Carsten Dietrich

High-Resolution Analysis of Gut Environment and Bacterial Microbiota Reveals Functional Compartmentation of the Gut in Wood-Feeding Higher Termites (Nasutitermes spp.)

ABSTRACT Higher termites are characterized by a purely prokaryotic gut microbiota and an increased compartmentation of their intestinal tract. In soil-feeding species, each gut compartment has different physicochemical conditions and is colonized by a specific microbial community. Although considerable information has accumulated also for wood-feeding species of the genus Nasutitermes, including cellulase activities and metagenomic data, a comprehensive study linking physicochemical gut conditions with the structure of the microbial communities in the different gut compartments is lacking. In this study, we measured high-resolution profiles of H2, O2, pH, and redox potential in the gut of Nasutitermes corniger termites, determined the fermentation products accumulating in the individual gut compartments, and analyzed the bacterial communities in detail by pyrotag sequencing of the V3-V4 region of the 16S rRNA genes. The dilated hindgut paunch (P3 compartment) was the only anoxic gut region, showed the highest density of bacteria, and accumulated H2 to high partial pressures (up to 12 kPa). Molecular hydrogen is apparently produced by a dense community of Spirochaetes and Fibrobacteres, which also dominate the gut of other Nasutitermes species. All other compartments, such as the alkaline P1 compartment (average pH, 10.0), showed high redox potentials and comprised small but distinct populations characteristic for each gut region. In the crop and the posterior hindgut compartments, the community was even more diverse than in the paunch. Similarities in the communities of the posterior hindgut and crop suggested that proctodeal trophallaxis or coprophagy also occurs in higher termites. The large sampling depths of pyrotag sequencing in combination with the determination of important physicochemical parameters allow cautious conclusions concerning the functions of particular bacterial lineages in the respective gut sections to be drawn.

The cockroach origin of the termite gut microbiota: patterns in bacterial community structure reflect major evolutionary events.

ABSTRACT Termites digest wood and other lignocellulosic substrates with the help of their intestinal microbiota. While the functions of the symbionts in the digestive process are slowly emerging, the origin of the bacteria colonizing the hindgut bioreactor is entirely unknown. Recently, our group discovered numerous representatives of bacterial lineages specific to termite guts in a closely related omnivorous cockroach, but it remains unclear whether they derive from the microbiota of a common ancestor or were independently selected by the gut environment. Here, we studied the bacterial gut microbiota in 34 species of termites and cockroaches using pyrotag analysis of the 16S rRNA genes. Although the community structures differed greatly between the major host groups, with dramatic changes in the relative abundances of particular bacterial taxa, we found that the majority of sequence reads belonged to bacterial lineages that were shared among most host species. When mapped onto the host tree, the changes in community structure coincided with major events in termite evolution, such as acquisition and loss of cellulolytic protists and the ensuing dietary diversification. UniFrac analysis of the core microbiota of termites and cockroaches and construction of phylogenetic tree of individual genus level lineages revealed a general host signal, whereas the branching order often did not match the detailed phylogeny of the host. It remains unclear whether the lineages in question have been associated with the ancestral cockroach since the early Cretaceous (cospeciation) or are diet-specific lineages that were independently acquired from the environment (host selection).

The Gut Microbiota of Termites: Digesting the Diversity in the Light of Ecology and Evolution

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. Whereas the intestinal communities in the closely related cockroaches seem 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.

Cartwheel Architecture of Trichonympha Basal Body

Electron microscopy provides a close-up view of the ninefold-symmetric stacked rings at the base of cilia and flagella. Centrioles and basal bodies are essential for the formation of cilia, flagella, and centrosomes. They exhibit a characteristic ninefold symmetry imparted by a cartwheel thought to contain rings of SAS-6 proteins. We used cryoelectron tomography to investigate the architecture of the exceptionally long cartwheel of the flagellate Trichonympha. We found that the cartwheel is a stack of central rings that exhibit a vertical periodicity of 8.5 nanometers and is able to accommodate nine SAS-6 homodimers. The spokes that emanate from two such rings associate into a layer, with a vertical periodicity of 17 nanometers on the cartwheel margin. Thus, by using the power of biodiversity, we unveiled the architecture of the cartwheel at the root of the ninefold symmetry of centrioles and basal bodies.

Diet is the primary determinant of bacterial community structure in the guts of higher termites.

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) harbour 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 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 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 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.

Honeybee gut microbiota promotes host weight gain via bacterial metabolism and hormonal signaling

Significance Honey bees are globally important plant pollinators. Guts of adult workers contain specialized bacteria not found outside bees. Experimental results show that gut bacteria increase weight gain in young adult bees, affect expression of genes governing insulin and vitellogenin levels, and increase sucrose sensitivity. Gut bacteria also shape the physicochemical conditions within the gut, lowering pH and oxygen levels. Peripheral resident bacteria consume oxygen, thus maintaining anoxia, as required for microbial activity. Additionally, gut bacteria produce short-chain fatty acids, with acetate and propionate as the major metabolites, as in guts of human and other animals. This study demonstrates how bacteria in the honey bee gut affect host weight gain and improves our understanding of how gut symbionts influence host health. Social bees harbor a simple and specialized microbiota that is spatially organized into different gut compartments. Recent results on the potential involvement of bee gut communities in pathogen protection and nutritional function have drawn attention to the impact of the microbiota on bee health. However, the contributions of gut microbiota to host physiology have yet to be investigated. Here we show that the gut microbiota promotes weight gain of both whole body and the gut in individual honey bees. This effect is likely mediated by changes in host vitellogenin, insulin signaling, and gustatory response. We found that microbial metabolism markedly reduces gut pH and redox potential through the production of short-chain fatty acids and that the bacteria adjacent to the gut wall form an oxygen gradient within the intestine. The short-chain fatty acid profile contributed by dominant gut species was confirmed in vitro. Furthermore, metabolomic analyses revealed that the gut community has striking impacts on the metabolic profiles of the gut compartments and the hemolymph, suggesting that gut bacteria degrade plant polymers from pollen and that the resulting metabolites contribute to host nutrition. Our results demonstrate how microbial metabolism affects bee growth, hormonal signaling, behavior, and gut physicochemical conditions. These findings indicate that the bee gut microbiota has basic roles similar to those found in some other animals and thus provides a model in studies of host–microbe interactions.

Agl16, a Thermophilic Glycosyltransferase Mediating the Last Step of N-Glycan Biosynthesis in the Thermoacidophilic Crenarchaeon Sulfolobus acidocaldarius

Recently, the S-layer protein of Sulfolobus acidocaldarius was shown to be N-linked with a tribranched hexasaccharide, composed of Man2Glc1GlcNAc2 and a sulfated sugar called sulfoquinovose. To identify genes involved in the biosynthesis and attachment of this glycan, markerless in-frame deletions of genes coding for predicted glycosyltransferases were created. The successful deletion of agl16, coding for a glycosyltransferase, resulted in the S-layer protein and archaellins having reduced molecular weights, as visualized by Coomassie staining or immunoblotting. This analysis indicated a change in the N-glycan composition. Nano-liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses confirmed that the glycan of the S-layer protein from the agl16 deletion mutant was a pentasaccharide, which was missing a terminal hexose residue. High-performance liquid chromatography (HPLC) analyses of the hydrolyzed N-glycan indicated that the missing hexose is a glucose residue. A physiological characterization of the agl16 deletion mutant revealed a significant effect on the growth at elevated salt concentrations. At 300 mM NaCl, the doubling time of the Δagl16 mutant was increased 2-fold compared to that of the background strain. Furthermore, the incomplete glycan structure of the Δagl16 deletion strain affected the assembly and function of the archaellum, as exemplified by semisolid Gelrite plate analysis, in which the motility is decreased according to the N-glycan size.

Population Structure of Endomicrobia in Single Host Cells of Termite Gut Flagellates (Trichonympha spp.)

The gut microbiota of many phylogenetically lower termites is dominated by the cellulolytic flagellates of the genus Trichonympha, which are consistently associated with bacterial symbionts. In the case of Endomicrobia, an unusual lineage of endosymbionts of the Elusimicrobia phylum that is also present in other gut flagellates, previous studies have documented strict host specificity, leading to the cospeciation of “Candidatus Endomicrobium trichonymphae” with their respective flagellate hosts. However, it currently remains unclear whether one Trichonympha species is capable of harboring more than one Endomicrobia phylotype. In the present study, we selected single Trichonympha cells from the guts of Zootermopsis nevadensis and Reticulitermes santonensis and characterized their Endomicrobia populations based on internal transcribed spacer (ITS) region sequences. We found that each host cell harbored a homogeneous population of symbionts that were specific to their respective host species, but phylogenetically distinct between each host lineage, corroborating cospeciation being caused by vertical inheritance. The experimental design of the present study also allowed for the identification of an unexpectedly large amount of tag-switching between samples, which indicated that any high-resolution analysis of microbial community structures using the pyrosequencing technique has to be interpreted with great caution.

Metagenomic analysis of the microbiota in the highly compartmented hindguts of six wood- or soil-feeding higher termites

BackgroundTermites are important contributors to carbon and nitrogen cycling in tropical ecosystems. Higher termites digest lignocellulose in various stages of humification with the help of an entirely prokaryotic microbiota housed in their compartmented intestinal tract. Previous studies revealed fundamental differences in community structure between compartments, but the functional roles of individual lineages in symbiotic digestion are mostly unknown.ResultsHere, we conducted a highly resolved analysis of the gut microbiota in six species of higher termites that feed on plant material at different levels of humification. Combining amplicon sequencing and metagenomics, we assessed similarities in community structure and functional potential between the major hindgut compartments (P1, P3, and P4). Cluster analysis of the relative abundances of orthologous gene clusters (COGs) revealed high similarities among wood- and litter-feeding termites and strong differences to humivorous species. However, abundance estimates of bacterial phyla based on 16S rRNA genes greatly differed from those based on protein-coding genes.ConclusionCommunity structure and functional potential of the microbiota in individual gut compartments are clearly driven by the digestive strategy of the host. The metagenomics libraries obtained in this study provide the basis for future studies that elucidate the fundamental differences in the symbiont-mediated breakdown of lignocellulose and humus by termites of different feeding groups. The high proportion of uncultured bacterial lineages in all samples calls for a reference-independent approach for the correct taxonomic assignment of protein-coding genes.

Epigenome-Wide Association Study Identifies Cardiac Gene Patterning and a Novel Class of Biomarkers for Heart Failure

Background: Biochemical DNA modification resembles a crucial regulatory layer among genetic information, environmental factors, and the transcriptome. To identify epigenetic susceptibility regions and novel biomarkers linked to myocardial dysfunction and heart failure, we performed the first multi-omics study in myocardial tissue and blood of patients with dilated cardiomyopathy and controls. Methods: Infinium human methylation 450 was used for high-density epigenome-wide mapping of DNA methylation in left-ventricular biopsies and whole peripheral blood of living probands. RNA deep sequencing was performed on the same samples in parallel. Whole-genome sequencing of all patients allowed exclusion of promiscuous genotype-induced methylation calls. Results: In the screening stage, we detected 59 epigenetic loci that are significantly associated with dilated cardiomyopathy (false discovery corrected P⩽0.05), with 3 of them reaching epigenome-wide significance at P⩽5×10−8. Twenty-seven (46%) of these loci could be replicated in independent cohorts, underlining the role of epigenetic regulation of key cardiac transcription regulators. Using a staged multi-omics study design, we link a subset of 517 epigenetic loci with dilated cardiomyopathy and cardiac gene expression. Furthermore, we identified distinct epigenetic methylation patterns that are conserved across tissues, rendering these CpGs novel epigenetic biomarkers for heart failure. Conclusions: The present study provides to our knowledge the first epigenome-wide association study in living patients with heart failure using a multi-omics approach.