The galatheid crab, Shinkaia crosnieri (Decapoda: Galatheidae), forms dense colonies in the Iheya North and Hatoma Knoll deep-sea hydrothermal fields and has numerous setae covered with filamentous epibiotic microorganisms. Molecular phylogenetic analyses revealed that the epibiotic communities in S. crosnieri consisted mainly of yet-uncultivated phylotypes within Epsilonproteobacteria and Gammaproteobacteria in both hydrothermal vent fields. Uptake experiments using 13 C-labeled tracers clearly demonstrated that both H 13 CO3 − and 13 CH4 were assimilated into not only the epibiotic microbial communities associated with the setae, but also the epibiont-free tissue of living S. crosnieri. In addition, the incorporation of H 13 CO3 − into the microbial cells was strongly stimulated by the presence of reduced sulfur compounds but not by H2. In conclusion, the uptake experiments suggested that sulfur-oxidizing chemolithoautotrophic and methanotrophic production by the epibionts provides the nutrition for S. crosnieri.

Symbiobacterium thermophilum is an uncultivable bacterium isolated from compost that depends on microbial commensalism. The 16S ribosomal DNA-based phylogeny suggests that this bacterium belongs to an unknown taxon in the Gram-positive bacterial cluster. Here, we describe the 3.57 Mb genome sequence of S.thermophilum. The genome consists of 3338 protein-coding sequences, out of which 2082 have functional assignments. Despite the high G + C content (68.7%), the genome is closest to that of Firmicutes, a phylum consisting of low G + C Gram-positive bacteria. This provides evidence for the presence of an undefined category in the Gram-positive bacterial group. The presence of both spo and related genes and microscopic observation indicate that S.thermophilum is the first high G + C organism that forms endospores. The S.thermophilum genome is also characterized by the widespread insertion of class C group II introns, which are oriented in the same direction as chromosomal replication. The genome has many membrane transporters, a number of which are involved in the uptake of peptides and amino acids. The genes involved in primary metabolism are largely identified, except those that code several biosynthetic enzymes and carbonic anhydrase. The organism also has a variety of respiratory systems including Nap nitrate reductase, which has been found only in Gram-negative bacteria. Overall, these features suggest that S.thermophilum is adaptable to and thus lives in various environments, such that its growth requirement could be a substance or a physiological condition that is generally available in the natural environment rather than a highly specific substance that is present only in a limited niche. The genomic information from S.thermophilum offers new insights into microbial diversity and evolutionary sciences, and provides a framework for characterizing the molecular basis underlying microbial commensalism.

Deep-sea vents harbor dense populations of various animals that have their specific symbiotic bacteria. Scaly-foot gastropods, which are snails with mineralized scales covering the sides of its foot, have a gammaproteobacterial endosymbiont in their enlarged esophageal glands and diverse epibionts on the surface of their scales. In this study, we report the complete genome sequencing of gammaproteobacterial endosymbiont. The endosymbiont genome displays features consistent with ongoing genome reduction such as large proportions of pseudogenes and insertion elements. The genome encodes functions commonly found in deep-sea vent chemoautotrophs such as sulfur oxidation and carbon fixation. Stable carbon isotope (13C)-labeling experiments confirmed the endosymbiont chemoautotrophy. The genome also includes an intact hydrogenase gene cluster that potentially has been horizontally transferred from phylogenetically distant bacteria. Notable findings include the presence and transcription of genes for flagellar assembly, through which proteins are potentially exported from bacterium to the host. Symbionts of snail individuals exhibited extreme genetic homogeneity, showing only two synonymous changes in 19 different genes (13 810 positions in total) determined for 32 individual gastropods collected from a single colony at one time. The extremely low genetic individuality in endosymbionts probably reflects that the stringent symbiont selection by host prevents the random genetic drift in the small population of horizontally transmitted symbiont. This study is the first complete genome analysis of gastropod endosymbiont and offers an opportunity to study genome evolution in a recently evolved endosymbiont.

S)mbiobacterium themaophilum is a strictly symbiotic thermophile, the growth of which is dependent on the coexjstence of an associating thermophilic Bacilins sp., strain S. S. thermophiinm grows only in mixed culture with the BacMtzs strajn in liquid media, and does not ferm visible colonies on solid inedia. To measure the growth of this symbiotic bacterium and to analyze its growth requirements, we developed a quantitative PCR method by uslng its specific sequences in a putatiye membralle translocator gene tnaT as primers. Aecording to this method, independent growth of S. thennophilum was first confirmed in a dialyzing culture physically separated frem Bacilins strain S with a ce}]ulose membra"e. Independent growth of SL thermqphiinm was alse managed by adding conditioned medium prepared from the cultllre Mtrate of the BaciUus strai", but the growth in the conditioned medium stopped at a very ]imited extent with appearance of filamentous cells, suggesting the llncoupling of cellular grewth and cell diyisioll. Formation ef micro-colonies of S. thermophilum was observed en the collditioned agar medium under both aerobic and anaerobic conditions, but the coleny-forming eMciencies remained below 1 %. Several other bacterial species, such as Bacilins stearothenmoph"tts, Bacillas subtiZis, 11hermus thermophiltzs, and eyen Escherichia coli, were also found to suppert the grewth of S. thermophiinm. These results indicate that S. thermophiinm essentially requires some ubiquitous metabolite(s) of low mo}ecular weight produced by various bacterial species as growth factor(s) but coexistence of the living partner cells is stil1 required, probably to maintain an effectiye level of the putative factor{s) in the medium.

In this study, we report experimental evidence of the thioautotrophic activity of the epibiotic microbial community associated with the setae of Shinkaia crosnieri, a galatheid crab that is endemic to deep-sea hydrothermal systems in the Okinawa Trough in Japan. Microbial consumption of reduced sulfur compounds under in situ hydrostatic and atmospheric pressure provided evidence of sulfur-oxidizing activity by the epibiotic microbial community; the rate of sulfur oxidation was similar under in situ and decompressed conditions. Results of the microbial consumption of reduced sulfur compounds and tracer experiments using 13C-labeled bicarbonate in the presence and absence of thiosulfate (used as a thioautotrophic substrate) convincingly demonstrated that the epibiotic microbial community on S. crosnieri drove primary production via an energy metabolism that was coupled with the oxidation of reductive sulfur compounds. A combination of tracer experiments, fluorescence in situ hybridization (FISH) and nano-scale secondary ion mass spectrometry (Nano-SIMS) indicated that the filamentous cells of the genus Sulfurovum belonging to the class Epsilonproteobacteria were thioautotrophs in the epibiotic community of S. crosnieri. In conclusion, our results strongly suggest that thioautotrophic production by Sulfurovum members present as the epibiotic microbial community play a predominant role in a probable nutritional ectosymbiosis with S. crosnieri.