Organic matter degradation in the deep, sulfidic waters of the Black Sea: insights into the ecophysiology of novel anaerobic bacteria.

BACKGROUND: Recent studies have reported the identity and functions of key anaerobes involved in the degradation of organic matter (OM) in deep (> 1000 m) sulfidic marine habitats. However, due to the lack of available isolates, detailed investigation of their physiology has been precluded. In this study, we cultivated and characterized the ecophysiology of a wide range of novel anaerobes potentially involved in OM degradation in deep (2000 m depth) sulfidic waters of the Black Sea. RESULTS: We have successfully cultivated a diverse group of novel anaerobes belonging to various phyla, including Fusobacteriota (strain S5), Bacillota (strains A1T and A2), Spirochaetota (strains M1T, M2, and S2), Bacteroidota (strains B1T, B2, S6, L6, SYP, and M2P), Cloacimonadota (Cloa-SY6), Planctomycetota (Plnct-SY6), Mycoplasmatota (Izemo-BS), Chloroflexota (Chflx-SY6), and Desulfobacterota (strains S3T and S3-i). These microorganisms were able to grow at an elevated hydrostatic pressure of up to 50 MPa. Moreover, this study revealed that different anaerobes were specialized in degrading specific types of OM. Strains affiliated with the phyla Fusobacteriota, Bacillota, Planctomycetota, and Mycoplasmatota were found to be specialized in the degradation of cellulose, cellobiose, chitin, and DNA, respectively, while strains affiliated with Spirochaetota, Bacteroidota, Cloacimonadota, and Chloroflexota preferred to ferment less complex forms of OM. We also identified members of the phylum Desulfobacterota as terminal oxidizers, potentially involved in the consumption of hydrogen produced during fermentation. These results were supported by the identification of genes in the (meta)genomes of the cultivated microbial taxa which encode proteins of specific metabolic pathways. Additionally, we analyzed the composition of membrane lipids of selected taxa, which could be critical for their survival in the harsh environment of the deep sulfidic waters and could potentially be used as biosignatures for these strains in the sulfidic waters of the Black Sea. CONCLUSIONS: This is the first report that demonstrates the cultivation and ecophysiology of such a diverse group of microorganisms from any sulfidic marine habitat. Collectively, this study provides a step forward in our understanding of the microbes thriving in the extreme conditions of the deep sulfidic waters of the Black Sea. Video Abstract.

Marine particle microbiomes during a spring diatom bloom contain active sulfate-reducing bacteria.

Phytoplankton blooms fuel marine food webs with labile dissolved carbon and also lead to the formation of particulate organic matter composed of living and dead algal cells. These particles contribute to carbon sequestration and are sites of intense algal-bacterial interactions, providing diverse niches for microbes to thrive. We analyzed 16S and 18S ribosomal RNA gene amplicon sequences obtained from 51 time points and metaproteomes from 3 time points during a spring phytoplankton bloom in a shallow location (6-10 m depth) in the North Sea. Particulate fractions larger than 10 µm diameter were collected at near daily intervals between early March and late May in 2018. Network analysis identified two major modules representing bacteria co-occurring with diatoms and with dinoflagellates, respectively. The diatom network module included known sulfate-reducing Desulfobacterota as well as potentially sulfur-oxidizing Ectothiorhodospiraceae. Metaproteome analyses confirmed presence of key enzymes involved in dissimilatory sulfate reduction, a process known to occur in sinking particles at greater depths and in sediments. Our results indicate the presence of sufficiently anoxic niches in the particle fraction of an active phytoplankton bloom to sustain sulfate reduction, and an important role of benthic-pelagic coupling for microbiomes in shallow environments. Our findings may have implications for the understanding of algal-bacterial interactions and carbon export during blooms in shallow-water coastal areas.

Functional Microorganisms Drive the Formation of Black-Odorous Waters.

Black-odorous waters are water bodies that are noticeably abnormal in color or emit unpleasant odors. River water pollution and ecological degradation have gradually emerged with urbanization and rapid economic development, and BOW has become frequent. The black-odorous evolution of urban water bodies is a serious environmental problem in many areas, posing a serious threat to both human health and the ecological environment. Functional microorganisms are closely related to the formation of black-odorous phenomena in water bodies, but the understanding of the mechanisms by which functional microorganisms influence the formation of BOW is very limited. In this study, water samples from the Guangdang River in Yantai, Shandong Province, China, were collected as the bacterial solution in the study, and how environmental factors and functional microorganisms affect the formation of black smelly water was investigated by artificially simulating black smelly water. The results indicated that different environmental factors have different effects on the formation of BOW. Anaerobic conditions accelerated the formation of BOW, and species diversity and species abundance were lowest under this condition. Hydraulic disturbance and nitrate effectively mitigated the BOW phenomenon, in which species diversity and species abundance were higher; controlling either of these variables was effective in mitigating the BOW phenomenon. Desulfobacterota played a key role in the formation of BOW, and reducing the proportion of Desulfobacterota in the microbial community could effectively improve the water quality. Possible directions of electron transfer in the process were hypothesized. This study contributes to identifying the biological driving factors for black-odorous evolution, presents insight for preventing BOW formation, and provides a scientific basis for subsequent BOW management.

Revisiting the mercury cycle in marine sediments: A potential multifaceted role for Desulfobacterota.

Marine sediments impacted by urban and industrial pollutants are typically exposed to reducing conditions and represent major reservoirs of toxic mercury species. Mercury methylation mediated by anaerobic microorganisms is favored under such conditions, yet little is known about potential microbial mechanisms for mercury detoxification. We used culture-independent (metagenomics, metabarcoding) and culture-dependent approaches in anoxic marine sediments to identify microbial indicators of mercury pollution and analyze the distribution of genes involved in mercury reduction (merA) and demethylation (merB). While none of the isolates featured merB genes, 52 isolates, predominantly affiliated with Gammaproteobacteria, were merA positive. In contrast, merA genes detected in metagenomes were assigned to different phyla, including Desulfobacterota, Actinomycetota, Gemmatimonadota, Nitrospirota, and Pseudomonadota. This indicates a widespread capacity for mercury reduction in anoxic sediment microbiomes. Notably, merA genes were predominately identified in Desulfobacterota, a phylum previously associated only with mercury methylation. Marker genes involved in the latter process (hgcAB) were also mainly assigned to Desulfobacterota, implying a potential central and multifaceted role of this phylum in the mercury cycle. Network analysis revealed that Desulfobacterota were associated with anaerobic fermenters, methanogens and sulfur-oxidizers, indicating potential interactions between key players of the carbon, sulfur and mercury cycling in anoxic marine sediments.

N-acetyl-L-leucine protects MPTP-treated Parkinson's disease mouse models by suppressing Desulfobacterota via the gut-brain axis.

Parkinson's disease (PD) is the second most common neurodegenerative disease, and communication between the gut and brain (the gut-brain axis) has been found to be essential in behavior and cognitive function. However, the exact mechanisms underlying microbiota dysbiosis in PD progression have not yet been elucidated. Our study aimed to investigate the correlation between gut microbiota disturbances and feces metabolic disorders in PD. We used 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to induce PD models and observed mice's motor symptoms, dopaminergic (DA) neuron death, and gastrointestinal dysfunction. To identify alterations in microbiota and metabolome, feces were collected from mice and analyzed using 16 S ribosomal RNA sequencing feces metabolomics. Pearson analysis was utilized to investigate correlations between the abundances of gut microbiota components and the levels of gut microbiota metabolites, displaying their interaction networks. Our findings revealed a significant increase in Desulfobacterota in the PD mouse model and 151 differentially expressed fecal metabolites between PD and vehicle mice. Moreover, Pearson correlation analysis suggested that the protective factor N-acetyl-L-leucine (NALL) may be associated with neuroinflammation in the striatum and substantia nigra, which also had a negative relationship with the concentration of Desulfobacterota. Additionally, we found that oral administration of NALL alleviated MPTP-induced Motor Impairments and DA neuronal deficits. All in all, we concluded that the decrease of NALL might lead to a significant increase of Desulfobacterota in the MPTP model mouse and subsequently result in the damage of DA neurons via the gut-brain aix pathway.

Baseline Gut Microbiome Signatures Correlate with Immunogenicity of SARS-CoV-2 mRNA Vaccines.

The powerful immune responses elicited by the mRNA vaccines targeting the SARS-CoV-2 Spike protein contribute to their high efficacy. Yet, their efficacy can vary greatly between individuals. For vaccines not based on mRNA, cumulative evidence suggests that differences in the composition of the gut microbiome, which impact vaccine immunogenicity, are some of the factors that contribute to variations in efficacy. However, it is unclear if the microbiome impacts the novel mode of immunogenicity of the SARS-CoV-2 mRNA vaccines. We conducted a prospective longitudinal cohort study of individuals receiving SARS-CoV-2 mRNA vaccines where we measured levels of anti-Spike IgG and characterized microbiome composition, at pre-vaccination (baseline), and one week following the first and second immunizations. While we found that microbial diversity at all timepoints correlated with final IgG levels, only at baseline did microbial composition and predicted function correlate with vaccine immunogenicity. Specifically, the phylum Desulfobacterota and genus Bilophila, producers of immunostimulatory LPS, positively correlated with IgG, while Bacteroides was negatively correlated. KEGG predicted pathways relating to SCFA metabolism and sulfur metabolism, as well as structural components such as flagellin and capsular polysaccharides, also positively correlated with IgG levels. Consistent with these findings, depleting the microbiome with antibiotics reduced the immunogenicity of the BNT162b2 vaccine in mice. These findings suggest that gut microbiome composition impacts the immunogenicity of the SARS-CoV-2 mRNA vaccines.

The response mechanism of microorganisms to the organic carbon-driven formation of black and odorous water.

The formation of black and odorous water is a complex process influenced by various factors such as organic matter and environmental conditions. However, there are limited studies on the role of microorganisms in water and sediment during the blackening and odorization process. In this study, we investigated the characteristics of black and odorous water formation by simulating organic carbon-driven black and odorous water through indoor experiments. The study revealed that the water turned black and odorous when DOC reached 50 mg/L and the microbial community structure in the water changed significantly during this process, with the relative abundance of Desulfobacterota increasing significantly and Desulfovibrio being the main dominant genus in Desulfobacterota. Additionally, we observed a notable decrease in the α-diversity of the microbial community in water and a considerable increase in microbial function of sulfur compounds respiration in water. In contrast, the sediment microbial community changed slightly, and the main functions of the sediment microbial community remained unchanged. The partial least squares path model (PLS-PM) suggested that organic carbon will drive the blackening and odorization process by affecting DO levels and microbial community structure and that the contribution of Desulfobacterota in water to the formation of black and odorous water was higher than that in sediment. Overall, our study provides insights into the characteristics of black and odorous water formation and suggests potential ways to prevent its formation by controlling DOC and inhibiting the growth of Desulfobacterota in water bodies.

Consortia cultivation of the Desulfobacterota from macrophyte periphyton: tool for increasing the cultivation of microorganisms involved in mercury methylation.

Invasive macrophytes are a persistent environmental problem in aquatic ecosystems. They also cause potential health issues, since periphyton colonizing their aquatic roots are hot spot of mercury methylation. Because periphytons are at the base of the trophic chain, the produced methylmercury is bioamplified through the food webs. In this work, a consortia cultivation approach was applied in order to investigate methylators in the periphyton of Ludwigia sp., an invasive macrophyte. Five growth conditions were used in order to favor the growth of different sulfate reducers, the major mercury methylators in this periphyton. A total of 33 consortia containing putative Hg methylators were obtained. Based on the amino acid sequences of HgcA (essential enzyme for Hg methylation), the obtained consortia could be subdivided into five main clusters, affiliated with Desulfovibrionaceae, Desulfobulbaceae and Syntrophobacteraceae. The main cluster, related to Desulfovibrionaceae, showed the highest sequence diversity; notwithstanding most of the sequences of this cluster showed no close representatives. Through the consortia approach, species thus far uncultivated were cultivated. The successful cultivation of these species was probably possible through the metabolites produced by other members of the consortium. The analysis of the microbial composition of the consortia uncover certain microbial interactions that may exist within this complex environment.

A Cristae-Like Microcompartment in Desulfobacterota.

Some Alphaproteobacteria contain intracytoplasmic membranes (ICMs) and proteins homologous to those responsible for the mitochondrial cristae, an observation which has given rise to the hypothesis that the Alphaproteobacteria endosymbiont had already evolved cristae-like structures and functions. However, our knowledge of microbial fine structure is still limited, leaving open the possibility of structurally homologous ICMs outside the Alphaproteobacteria. Here, we report on the detailed characterization of lamellar cristae-like ICMs in environmental sulfate-reducing Desulfobacterota that form syntrophic partnerships with anaerobic methane-oxidizing (ANME) archaea. These structures are junction-bound to the cytoplasmic membrane and resemble the form seen in the lamellar cristae of opisthokont mitochondria. Extending these observations, we also characterized similar structures in Desulfovibrio carbinolicus, a close relative of the magnetotactic D. magneticus, which does not contain magnetosomes. Despite a remarkable structural similarity, the key proteins involved in cristae formation have not yet been identified in Desulfobacterota, suggesting that an analogous, but not a homologous, protein organization system developed during the evolution of some members of Desulfobacterota. IMPORTANCE Working with anaerobic consortia of methane oxidizing ANME archaea and their sulfate-reducing bacterial partners recovered from deep sea sediments and with the related sulfate-reducing bacterial isolate D. carbinolicus, we discovered that their intracytoplasmic membranes (ICMs) appear remarkably similar to lamellar cristae. Three-dimensional electron microscopy allowed for the novel analysis of the nanoscale attachment of ICMs to the cytoplasmic membrane, and these ICMs are structurally nearly identical to the crista junction architecture seen in metazoan mitochondria. However, the core junction-forming proteins must be different. The outer membrane vesicles were observed to bud from syntrophic Desulfobacterota, and darkly stained granules were prominent in both Desulfobacterota and D. carbinolicus. These findings expand the taxonomic breadth of ICM-producing microorganisms and add to our understanding of three-dimensional microbial fine structure in environmental microorganisms.

Proposal to reclassify the proteobacterial classes Deltaproteobacteria and Oligoflexia, and the phylum Thermodesulfobacteria into four phyla reflecting major functional capabilities.

The class Deltaproteobacteria comprises an ecologically and metabolically diverse group of bacteria best known for dissimilatory sulphate reduction and predatory behaviour. Although this lineage is the fourth described class of the phylum Proteobacteria, it rarely affiliates with other proteobacterial classes and is frequently not recovered as a monophyletic unit in phylogenetic analyses. Indeed, one branch of the class Deltaproteobacteria encompassing Bdellovibrio-like predators was recently reclassified into a separate proteobacterial class, the Oligoflexia. Here we systematically explore the phylogeny of taxa currently assigned to these classes using 120 conserved single-copy marker genes as well as rRNA genes. The overwhelming majority of markers reject the inclusion of the classes Deltaproteobacteria and Oligoflexia in the phylum Proteobacteria. Instead, the great majority of currently recognized members of the class Deltaproteobacteria are better classified into four novel phylum-level lineages. We propose the names Desulfobacterota phyl. nov. and Myxococcota phyl. nov. for two of these phyla, based on the oldest validly published names in each lineage, and retain the placeholder name SAR324 for the third phylum pending formal description of type material. Members of the class Oligoflexia represent a separate phylum for which we propose the name Bdellovibrionota phyl. nov. based on priority in the literature and general recognition of the genus Bdellovibrio. Desulfobacterota phyl. nov. includes the taxa previously classified in the phylum Thermodesulfobacteria, and these reclassifications imply that the ability of sulphate reduction was vertically inherited in the Thermodesulfobacteria rather than laterally acquired as previously inferred. Our analysis also indicates the independent acquisition of predatory behaviour in the phyla Myxococcota and Bdellovibrionota, which is consistent with their distinct modes of action. This work represents a stable reclassification of one of the most taxonomically challenging areas of the bacterial tree and provides a robust framework for future ecological and systematic studies.