Digital DNA-DNA Hybridization Research Articles

Campylobacter sputorum subsp. bovis subsp. nov., isolated from cattle, and an emended description of Campylobacter sputorum.

Six urease-negative Campylobacter strains were isolated from cattle faeces over a 19-month period from 2009 to 2010. These strains were initially identified as Campylobacter sputorum by 16S rRNA gene and atpA typing. Initial studies characterizing these strains by multilocus sequence typing and genome sequencing further supported their classification as C. sputorum but indicated that these strains form a divergent clade within the species. A polyphasic study was undertaken here to clarify their taxonomic position. Phylogenetic analyses were performed based on 16S rRNA gene sequences and the concatenated sequences of 330 core genes, with the latter analysis also placing the six strains into a clade distinct from the three C. sputorum biovars. Pairwise digital DNA-DNA hybridization values identified these strains as C. sputorum, and the pairwise average nucleotide identity values were consistent with those observed between current Campylobacter subspecies pairs. Standard phenotypic testing was also performed. All strains are microaerobic, anaerobic, motile, Gram-negative and oxidase- and catalase-positive; cells are curved rods or spirals. Strains can be distinguished from the C. sputorum biovars by the presence of alkaline phosphatase activity and triphenyltetrazolium chloride reduction and absence of nitrate reduction. The data presented here show that these strains represent a novel subspecies within C. sputorum, for which the name C. sputorum subsp. bovis subsp. nov. (type strain RM8705T=LMG 32300T=CCUG 75470T) is proposed.

Description of Olsenella absiana sp. nov., an anaerobic bacterium isolated from pig faeces.

A short-rod-shaped, obligate anaerobic, Gram-positive bacterium isolated from the faeces of a pig was designated as the strain YH-ols2223T. Analysis of the 16S rRNA gene sequence revealed that the isolate was most closely related to Olsenella urininfantis KCTC 25755T with 97.7% similarity. The multi-locus sequence tree revealed that the isolate formed a sub-cluster within the genus Olsenella. The average nucleotide identity and the digital DNA-DNA hybridization values for the isolate and the most closely related strains were 74.9-76.6% and 20.8-22.5%, respectively. The end products of the fermentation were lactic acid and acetic acid. The main fatty acids identified were C18 : 0 DMA, C18 : 1 ω9c DMA and C18 : 1 ω9c. The cell wall contained the meso-diaminopimelic acid-based peptidoglycan. Given the chemotaxonomic, phenotypic and phylogenetic properties, YH-ols2223T (= KCTC 25800T = NBRC 116680T) represents a novel taxon. The name Olsenella absiana sp. nov. was proposed.

Roseovarius phycicola sp. nov. and Roseovarius rhodophyticola sp. nov., isolated from marine red algae.

Two Gram-stain-negative, strictly aerobic, non-motile, rod-shaped bacteria, designated as strains S88T and W115T, exhibiting catalase- and oxidase-positive reactions, were isolated from marine red algae in South Korea. Strain S88T exhibited growth at 20-30 °C, pH 6.0-9.0 and 2.0-5.0% (w/v) NaCl, while strain W115T grew at 20-30 °C, pH 7.0-9.0 and 2.0-5.0% (w/v) NaCl. Strain S88T contained summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), C16 : 0 and C16 : 0 2-OH as major fatty acids (>5%), with major polar lipids being phosphatidylglycerol, phosphatidylcholine, an unidentified phospholipid, an unidentified aminolipid and two unidentified lipids. Strain W115T contained summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c) and C16 : 0 as major fatty acids (>5%), with major polar lipids including phosphatidylglycerol, phosphatidylcholine, an unidentified phospholipid, an unidentified aminolipid and unidentified lipids. Ubiquinone-10 was the sole respiratory quinone in both strains, and the genomic DNA G+C contents were 57.0% for strain S88T and 56.5% for strain W115T. Despite 99.86% 16S rRNA gene sequence similarity, strains S88T and W115T shared 88.8% average nucleotide identity (ANI) and 36.8% digital DNA-DNA hybridization (dDDH) value, indicating different species. Phylogenetic and phylogenomic analyses based on 16S rRNA gene and genome sequences, respectively, revealed that strains S88T and W115T formed a phylogenetic lineage within the genus Roseovarius. ANI and dDDH values of both strains with other type strains were less than 73.7 and 20.3%, respectively, confirming that they represent novel species. Based on phenotypic, chemotaxonomic and molecular characteristics, strains S88T and W115T represent two novel species of the genus Roseovarius, for which the names Roseovarius phycicola sp. nov. (S88T =KACC 23423T =JCM 36647T) and Roseovarius rhodophyticola sp. nov. (W115T =KACC 23690T =JCM 36651T) are proposed, respectively.

Aurantiacibacter flavus sp. nov. and Aurantiacibacter gilvus sp. nov., isolated from the mudflat of Suaeda japonica colonies.

Two novel strains were isolated from the mudflat of Suaeda japonica colonies in Incheon, Republic of Korea. Designated as DGU5T and DGU6T, these strains were Gram-stain-negative, facultatively anaerobic and rod-shaped and had yellowish colonies. Both strains were determined to belong to the genus Aurantiacibacter through phylogenetic analysis of their 16S rRNA sequences and draft genomes. The cells of strain DGU5T were non-motile and grew at temperatures ranging between 7-45°C (optimum, 25-30°C), pH 6.0-10.0 (optimum, 7.0-8.0) and in the presence of 0-11.0% NaCl (optimum, 2.0%). The cells of strain DGU6T were non-motile and grew in temperatures ranging from 10-45 °C (optimum, 30-35°C), pH 3.0-10.0 (optimum, 7.0-8.0) and in the presence of 0-11.0% NaCl (optimum, 2.0%). Overall genome relatedness index calculations revealed average nucleotide identity values (72.3-88.6%) and digital DNA-DNA hybridization values (18.8-35.9%) aligning with those of the genus Aurantiacibacter. The major fatty acids in both strains were C17:1 ω6c and summed feature 8 (C18:1 ω6c/C18:1 ω7c), while the predominant polar lipids were sphingoglycolipid, phosphatidylglycerol, and diphosphatidylglycerol. Phylogenetic, average nucleotide identity, digital DNA-DNA hybridization, physiological, and biochemical data collectively demonstrated the distinctiveness of the novel strains from other members within the family Erythrobacteraceae. We propose the names A. flavus sp. nov. (type strain DGU5T = KACC 23720T = TBRC 19015T) and A. gilvus sp. nov. (type strain DGU6T = KACC 23721T = TBRC 19016T) for the two strains.

Microbispora maris sp. nov., a novel actinobacterium isolated from the gill of the leopard coral grouper (Plectropomus leopardus).

A novel actinomycete, designated strain ZYX-F-249T, was isolated from the gill of a leopard coral grouper in Yongxing Island, Hainan Province, China. Based on 16S rRNA gene sequence analysis, strain ZYX-F-249T belonged to the genus Microbispora, with high similarities to Microbispora rosea ATCC 12950T (98.7 %), Microbispora hainanensis 211020T (98.5 %), Microbispora clausenae CLES2T (98.4 %), Microbispora bryophytorum NEAU-TX2-2T (98.1 %) and Microbispora oryzaeRL4-1ST (98.0 %). Phylogenetic analysis of the 16S rRNA gene sequence of strain ZYX-F-249T showed that the strain formed a stable subclade with Microbispora rosea ATCC 12950T in the genus Microbispora. The cell wall of the novel isolate contained meso-diaminopimelic acid, while whole-cell sugars were madurose, glucose and ribose. The predominant menaquinones were MK-9(H6), MK-9(H4), MK-9(H2) and MK-9(H0). The characteristic phospholipids (PLs) were phosphatidylethanolamine, phosphatidylinositol, diphosphatidylglycerol, phosphatidylmethylethanolamine, phosphatidylinositol mannoside, glycophospholipids and an unknown PL. The major cellular fatty acids (>10%) were iso-C16 : 0 and iso-C18 : 0. Genome sequencing showed a DNA G+C content of 71.6%. The low average nucleotide identity, digital DNA-DNA hybridization and average amino acid identity values demonstrated that strain ZYX-F-249T could be readily distinguished from closely related species. Based on data from these phylogenetic, chemotaxonomic and physiological characteristics, strain ZYX-F-249T represents a novel species of the genus Microbispora, for which the name Microbispora maris sp. nov. is proposed. The type strain is ZYX-F-249T (=CCTCC AA 2023030T =JCM 36778T).

Bifidobacterium favimelis sp. nov., isolated from black comb honey.

A strain of the bacterial genus Bifidobacterium, IMAU50987T, that was isolated from black comb honey from Lianghe County, Dehong Autonomous Prefecture, Yunnan Province, China, could not be accurately identified as a known species using 16S rRNA gene sequencing. In this study, the taxonomic status of strain IMAU50987T was evaluated using whole-genome sequencing combined with phenotypic analyses. Strain IMAU50987T was Gram-positive, non-motile and non-spore-forming. Comparative of 16S rRNA gene sequence similarity analysis indicated that strain IMAU50987T could be attributed to the genus Bifidobacterium and most closely related to the B. mellis Bin7NT (98.5% identity). Comparative sequence analysis of eight housekeeping genes (pyrG, thrS, glnA1, recA, tuf, dnaB, rpoC and xfp) showed that strain IMAU50987T was different from closely related species. Phylogenomic analysis based on the 88 single-copy genes in the phylogenetic tree showed that strain IMAU50987T belonged to the B. asteroides group. The highest average nucleotide identity value for strain IMAU50987T was 77.0% in comparison with B. choladohabitans B14384H11T and B. apis F753-1T. However, the highest digital DNA-DNA hybridization value was 23.1% compared with B. apis F753-1T and B. xylocopae XV2T; the highest average amino acid identity value was 73.2% compared with B. polysaccharolyticum W8117T. These values are below the accepted threshold for novel species boundaries. The draft genome size of strain IMAU50987T was 1.9 Mb, and the DNA G+C content was 63.5%. The major cellular fatty acids of strain IMAU50987T were C16 : 0, C18 : 1 ω9c and Sum in Feature 7 (C19 : 1 ω7c/C19 : 1 ω6c). The cell-wall peptidoglycan was of the A3α l-Lys-Gly type. Based on data from this study, a novel species, Bifidobacterium favimelis sp. nov., is proposed with the type strain IMAU50987T (= GDMCC 1.4189T = JCM 36315T).

Cultivation of deep-sea bacteria from the Northwest Pacific Ocean and characterization of Limnobacter profundi sp. nov., a phenol-degrading bacterium

Despite previous culture-independent studies highlighting the prevalence of the order Burkholderiales in deep-sea environments, the cultivation and characterization of deep-sea Burkholderiales have been infrequent. A total of 243 deep-sea bacterial strains were isolated from various depths in the Northwest Pacific Ocean, with 33 isolates (13.6%) from a depth of 4000 m classified into Burkholderiales. Herein, we report the isolation and genome characteristics of strain SAORIC-580T, from a depth of 4000 m in the Northwest Pacific Ocean. The strain showed a close phylogenetic relationship with Limnobacter thiooxidans CS-K2T, sharing 99.9% 16S rRNA gene sequence identity. The complete whole-genome sequence of strain SAORIC-580T comprised 3.3 Mbp with a DNA G+C content of 52.5%. Comparative genomic analysis revealed average nucleotide identities between 79.4–85.7% and digital DNA-DNA hybridization values of 19.9–29.5% when compared to other Limnobacter genomes, indicating that the strain represents a novel species within the genus. Genomic analysis revealed unique adaptations to deep-sea conditions, including genes associated with phenol degradation, stress responses, cold adaptation, heavy metal resistance, signal transduction, and carbohydrate metabolism. The SAORIC-580T genome was found to be more abundant in the deep sea than at the surface in the trenches of the Northwest Pacific Ocean, suggesting adaptations to the deep-sea environment. Phenotypic characterization highlighted distinct differences from other Limnobacter species, including variations in growth conditions, enzyme activities, and phenol degradation capabilities. Chemotaxonomic markers of the strain included ubiquinone-10, major fatty acids such as C16:0, C16:1, and C18:1, and major polar lipids including phosphatidylethanolamine, phosphatidylglycerol, and diphosphatidylglycerol. Based on the polyphasic taxonomic data, it is concluded that strain SAORIC-580T (= KACC 21440T = NBRC 114111T) represents a novel species, for which the name Limnobacter profundi sp. nov. is proposed.

Polyphasic Identification of Rhizomonospora bruguierae gen. nov., sp. nov., Isolated from Mangrove Rhizosphere Soil.

A novel yellowish-white actinobacterial strain, designated as TS60-4CT, was isolated from the rhizosphere soil of the mangrove Bruguiera gymnorhiza (L.) Lam. in Okinawa, Japan, and was subjected to a polyphasic assessment. The strain could grow at 0-3.0% NaCl concentrations (w/v), pH 6.0-10.0, and 20-37°C. The 16S rRNA gene sequences-based phylogeny showed that the novel isolate belongs to the family Micromonosporaceae and that it shared the highest sequence similarity (98.4%) with Micromonospora craniellae LHW63014T. The hydrolysate of cell-wall of strain TS60-4CT contained alanine, glutamic acid, glycine, and meso-A2pm. The acyl type of muramic acid was N-glycolyl. The main (> 10%) fatty acids were anteiso-C17:0 (22.1%), iso-C16:0 (17.8%), iso-C15:0 (14.7%), and anteiso-C15:0 (10.3%) and the predominant menaquinones of the isolate were MK-9(H6) and MK-9(H4). The polar lipids of the isolate were phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, an unidentified aminolipid, and three unidentified lipids. The genome size of strain TS60-4CT was 6.7 Mbp with a DNA G + C content of 72.4%. The average nucleotide identity, digital DNA-DNA hybridization, and average amino acid identity value could differentiate strain TS60-4CT from its closely related taxa. The genome of the strain had 22 putative biosynthetic gene clusters for secondary metabolites, indicating a significant potential as a producer of bioactive chemicals. Consequently, the strain is considered to represent a novel genus and a new species of the family Micromonosporaceae, for which the name Rhizomonospora bruguierae gen. nov., sp. nov. is proposed. The type species is R. bruguierae, with the type strain TS60-4CT (= NBRC 107566T = TBRC 2024T).

Inconsistent identification of Apilactobacillus kunkeei-related strains obtained by well-developed overall genome-related indices

During a decade, overall genome related indices, including average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH), have been used as standards for the classification and identification of bacteria. On the other hand, a former study suggested that ANI-based classification is difficult for fructophilic lactic acid bacterium Apilactobacillus kunkeei. In the present study, the classification of Apilactobacillus spp., including A. kunkeei, was evaluated by multiple genome-based analyses. ANIb-based classification appropriately identified strains of Apilactobacillus spp., except for A. kunkeei-related strains. A number of strain pairings in A. kunkeei-related strains showed ANIb values around the threshold value of 95 %, based on which they were unable to be identified. On the other hand, dDDH provided clearer identification results for A. kunkeei-related strains but segmentalized them into a number of groups, while the validity of this segmentation was unclear. Certain strains shared similarities over the threshold with multiple species-level taxonomic groups in ANIb and dDDH. GTDB-Tk classifies the A. kunkeei-related strains into six species-level taxonomic groups without marked confusion, while the classification results differed from those obtained by ANIb and dDDH. The present study highlighted the inconsistent identification of A. kunkeei-related strains by the well-developed overall genome related indices, which would be a significant concern for bacterial taxonomy. Moreover, the rule adopted in GTDB-Tk, i.e., the classification of strains to taxa containing type strains showing the highest similarity, is recommended for introduction into ANIb- and GGDC-based classifications

Comparative genomic analysis and characterization of novel high-quality draft genomes from the coal metagenome.

Coal, a sedimentary rock harbours a complex microbial community that plays a significant role in its formation and characteristics. However, coal metagenome sequencing and studies were less, limiting our understanding of this complex ecosystem. This study aimed to reconstruct high-quality metagenome-assembled genomes (MAGs) from the coal sample collected in the Neyveli mine to explore the unrevealed diversity of the coal microbiome. Using Illumina sequencing, we obtained high-quality raw reads in FASTQ format. Subsequently, de novo assembly and binning with metaWRAP software facilitated the reconstruction of coal MAGs. Quality assessment using CheckM identified 10 High-Quality MAGs (HQ MAGs), 7 medium-quality MAGs (MQ MAGs), and 6 low-quality MAGs (LQ MAGs). Further analysis using GTDB-Tk revealed four HQ MAGs as known species like Dermacoccus abyssi, Sphingomonas aquatilis, Acinetobacter baumannii, and Burkholderia cenocepacia. The remaining six HQ MAGs were classified as Comamonas, Arthrobacter, Noviherbaspirillum, Acidovorax, Oxalicibacterium, and Bordetella and designated as novel genomes by the validation of digital DNA-DNA hybridization (dDDH). Phylogenetic analysis and further pangenome analysis across the phylogenetic groups revealed a similar pattern with a high proportion of cloud genes. We further analysed the functional potential of these MAGs and closely related genomes using COG. The comparative functional genomics revealed that novel genomes are highly versatile, potentially reflecting adaptations to the coal environment. BlastKOALA was used to conduct a detailed analysis of the metabolic pathways associated with the MAGs. This study highlights the comparative genomic analysis of novel coal genomes with their closely related genomes to understand the evolutionary relationships and functional properties.

Pseudomonas huanghezhanensis sp. nov. and Pseudomonas fjordensis sp. nov., two novel species isolated from seawater of Arctic Kongsfjorden in the Ny-Ålesund area, Svalbard.

Two Gram-stain-negative bacterial strains, BSw22131T and DMSP-1T, were isolated from seawater of Arctic Kongsfjorden in the Ny-Ålesund area, Svalbard. Strains BSw22131T and DMSP-1T were within the genus Pseudomonas and presented 98.1 and 99.8% 16S rRNA gene sequence similarity to Pseudomonas piscicola P50T and Pseudomonas paracarnis V5/DAB/2/5T, respectively. However, the results of multilocus sequence analysis (concatenated 16S rRNA, gyrB, rpoB and rpoD gene sequences) and whole-genome sequence analysis indicated that BSw22131T and DMSP-1T were most closely related to Pseudomonas petrae P2653T and Pseudomonas lactis DSM 29167T, respectively. The genome G+C contents of strains BSw22131T and DMSP-1T were 58.2 and 60.0%, respectively. The two isolates presented digital DNA-DNA hybridization and average nucleotide identity values of less than 70 and 96%, respectively, with type strain genomes of their close relatives. BSw22131T contained summed feature 3 (C16 : 1 ω6c/C16 : 1 ω7c), C16 : 0 and summed feature 8 (C18 : 1 ω6c/C18 : 1 ω7c) as the major cellular fatty acids, whereas DMSP-1T contained an additional C17 : 0-cyclo as the major fatty acid. The major quinones of both strains were Q-9 (57.5-77.2%) and Q-8 (22.8-37.2%), and their major cellular polar lipids were diphosphatidylglycerol, phosphatidylethanolamine and phosphatidylglycerol. BSw22131T could be differentiated from its related species by positive ornithine decarboxylase, esterase (C8) and l-rhamnose and d-fructose-6-PO4 utilization, whereas DMSP-1T could be differentiated by positive acid phosphatase but negative p-hydroxyphenylacetic acid utilization. The two strains were negative for the reduction of nitrates but contained genes related to dimethylsulfoniopropionate catabolism. The results of the polyphasic taxonomy analysis revealed that strains BSw22131T and DMSP-1T represent two novel species of the genus Pseudomonas, for which the names Pseudomonas huanghezhanensis sp. nov. (type strain BSw22131T = CCTCC AB 2019135T = JCM 36635T = DSM 117336T) and Pseudomonas fjordensis sp. nov. (type strain DMSP-1T = CCTCC AB 2019124T = JCM 36636T = DSM 117337T) are proposed.

Bartonella Species in Small Mammals in Turkey: Bartonella bilalgolemii sp. nov. Isolated from a Ural Field Mouse (Apodemus uralensis).

Background: The genus Bartonella is composed of Gram-negative, fastidious, facultative intracellular bacteria that can cause bacteremia in mammals and various disorders in humans. Rodents have been reported as reservoirs of more than 30 Bartonella species, seven of which cause zoonotic infections. Materials and Methods: In the present study, the isolation of Bartonella sp. was attempted from 150 spleen samples from 13 rodent species (mostly Apodemus species) from three geographically different regions in Turkey. Results: Bartonella sp. was successfully isolated from 65 of these 150 samples (43%). The prevalences of Bartonella sp. in tested rodents in the regions of Giresun, Yozgat, and Burdur were 68%, 44%, and 16%, respectively. Using polymerase chain reaction/sequence analysis of the citrate synthase-coding gene (gltA), Bartonellaisolates were classified seven species including B. taylorii, B. grahamii, B. birtlesii, B. mastomydis, and three putatively new Bartonella species. We performed further identification techniques for one of the three Bartonella species that were different from the validated Bartonella species according to the gltA sequence analysis. Conclusion: Here, we report the genomic and phenotypic characterization of Bartonella sp. strain G70 that was isolated from the splenic tissue of an Apodemus uralensis (Pallas 1881), the Ural field mouse, captured in the Giresun region of northeastern Turkey. Bartonella sp. strainG70 (RSKK 22001) was characterized by whole genome and partial gene (gltA, 16S ribosomal RNA) sequencing and comparison, scanning electron microscopy, biochemical tests, and matrix-assisted laser desorption ionization time-of-flight mass spectrometry. This novel Bartonella is a Gram-negative, rod-shaped bacterium and has neither flagella nor pili. The genome from strain G70 was 1,606,969-bp-long with a G + C content of 35.7%. Bartonella rochalimae was found to be the closest phylogenetic relative of strain G70 (OrthoANI = 90.5%, digital DNA-DNA hybridization = 41.4%). We therefore propose that this new species be named Bartonella bilalgolemii sp. nov. with strain G70T as the type strain.

Efficient low-temperature wastewater treatment by Pseudomonas zhanjiangensis sp. nov.: a novel cold-tolerant bacterium isolated from mangrove sediment.

A novel heterotrophic, cold-tolerant bacterium, designated Pseudomonas zhanjiangensis 25A3ET, was isolated from mangrove sediment and demonstrated excellent efficiency in cold wastewater treatment. Phylogenetic analysis based on 16S rRNA gene sequences positioned strain 25A3ET within the genus Pseudomonas, showing the highest similarity (98.7%) with Pseudomonas kurunegalensis LMG 32023T. Digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values were below the species delineation thresholds (70% for dDDH, 95% for ANI), indicating that strain 25A3ET represents a novel species. This strain demonstrated high efficiency in removing nitrogen (N) and organic pollutants under low-temperature conditions. Specifically, it achieved 72.9% removal of chemical oxygen demand (COD), 70.6% removal of ammoniacal nitrogen (NH4 +-N), and 69.1% removal of total nitrogen (TN) after 96 h at 10°C. Genomic analysis identified key genes associated with cold adaptation, nitrogen removal and organic matter degradation. These findings indicate that Pseudomonas zhanjiangensis 25A3ET holds significant potential for application in cold temperature wastewater treatment, offering a promising solution for environmental remediation in regions with low ambient temperatures.

Paracoccus spongiarum sp. nov., isolated from the marine sponge, Phakellia elegans.

A facultative anaerobic Gram-negative bacterium, designated as strain 2205BS29-5T, was isolated from a marine sponge, Phakellia elegans, in Beomseom on Jeju Island, Republic of Korea, and taxonomically characterized. The cells were catalase and oxidase positive, non-motile, coccoid-rod shaped and capable of poly-β-hydroxybutyrate production. Growth was observed at 10-37 °C (optimum, 25 °C) and pH 5.0-8.0 (optimum, pH 7.0), and in the presence of 0-9% NaCl (w/v) (optimum, 3.0-4.0%). The major cellular fatty acids and respiratory quinone were identified as summed feature 8 (C18 : 1 ω7c/C18 : 1 ω6c) and Q-10, respectively. The major polar lipids comprised diphosphatidylglycerol, phosphatidylglycerol, four phosphoglycolipids, two unidentified amino lipids and eight unidentified lipids. The DNA G+C content was 67.8%. Strain 2205BS29-5T was most closely represented by Paracoccus amoyensis 11-3T and P. caeni MJ17T with 97.8 and 97.5% 16S rRNA gene sequence similarities, respectively. Phylogenetic analyses based on 16S rRNA genes and whole-genome sequences showed that strain 2205BS29-5T was affiliated with the genus Paracoccus. Genomic analysis showed that strain 2205BS29-5T could synthesize vitamin B family (folate and cobalamin) and ectoine. The average nucleotide identity and digital DNA-DNA hybridization values between strain 2205BS29-5T and P. amoyensis 11-3T were 77.1% and 18.8%, respectively, and with P. caeni MJ17T were 78.4 and 21.2%, respectively. Based on phylogenetic, chemotaxonomic and genome relatedness analyses, strain 2205BS29-5T represents a novel species of the genus Paracoccus, for which the name Paracoccus spongiarum sp. nov. is proposed. The type strain is 2205BS29-5T (=LMG 33062T =KACC 23240T).

Amycolatopsis melonis sp. nov., a novel protease-producing and cellulose-degrading actinobacterium isolated from soil.

A novel protease-producing and cellulose-degrading actinobacterium, designated strain NEAU-NG30T, was isolated from a melon rhizosphere soil sample collected in Harbin, Heilongjiang Province, China, and established its status using a polyphasic taxonomic study. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain NEAU-NG30T was closely related to Amycolatopsis bullii DSM 45802T (98.7%) and Amycolatopsis vancoresmycina DSM 44592T (98.3%). The phospholipid profile contained diphosphatidylglycerol, phosphatidyl methylethanolamine, phosphatidylethanolamine and phosphatidylinositol. The diagnostic sugars in cell hydrolysates were determined to be galactose and arabinose. Cell walls contained meso-diaminopimelic acid as the diagnostic diamino acid. The predominant menaquinone was MK-9(H4). The major fatty acids were iso-C15:0 and iso-C16:0. Meanwhile, genome analysis of sequences revealed a genome size of 9 338 250 bp and a DNA G+C content of 71.6%. In addition, the average nucleotide identity values and the level of digital DNA-DNA hybridization between strain NEAU-NG30T and its reference strains fall below the thresholds typically used for delineating prokaryote species. According to phenotypic, chemotaxonomic and genotypic studies, it is indicated that strain NEAU-NG30T is considered to be a novel species of the genus Amycolatopsis, for which the name Amycolatopsis melonis sp. nov. is proposed, with NEAU-NG30T (=MCCC 1K08677T=JCM 35654T) as the type strain.

Janibacter alittae sp. nov., an actinobacterium isolated from the gut of marine sandworm (Alitta virens).

A novel Gram-stain-positive, facultatively anaerobic, non-motile, catalase-positive, oxidase-negative and ovoid cocci, designated as A1S7T, was isolated from the gut of a marine sandworm (Alitta virens). Strain A1S7T exhibited optimal growth at temperatures of 20-30 ℃, pH 6-8 and in the presence of 2-4% (w/v) NaCl. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain A1S7T belonged to the genus Janibacter, exhibiting a similarity of 99.0% to Janibacter cremeus KCTC 49873T, followed by Janibacter anophelis KCTC 19282T (98.8%), Janibacter hoylei KCTC 49872T (98.4%), Janibacter limosus KACC 20518T (98.2%) and Janibacter corallicola KACC 21120T (97.2%). The complete genome sequence of strain A1S7T revealed a genome size of 3360920 bp with a genomic G+C content of 70.1 mol%. The orthologous average nucleotide identity and the digital DNA-DNA hybridization values between strain A1S7T and Janibacter cremeus KCTC 49873T were determined to be 89.5 and 37.2%, respectively. The major respiratory quinone was MK-8(H4). The predominant fatty acids (>10%) included iso-C16:0, C17:1 ω8c, C18:1 ω9c and C17:0. Polar lipids comprised diphosphatidylglycerol, phosphatidylglycerol, phosphatidylinositol, one unknown phosphoglycolipid and three unknown polar lipids. The cell-wall peptidoglycan type was A1γ. The major whole-cell sugars were ribose, mannose and glucose. Based on phenotypic, phylogenetic, genotypic and chemotaxonomic properties, strain A1S7T represents a novel species in the genus Janibacter, for which the name Janibacter alittae sp. nov. is proposed. The type strain is A1S7T (=KCTC 49714T = JCM 36706T).

Paracoccus actinidiae sp.nov., a novel bacterium isolated from kiwi tree rhizosphere soil.

Strain M09T was isolated from the rhizoshere of kiwi fruit trees from an orchard located in Fangshan, Beijing, PR China (39° 49' 25.1″ N, 116° 4' 44.5″ E,). It is a short rod-shaped, Gram-stain-negative, facultatively anaerobic bacterium that tests positive for both oxidase and catalase. The strain exhibited growth within the temperature range of 15-45 °C (optimal growth at 30 °C) and the pH range of 4.0-10.0 (optimal growth at pH 7.0) and without NaCl. It also grew in a sodium chloride-free nutrient agar (NA) medium. The results of phylogenetic analysis of the 16S rRNA gene sequences indicated that M09T represents a member of the genus Paracoccus and shares high similarity with Paracoccus everestensis S8-55T (98.46%) and Paracoccus aerius 011410T (97.58%). The average nucleotide identity values between M09T and P. everestensis S8-55T, P. aerius 011410T, Paracoccus marinaquae X HP0099T and Paracoccus fontiphilus MVW-1T were 95.56, 84.51, 79.83 and 83.68%, respectively. The digital DNA-DNA hybridisation values between between M09T and P. everestensis S8-55T, P. aerius 011410T, P. marinaquae X HP0099T and P. fontiphilus MVW-1T were 56.40, 29.30, 21.60 and 28.60%, respectively. The major fatty acids identified were C10 : 0 3-OH (51.8%) and C18 : 1ω7c (35.5%). The major respiratory quinone was Q-10, with Q-8 present as a minor component. Polar lipids were mainly comprised of diphosphatidylglycerol (DPG), phosphatidylglycerol (PG), phosphatidylcholine (PC), and phosphatidylethanolamine (PE). Genome sequencing revealed that the strain has a DNA G+C content of 64.31 mol%. On the basis of this comprehensive taxonomic characterisation data, M09T represents a novel species within the genus Paracoccus and has been named Paracoccus actinidiae sp. nov. The type strain is designated as M09T (=GDMCC 1.4157T=KCTC 8143T).

Luteibacter sahnii sp. nov., A Novel Yellow-Colored Xanthomonadin Pigment Producing Probiotic Bacterium from Healthy Rice Seed Microbiome.

To explore the rice seed microbiome, our objective was to isolate novel strains of Xanthomonas, a plant-associated bacterium with diverse lifestyles. Four isolates, anticipated to be Xanthomonas based on morphological features of yellow colonies, were obtained from healthy rice seeds. Phylo-taxono-genomic analysis revealed that these isolates formed monophyletic lineages belonging to a novel species within the genus Luteibacter. Pairwise ortho Average Nucleotide Identity and digital DNA-DNA hybridization confirmed their distinct species status. We propose Luteibacter sahnii sp. nov. as a novel species, with PPL193T = MTCC 13290T = ICMP 24807T = CFBP 9144T as the type strain and PPL201, PPL552, and PPL554 as other constituent members. The fatty acid profile of the type strain is dominated by branched fatty acids like Iso-C15:0, consistent with other members of the genus. The novel species displays non-pathogenic attributes and exhibits plant probiotic properties, protecting rice plants from the leaf blight pathogen X. oryzae pv. oryzae. Production of Indole-3-Acetic Acid (IAA) and genomic regions encoding anti-microbial peptides emphasize its potential contributions to plant hosts. This study underscores the importance of employing a combination of phenotypic and genotypic methods in culturomics to enhance our understanding of rice seed microbiome diversity.

Flavobacterium capsici sp. nov., isolated from the rhizospheric soils of bell pepper (Capsicum annuum L.).

Two separate bacterial strains, PMTSA4T and PMR2A8, were isolated from the rhizospheric soils of bell pepper plants grown in a plant nursery. These strains are Gram-negative, non-motile and rod-shaped and grow in aerobic conditions. They exhibit a positive reaction for catalase activity but negative results for oxidase activity. Phylogenetic analysis of the 16S rRNA gene sequences revealed that the strains PMTSA4T and PMR2A8 are closely related to Flavobacterium piscinae ICH-30T (95.6%, respectively), Flavobacterium ahnfeltiae 10Alg 130T (95.5%) and Flavobacterium maris KMM 9535T (95.3%), aligning them within the genus Flavobacterium. Digital DNA-DNA hybridization (dDDH) values and average nucleotide identities (ANIs) of the whole-genome sequences for the two strains and related Flavobacterium species were significantly below the established thresholds for prokaryotic species delineation (<70% for dDDH and <95% for ANI). The observed values were as follows: Flavobacterium aquatile LMG 4008T (dDDH: 19.8% and ANI: 75.5%), F. piscinae ICH-30T (dDDH: 18.6% and ANI: 73.3%) and F. stagni WWJ 16T (dDDH: 18.5% and ANI: 72.0%). The strains have genome sizes of 3 068 185 bp and 3 068 330 bp, with a G+C content of 32.5 mol%. In phenotypic characterization, the new strains grew at 10-35 °C and tolerated up to 4% NaCl at pH 5-9 (optimum pH 8). The predominant cellular fatty acids were observed to be iso-C15:0, iso-C17:0 3-OH and iso-C15:0 3-OH. Menaquinone-6 was the predominant quinone. Considering the results from phenotypic, chemotaxonomic, phylogenetic and genomic analyses, it is proposed that the strains PMTSA4T and PMR2A8 represent a novel species within the genus Flavobacterium.

Streptomyces yaizuensis sp. nov., a berninamycin C-producing actinomycete isolated from sponge.

While screening for antibiotics in a marine sample, we discovered a berninamycin C-producing actinomycete, designated YSPA8T, isolated from a sponge. A polyphasic approach was used to determine the taxonomic position of the strain. Strain YSPA8T formed sympodially branched aerial mycelia that ultimately segment into chains of spores. Comparative and phylogenetic analyses of the 16S rRNA gene sequence showed that strain YSPA8T were closely related to Streptomyces clavuligerus ATCC 27064T (99.66%), Streptomyces amakusaensis NRRL B-3351T (98.69%), Streptomyces inusitatus NBRC 13601T (98.48%), and 'Streptomyces jumonjinensis' JCM 4947 (98.41%). The phylogenetic tree using the 16S rRNA gene sequences, and both phylogenomic trees suggested that the closest relative of strain YSPA8T was S. clavuligerus ATCC 27064T. The average nucleotide identity, digital DNA-DNA hybridization, and average amino acid identity values between strain YSPA8T and S. clavuligerus ATCC 27064T were 84.1%, 28.9%, and 82.5%, respectively, which were below the thresholds of 95%, 70%, and 95% for a prokaryotic conspecific assignment. The G + C of the strain YSPA8T was 72.6%. Whole-cell hydrolysates of strain YSPA8T contained LL-diaminopimelic acid. The predominant menaquinones were MK-9(H6) (49%) and MK-9(H8) (48%), and the major fatty acids were C16:0 (26.8%), C16:1 ω7c/ω6c (17.2%), iso-C16:0 (16.0%), and iso-C15:0 (12.5%). The major phospholipids were diphosphatidylglycerol, phosphatidylethanolamine, and other unidentified phospholipids. Based on the phenotypic, phylogenetic, genomic, and chemotaxonomic data, strain YSPA8T represents a novel species of the genus Streptomyces, and the proposed name for this species is Streptomyces yaizuensis sp. nov. The type strain is YSPA8T (=NBRC 115866T = TBRC 17196T).