Termites digest wood using Carbohydrate-Active Enzymes (CAZymes) produced by gut bacteria with whom they have cospeciated at geological timescales. Whether CAZymes were encoded in the genomes of their ancestor's gut bacteria and transmitted to modern termites or acquired more recently from bacteria not associated with termites is unclear. We used gut metagenomes from 195 termites and one Cryptocercus, the sister group of termites, to investigate the evolution of termite gut bacterial CAZymes. We found 420 termite-specific clusters in 81 bacterial CAZyme gene trees, including 404 clusters showing strong cophylogenetic patterns with termites. Of the 420 clusters, 131 included at least one bacterial CAZyme sequence associated with Cryptocercus or Mastotermes, the sister group of all other termites. Our results suggest many bacterial CAZymes have been encoded in the genomes of termite gut bacteria since termite origin, indicating termites rely upon many bacterial CAZymes endemic to their guts to digest wood.

• MeSH
• Bacteria * enzymologie   genetika   MeSH
• bakteriální proteiny metabolismus   genetika   MeSH
• fylogeneze * MeSH
• Isoptera * mikrobiologie   enzymologie   MeSH
• metabolismus sacharidů MeSH
• metagenom MeSH
• molekulární evoluce MeSH
• střevní mikroflóra MeSH
• švábi mikrobiologie   enzymologie   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• bakteriální proteiny MeSH

Millipedes are thought to depend on their gut microbiome for processing plant-litter-cellulose through fermentation, similar to many other arthropods. However, this hypothesis lacks sufficient evidence. To investigate this, we used inhibitors to disrupt the gut microbiota of juvenile Epibolus pulchripes (tropical, CH4-emitting) and Glomeris connexa (temperate, non-CH4-emitting) and isotopic labelling. Feeding the millipedes sterile or antibiotics-treated litter reduced faecal production and microbial load without major impacts on survival or weight. Bacterial diversity remained similar, with Bacteroidota dominant in E. pulchripes and Pseudomonadota in G. connexa. Sodium-2-bromoethanesulfonate treatment halted CH4 emissions in E. pulchripes, but it resumed after returning to normal feeding. Employing 13C-labeled leaf litter and RNA-SIP revealed a slow and gradual prokaryote labelling, indicating a significant density shift only by day 21. Surprisingly, labelling of the fungal biomass was somewhat quicker. Our findings suggest that fermentation by the gut microbiota is likely not essential for the millipede's nutrition.

• MeSH
• Bacteria metabolismus   genetika   MeSH
• členovci * mikrobiologie   metabolismus   MeSH
• feces mikrobiologie   MeSH
• fermentace * MeSH
• listy rostlin metabolismus   mikrobiologie   MeSH
• střevní mikroflóra * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

Bacterial endosymbionts of eukaryotic hosts typically experience massive genome reduction, but the underlying evolutionary processes are often obscured by the lack of free-living relatives. Endomicrobia, a family-level lineage of host-associated bacteria in the phylum Elusimicrobiota that comprises both free-living representatives and endosymbionts of termite gut flagellates, are an excellent model to study evolution of intracellular symbionts. We reconstructed 67 metagenome-assembled genomes (MAGs) of Endomicrobiaceae among more than 1,700 MAGs from the gut microbiota of a wide range of termites. Phylogenomic analysis confirmed a sister position of representatives from termites and ruminants, and allowed to propose eight new genera in the radiation of Endomicrobiaceae. Comparative genome analysis documented progressive genome erosion in the new genus Endomicrobiellum, which comprises all flagellate endosymbionts characterized to date. Massive gene losses were accompanied by the acquisition of new functions by horizontal gene transfer, which led to a shift from a glucose-based energy metabolism to one based on sugar phosphates. The breakdown of glycolysis and many anabolic pathways for amino acids and cofactors in several subgroups was compensated by the independent acquisition of new uptake systems, including an ATP/ADP antiporter, from other gut microbiota. The putative donors are mostly flagellate endosymbionts from other bacterial phyla, including several, hitherto unknown lineages of uncultured Alphaproteobacteria, documenting the importance of horizontal gene transfer in the convergent evolution of these intracellular symbioses. The loss of almost all biosynthetic capacities in some lineages of Endomicrobiellum suggests that their originally mutualistic relationship with flagellates is on its decline.IMPORTANCEUnicellular eukaryotes are frequently colonized by bacterial and archaeal symbionts. A prominent example are the cellulolytic gut flagellates of termites, which harbor diverse but host-specific bacterial symbionts that occur exclusively in termite guts. One of these lineages, the so-called Endomicrobia, comprises both free-living and endosymbiotic representatives, which offers the unique opportunity to study the evolutionary processes underpinning the transition from a free-living to an intracellular lifestyle. Our results revealed a progressive gene loss in energy metabolism and biosynthetic pathways, compensated by the acquisition of new functions via horizontal gene transfer from other gut bacteria, and suggest the eventual breakdown of an initially mutualistic symbiosis. Evidence for convergent evolution of unrelated endosymbionts reflects adaptations to the intracellular environment of termite gut flagellates.

• Klíčová slova
• Endomicrobiaceae, Parabasalia, convergent evolution, endosymbionts, lateral gene transfer, termites,
• MeSH
• Bacteria * genetika   klasifikace   MeSH
• fylogeneze * MeSH
• genom bakteriální * MeSH
• Isoptera * mikrobiologie   parazitologie   MeSH
• metagenom MeSH
• molekulární evoluce MeSH
• přenos genů horizontální * MeSH
• střevní mikroflóra * MeSH
• symbióza * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH

BACKGROUND: Many arthropods rely on their gut microbiome to digest plant material, which is often low in nitrogen but high in complex polysaccharides. Detritivores, such as millipedes, live on a particularly poor diet, but the identity and nutritional contribution of their microbiome are largely unknown. In this study, the hindgut microbiota of the tropical millipede Epibolus pulchripes (large, methane emitting) and the temperate millipede Glomeris connexa (small, non-methane emitting), fed on an identical diet, were studied using comparative metagenomics and metatranscriptomics. RESULTS: The results showed that the microbial load in E. pulchripes is much higher and more diverse than in G. connexa. The microbial communities of the two species differed significantly, with Bacteroidota dominating the hindguts of E. pulchripes and Proteobacteria (Pseudomonadota) in G. connexa. Despite equal sequencing effort, de novo assembly and binning recovered 282 metagenome-assembled genomes (MAGs) from E. pulchripes and 33 from G. connexa, including 90 novel bacterial taxa (81 in E. pulchripes and 9 in G. connexa). However, despite this taxonomic divergence, most of the functions, including carbohydrate hydrolysis, sulfate reduction, and nitrogen cycling, were common to the two species. Members of the Bacteroidota (Bacteroidetes) were the primary agents of complex carbon degradation in E. pulchripes, while members of Proteobacteria dominated in G. connexa. Members of Desulfobacterota were the potential sulfate-reducing bacteria in E. pulchripes. The capacity for dissimilatory nitrate reduction was found in Actinobacteriota (E. pulchripes) and Proteobacteria (both species), but only Proteobacteria possessed the capacity for denitrification (both species). In contrast, some functions were only found in E. pulchripes. These include reductive acetogenesis, found in members of Desulfobacterota and Firmicutes (Bacillota) in E. pulchripes. Also, diazotrophs were only found in E. pulchripes, with a few members of the Firmicutes and Proteobacteria expressing the nifH gene. Interestingly, fungal-cell-wall-degrading glycoside hydrolases (GHs) were among the most abundant carbohydrate-active enzymes (CAZymes) expressed in both millipede species, suggesting that fungal biomass plays an important role in the millipede diet. CONCLUSIONS: Overall, these results provide detailed insights into the genomic capabilities of the microbial community in the hindgut of millipedes and shed light on the ecophysiology of these essential detritivores. Video Abstract.

• Klíčová slova
• Acetogens, Ecosystem engineers, Glycoside hydrolases, Hindgut microbiota, Millipede holobiont, Nutrient cycling, Polysaccharide degradation, Symbiosis,
• MeSH
• Bacteria MeSH
• Bacteroidetes genetika   MeSH
• členovci * genetika   MeSH
• dusík metabolismus   MeSH
• fylogeneze MeSH
• metagenom MeSH
• metagenomika MeSH
• Proteobacteria genetika   MeSH
• sacharidy MeSH
• sírany metabolismus   MeSH
• střevní mikroflóra * genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• audiovizuální média MeSH
• časopisecké články MeSH
• Názvy látek
• dusík MeSH
• sacharidy MeSH
• sírany MeSH

Methane emission by terrestrial invertebrates is restricted to millipedes, termites, cockroaches, and scarab beetles. The arthropod-associated archaea known to date belong to the orders Methanobacteriales, Methanomassiliicoccales, Methanomicrobiales, and Methanosarcinales, and in a few cases also to non-methanogenic Nitrososphaerales and Bathyarchaeales. However, all major host groups are severely undersampled, and the taxonomy of existing lineages is not well developed. Full-length 16S rRNA gene sequences and genomes of arthropod-associated archaea are scarce, reference databases lack resolution, and the names of many taxa are either not validly published or under-classified and require revision. Here, we investigated the diversity of archaea in a wide range of methane-emitting arthropods, combining phylogenomic analysis of isolates and metagenome-assembled genomes (MAGs) with amplicon sequencing of full-length 16S rRNA genes. Our results allowed us to describe numerous new species in hitherto undescribed taxa among the orders Methanobacteriales (Methanacia, Methanarmilla, Methanobaculum, Methanobinarius, Methanocatella, Methanoflexus, Methanorudis, and Methanovirga, all gen. nova), Methanomicrobiales (Methanofilum and Methanorbis, both gen. nova), Methanosarcinales (Methanofrustulum and Methanolapillus, both gen. nova), Methanomassiliicoccales (Methanomethylophilaceae fam. nov., Methanarcanum, Methanogranum, Methanomethylophilus, Methanomicula, Methanoplasma, Methanoprimaticola, all gen. nova), and the new family Bathycorpusculaceae (Bathycorpusculum gen. nov.). Reclassification of amplicon libraries from this and previous studies using this new taxonomic framework revealed that arthropods harbor only CO2 and methyl-reducing hydrogenotrophic methanogens. Numerous genus-level lineages appear to be present exclusively in arthropods, suggesting long evolutionary trajectories with their termite, cockroach, and millipede hosts, and a radiation into various microhabitats and ecological niches provided by their digestive tracts (e.g., hindgut compartments, gut wall, or anaerobic protists). The distribution patterns among the different host groups are often complex, indicating a mixed mode of transmission and a parallel evolution of invertebrate and vertebrate-associated lineages.

• Klíčová slova
• Bathyarchaeia, Nitrososphaerales, archaea, cockroaches, gut microbiota, methanogens, millipedes, termites,
• Publikační typ
• časopisecké články MeSH

Termites host diverse communities of gut microbes, including many bacterial lineages only found in this habitat. The bacteria endemic to termite guts are transmitted via two routes: a vertical route from parent colonies to daughter colonies and a horizontal route between colonies sometimes belonging to different termite species. The relative importance of both transmission routes in shaping the gut microbiota of termites remains unknown. Using bacterial marker genes derived from the gut metagenomes of 197 termites and one Cryptocercus cockroach, we show that bacteria endemic to termite guts are mostly transferred vertically. We identified 18 lineages of gut bacteria showing cophylogenetic patterns with termites over tens of millions of years. Horizontal transfer rates estimated for 16 bacterial lineages were within the range of those estimated for 15 mitochondrial genes, suggesting that horizontal transfers are uncommon and vertical transfers are the dominant transmission route in these lineages. Some of these associations probably date back more than 150 million years and are an order of magnitude older than the cophylogenetic patterns between mammalian hosts and their gut bacteria. Our results suggest that termites have cospeciated with their gut bacteria since first appearing in the geological record.

• Klíčová slova
• cophylogeny, endosymbionts, isoptera, metagenomics, vertical inheritance,
• MeSH
• Bacteria genetika   MeSH
• fylogeneze MeSH
• Isoptera * MeSH
• savci MeSH
• střevní mikroflóra * MeSH
• symbióza MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH

The decomposition of wood and detritus is challenging to most macroscopic organisms due to the recalcitrant nature of lignocellulose. Moreover, woody plants often protect themselves by synthesizing toxic or nocent compounds which infuse their tissues. Termites are essential wood decomposers in warmer terrestrial ecosystems and, as such, they have to cope with high concentrations of plant toxins in wood. In this paper, we evaluated the influence of wood age on the gut microbial (bacterial and fungal) communities associated with the termites Reticulitermes flavipes (Rhinotermitidae) (Kollar, 1837) and Microcerotermes biroi (Termitidae) (Desneux, 1905). We confirmed that the secondary metabolite concentration decreased with wood age. We identified a core microbial consortium maintained in the gut of R. flavipes and M. biroi and found that its diversity and composition were not altered by the wood age. Therefore, the concentration of secondary metabolites had no effect on the termite gut microbiome. We also found that both termite feeding activities and wood age affect the wood microbiome. Whether the increasing relative abundance of microbes with termite activities is beneficial to the termites is unknown and remains to be investigated. IMPORTANCE Termites can feed on wood thanks to their association with their gut microbes. However, the current understanding of termites as holobiont is limited. To our knowledge, no studies comprehensively reveal the influence of wood age on the termite-associated microbial assemblage. The wood of many tree species contains high concentrations of plant toxins that can vary with their age and may influence microbes. Here, we studied the impact of Norway spruce wood of varying ages and terpene concentrations on the microbial communities associated with the termites Reticulitermes flavipes (Rhinotermitidae) and Microcerotermes biroi (Termitidae). We performed a bacterial 16S rRNA and fungal ITS2 metabarcoding study to reveal the microbial communities associated with R. flavipes and M. biroi and their impact on shaping the wood microbiome. We noted that a stable core microbiome in the termites was unaltered by the feeding substrate, while termite activities influenced the wood microbiome, suggesting that plant secondary metabolites have negligible effects on the termite gut microbiome. Hence, our study shed new insights into the termite-associated microbial assemblage under the influence of varying amounts of terpene content in wood and provides a groundwork for future investigations for developing symbiont-mediated termite control measures.

• Klíčová slova
• Microcerotermes biroi, Reticulitermes flavipes, bacteria, core-microbiome, ectosymbionts, endosymbionts, fungi, plant defenses, terpenoids, wood-feeding termites,
• MeSH
• Bacteria genetika   MeSH
• dřevo * metabolismus   MeSH
• ekosystém MeSH
• Isoptera * mikrobiologie   MeSH
• RNA ribozomální 16S genetika   metabolismus   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• RNA ribozomální 16S MeSH

Termites are dominant animals of tropical terrestrial ecosystems. Their success is due to their eusocial organization as well as their ability to digest dead plant tissues. While being extremely abundant, the termite diet is poor in crucial nutrients, such as fatty acids. Linoleic acid (LA) is a precursor for many vital biomolecules, and most animals depend on its dietary supply. Termites count among the exceptions known to produce LA de novo, presumably via the action of an unknown Δ12 fatty acyl desaturase (FAD) introducing the second double bond into monounsaturated oleic acid. Here, we search for the evolutionary origin of LA biosynthesis in termites. To this end, we compile the repertoire of FAD homologs from 57 species of termites and their closest relatives, the cockroaches, analyze FAD phylogeny, and identify a potential Δ12 FAD branch, which arose through duplication of a likely Δ9 FAD. We functionally characterize both paralogs and identify the Δ9 activity in the ancestral FAD-A1a and the Δ12 activity responsible for LA biosynthesis in FAD-A1b. Through the combination of homology modeling and site-directed mutagenesis, we pinpoint structural features possibly contributing to the distinct functions, regiospecificities, and substrate preferences of the two enzymes. We confirm the presence of both paralogs in all 36 studied species of the Blattoidea lineage (Blattidae, Lamproblattidae, Cryptocercidae, and termites) and conclude that we identified an evolutionary event important for the ecological success of termites, which took place in their cockroach ancestors roughly 160 My and remained conserved throughout termite diversification into 3,000 extant species.

• Klíčová slova
• Blattodea, Isoptera, biosynthesis, fatty acyl desaturases, linoleic acid, termites,
• MeSH
• ekosystém MeSH
• fylogeneze MeSH
• Isoptera * genetika   MeSH
• kyselina linolová MeSH
• mastné kyseliny MeSH
• švábi * MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• kyselina linolová MeSH
• mastné kyseliny MeSH

Collembola are a group of globally distributed microarthropods that can tolerate low temperature and are active in extremely cold environments. While it is well known that animal diets can shape their microbiota, the microbiota of soil animals is not well described, particularly for animals with limited food resources, such as Collembola active in winter at low temperatures. In this study, we explored the effects of three different food sources; corn litter (agriculture grain residuals), Mongolian oak litter (natural plant residuals), and yeast (common food for Collembola culture), on the microbiota of a winter-active Collembola species, Desoria ruseki. We found that microbial diversity and community composition of the Collembola were strongly altered after feeding with different food sources for 30 days. Collembola individuals fed on corn litter harbored the highest bacterial richness and were dominated by a representative of Microbacteriaceae. In contrast, those fed on yeast exhibited the lowest bacterial richness and were primarily colonized by Pseudomonas. The microbial communities associated with the winter-active Collembola differed significantly from those observed in the food. Collembola nutrient turnover also differed when cultured with different food sources, as indicated by the C and N stable isotopic signatures. Our study highlights microbial associations with stable isotopic enrichments of the host. Specifically, the Arthrobacter was positively correlated with δ13C enrichment in the host. Representatives of Microbacteriaceae, Micrococcaceae, TM7a, Devosia, and Rathayibacter were positively correlated with δ15N enrichment of the host. Our study indicates that food sources are major determinants for Collembola microbiota that simultaneously alter consumers' isotopic niches, thereby improving our understanding of the roles played by host-microbiota interactions in sustaining soil biodiversity during the winter.

• Klíčová slova
• feeding ecology, host-microbiota interaction, soil arthropods, springtail, stable isotope, winter biodiversity,
• Publikační typ
• časopisecké články MeSH

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. Video Abstract.

• Klíčová slova
• Endosymbionts, Isoptera, Metagenomics, Vertical inheritance,
• MeSH
• fylogeneze MeSH
• Isoptera * MeSH
• metagenom MeSH
• půda MeSH
• střevní mikroflóra * genetika   MeSH
• zvířata MeSH
• Check Tag
• zvířata MeSH
• Publikační typ
• audiovizuální média MeSH
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• půda MeSH