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Andreas Brune - One of the best experts on this subject based on the ideXlab platform.
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the fibre associated cellulolytic bacterial community in the Hindgut of wood feeding higher termites nasutitermes spp
Environmental Microbiology, 2014Co-Authors: Aram Mikaelyan, Gaku Tokuda, Jurgen F H Strassert, Andreas BruneAbstract:Summary Termites digest lignocellulose with the help of their symbiotic gut microbiota. In the Hindgut of evolutionary lower termites, a dense community of cellulolytic flagellates sequesters wood particles from the Hindgut content into their digestive vacuoles. In higher termites (family Termitidae), which possess an entirely prokaryotic microbiota, the wood particles are available for bacterial colonization. Substantial particle-associated cellulase activities have been detected in the Hindgut of Nasutitermes species, but the microorganisms responsible for these activities and their potential association with the wood fibres remain to be studied. Here, we used density-gradient centrifugation to separate wood fibres and adherent bacterial cells from cells freely suspended in the Hindgut fluid. In Nasutitermes corniger, the fibre fraction contained 28% of the DNA and 45% of the cellulase activity in the luminal contents (P3 region). Community fingerprinting (terminal restriction fragment length polymorphism) and pyrotag sequencing analysis of the bacterial 16S rRNA genes demonstrated that the wood fibres in the Hindgut of both N. corniger and N. takasagoensis are specifically colonized by members of Fibrobacteres, the TG3 phylum, and certain lineages of Spirochaetes characteristic of the gut microbiota of wood-feeding higher termites. We propose that the loss of flagellates in higher termites provided a new niche for fibre-associated cellulolytic bacteria.
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inter and intraspecific comparison of the bacterial assemblages in the Hindgut of humivorous scarab beetle larvae pachnoda spp
FEMS Microbiology Ecology, 2010Co-Authors: Janet Andert, Andreas Marten, Roland Brandl, Andreas BruneAbstract:The larvae of scarab beetles are model organisms for studying the role of physicochemical gut conditions and intestinal microbiota in symbiotic digestion, particularly of humus. Here, we address the question of whether the enlarged Hindgut paunch of Pachnoda ephippiata and Pachnoda marginata, two closely related, but allopatric species, harbors a specific bacterial microbiota. Terminal restriction length fragment polymorphism (T-RFLP) analysis revealed that in both species, the bacterial Hindgut community differs strongly from that in the midgut, food soil, and fecal pellets. High intra- and interspecific similarities between the T-RFLP profiles of different larvae indicate the presence of a Hindgut-specific microbiota. Nevertheless, we found a clear separation of the two species. A 16S rRNA gene clone library from the Hindgut of P. ephippiata identified the major phylogenetic groups as members of the Clostridia, Betaproteobacteria, and Bacteroidetes, followed by Bacillales and Deltaproteobacteria. A comparison with a previously obtained clone library of the same species corroborates both the similarities and the intraspecific variance of the Hindgut microbiota.
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Methane oxidation in termite Hindguts: absence of evidence and evidence of absence.
Applied and Environmental Microbiology, 2007Co-Authors: Michael Pester, Michael W Friedrich, Anne Tholen, Andreas BruneAbstract:A steep oxygen gradient and the presence of methane render the Hindgut internal periphery of termites a potential habitat for aerobic methane-oxidizing bacteria. However, methane emissions of various termites increased, if at all, only slightly when termites were exposed to an anoxic (nitrogen) atmosphere, and 14CH4 added to the air headspace over live termites was not converted to 14CO2. Evidence for the absence of methane oxidation in living termites was corroborated by the failure to detect pmoA, the marker gene for particulate methane monooxygenase, in Hindgut DNA extracts of all termites investigated. This adds robustness to our concept of the degradation network in the termite Hindgut and eliminates the gut itself as a potential sink of this important greenhouse gas.
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microbial community structure in midgut and Hindgut of the humus feeding larva of pachnoda ephippiata coleoptera scarabaeidae
Applied and Environmental Microbiology, 2003Co-Authors: Markus Egert, Andreas Brune, Bianca Wagner, Thorsten Lemke, Michael W FriedrichAbstract:The guts of soil-feeding macroinvertebrates contain a complex microbial community that is involved in the transformation of ingested soil organic matter. In a companion paper (T. Lemke, U. Stingl, M. Egert, M. W. Friedrich, and A. Brune, Appl. Environ. Microbiol. 69:6650-6658, 2003), we show that the gut of our model organism, the humivorous larva of the cetoniid beetle Pachnoda ephippiata, is characterized by strong midgut alkalinity, high concentrations of microbial fermentation products, and the presence of a diverse, yet unstudied microbial community. Here, we report on the community structure of bacteria and archaea in the midgut, Hindgut, and food soil of P. ephippiata larvae, determined with cultivation-independent techniques. Clone libraries and terminal restriction fragment length polymorphism analysis of 16S rRNA genes revealed that the intestines of P. ephippiata larvae contain a complex gut microbiota that differs markedly between midgut and Hindgut and that is clearly distinct from the microbiota in the food soil. The bacterial community is dominated by phylogenetic groups with a fermentative metabolism (Lactobacillales, Clostridiales, Bacillales, and Cytophaga-Flavobacterium-Bacteroides [CFB] phylum), which is corroborated by high lactate and acetate concentrations in the midgut and Hindgut and by the large numbers of lactogenic and acetogenic bacteria in both gut compartments reported in the companion paper. Based on 16S rRNA gene frequencies, Actinobacteria dominate the alkaline midgut, while the Hindgut is dominated by members of the CFB phylum. The archaeal community, however, is less diverse. 16S rRNA genes affiliated with mesophilic Crenarchaeota, probably stemming from the ingested soil, were most frequent in the midgut, whereas Methanobacteriaceae-related 16S rRNA genes were most frequent in the Hindgut. These findings agree with the reported restriction of methanogenesis to the Hindgut of Pachnoda larvae.
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impact of oxygen on metabolic fluxes and in situ rates of reductive acetogenesis in the Hindgut of the wood feeding termite reticulitermes flavipes
Environmental Microbiology, 2000Co-Authors: Anne Tholen, Andreas BruneAbstract:: The symbiotic digestion of lignocellulose in the Hindgut of the wood-feeding termite Reticulitermes flavipes is characterized by two major metabolic pathways: (i) the oxidation of polysaccharides to acetate by anaerobic hydrogen-producing protozoa; and (ii) the reduction of CO2 by hydrogenotrophic acetogenic bacteria. Both reactions together would render the Hindgut largely homoacetogenic. However, the results of this study show that the situation is more complex. By microinjection of radiolabelled metabolites into intact agarose-embedded Hindguts, we showed that the in situ rates of reductive acetogenesis (3.3 nmol termite(-1) h(-1)) represent only 10% of the total carbon flux in the living termite, whereas 30% of the carbon flux proceeds via lactate. The rapid turnover of the lactate pool (7.2 nmol termite(-1) h(-1)) consolidates the previously reported presence of lactic acid bacteria in the R. flavipes Hindgut and the low lactate concentrations in the Hindgut fluid. However, the immediate precursor of lactate remains unknown; the low turnover rates of injected glucose (< 0.5 nmol termite(-1) h(-1)) indicate that free glucose is not an important intermediate under in situ conditions. The influence of the incubation atmosphere on the turnover rate and the product pattern of glucose and lactate confirmed that the influx of oxygen via the gut epithelium and its reduction in the Hindgut periphery have a significant impact on carbon and electron flow within the Hindgut microbial community. The in situ rates of reductive acetogenesis were not significantly affected by the presence of oxygen or exogenous H2, which is in agreement with a localization of homoacetogens in the anoxic gut lumen rather than in the oxic periphery. This adds strong support to the hypothesis that the co-existence of methanogens and homoacetogens in this termite is based on the spatial arrangement of the different populations of the gut microbiota. A refined model of metabolic fluxes in the Hindgut of R. flavipes is presented.
Helmut König - One of the best experts on this subject based on the ideXlab platform.
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differential enumeration and in situ localization of microorganisms in the Hindgut of the lower termite mastotermes darwiniensis by hybridization with rrna targeted probes
Archives of Microbiology, 1999Co-Authors: Manfred Berchtold, Wilhelm Schonhuber, D Hahn, Rudolf Amann, Andreas Brune, Antonis Chatzinotas, Helmut KönigAbstract:We examined the abundance and spatial distribution of major phylogenetic groups of the domain Bacteria in Hindguts of the Australian lower termite Mastotermes darwiniensis by using in situ hybridization with group-specific, fluorescently labeled, rRNA-targeted oligonucleotide probes. Between 32.0 ± 7.2% and 52.3 ± 8.2% of the DAPI-stained cells in different Hindgut fractions were detected with probe EUB338, specific for members of the domain Bacteria. About 85% of the prokaryotic cells were associated with the flagellates of the thin-walled anterior region (P3a) and the thick wall of the posterior region (P3b/P4) of the Hindgut, as shown by DAPI staining. At most, half of the EUB338-detected cells hybridized with one of the other probes that targeted a smaller assemblage within the bacterial domain. In most fractions, cells were found in varying numbers with probe ALF1b, which targeted members of the α-Proteobacteria, whereas substantial amounts of sulfate-reducing bacteria, gram-positive bacteria with a high DNA G+C content and members of the Cytophaga-Flavobacterium cluster of the Cytophaga-Flavobacterium-Bacteroides (CFB) phylum could be detected only in the wall fraction of P3b/P4. This clearly indicates that the Hindgut microhabitats differ in the composition of their microbial community. In situ hybridization of cryosections through the Hindgut showed only low numbers of bacteria attached to the P3a wall. In contrast, the wall of P3b was densely colonized by rod- and coccus-shaped bacteria, which could be assigned to the Cytophaga-Flavobacterium cluster of the CFB phylum and to the group of gram-positive bacteria with a high DNA G+C content, respectively. Oxygen concentration profiles determined with microelectrodes revealed steep oxygen gradients both in P3a and P3b. Oxygen was consumed within 100 μm below the gut surface, and anoxic conditions prevailed in the central portions of both gut regions, indicating that oxygen consumption in the Hindgut does not depend on the presence of a biofilm on the Hindgut wall.
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phylogenetic analysis and in situ identification of uncultivated spirochetes from the Hindgut of the termite mastotermes darwiniensis
Systematic and Applied Microbiology, 1996Co-Authors: Manfred Berchtold, Helmut KönigAbstract:Summary Different small subunit ribosomal DNA (SSU rDNA) sequences of uncultivated spirochetes from the Hindgut of the termite Mastotermes darwiniensis were determined and phylogenetically analysed. Starting with DNA isolated from Hindgut contents we amplified SSU rDNA coding regions by PCR using bacteria- and spirochete-specific primers, respectively. Cloning and sequencing of the amplification products re-sulted in 11 different spirochetal SSU rDNA sequences with similarity values of 81.8% to 99.0% to each other. Phylogenetic trees showed that the obtained sequences are related to the genus Treponema with 84.0% to 91.1% similarity to Treponema spec. H1 and Spirochaeta stenostrepta as their nearest relatives among the cultivated spirochetes. Using fluorescence in situ hybridisation one of the SSU rDNA sequences could be assigned to a large spirochete. So at least one of the large Hindgut spirochetes unique in termite Hindguts and assigned to the genera Hollandina , Pillotina , Clevelandina or Diplocalyx phylogenetically belongs to the treponeme cluster of spirochetes.
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16s rdna sequence and phylogenetic position of an uncultivated spirochete from the Hindgut of the termite mastotermes darwiniensis froggatt
Fems Microbiology Letters, 1994Co-Authors: Manfred Berchtold, Wolfgang Ludwig, Helmut KönigAbstract:We have analyzed the 16S rDNA sequence and the phylogenetic position of an uncultivated spirochete from the Hindgut contents of the Australian termite Mastotermes darwiniensis Froggatt. The 16S rRNA genes of bacteria from the Hindgut contents of Mastotermes darwiniensis were amplified by polymerase chain reaction. The amplification products were cloned and sequenced. The sequences were compared to known homologous primary structures. Two of the clones (MDS1 and MDS3) had an insert of 1498 nucleotides showing typical signatures of spirochete 16S rRNA sequences. The sequences of the two clones were most similar to the 16S rRNA sequence of Spirochaeta stenostrepta (89.8%) and Treponema sp. strain H1 (90.7%). Phylogenetical analysis positioned the Hindgut spirochete sequence with that of the free-living anaerobic Spirochaeta stenostrepta and Treponema sp. strain H1 as its nearest relatives within the cluster of the spirochetes. We conclude that the analyzed SSU rDNA sequences originate from a spirochete related to the genus Treponema. It is possibly one of the uncultivated unique spirochetes symbiotic in termite Hindguts.
Manfred Berchtold - One of the best experts on this subject based on the ideXlab platform.
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differential enumeration and in situ localization of microorganisms in the Hindgut of the lower termite mastotermes darwiniensis by hybridization with rrna targeted probes
Archives of Microbiology, 1999Co-Authors: Manfred Berchtold, Wilhelm Schonhuber, D Hahn, Rudolf Amann, Andreas Brune, Antonis Chatzinotas, Helmut KönigAbstract:We examined the abundance and spatial distribution of major phylogenetic groups of the domain Bacteria in Hindguts of the Australian lower termite Mastotermes darwiniensis by using in situ hybridization with group-specific, fluorescently labeled, rRNA-targeted oligonucleotide probes. Between 32.0 ± 7.2% and 52.3 ± 8.2% of the DAPI-stained cells in different Hindgut fractions were detected with probe EUB338, specific for members of the domain Bacteria. About 85% of the prokaryotic cells were associated with the flagellates of the thin-walled anterior region (P3a) and the thick wall of the posterior region (P3b/P4) of the Hindgut, as shown by DAPI staining. At most, half of the EUB338-detected cells hybridized with one of the other probes that targeted a smaller assemblage within the bacterial domain. In most fractions, cells were found in varying numbers with probe ALF1b, which targeted members of the α-Proteobacteria, whereas substantial amounts of sulfate-reducing bacteria, gram-positive bacteria with a high DNA G+C content and members of the Cytophaga-Flavobacterium cluster of the Cytophaga-Flavobacterium-Bacteroides (CFB) phylum could be detected only in the wall fraction of P3b/P4. This clearly indicates that the Hindgut microhabitats differ in the composition of their microbial community. In situ hybridization of cryosections through the Hindgut showed only low numbers of bacteria attached to the P3a wall. In contrast, the wall of P3b was densely colonized by rod- and coccus-shaped bacteria, which could be assigned to the Cytophaga-Flavobacterium cluster of the CFB phylum and to the group of gram-positive bacteria with a high DNA G+C content, respectively. Oxygen concentration profiles determined with microelectrodes revealed steep oxygen gradients both in P3a and P3b. Oxygen was consumed within 100 μm below the gut surface, and anoxic conditions prevailed in the central portions of both gut regions, indicating that oxygen consumption in the Hindgut does not depend on the presence of a biofilm on the Hindgut wall.
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phylogenetic analysis and in situ identification of uncultivated spirochetes from the Hindgut of the termite mastotermes darwiniensis
Systematic and Applied Microbiology, 1996Co-Authors: Manfred Berchtold, Helmut KönigAbstract:Summary Different small subunit ribosomal DNA (SSU rDNA) sequences of uncultivated spirochetes from the Hindgut of the termite Mastotermes darwiniensis were determined and phylogenetically analysed. Starting with DNA isolated from Hindgut contents we amplified SSU rDNA coding regions by PCR using bacteria- and spirochete-specific primers, respectively. Cloning and sequencing of the amplification products re-sulted in 11 different spirochetal SSU rDNA sequences with similarity values of 81.8% to 99.0% to each other. Phylogenetic trees showed that the obtained sequences are related to the genus Treponema with 84.0% to 91.1% similarity to Treponema spec. H1 and Spirochaeta stenostrepta as their nearest relatives among the cultivated spirochetes. Using fluorescence in situ hybridisation one of the SSU rDNA sequences could be assigned to a large spirochete. So at least one of the large Hindgut spirochetes unique in termite Hindguts and assigned to the genera Hollandina , Pillotina , Clevelandina or Diplocalyx phylogenetically belongs to the treponeme cluster of spirochetes.
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16s rdna sequence and phylogenetic position of an uncultivated spirochete from the Hindgut of the termite mastotermes darwiniensis froggatt
Fems Microbiology Letters, 1994Co-Authors: Manfred Berchtold, Wolfgang Ludwig, Helmut KönigAbstract:We have analyzed the 16S rDNA sequence and the phylogenetic position of an uncultivated spirochete from the Hindgut contents of the Australian termite Mastotermes darwiniensis Froggatt. The 16S rRNA genes of bacteria from the Hindgut contents of Mastotermes darwiniensis were amplified by polymerase chain reaction. The amplification products were cloned and sequenced. The sequences were compared to known homologous primary structures. Two of the clones (MDS1 and MDS3) had an insert of 1498 nucleotides showing typical signatures of spirochete 16S rRNA sequences. The sequences of the two clones were most similar to the 16S rRNA sequence of Spirochaeta stenostrepta (89.8%) and Treponema sp. strain H1 (90.7%). Phylogenetical analysis positioned the Hindgut spirochete sequence with that of the free-living anaerobic Spirochaeta stenostrepta and Treponema sp. strain H1 as its nearest relatives within the cluster of the spirochetes. We conclude that the analyzed SSU rDNA sequences originate from a spirochete related to the genus Treponema. It is possibly one of the uncultivated unique spirochetes symbiotic in termite Hindguts.
Gaku Tokuda - One of the best experts on this subject based on the ideXlab platform.
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the fibre associated cellulolytic bacterial community in the Hindgut of wood feeding higher termites nasutitermes spp
Environmental Microbiology, 2014Co-Authors: Aram Mikaelyan, Gaku Tokuda, Jurgen F H Strassert, Andreas BruneAbstract:Summary Termites digest lignocellulose with the help of their symbiotic gut microbiota. In the Hindgut of evolutionary lower termites, a dense community of cellulolytic flagellates sequesters wood particles from the Hindgut content into their digestive vacuoles. In higher termites (family Termitidae), which possess an entirely prokaryotic microbiota, the wood particles are available for bacterial colonization. Substantial particle-associated cellulase activities have been detected in the Hindgut of Nasutitermes species, but the microorganisms responsible for these activities and their potential association with the wood fibres remain to be studied. Here, we used density-gradient centrifugation to separate wood fibres and adherent bacterial cells from cells freely suspended in the Hindgut fluid. In Nasutitermes corniger, the fibre fraction contained 28% of the DNA and 45% of the cellulase activity in the luminal contents (P3 region). Community fingerprinting (terminal restriction fragment length polymorphism) and pyrotag sequencing analysis of the bacterial 16S rRNA genes demonstrated that the wood fibres in the Hindgut of both N. corniger and N. takasagoensis are specifically colonized by members of Fibrobacteres, the TG3 phylum, and certain lineages of Spirochaetes characteristic of the gut microbiota of wood-feeding higher termites. We propose that the loss of flagellates in higher termites provided a new niche for fibre-associated cellulolytic bacteria.
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symbiotic archaezoa of the primitive termite mastotermes darwiniensis still play a role in cellulase production
Eukaryotic Cell, 2006Co-Authors: Hirofumi Watanabe, Aya Takase, Gaku Tokuda, Akinori Yamada, Nathan LoAbstract:The relictual Mastotermes darwiniensis is one of the world’s most destructive termites. Like all phylogenetically basal termites, it possesses protozoa in its Hindgut, which are believed to help it digest wood. L. Li, J. Frohlich, P. Pfeiffer, and H. Konig (Eukaryot. Cell 2:1091–1098, 2003) recently cloned the genes encoding cellulases from the protozoa of M. darwiniensis; however, they claimed that these genes are essentially inactive, not contributing significantly to cellulose digestion. Instead, they suggested that the protozoa sequester enzymes produced by the termite in its salivary glands and use these to degrade cellulose in the Hindgut. We tested this idea by performing gel filtration of enzymes in extracts of the Hindgut, as well as in a combination of the salivary glands, foregut, and midgut. Three major cellulases were found in the Hindgut, each of which had a larger molecular size than termite-derived salivary gland enzymes. N-terminal amino acid sequencing of one of the Hindgut-derived enzymes showed that it was identical to the putative amino acid sequence of one mRNA sequence isolated by Li et al. (Eukaryot. Cell 2:1091–1098, 2003). The overall activity of the Hindgut cellulases was found to be of approximately equal magnitude to the termite-derived cellulases detected in the mixture of salivary gland, foregut, and midguts. Based on these results, we conclude that, contrary to Li et al. (Eukaryot. Cell 2:1091–1098, 2003), the Hindgut protozoan fauna of M. darwiniensis actively produce cellulases, which play an important role in cellulose digestion of the host termite.
Anne Tholen - One of the best experts on this subject based on the ideXlab platform.
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Methane oxidation in termite Hindguts: absence of evidence and evidence of absence.
Applied and Environmental Microbiology, 2007Co-Authors: Michael Pester, Michael W Friedrich, Anne Tholen, Andreas BruneAbstract:A steep oxygen gradient and the presence of methane render the Hindgut internal periphery of termites a potential habitat for aerobic methane-oxidizing bacteria. However, methane emissions of various termites increased, if at all, only slightly when termites were exposed to an anoxic (nitrogen) atmosphere, and 14CH4 added to the air headspace over live termites was not converted to 14CO2. Evidence for the absence of methane oxidation in living termites was corroborated by the failure to detect pmoA, the marker gene for particulate methane monooxygenase, in Hindgut DNA extracts of all termites investigated. This adds robustness to our concept of the degradation network in the termite Hindgut and eliminates the gut itself as a potential sink of this important greenhouse gas.
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impact of oxygen on metabolic fluxes and in situ rates of reductive acetogenesis in the Hindgut of the wood feeding termite reticulitermes flavipes
Environmental Microbiology, 2000Co-Authors: Anne Tholen, Andreas BruneAbstract:: The symbiotic digestion of lignocellulose in the Hindgut of the wood-feeding termite Reticulitermes flavipes is characterized by two major metabolic pathways: (i) the oxidation of polysaccharides to acetate by anaerobic hydrogen-producing protozoa; and (ii) the reduction of CO2 by hydrogenotrophic acetogenic bacteria. Both reactions together would render the Hindgut largely homoacetogenic. However, the results of this study show that the situation is more complex. By microinjection of radiolabelled metabolites into intact agarose-embedded Hindguts, we showed that the in situ rates of reductive acetogenesis (3.3 nmol termite(-1) h(-1)) represent only 10% of the total carbon flux in the living termite, whereas 30% of the carbon flux proceeds via lactate. The rapid turnover of the lactate pool (7.2 nmol termite(-1) h(-1)) consolidates the previously reported presence of lactic acid bacteria in the R. flavipes Hindgut and the low lactate concentrations in the Hindgut fluid. However, the immediate precursor of lactate remains unknown; the low turnover rates of injected glucose (< 0.5 nmol termite(-1) h(-1)) indicate that free glucose is not an important intermediate under in situ conditions. The influence of the incubation atmosphere on the turnover rate and the product pattern of glucose and lactate confirmed that the influx of oxygen via the gut epithelium and its reduction in the Hindgut periphery have a significant impact on carbon and electron flow within the Hindgut microbial community. The in situ rates of reductive acetogenesis were not significantly affected by the presence of oxygen or exogenous H2, which is in agreement with a localization of homoacetogens in the anoxic gut lumen rather than in the oxic periphery. This adds strong support to the hypothesis that the co-existence of methanogens and homoacetogens in this termite is based on the spatial arrangement of the different populations of the gut microbiota. A refined model of metabolic fluxes in the Hindgut of R. flavipes is presented.
