Metabolic diversification of anaerobic methanotrophic archaea in a deep-sea cold seep

Anaerobic methanotrophic archaea (ANME) can assimilate methane and govern the greenhouse effect of deep-sea cold seeps. In this study, a total of 13 ANME draft genomes representing five ANME types (ANME-1a, ANME-1b, ANME-2a, ANME-2b and ANME-2c), in size between 0.8 and 1.8 Mbp, were obtained from the Jiaolong cold seep in the South China Sea. The small metagenome-assembled genomes (MAGs) contained all the essential pathways for methane oxidization and carbon dioxide fixation. All genes related to nitrate and sulfate reduction were absent from the MAGs, indicating their syntrophic dependence on partner organisms. Aside from acetate secretion and sugar storage, propanoate synthesis pathway, as an alternative novel carbon flow, was identified in all the MAGs and transcriptionally active. Regarding type-specific features of the MAGs, the genes encoding archaellum and bacteria-derived chemotaxis were specific to ANME-2, perhaps for fitness under fluctuation of methane and sulfate concentration flux. Our genomic and transcriptomic results strongly suggested that ANME could carry out simple carbon metabolism from C1 assimilation to C3 biosynthesis in the SCS cold seep, which casts light on a novel approach for synthetic biology.

Molecular identification of methane monooxygenase and quantitative analysis of methanotrophic endosymbionts under laboratory maintenance inBathymodiolus platifronsfrom the South China Sea

Deep-sea mussels of the genusBathymodiolusare numerically dominant macrofauna in many cold seep and hydrothermal vent ecosystems worldwide, and they depend on organic carbon produced by symbionts present in the epithelial cells of the gills. AlthoughBathymodiolus platifronsrepresents typical methanotrophic endosymbiosis, our understanding of molecular mechanisms of methane oxidization and carbon fixation is still in its infancy. Moreover, the laboratory maintenance ofB. platifronsand the symbiont abundance dynamics during maintenance has not been reported. In the present study, we report the first systematic identification and phylogenetic analysis of three subunits of methane monooxygenase (pmoA, pmoB, and pmoC) obtained from the endosymbiotic bacteria found inB. platifrons. The coding sequences (CDS) of the three genes in theB. platifronsendosymbiont were 750, 1,245, and 753 bp, encoding 249, 414, and 250 amino acids, respectively. Sequence alignment and phylogenetic analysis revealed that the symbiont ofB. platifronsbelongs to the type I methanotrophs. In order to clarify the impact of environmental methane on symbiont abundance, a 34-day laboratory maintenance experiment was conducted in whichB. platifronsindividuals were acclimatized to methane-present and methane-absent environments. Symbiont abundance was evaluated by calculating the relative DNA content of the methane monooxygenase gene using quantitative real-time PCR. We found that symbiont quantity immediately decreased from its initial level, then continued to gradually decline during maintenance. At 24 and 34 days of maintenance, symbiont abundance in the methane-absent environment had significantly decreased compared to that in the methane-present environment, indicating that the maintenance of symbionts relies on a continuous supply of methane. Our electron microscopy results validated the qPCR analysis. This study enriches our knowledge of the molecular basis and the dynamic changes of the methanotrophic endosymbiosis inB. platifrons, and provides a feasible model biosystem for further investigation of methane oxidization, the carbon fixation process, and environmental adaptations of deep-sea mussels.