物与二氧化碳的还原和氮的循环等代谢有关。但肠道共生微生物的具体功能和作用机制还有待进一步的揭示。     

Differences between bacterial communities in the gut of a soil-feeding termite (Cubitermes niokoloensis) and its mounds

In tropical ecosystems, termite mound soils constitute an important soil compartment covering around 10% of African soils. Previous studies have shown (S. Fall, S. Nazaret, J. L. Chotte, and A. Brauman, Microb. Ecol. 28:191-199, 2004) that the bacterial genetic structure of the mounds of soil-feeding termites (Cubitermes niokoloensis) is different from that of their surrounding soil. The aim of this study was to characterize the specificity of bacterial communities within mounds with respect to the digestive and soil origins of the mound. We have compared the bacterial community structures of a termite mound, termite gut sections, and surrounding soil using PCR-denaturing gradient gel electrophoresis (DGGE) analysis and cloning and sequencing of PCR-amplified 16S rRNA gene fragments. DGGE analysis revealed a drastic difference between the genetic structures of the bacterial communities of the termite gut and the mound. Analysis of 266 clones, including 54 from excised bands, revealed a high level of diversity in each biota investigated. The soil-feeding termite mound was dominated by the Actinobacteria phylum, whereas the Firmicutes and Proteobacteria phyla dominate the gut sections of termites and the surrounding soil, respectively. Phylogenetic analyses revealed a distinct clustering of Actinobacteria phylotypes between the mound and the surrounding soil. The Actinobacteria clones of the termite mound were diverse, distributed among 10 distinct families, and like those in the termite gut environment lightly dominated by the Nocardioidaceae family. Our findings confirmed that the soil-feeding termite mound (C. niokoloensis) represents a specific bacterial habitat in the tropics.     

Influence of feed components on symbiotic bacterial community structure in the gut of the wood-feeding higher termite Nasutitermes takasagoensis

The influence of carbon sources on bacterial community structure in the gut of the wood-feeding higher termite Nasutitermes takasagoensis was investigated. 16S rRNA gene sequencing and terminal-restriction fragment length polymorphism (T-RFLP) analyses revealed that the bacterial community structure changed markedly depending on feed components at the phylum level. Spirochaetes was predominant in the clone libraries from wood- and wood powder-fed termites, whereas Bacteroidetes was the largest group in the libraries from xylan-, cellobiose-, and glucose-fed termites, and Firmicutes was predominant in the library from xylose-fed termites. In addition, clones belonging to the phylum Termite Group I (TG1) were found in the library from xylose-fed termites. Our results indicate that the symbiotic relationship between termite and gut microorganisms is not very strong or stable over a short time, and that termite gut microbial community structures vary depending on components of the feeds.     

The motility symbiont of the termite gut flagellate Caduceia versatilis is a member of the "Synergistes" group

The flagellate Caduceia versatilis in the gut of the termite Cryptotermes cavifrons reportedly propels itself not by its own flagella but solely by the flagella of ectosymbiotic bacteria. Previous microscopic observations have revealed that the motility symbionts are flagellated rods partially embedded in the host cell surface and that, together with a fusiform type of ectosymbiotic bacteria without flagella, they cover almost the entire surface. To identify these ectosymbionts, we conducted 16S rRNA clone analyses of bacteria physically associated with the Caduceia cells. Two phylotypes were found to predominate in the clone library and were phylogenetically affiliated with the "Synergistes" phylum and the order Bacteroidales in the Bacteroidetes phylum. Probes specifically targeting 16S rRNAs of the respective phylotypes were designed, and fluorescence in situ hybridization (FISH) was performed. As a result, the "Synergistes" phylotype was identified as the motility symbiont; the Bacteroidales phylotype was the fusiform ectobiont. The "Synergistes" phylotype was a member of a cluster comprising exclusively uncultured clones from the guts of various termite species. Interestingly, four other phylotypes in this cluster, including the one sharing 95% sequence identity with the motility symbiont, were identified as nonectosymbiotic, or free-living, gut bacteria by FISH. We thus suggest that the motility ectosymbiont has evolved from a free-living gut bacterium within this termite-specific cluster. Based on these molecular and previous morphological data, we here propose a novel genus and species, "Candidatus Tammella caduceiae," for this unique motility ectosymbiont of Caducaia versatilis.     

A molecular phylogenetic survey of sea-ice microbial communities (SIMCO)

16S rDNA clone library analysis was used to identify bacterial biodiversity in a variety of sea-ice microbial communities (SIMCO). DNA was extracted from seven Antarctic sea-ice samples and one Arctic sea-ice sample and 16S rDNA PCR-amplified using universal and Archaea-specific primers. Recombinant 16S rDNA clones were obtained and dereplicated using restriction fragment length polymorphism analysis (RFLP). After RFLP analysis, 100 distinct phylotypes (a unique clone or group of clones with sequence similarity of >0.98) were defined. From the clone libraries 16S rDNA sequences of bacterial and eukaryotic origin were detected, however Archaea were not detected either with universal or Archaea-specific 16S rDNA primer sets. Bacterial phylotypes grouped within the alpha and gamma proteobacteria, the Cytophaga-Flavobacterium-Bacteroides division, the Gram-Positive bacteria and the orders Chlamydiales and Verrucomicrobiales. The majority of bacterial phylotypes were affiliated with heterotrophic taxa and many grouped closely with cultivated genera and species. Eukaryotic clones were affiliated with a variety of autotrophic and heterotrophic nanoplankton and included a large number of chloroplast 16S rDNA genes. The findings of this investigation corroborated culture data indicating bacterial biodiversity increased in SIMCO displaying high levels of primary production, however the bacterial communities within SIMCO were highly heterogeneous at the genus/species-level between different samples. A comparison of Antarctic and Arctic SIMCO revealed certain sea-ice dwelling bacterial genera are common at both poles.     

Influence of Feed Components on Symbiotic Bacterial Community Structure in the Gut of the Wood-Feeding Higher Termite Nasutitermes takasagoensis

The influence of carbon sources on bacterial community structure in the gut of the wood-feeding higher termite Nasutitermes takasagoensis was investigated. 16S rRNA gene sequencing and terminal-restriction fragment length polymorphism (T-RFLP) analyses revealed that the bacterial community structure changed markedly depending on feed components at the phylum level. Spirochaetes was predominant in the clone libraries from wood- and wood powder-fed termites, whereas Bacteroidetes was the largest group in the libraries from xylan-, cellobiose-, and glucose-fed termites, and Firmicutes was predominant in the library from xylose-fed termites. In addition, clones belonging to the phylum Termite Group I (TG1) were found in the library from xylose-fed termites. Our results indicate that the symbiotic relationship between termite and gut microorganisms is not very strong or stable over a short time, and that termite gut microbial community structures vary depending on components of the feeds.     

Differences between Bacterial Communities in the Gut of a Soil-Feeding Termite (Cubitermes niokoloensis) and Its Mounds

In tropical ecosystems, termite mound soils constitute an important soil compartment covering around 10% of African soils. Previous studies have shown (S. Fall, S. Nazaret, J. L. Chotte, and A. Brauman, Microb. Ecol. 28:191-199, 2004) that the bacterial genetic structure of the mounds of soil-feeding termites (Cubitermes niokoloensis) is different from that of their surrounding soil. The aim of this study was to characterize the specificity of bacterial communities within mounds with respect to the digestive and soil origins of the mound. We have compared the bacterial community structures of a termite mound, termite gut sections, and surrounding soil using PCR-denaturing gradient gel electrophoresis (DGGE) analysis and cloning and sequencing of PCR-amplified 16S rRNA gene fragments. DGGE analysis revealed a drastic difference between the genetic structures of the bacterial communities of the termite gut and the mound. Analysis of 266 clones, including 54 from excised bands, revealed a high level of diversity in each biota investigated. The soil-feeding termite mound was dominated by the Actinobacteria phylum, whereas the Firmicutes and Proteobacteria phyla dominate the gut sections of termites and the surrounding soil, respectively. Phylogenetic analyses revealed a distinct clustering of Actinobacteria phylotypes between the mound and the surrounding soil. The Actinobacteria clones of the termite mound were diverse, distributed among 10 distinct families, and like those in the termite gut environment lightly dominated by the Nocardioidaceae family. Our findings confirmed that the soil-feeding termite mound (C. niokoloensis) represents a specific bacterial habitat in the tropics.     

Coevolution of symbiotic spirochete diversity in lower termites.

The phylogenetic relationships of symbiotic spirochetes from five dry-wood feeding lower termites (Cryptotermes cavifrons, Heterotermes tenuis, Kalotermes flavicollis, Neotermes mona, and Reticulitermes grassei) was compared to those described in previous reports. The 16S rDNA bacterial genes were PCR-amplified from DNA isolated from intestinal samples using a spirochete-selective primer, and the 16S amplicons were cloned into Escherichia coli. Sequences of the cloned inserts were then used to determine closest relatives by comparison with published sequences. Clones sharing more than 97% sequence identity were grouped into the same phylotype. Forty-three new phylotypes were identified. These termite whole-gut-spirochetes fell into two previous defined clusters, designated as Treponema Clusters I and II, and one new Cluster III. Thirty-seven phylotypes were grouped in Cluster I. Cluster II comprised three phylotypes, two from Reticulitermes grassei (LJ029 and LJ012) and one from Heterotermes tenuis (LQ016). Three phylotypes, LK057, LK050 and LK028, were affiliated to Cluster III. Members of Cluster I showed the following characteristics: (i) spirochete phylotypes from a particular species of termite were more closely related to each other than to phylotypes of other termite species; (ii) spirochetes obtained from different genera of the same family, such as Cryptotermes sp., Kalotermes sp., and Neotermes sp., all from the family Kalotermitidae, were also related to each other. It was therefore concluded that spirochetes are specific symbionts that have coevolved with their respective species of termites, are stably harbored, and are closely related to members of the same termite family.     

Diversity and structure of the archaeal community in the leachate of a full-scale recirculating landfill as examined by direct 16S rRNA gene sequence retrieval

The diversity and structure of the archaeal community in the effluent leachate from a full-scale recirculating landfill was characterized by direct 16S rRNA gene (16S rDNA) retrieval. Total-community DNA was extracted from the microbial assemblages in the landfill leachate, and archaeal 16S rDNAs were amplified with a universally conserved primer and an Archaea-specific primer. The amplification product was then used to construct a 16S rDNA clone library, and 70 randomly selected archaeal clones in the library were grouped by restriction fragment length polymorphism (RFLP) analysis. Sequencing and phylogenetic analysis of representatives from each unique RFLP type showed that the archaeal library was dominated by methanogen-like rDNAs. Represented in the kingdom of Euryarchaeota were phylotypes highly similar to the methanogenic genera Methanoculleus, Methanosarcina, Methanocorpusculum, Methanospirillum and Methanogenium, where the clone distribution was 48, 11, 3, 1 and 1, respectively. No sequences related to known Methanosaeta spp. were retrieved. Four rDNA clones were not affiliated with the known methanogenic Archaea, but instead, they were clustered with the uncultured archaeal sequences recently recovered from anaerobic habitats. Two chimeric sequences were identified among the clones analyzed.     

Identifying the core microbial community in the gut of fungus‐growing termites

Gut microbes play a crucial role in decomposing lignocellulose to fuel termite societies, with protists in the lower termites and prokaryotes in the higher termites providing these services. However, a single basal subfamily of the higher termites, the Macrotermitinae, also domesticated a plant biomass‐degrading fungus (Termitomyces), and how this symbiont acquisition has affected the fungus‐growing termite gut microbiota has remained unclear. The objective of our study was to compare the intestinal bacterial communities of five genera (nine species) of fungus‐growing termites to establish whether or not an ancestral core microbiota has been maintained and characterizes extant lineages. Using 454‐pyrosequencing of the 16S rRNA gene, we show that gut communities have representatives of 26 bacterial phyla and are dominated by Firmicutes, Bacteroidetes, Spirochaetes, Proteobacteria and Synergistetes. A set of 42 genus‐level taxa was present in all termite species and accounted for 56–68% of the species‐specific reads. Gut communities of termites from the same genus were more similar than distantly related species, suggesting that phylogenetic ancestry matters, possibly in connection with specific termite genus‐level ecological niches. Finally, we show that gut communities of fungus‐growing termites are similar to cockroaches, both at the bacterial phylum level and in a comparison of the core Macrotermitinae taxa abundances with representative cockroach, lower termite and higher nonfungus‐growing termites. These results suggest that the obligate association with Termitomyces has forced the bacterial gut communities of the fungus‐growing termites towards a relatively uniform composition with higher similarity to their omnivorous relatives than to more closely related termites.     

Intracolony variation of bacterial gut microbiota among castes and ages in the fungus-growing termite Macrotermes gilvus

The fungus-growing termites Macrotermes cultivate the obligate ectosymbiontic fungi, Termitomyces. While their relationship has been extesively studied, little is known about the gut bacterial symbionts, which also presumably play a crucial role for the nutrition of the termite host. In this study, we investigated the bacterial gut microbiota in two colonies of Macrotermes gilvus, and compared the diversity and community structure of bacteria among nine termite morphotypes, differing in caste and/or age, using terminal restriction fragment length polymorphism (T-RFLP) and clonal analysis of 16S rRNA. The obtained molecular community profiles clustered by termite morphotype rather than by colony, and the clustering pattern was clearly more related to a difference in age than to caste. Thus, we suggest that the bacterial gut microbiota change in relation to the food of the termite, which comprises fallen leaves and the fungus nodules of Termitomyces in young workers, and leaves degraded by the fungi, in old workers. Despite these intracolony variations in bacterial gut microbiota, their T-RFLP profiles formed a distinct cluster against those of the fungus garden, adjacent soil and guts of sympatric wood-feeding termites, implying a consistency and uniqueness of gut microbiota in M. gilvus. Since many bacterial phylotypes from M. gilvus formed monophyletic clusters with those from distantly related termite species, we suggest that gut bacteria have co-evolved with the termite host and form a microbiota specific to a termite taxonomic and/or feeding group, and furthermore, to caste and age within a termite species.