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The Yessentuki mineral water deposit (YMWD) is a well-known source of balneologically valuable drinking mineral water, but it has rarely been investigated in terms of the microbes inhabiting it. In this work, we have studied the microbial communities of the continuously operating production well 9, penetrating the Lower Cretaceous aquifer of the YMWD, and characterized, in detail, two novel representatives of class Ignavibacteria (Bacteroidota). One representative of the so-called XYB12-FULL-38-5 group within the family Melioribacteraceae has been isolated in pure culture, designated strain 09-Me, and physiologically characterized. It is a facultatively anaerobic thermotolerant microorganism capable of fermentation and respiration on simple and complex sugars (lichenan, xanthan gum, glucomannan, curdlan, pachyman). In addition to oxygen, ferric iron, arsenate, and elemental sulfur were also used as electron acceptors. Phylogenomic and physiological analyses reveal this novel isolate to represent a novel genus and species for which the name Stygiobacter electus gen. nov., sp. nov. is proposed. The second representative of the family Melioribacteraceae described here belonged to the so-called DSXH01 group, which comprises the dominant group (up to 28%) of the microbial community of well 9 water. The organism was characterized through the analysis of its genome, assembled from metagenome of well 9 (Ess09-04 MAG). Genes encoding enzymes of carbohydrate utilization and genes responsible for aerobic and anaerobic respiration have been identified in the genomes of both bacteria. The investigation of the environmental distribution of Stygiobacter genus-related bacteria and representatives of the lineage DSXH01 has shown that they all are typical inhabitants of the subsurface biosphere, and are often found in bioreactors. These data significantly expand our knowledge on the microbes of subsurface water basins and pave the way for future studies of the novel members of Ignavibacteria class. [ABSTRACT FROM AUTHOR]

A novel facultatively anaerobic moderately thermophilic bacteria, strains 4137-MeT and 4148-MeT, were isolated from hot springs of Karmadon and Ursdon, respectively (North Ossetia, Russian Federation). Gram-negative, motile rods were present singly, in pairs, rosettes, and aggregates, or formed biofilms. Both strains grew optimally at 50–55 °C, pH 7.0 and did not require sodium chloride or yeast extract for growth. They were chemoorganoheterotrophs, growing on mono-, di- and polysaccharides (cellulose, starch, xylan, lichenan, galactan, xyloglucan, mannan, xanthan gum, guar gum) as well as proteinaceous substrates (gelatin, peptone, beef and yeast extract). Growth under anaerobic conditions was observed in presence and absence of external electron acceptors. Sulfur, thiosulfate, arsenate, Fe-citrate, and ferrihydrite were reduced with acetate, starch, or yeast extract as electron donors. The respiratory quinone was MK-7. Major cellular fatty acids of both strains were iso -C 15:0 , anteiso -C 17:0 , C 15:0 , iso -C 16:0 and additionally iso -C 17:0 for strain 4137-MeT. The size of the genome and genomic DNA G + C content of strain 4137-MeT were 3.24 Mb. and 29.9 %, respectively; for strain 4148-MeT – 3.33 Mb and 30.7 %. According to the 16S rRNA gene sequence and conserved protein sequences phylogenies, strains 4137-MeT and 4148-MeT represented a distinct lineage of the family Melioribacteraceae within the class Ignavibacteria. Based on phylogenetic analysis and phenotypic features, the novel isolates were assigned to a novel genus, for which the name Rosettibacter gen. nov. is proposed. Strain 4148-MeT represents its type species Rosettibacter primus sp. nov., while strain 4137-MeT represents a new species Rosettibacter firmus sp. nov. [ABSTRACT FROM AUTHOR]

The Yessentuki mineral water deposit (YMWD) is a well-known source of balneologically valuable drinking mineral water, but it has rarely been investigated in terms of the microbes inhabiting it. In this work, we have studied the microbial communities of the continuously operating production well 9, penetrating the Lower Cretaceous aquifer of the YMWD, and characterized, in detail, two novel representatives of class Ignavibacteria (Bacteroidota). One representative of the so-called XYB12-FULL-38-5 group within the family Melioribacteraceae has been isolated in pure culture, designated strain 09-Me, and physiologically characterized. It is a facultatively anaerobic thermotolerant microorganism capable of fermentation and respiration on simple and complex sugars (lichenan, xanthan gum, glucomannan, curdlan, pachyman). In addition to oxygen, ferric iron, arsenate, and elemental sulfur were also used as electron acceptors. Phylogenomic and physiological analyses reveal this novel isolate to represent a novel genus and species for which the name Stygiobacter electus gen. nov., sp. nov. is proposed. The second representative of the family Melioribacteraceae described here belonged to the so-called DSXH01 group, which comprises the dominant group (up to 28%) of the microbial community of well 9 water. The organism was characterized through the analysis of its genome, assembled from metagenome of well 9 (Ess09-04 MAG). Genes encoding enzymes of carbohydrate utilization and genes responsible for aerobic and anaerobic respiration have been identified in the genomes of both bacteria. The investigation of the environmental distribution of Stygiobacter genus-related bacteria and representatives of the lineage DSXH01 has shown that they all are typical inhabitants of the subsurface biosphere, and are often found in bioreactors. These data significantly expand our knowledge on the microbes of subsurface water basins and pave the way for future studies of the novel members of Ignavibacteria class.

Evolutionary relationships amongst Chlorobia and Ignavibacteria species/strains were examined using phylogenomic and comparative analyses of genome sequences. In a phylogenomic tree based on 282 conserved proteins, the named Chlorobia species formed a monophyletic clade containing two distinct subclades. One clade, encompassing the genera Chlorobaculum, Chlorobium, Pelodictyon, and Prosthecochloris, corresponds to the family Chlorobiaceae, whereas another clade, harboring Chloroherpeton thalassium, Candidatus Thermochlorobacter aerophilum, Candidatus Thermochlorobacteriaceae bacterium GBChlB, and Chlorobium sp. 445, is now proposed as a new family (Chloroherpetonaceae fam. nov). In parallel, our comparative genomic analyses have identified 47 conserved signature indels (CSIs) in diverse proteins that are exclusively present in members of the class Chlorobia or its two families, providing reliable means for identification. Two known Ignavibacteria species in our phylogenomic tree are found to group within a larger clade containing several Candidatus species and uncultured Chlorobi strains. A CSI in the SecY protein is uniquely shared by the species/strains from this "larger Ignavibacteria clade". Two additional CSIs, which are commonly shared by Chlorobia species and the "larger Ignavibacteria clade", support a specific relationship between these two groups. The newly identified molecular markers provide novel tools for genetic and biochemical studies and identification of these organisms. [ABSTRACT FROM AUTHOR]

Evolutionary relationships amongst Chlorobia and Ignavibacteria species/strains were examined using phylogenomic and comparative analyses of genome sequences. In a phylogenomic tree based on 282 conserved proteins, the named Chlorobia species formed a monophyletic clade containing two distinct subclades. One clade, encompassing the genera Chlorobaculum, Chlorobium, Pelodictyon, and Prosthecochloris, corresponds to the family Chlorobiaceae, whereas another clade, harboring Chloroherpeton thalassium, Candidatus Thermochlorobacter aerophilum, Candidatus Thermochlorobacteriaceae bacterium GBChlB, and Chlorobium sp. 445, is now proposed as a new family (Chloroherpetonaceae fam. nov). In parallel, our comparative genomic analyses have identified 47 conserved signature indels (CSIs) in diverse proteins that are exclusively present in members of the class Chlorobia or its two families, providing reliable means for identification. Two known Ignavibacteria species in our phylogenomic tree are found to group within a larger clade containing several Candidatus species and uncultured Chlorobi strains. A CSI in the SecY protein is uniquely shared by the species/strains from this “larger Ignavibacteria clade”. Two additional CSIs, which are commonly shared by Chlorobia species and the “larger Ignavibacteria clade”, support a specific relationship between these two groups. The newly identified molecular markers provide novel tools for genetic and biochemical studies and identification of these organisms.