Carbon metabolism and biogeography of candidate phylum “Candidatus Bipolaricaulota” in geothermal environments of Biga Peninsula, Turkey

Abstract

Terrestrial hydrothermal springs and aquifers are excellent sites to study microbial biogeography because of their high physicochemical heterogeneity across relatively limited geographic regions. In this study, we performed 16S rRNA gene sequencing and metagenomic analyses of the microbial diversity of 11 different geothermal aquifers and springs across the tectonically active Biga Peninsula (Turkey). Across geothermal settings ranging in temperature from 43 to 79°C, one of the most highly represented groups in both 16S rRNA gene and metagenomic datasets was affiliated with the uncultivated phylum “Candidatus Bipolaricaulota” (former “Ca. Acetothermia” and OP1 division). The highest relative abundance of “Ca. Bipolaricaulota” was observed in a 68°C geothermal brine sediment, where it dominated the microbial community, representing 91% of all detectable 16S rRNA genes. Correlation analysis of “Ca. Bipolaricaulota” operational taxonomic units (OTUs) with physicochemical parameters indicated that salinity was the strongest environmental factor measured associated with the distribution of this novel group in geothermal fluids. Correspondingly, analysis of 23 metagenome-assembled genomes (MAGs) revealed two distinct groups of “Ca. Bipolaricaulota” MAGs based on the differences in carbon metabolism: one group encoding the bacterial Wood-Ljungdahl pathway (WLP) for H2 dependent CO2 fixation is selected for at lower salinities, and a second heterotrophic clade that lacks the WLP that was selected for under hypersaline conditions in the geothermal brine sediment. In conclusion, our results highlight that the biogeography of “Ca. Bipolaricaulota” taxa is strongly correlated with salinity in hydrothermal ecosystems, which coincides with key differences in carbon acquisition strategies. The exceptionally high relative abundance of apparently heterotrophic representatives of this novel candidate Phylum in geothermal brine sediment observed here may help to guide future enrichment experiments to obtain representatives in pure culture.

Quantifying genome-specific carbon fixation in a 750-meter deep subsurface hydrothermal microbial community

“…In the samples collected in 2021, Gammaproteobacteria (53%), Nitrospirae (20%), and Aquificae (12%) predominated the initial microbial community of the subsurface alkaline hydrothermal fluids, which were used for the stable isotope probing (SIP) incubations. This overall microbial community structure is similar to thermophilic microbial communities found in other alkaline hot springs such as those in the Tibetan Plateau (Wang et al 2013 ), Yellowstone National Park (Schubotz et al 2015 ), Costa Rican convergent margin (Fullerton et al 2021 ), deep-subsurface aquifers (Spanevello and Patel 2004 ), and hydrothermal fluids in Bardakçılar geothermal field (Coskun et al 2023 ).…”

“…We sampled fluids for geochemical measurements from numerous hydrothermal aquifers ranging from surface hot spring pools to fluids from 1350-m subsurface deep boreholes, across the Biga Peninsula (Coskun et al 2023 ) ( Fig. S1 , Supporting Information ).…”

“…Filters were stored in 15 ml sterile falcon tubes and immediately placed on dry ice, shipped back to Germany, and stored at −80°C until DNA extractions were performed at the Geomicrobiology Lab at the University of Munich (LMU Munich). Geochemical analysis of the fluids was performed as described previously (Einsiedl et al 2020 , Coskun et al 2023 ). Briefly, samples for laboratory-based measurements of major anions and cations concentrations and water isotopes (δ 2 H and δ 18 O) were collected in 1.5 ml glass flasks after filtering with 0.2 µm PES filters.…”

“…The nonmetric multidimensional scaling (NMDS) using Bray–Curtis matrices was generated in vegan package (Oksanen 2010 ) and pairwise Tukey’s Honest Significant Difference (HSD) post hoc test was performed to distinguish dissimilarities among the samples. Furthermore, the studied geothermal field was compared to other hydrothermal locations in the vicinity using already published data (Coskun et al 2023 ). The 16S sequence data were entered in the NCBI Short Read Archive under BioProject ID PRJNA837050.…”

“…KEGGdecoder (Graham et al 2018 ), anvi’o and COG annotations were used to estimate the completeness of the predicted metabolisms. MAGSIPTR2 and MAGSIPTR39 were published previously with MAG names Candidatus Bipolaricaulota CK101 and CK84 (Coskun et al 2023 ). The metagenomic sequences were stored in the NCBI Short Read Archive under BioProject ID PRJNA837050.…”