Masahiro Yuki

Acetogenesis from H2 plus CO2 and nitrogen fixation by an endosymbiotic spirochete of a termite-gut cellulolytic protist.

Symbiotic associations of cellulolytic eukaryotic protists and diverse bacteria are common in the gut microbial communities of termites. Besides cellulose degradation by the gut protists, reductive acetogenesis from H2 plus CO2 and nitrogen fixation by gut bacteria play crucial roles in the host termites’ nutrition by contributing to the energy demand of termites and supplying nitrogen poor in their diet, respectively. Fractionation of these activities and the identification of key genes from the gut community of the wood-feeding termite Hodotermopsis sjoestedti revealed that substantial activities in the gut—nearly 60% of reductive acetogenesis and almost exclusively for nitrogen fixation—were uniquely attributed to the endosymbiotic bacteria of the cellulolytic protist in the genus Eucomonympha. The rod-shaped endosymbionts were surprisingly identified as a spirochete species in the genus Treponema, which usually exhibits a characteristic spiral morphology. The endosymbionts likely use H2 produced by the protist for these dual functions. Although H2 is known to inhibit nitrogen fixation in some bacteria, it seemed to rather stimulate this important mutualistic process. In addition, the single-cell genome analyses revealed the endosymbionts potentials of the utilization of sugars for its energy requirement, and of the biosynthesis of valuable nutrients such as amino acids from the fixed nitrogen. These metabolic interactions are suitable for the dual functions of the endosymbiont and reconcile its substantial contributions in the gut.

Comparison of intracellular “Ca. Endomicrobium trichonymphae” genomovars illuminates the requirement and decay of defense systems against foreign DNA

“Candidatus Endomicrobium trichonymphae” (Bacteria; Elusimicrobia) is an obligate intracellular symbiont of the cellulolytic protist genus Trichonympha in the termite gut. A previous genome analysis of “Ca. Endomicrobium trichonymphae” phylotype Rs-D17 (genomovar Ri2008), obtained from a Trichonympha agilis cell in the gut of the termite Reticulitermes speratus, revealed that its genome is small (1.1 Mb) and contains many pseudogenes; it is in the course of reductive genome evolution. Here we report the complete genome sequence of another Rs-D17 genomovar, Ti2015, obtained from a different T. agilis cell present in an R. speratus gut. These two genomovars share most intact protein-coding genes and pseudogenes, showing 98.6% chromosome sequence similarity. However, characteristic differences were found in their defense systems, which comprised restriction-modification and CRISPR/Cas systems. The repertoire of intact restriction-modification systems differed between the genomovars, and two of the three CRISPR/Cas loci in genomovar Ri2008 are pseudogenized or missing in genomovar Ti2015. These results suggest relaxed selection pressure for maintaining these defense systems. Nevertheless, the remaining CRISPR/Cas system in each genomovar appears to be active; none of the “spacer” sequences (112 in Ri2008 and 128 in Ti2015) were shared whereas the “repeat” sequences were identical. Furthermore, we obtained draft genomes of three additional endosymbiotic Endomicrobium phylotypes from different host protist species, and discovered multiple, intact CRISPR/Cas systems in each genome. Collectively, unlike bacteriome endosymbionts in insects, the Endomicrobium endosymbionts of termite-gut protists appear to require defense against foreign DNA, although the required level of defense has likely been reduced during their intracellular lives.

Draft Genome Sequences of Psychrobacter Strains JCM 18900, JCM 18901, JCM 18902, and JCM 18903, Isolated Preferentially from Frozen Aquatic Organisms

ABSTRACT Four Psychrobacter strains, JCM 18900, JCM 18901, JCM 18902, and JCM 18903, related to either Psychrobacter nivimaris or Psychrobacter cibarius, were isolated from frozen marine animals. The genome information of these four strains will be useful for studies of their physiology and adaptation properties to frozen conditions.

Phylogenetic Diversity and Single-Cell Genome Analysis of “Melainabacteria”, a Non-Photosynthetic Cyanobacterial Group, in the Termite Gut

Termite guts harbor diverse yet-uncultured bacteria, including a non-photosynthetic cyanobacterial group, the class “Melainabacteria”. We herein reported the phylogenetic diversity of “Melainabacteria” in the guts of diverse termites and conducted a single-cell genome analysis of a melainabacterium obtained from the gut of the termite Termes propinquus. We performed amplicon sequencing of 16S rRNA genes from the guts of 60 termite and eight cockroach species, and detected melainabacterial sequences in 48 out of the 68 insect species, albeit with low abundances (0.02–1.90%). Most of the melainabacterial sequences obtained were assigned to the order “Gastranaerophilales” and appeared to form clusters unique to termites and cockroaches. A single-cell genome of a melainabacterium, designated phylotype Tpq-Mel-01, was obtained using a fluorescence-activated cell sorter and whole genome amplification. The genome shared basic features with other melainabacterial genomes previously reconstructed from the metagenomes of human and koala feces. The bacterium had a small genome (~1.6 Mb) and possessed fermentative pathways possibly using sugars and chitobiose as carbon and energy sources, while the pathways for photosynthesis and carbon fixation were not found. The genome contained genes for flagellar components and chemotaxis; therefore, the bacterium is likely motile. A fluorescence in situ hybridization analysis showed that the cells of Tpq-Mel-01 and/or its close relatives are short rods with the dimensions of 1.1±0.2 μm by 0.5±0.1 μm; for these bacteria, we propose the novel species, “Candidatus Gastranaerophilus termiticola”. Our results provide fundamental information on “Melainabacteria” in the termite gut and expand our knowledge on this underrepresented, non-photosynthetic cyanobacterial group.

Lactococcus reticulitermitis sp. nov., isolated from the gut of the subterranean termite Reticulitermes speratus

Strain Rs-Y01T was isolated from the gut of the wood-feeding subterranean termite Reticulitermes speratus. Phylogenetic analysis based on 16S rRNA gene sequence indicated that the strain Rs-Y01T belonged to the genus Lactococcus and was most closely related to Lactococcus raffinolactis JCM 5706T with 98.1 % similarity in the 16S rRNA gene, followed by Lactococcus piscium JCM 16647T (97.2 %). Genomic comparisons of strain Rs-Y01T with L. raffinolactis JCM 5706Twere made using the Genome-to-Genome Distance Calculator and average nucleotide identity analysis (values indicated 29.2 and 84.6 %, respectively). Strain Rs-Y01T was a Gram-stain-positive, facultatively anaerobic, non-motile coco-bacilli and formed l-lactic acid. The sugar fermentation and enzyme reactions of strain Rs-Y01T differed from those of other species of the genus Lactococcus. The major cellular fatty acids were summed feature 8 (C18 : 1ω7c/C18 : 1ω6c; 32.0 %), C16 : 0 (29.7 %) and C14 : 0 (18.1 %). Based on these characteristics, strain Rs-Y01T represents a novel species of the genus Lactococcus, for which the name Lactococcusreticulitermitis sp. nov. is proposed. The type strain is Rs-Y01T (=JCM 32106T=DSM 105715T).

Genome of |[lsquo]|Ca. Desulfovibrio trichonymphae|[rsquo]|, an H2-oxidizing bacterium in a tripartite symbiotic system within a protist cell in the termite gut

The cellulolytic protist Trichonympha agilis in the termite gut permanently hosts two symbiotic bacteria, ‘Candidatus Endomicrobium trichonymphae’ and ‘Candidatus Desulfovibrio trichonymphae’. The former is an intracellular symbiont, and the latter is almost intracellular but still connected to the outside via a small pore. The complete genome of ‘Ca. Endomicrobium trichonymphae’ has previously been reported, and we here present the complete genome of ‘Ca. Desulfovibrio trichonymphae’. The genome is small (1 410 056 bp), has many pseudogenes, and retains biosynthetic pathways for various amino acids and cofactors, which are partially complementary to those of ‘Ca. Endomicrobium trichonymphae’. An amino acid permease gene has apparently been transferred between the ancestors of these two symbionts; a lateral gene transfer has affected their metabolic capacity. Notably, ‘Ca. Desulfovibrio trichonymphae’ retains the complex system to oxidize hydrogen by sulfate and/or fumarate, while genes for utilizing other substrates common in desulfovibrios are pseudogenized or missing. Thus, ‘Ca. Desulfovibrio trichonymphae’ is specialized to consume hydrogen that may otherwise inhibit fermentation processes in both T. agilis and ‘Ca. Endomicrobium trichonymphae’. The small pore may be necessary to take up sulfate. This study depicts a genome-based model of a multipartite symbiotic system within a cellulolytic protist cell in the termite gut.

Draft Genome Sequence of Cytophaga fermentans JCM 21142T, a Facultative Anaerobe Isolated from Marine Mud

ABSTRACT Cytophaga fermentans strain JCM 21142T is a marine-dwelling facultative anaerobe. The draft genome sequence of this strain revealed its diverse chemoorganotrophic potential, which makes it capable of metabolizing various polysaccharide substrates. The genome data will facilitate further studies on its taxonomic reclassification, its metabolism, and the mechanisms pertaining to bacterial gliding.

Host-Symbiont Cospeciation of Termite-Gut Cellulolytic Protists of the Genera Teranympha and Eucomonympha and their Treponema Endosymbionts

Cellulolytic flagellated protists inhabit the hindgut of termites. They are unique and essential to termites and related wood-feeding cockroaches, enabling host feeding on cellulosic matter. Protists of two genera in the family Teranymphidae (phylum Parabasalia), Eucomonympha and Teranympha, are phylogenetically closely related and harbor intracellular endosymbiotic bacteria from the genus Treponema. In order to obtain a clearer understanding of the evolutionary history of this triplex symbiotic relationship, the molecular phylogenies of the three symbiotic partners, the Teranymphidae protists, their Treponema endosymbionts, and their host termites, were inferred and compared. Strong congruence was observed in the tree topologies of all interacting partners, implying their cospeciating relationships. In contrast, the coevolutionary relationship between the Eucomonympha protists and their endosymbionts was more complex, and evidence of incongruence against cospeciating relationships suggested frequent host switches of the endosymbionts, possibly because multiple Eucomonympha species are present in the same gut community. Similarities in the 16S rRNA and gyrB gene sequences of the endosymbionts were higher among Teranympha spp. (>99.25% and >97.2%, respectively), whereas those between Teranympha and Eucomonympha were lower (<97.1% and <91.9%, respectively). In addition, the endosymbionts of Teranympha spp. formed a phylogenetic clade distinct from those of Eucomonympha spp. Therefore, the endosymbiont species of Teranympha spp., designated here as “Candidatus Treponema teratonymphae”, needs to be classified as a species distinct from the endosymbiont species of Eucomonympha spp.

Lawsonibacter asaccharolyticus gen. nov., sp. nov., a butyrate-producing bacterium isolated from human faeces

An obligately anaerobic, Gram-positive, non-spore-forming, straight rod-shaped bacterium, designated strain 3BBH22T, was isolated from a faecal sample of a healthy Japanese woman. The 16S rRNA gene sequence analysis showed that strain 3BBH22T formed a monophyletic cluster with species in the genera Pseudoflavonifractor and Flavonifractor within the family Ruminococcaceae and had highest similarity to Pseudoflavonifractor capillosus ATCC 29799T (96.7 % sequence similarity), followed by Flavonifractor plautii ATCC 29863T (96.4 %). Acetate and butyrate were produced by strain 3BBH22T as metabolic end-products. The major cellular fatty acids were C14 : 0, C16 : 0, C18 : 1ω9c, C16 : 0 dimethyl acetal, C18 : 0 and C18 : 2ω6,9c. No respiratory quinones were detected. In contrast to F. plautii JCM 32125T, strain 3BBH22T did not degrade quercetin, one of the flavonoids. P. capillosus JCM 32126T also did not. Strain 3BBH22T was differentiated from P. capillosus JCM 32126T by its inability to hydrolyse aesculin. The G+C content of the genomic DNA was 61.2±1.0 mol%. On the basis of these data and the phylogenetic tree based on 89 proteins, strain 3BBH22T represents a novel species in a novel genus of the family Ruminococcaceae, for which the name Lawsonibacter asaccharolyticus gen. nov., sp. nov. is proposed. The type strain of L. asaccharolyticus is 3BBH22T (=JCM 32166T=DSM 106493T).

Different transferability of incompatibility (Inc) P-7 plasmid pCAR1 and IncP-1 plasmid pBP136 in stirring liquid conditions

[This corrects the article DOI: 10.1371/journal.pone.0186248.].