A Gram-stain-negative, oxidase-positive, catalase-negative, motile, rod-shaped bacterium, designated strain M5-1[T], was isolated from a freshwater green alga. Strain M5-1[T] was obligately aerobic and contained ubiquinone-8 (Q-8) as the sole respiratory quinone. The major fatty acids (>10%) were C16:0, C12:0, summed feature 3 (C16:1ω7c and/or C16:1ω6c), and summed feature 8 (C18:1ω7c and/or C18:1ω6c), and the major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, and diphosphatidylglycerol. The genomic DNA G+C content was 67.3 mol%. Based on 16S rRNA gene sequence analysis, strain M5-1[T] was most closely related to Roseateles cavernae (proposed herein as Kinneretia cavernae) YIM B04394[T] (99.3%) and Kinneretia oligotropha CHU3[T] (98.3%). Genome-based phylogenomic analyses consistently placed strain M5-1[T] together with R. cavernae within the genus Kinneretia. Pairwise average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between strain M5-1[T] and R. cavernae were 96.1% and 66.9%, respectively, whereas comparisons with all other type strains yielded ANI and dDDH values of ≤81.1% and ≤23.6%, respectively, supporting its recognition as a novel species. Based on its phylogenomic, genomic, phenotypic, and chemotaxonomic characteristics, strain M5-1[T] (=KACC 22491[T]=JCM 34976[T]=FBCC-B22965[T]) represents a novel species of the genus Kinneretia, for which the name Kinneretia algicola sp. nov. is proposed. In addition, comprehensive phylogenomic and genome-relatedness analyses support the transfer of six Roseateles and Paucibacter species to Kinneretia, two Roseateles species to Pelomonas, and Aquabacterium humicola to Pseudaquabacterium, providing a revised genome-based taxonomic framework for the family Sphaerotilaceae.

Many species of family Sphaerotilaceae have been isolated from aquatic environments. In this study, we conducted the taxonomic characterization of five novel strains Y2R2-4[T], B2R-40[T], DJ2R-2[T], DJ1R-11 and DJ4R-1, isolated from crater lakes in Da Hinggan Mountain region of northern China. A polyphasic taxonomic approach was employed to determine the taxonomic affiliation of these strains. Based on the 16S rRNA gene sequence analysis, these novel strains showed the highest similarities (98.9-99.9%) to Kinneretia aquatilis CR182[T] and Kinneretia alba hw1[T]. While the digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) between the five novel strains and their closely related species Kinneretia were significantly below the established taxonomic thresholds for species delineation, further supporting that these strains represent three novel species within the genus Kinneretia. Based on genomic annotation results, their metabolic potentials to produce terpene and RiPP-like compounds were further explored. In summary, the morphological, physiological, and biochemical characteristics of these novel strains indicated that the five isolates represent three novel species within the genus Kinneretia. The names, Kinneretia craterilacus sp. nov., Kinneretia daxingganlingensis sp. nov. and Kinneretia lacustris sp. nov., were proposed for them, with type strain Y2R2-4[T] (= CGMCC 1.19001[T] = KCTC 92188[T]), B2R-40[T] (= CGMCC 1.19003[T] = KCTC 92190[T]) and DJ2R-2[T] (= CGMCC 1.19002[T] = KCTC 92189[T]), respectively.

Genome-resolved phylogenomics reveals widespread misclassification of metal-reducing bacteria historically assigned to Geobacter based on 16S rRNA gene phylogeny, and highlights species that persist only as 16S rRNA entries without genomes for robust taxonomic resolution. Here, we resolve two such lineages by integrating whole-genome phylogeny with average amino acid identity (AAI) and percentage of conserved proteins (POCP) across 418 dereplicated genomes of Desulfuromonadia. We report a draft genome of the psychrophilic iron-reducing bacterium Geobacter psychrophilus (100% completeness). Phylogenomic analyses place both Geobacter psychrophilus and the GTDB placeholder genus g__JACRCG01 within the family 'Pseudopelobacteraceae', outside Geobacteraceae sensu stricto. Within this framework, G. psychrophilus forms a distinct, well-supported lineage separated from neighbouring genera by discontinuities in AAI and POCP, supporting its reclassification as Irobacter psychrophilus comb. nov. Additionally, we show that Geosyntrophus acetoxidans, a non-axenic syntrophic bacterium, forms a coherent genus with 51 other environmental genomes (placeholder genus g__JACRCG01), for which we propose the replacement name Geosyntrophus gen. nov. Comparative genome analysis revealed conserved family-level metabolic traits together with genus-specific differences in respiratory metabolism, while ANI-based clustering identified substantial species-level diversity within both proposed genera. Metagenome and 16S rRNA-gene survey data further show that Geosyntrophus and Irobacter occur in broadly similar aquatic and subsurface habitats spanning from the Arctic to the Antarctic. Together, these results resolve the taxonomy of two previously ambiguous Desulfuromonadales lineages and shed light on their environmental distribution.

Novel facultatively anaerobic moderately thermophilic bacteria, strains OK-6-Me[T] and OK-1-Me, were isolated from the hot springs of Karmadon (North Ossetia, Russian Federation). Gram-stain-negative, motile rods were present singly, in rosettes, and formed biofilms. Both strains grew optimally at 55 °C, pH 7.0 and did not require sodium chloride. They were chemoorganoheterotrophs, growing on mono-, di- and polysaccharides (cellulose, xylan, lichenan, xyloglucan, mannan, locust bean gum, pectin) as well as proteinaceous substrates (gelatin, casein). Growth under anaerobic conditions was observed both in the presence and absence of external electron acceptors (sulfur, thiosulfate, nitrite, arsenate, Fe-citrate, ferrihydrite). Major cellular fatty acids of both strains were iso-C15:0, anteiso-C15:0, and anteiso-C17:0. The size of the genomes were 3.3 and 3.2 Mb for strain OK-6-Me[T] and OK-1-Me, respectively. Genomic DNA G + C content was 37% for both strains. According to the 16S rRNA gene sequence and conserved protein sequences phylogenies, the strains represented a new species of the genus Melioribacter of family Melioribacteraceae within the class Ignavibacteria, for which the name Melioribacter sulfuriphilus sp. nov. is proposed, with type strain OK-6-Me[T] (= B-3972[T] = CGMCC 1.18264 [T] = BIM B-2154[T] = UQM 41932[T]). Analysis of OK-1 and OK-6 metagenomes revealed presence of various genes involved in carbon (CO2 fixation, carbohydrate hydrolysis, hydrocarbons degradation, fermentation), nitrogen (nitrate, nitrite, NO and N2O reduction) and sulfur cycles (sulfate reduction, sulfur or thiosulfate reduction, oxidation of sulfur compounds). MAGs OK-1-035 and OK-6-024 almost identical to genomes of strain OK-1-Me and OK-6-Me[T] presumably are integral part of these complex trophic chains.

Coastal tidal flats represent dynamic saline environments that harbor largely unexplored haloarchaeal communities. In this study, amplicon sequencing, metagenomic analyses, and cultivation-based approaches revealed substantial haloarchaeal diversity in tidal flats from four provinces of eastern China despite their relatively low salinity. Five haloarchaeal strains, designated YSMS36[T], DYSN1, QDMS2, CMSO5[T], and ZSTT2, were isolated from diverse tidal flats. Theses strains shared 16S rRNA gene sequence similarities of 92.1-92.2% with their closest validly named relative, Salinilacihabitans rarus AD-4[T]. Phylogenetic analyses based on 16S rRNA and rpoB' gene sequences showed that the five strains formed a distinct and well-supported monophyletic lineage, separated from currently recognized members of the class Halobacteria. Average amino acid identity (AAI), average nucleotide identity (ANI), and digital DNA-DNA hybridization (dDDH) values between these five strains and the related Halobacteria representatives were 49.3-62.7%, 66.9-74.9%, and 16.2-29.5%, respectively, and well below the accepted thresholds for species and genus delineation. Phylogenomic analyses further supported their placement within a novel family of the order Halobacteriales. Based on phylogenetic, genomic, chemotaxonomic, and phenotypic analyses, these five strains represent two novel species of a novel genus within a novel family. The names, Halolitoreaceae fam. nov., Halolitoreus marinus gen. nov., sp. nov., and Halolitoreus rarus sp. nov. are herein proposed.

Bacterial strains were isolated from different locations and screened for plant growth promoting (PGP) features. Comparative analyses of the 16S gene sequences of 5 of the strains indicated taxonomic relatedness within the genus Paenibacillus. A polyphasic taxonomic approach was employed to study the strains in detail to clarify their phylogenetic position. Genome-based analyses, including digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI), revealed values consistently below the accepted species delineation thresholds when compared with closest relatives of each strain. Additionally, the strains showed clear differences in their physiological and biochemical profiles to the type strains of the closest related species. A notably diverse set of genes potentially involved in plant growth promotion was detected in all strains. With respect to the analyses reported here, the following new names are proposed: Paenibacillus corni sp. nov., with AK-167[T] as the type strain (= LMG 34403[T] = DSM 121689[T]); Paenibacillus vaccinii sp. nov., with AK-264[T] as the type strain (= LMG 34404[T] = DSM 121714[T]); Paenibacillus ericacearum sp. nov., with AK-265[T] as the type strain (= CCM 6913[T] = LMG 34405[T] = DSM 121715[T]); Paenibacillus polytrichii sp. nov., with AK-286[T] as the type strain (= CCM 9614[T] = LMG 34406[T] = DSM 121716[T]); and Paenibacillus artemisiae sp. nov., with DT-106[T] as the type strain (= LMG 34409[T] = CCM 9610[T] = DSM 121690[T]).

Four Gram-staining negative, non-motile, rod-shape bacteria, named strains GN3S48[T], HN1S83, LN3S78[T], and M1N3S26, were isolated from the bulk saline soils, in Baotou, China. Among them, strains GN3S48[T] and HN1S83 could degrade 100 mg l[-1]n-hexadecane as sole carbon and energy source for their growth. Phylogenetic analyses showed that the four strains always formed two distinct clades: Strain LN3S78[T] clustered with strain M1N3S26, and strain GN3S48[T] clustered with strain HN1S83. Nonetheless, all four strains tightly clustered and shared the highest 16S rRNA gene similarities with Marinobacter species. Specifically, clade of strains LN3S78[T] and M1N3S26 cluster with Marinobacter lipolyticus CGMCC 1.7282[T], while clade of strains GN3S48[T] and HN1S83 clustered with Marinobacter zhanjiangensis CCTCC AB 208029[T]. The ANIb and AAI values between strains GN3S48[T] and HN1S83 were 96.4% and 94.6%, respectively, while those between strains LN3S78[T] and M1N3S26 were 99.3% and 99.1%, respectively. All ANI and AAI values between the four strains and their closest relatives were below the 95.0% species delineation threshold. The predominant respiratory quinone of the four strains was Q-9. Based on this polyphasic result, the two clades should be identified as two novel species within the genus Marinobacter. Thus, Marinobacter alkalisoli sp. nov. (type strain GN3S48[T] = CGMCC 1.62232[T] = KCTC 8701[T] = JCM 37359 [T]) and Marinobacter shunpengi sp. nov. (type strain LN3S78[T] = CGMCC 1.62233[T] = KCTC 8702[T] = JCM 37360[T]) are proposed. The metagenomic analysis revealed that the two new species are globally distributed in high-salt habitats. In addition, comparative genomic analysis confirmed that alkane-degrading genes are ubiquitous in Marinobacter strains.

A polyphasic study was conducted to establish the taxonomic status of strain T060[T], an orange, aerobic, coccoid and non-motile actinomycete, isolated from a marine bryozoan (Dendrobeania sp.) collected in the Barents Sea. Phylogenetic analysis of the 16S rRNA gene sequences revealed Rhodococcus sovatensis DSM 102881[T] as the closest related species to strain T060[T] with a similarity of 99.54%. Phylogenomic analysis confirmed a close relationship between T060[T] and R. sovatensis DSM 102881[T], while supporting their distinction. Digital DNA-DNA hybridization and average nucleotide identity values between strain T060[T] and R. sovatensis DSM 102881[T] were 26.4 and 84.3%, respectively, supporting the delineation of the isolate as a new species. Genomic characterization of the assembled genomes of T060[T] and R. sovatensis DSM 102881[T] showed genome sizes of 5.3 and 4.2 Mbp, with a G + C content of 64.38 and 65.01%, respectively. Genome analysis of strain T060[T] identified 15 biosynthetic gene clusters (BGCs) with low sequence similarity to known BGCs, indicating its capacity to produce unknown, potentially bioactive secondary metabolites. Furthermore, growth of T060[T] in eight different media revealed condition-dependent cytotoxic activity. The strongest cytotoxicity was observed for fractionated extracts from T060[T] grown in half-strength ISP 2 with filtered seawater against the human malignant cell lines MCF7 and A2058, and to a lesser extent against the non-malignant MRC5 cell line, highlighting its biotechnological potential. Based on the data from polyphasic taxonomy studies, it is proposed that strain T060[T] be classified in the genus Rhodococcus as Rhodococcus dendrobeaniae sp. nov. Additionally, the acquired data of R. sovatensis were used to amend its original description.

Strains MA13-6[T] and MA13-13, two Gram-stain-positive, aerobic, short rod-shaped actinobacteria, were isolated from a saline lake in Ngari Prefecture, Xizang Autonomous Region, China. Phylogenetic analysis based on 16S rRNA gene sequences indicated that these two strains belonged to the genus Aquipuribacter, with the closest relationship to Aquipuribacter hungaricus IV-75[T] (98.47% sequence similarity) and Aquipuribacter nitratireducens AMV4[T] (97.36% sequence similarity). Phylogenetic analysis based on genomes further confirmed their classification as a distinct cluster within the genus Aquipuribacter. The average nucleotide identity and digtal DNA-DNA hybridization values between these two strains and their closest relative Aquipuribacter hungaricus IV-75[T], were 82.44-82.49% and 23.00%, respectively, clearly indicating that strains MA13-6[T] and MA13-13 represent a novel species. The 16S rRNA gene sequence similarity, average nucleotide identity and digital DNA-DNA hybridization values between these two strains were 99.79%, 99.97% and 99.40%, respectively, unequivocally confirming their classification within the same species. However, DNA fingerprinting analysis distinguished them as non-clonal variants. The polar lipids comprised phosphatidylglycerol, two unidentified phospholipids, two unidentified glycolipids, and two unidentified lipids. The predominant respiratory quinone was MK-10 (H4). The major fatty acids were anteiso-C15:0, C18:1ω9c, isoC16:0 and anteiso-C17:0. The cell wall diagnostic diamino acid was meso-diaminopimelic acid. Based on phylogenetic analyses combined with phenotypic and chemotaxonomic characterization, strains MA13-6[T] and MA13-13 represent a novel species of the genus Aquipuribacter, for which the name Aquipuribacter aurantiacus sp. nov. is proposed. The type strain is MA13-6[T] (=MCCC 1K10045[T] = KCTC 59572[T]).

Four marine bacterial strains of the genus Vibrio, designated WJH972[T], RC27, FJH11[T], and HA2012[T], were isolated from coastal habitats in China. Phenotypic and physiological characterization showed that all four strains are Gram-stain-negative, facultatively anaerobic, short rod-shaped cells, and motile. The 16S rRNA gene sequence similarities between the isolates and their closest relatives were 96.5% - 98.2%, above the genus-level threshold but below the species delineation cutoff. Genomic analysis revealed that strains WJH972[T] and RC27 shared an average nucleotide identity (ANI) of 98.7% and a digital DNA-DNA hybridization (dDDH) value of 89.9%, confirming their conspecificity. In contrast, the ANI and dDDH values between the four strains and known Vibrio species ranged from 78.6% to 85.8% and from 13.7% to 59.3%, respectively, both falling below the thresholds for species delineation. The biosynthetic gene clusters support bacterial potentials for stress tolerance and antibiotic synthesis, consistent with strain FJH11[T]'s resistance to all 20 antibiotics and other strains' sensitivity to chloramphenicol and penicillin. Based on these polyphasic taxonomic evidence above, three novel species are thus proposed: Vibrio limimangrovi sp. nov. (type strain FJH11[T]), Vibrio phycocola sp. nov. (type strain WJH972[T]), and Vibrio sediminis sp. nov. (type strain HA2012[T]). This study provides novel insights into the systematics, metabolic diversity, antibiotic resistance, and ecological distribution of the genus Vibrio.

A novel strictly anaerobic moderately thermophilic bacterium, strain OB-3-ph[T], was isolated from a fouling under the flow of thermal well water (North Ossetia, Russian Federation). Gram-negative cells were nonspore-forming, straight motile rods, occasionally forming aggregates. The strain grew at 30-55 °C, pH range of 6.0-8.7, NaCl range 0-1.5%, with an optimum at 50 °C, pH 7.1 and 0.4% NaCl. It was a chemoorganoheterotroph, growing on proteinaceous substrates (albumin, gelatin, tryptone, casein, alpha-keratin) and carbohydrates (locust bean gum, guar gum, tragacanth, rhamnogalacturonan). Major products of glucose fermentation were acetate, propionate, hydrogen, and carbon dioxide. The genome size of strain OB-3-ph[T] was 2.462 Mbp; DNA G + C content was 36%. Genome analysis enabled the identification of genes encoding a limited set of carbohydrate-active enzymes, as well as numerous extracellular metalloproteases and serine proteases. In addition, central metabolism and energy conservation pathways of strain OB-3-ph[T] were reconstructed. Genes for respiration with oxygen and nitrite were identified, but due to the incompleteness of the menaquinone synthesis pathway, the strain obtains energy exclusively through fermentation. According to phylogenetic analyses based on 120 concatenated protein markers, strain OB-3-ph[T] represents the first cultivated member of the deep phylogenetic lineage known as "Candidatus Kapaibacteriia" (lineage OPB56). Here we propose a novel genus and species Eximiradius proteiniborus with type strain OB-3-ph[T] (=VKM B-3974[T] = UQM 42056[T]) within Eximiradiaceae fam. nov., Eximiradiales ord. nov. and Eximiradiia class. nov., respectively.

The methyl-coenzyme M reductase subunit alpha gene (mcrA) is an important phylogenetic marker for high throughput ecological profiling of methanogenic archaea, central to industrial biological methane production and greenhouse gas emissions. Yet, dedicated reference databases predate current relevant NCBI sequence accumulation and archaeal taxonomic revision. We present three updated mcrA reference databases: (i) one derived from NCBI-catalogued methanogen genomes (1572 sequences); (ii) a database built by expansion of a previously published reference dataset, leveraging the NCBI nucleotide collection (27,942 sequences); (iii) a curated-taxonomy version of the latter. The updated amplicon databases provide a ∼ 3.5-fold sequence richness expansion, extend genus-level richness from 31 to 83 taxa, more than 4-fold species-level richness, and incorporate novel lineages compared with the previous reference dataset (e.g. Thermoplasmatota-encompassed). All databases were formatted to support analysis with relevant contemporary software pipelines and packages. Overall, the generated databases facilitate a highly improved characterization of methanogen diversity and ecology.

Candidatus Saccharimonadia is a core lineage within the phylum Patescibacteriota (formerly the bacterial candidate phyla radiation, CPR), yet the class has long lacked a standardized, complete taxonomic framework. This nomenclatural gap severely hinders consistent academic exchange and global research into its diversity, evolutionary history, and ecological roles. Here, we recovered 29 medium- to high-quality Ca. Saccharimonadia metagenome-assembled genomes (MAGs) from groundwater, rhizosphere soil, and saline-alkali soil in the Hetao Irrigation District, Inner Mongolia, China, and performed integrated phylogenomic, genome size evolution, and metabolic analyses alongside reference genomes from the GTDB r220 database. Based on robust polyphasic taxonomic evidence (multi-dimensional phylogenetic analyses, widely accepted genome-wide ANI/AAI thresholds) and SeqCode rules, we formally propose two novel taxa: Parviradicicola hetaonensis gen. nov., sp. nov. (type material: txb011_bin.8.strict[TS]) and Parviputeicola dengkouensis gen. nov., sp. nov. (type material: sgl022_bin.19.orig[TS]), plus two novel families and one novel order. We further identified potential drivers and important associations related to Ca. Saccharimonadia genome size evolution and adaptive metabolic traits. This work refines the Ca. Saccharimonadia taxonomic framework, providing critical genomic references for follow-up research.

Archaeological excavation sites represent underexplored microbial habitats with the potential to recover taxonomically and biotechnologically valuable actinomycetes. In this study, two Saccharopolyspora strains, 5N708[T] and 5N102, were isolated from soil samples collected from the Gaziantep-Doliche-Dülük and Bitlis-Ahlat-Selçuklu Cemetery archaeological excavation sites in Türkiye. A polyphasic taxonomic approach, including 16S rRNA gene sequencing, phylogenetic and phylogenomic analyses, average nucleotide identity, digital DNA-DNA hybridization, phenotypic characterization, and chemotaxonomic analyses, showed that strain 5N708[T] represents a novel species of the genus Saccharopolyspora, for which the name Saccharopolyspora antiqui sp. nov. is proposed, whereas strain 5N102 was assigned to Saccharopolyspora elongata. Both isolates were further evaluated for their antimicrobial, antioxidant, and cytotoxic activities, and their biosynthetic potential was investigated by genome mining. Both strains showed activity against Staphylococcus aureus, with strain 5N708[T] producing the larger inhibition zone. Strain 5N102 exhibited markedly stronger antioxidant activity than strain 5N708[T] in radical scavenging, ferric reducing antioxidant power, and reducing power assays. In contrast, strain 5N708[T] showed more promising cytotoxic activity, with relative selectivity toward MIA PaCa-2 pancreatic cancer cells compared with HEK293 cells after prolonged incubation. Genome mining revealed multiple biosynthetic gene clusters in both isolates, supporting their capacity to produce secondary metabolites. These findings indicate that archaeological soils are promising reservoirs of taxonomically novel and biologically active Saccharopolyspora strains.