Microbacterium humicola sp. nov., Microbacterium terrisoli sp. nov., Paenibacillus pedocola sp. nov., Paenibacillus silviterrae sp. nov., Flavobacterium terrisoli sp. nov., and Aquabacterium humicola sp. nov., isolated from soil.

Four Gram-stain-positive and two Gram-stain-negative bacterial strains, designated as W4T, FW7T, TW48T, UW52T, PT-3T, and RJY3T, were isolated from soil samples collected from the Republic of Korea. The 16S rRNA gene sequence analysis showed that strains W4T and FW7T belonged to the genus Microbacterium, strains TW48T and UW52T were affiliated to the genus Paenibacillus, strain PT-3T was related to the genus Flavobacterium, and strain RJY3T was associated with the genus Aquabacterium. The closest phylogenetic taxa to W4T, FW7T, TW48T, UW52T, PT-3T, and RJY3T were Microbacterium bovistercoris NEAU-LLET (97.7 %), Microbacterium protaetiae DFW100M-13T (97.9 %), Paenibacillus auburnensis JJ-7T (99.6 %), Paenibacillus allorhizosphaerae JJ-447T (95.7 %), Flavobacterium buctense T7T (97.1 %), and Aquabacterium terrae S2T (99.5 %), respectively. Average nucleotide identity and digital DNA-DNA hybridization values between the novel strains and related reference type strains were <95.0 % and <70.0 %, respectively. The major cellular fatty acid in strains W4T, FW7T TW48T, and UW52T was antiso-C15 : 0. Similarly, strain PT-3T revealed iso-C15 : 0, iso-C15 : 1 G, iso-C17 : 0 3-OH, and iso-C15 : 0 3-OH as its principal fatty acids. On the other hand, RJY3T exhibited summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C16 : 0, summed feature 8 (C18 : 1 ω7c and/or C18 : 1 ω6c), and C12 : 0 as its predominant fatty acids. Overall, the polyphasic taxonomic data indicated that strains W4T, FW7T, TW48T, UW52T, PT-3T, and RJY3T represent novel species within the genera Microbacterium, Paenibacillus, Flavobacterium, and Aquabacterium. Accordingly, we propose the names Microbacterium humicola sp. nov., with the type strain W4T (=KCTC 49888T=NBRC 116001T), Microbacterium terrisoli sp. nov., with the type strain FW7T (=KCTC 49859T=NBRC 116000T), Paenibacillus pedocola sp. nov., with the type strain TW48T (=KCTC 43470T=NBRC 116017T), Paenibacillus silviterrae sp. nov., with the type strain UW52T (=KCTC 43477T=NBRC 116018T), Flavobacterium terrisoli sp. nov., with the type strain PT-3T (=KCTC 92106T=NBRC 116012T), and Aquabacterium humicola sp. nov., with the type strain RJY3T (=KCTC 92105T=NBRC 115831T).

Flavobacterium facile sp. nov., isolated from water system of Atlantic salmon (Salmo salar) fry cultured in Chile.

Strain T-12T, an orange, Gram-stain-negative, non-motile, rod-shaped strain, was isolated in November 2013 from water samples collected from an Atlantic salmon (Salmo salar) fry culturing system at a fish farm in Chile. Phylogenetic analysis based on 16S rRNA sequences (1394 bp) revealed that strain T-12T belonged to the genus Flavobacterium, showing close relationships to Flavobacterium bernardetii F-372T (99.48 %) and Flavobacterium terrigena DS-20T (98.50 %). The genome size of strain T-12T was 3.28 Mb, with a G+C content of 31.1 mol%. Genome comparisons aligned strain T-12T with Flavobacterium bernardetii F-372T (GCA_011305415) and Flavobacterium terrigena DSM 17934T (GCA_900108955). The highest digital DNA-DNA hybridization (dDDH) values were 42.6 % with F. bernardetii F-372T (GCA_011305415) and 33.9 % with F. terrigena DSM 17934T (GCA_900108955). Pairwise average nucleotide identity (ANI) calculations were below the species cutoff, with the best results with F. bernardetii F-372T being: ANIb, 90.33 %; ANIm, 91.85 %; and TETRA, 0.997 %. These dDDH and ANI results confirm that strain T-12T represents a new species. The major fatty acids were iso-C15 : 0 and C15 : 1ω6с. Detected polar lipids included phospholipids (n=2), aminophospholipid (n=1), aminolipid (n=1) and unidentified lipids (n=2). The predominant respiratory quinone was menaquinone MK7 (80 %) followed by MK-6 (20 %). Phenotypic, chemotaxonomic, and genomic data support the classification of strain T-12T (=CECT 30410T=RGM 3222T) as representing a novel species of Flavobacterium, for which the name Flavobacterium facile sp. nov. is proposed.

Flavobacterium adhaerens sp. nov. and Flavobacterium maritimum sp. nov., two novel flavobacteria isolated from the Pearl River Estuary.

Two strains, designated as SYSU M80004T and SYSU M80005T, were isolated from water sampled in the Pearl River Estuary, Guangzhou, Guangdong, PR China. The strains were Gram-stain-negative and aerobic. Strain SYSU M80004T could grow at pH 6.0-8.0 (optimum, pH 7.0), 22-30 °C (optimum, 28 °C) and in the presence of 0-1 % NaCl (w/v; optimum 0 %). Strain SYSU M80005T could grow at pH 6.0-8.0 (optimum, pH 7.0), 4-37 °C (optimum, 28 °C) and in the presence of 0-1 % NaCl (w/v; optimum 0%). Both strains contained MK-6 as the predominant menaquinone. C16 : 0 and iso-C15 : 0 were identified as the major fatty acids (>10 %) of strain SYSU M80004T while strain SYSU M80005T contained iso-C15 : 0 and iso-C17 : 0 3-OH as major fatty acids. Phosphatidylethanolamine was present as the major polar lipid in both strains. The average nucleotide identity and digital DNA-DNA hybridization values between these two strains and their closest relatives were 73.5-79.3 % and 19.6-23.2 %, respectively. Phylogenetic analysis based on the 16S rRNA gene and genomic sequences indicated they belonged to the genus Flavobacterium. Therefore, on the basis of phenotypic, physiological, chemotaxonomic and genomic evidence, two novel species, Flavobacterium adhaerens sp. nov. (type strain=SYSU M80004T=CDMCC 1.4522T=KCTC 102268T) and Flavobacterium maritimum sp. nov. (type strain=SYSU M80005T=CGMCC 1.4523T= KCTC 102269T) are proposed.

Flavobacterium rivulicola sp. nov., Isolated from a Freshwater Stream.

A Gram-strain-negative, aerobic, yellow-colored, non-motile, and rod-shaped bacterial strain, designated IMCC34852T, was isolated from a freshwater stream in the Republic of Korea. Cellular growth occurred at 10-37 °C, pH 6.0-9.0, and with 0-0.5% (w/v) NaCl. The 16S rRNA gene sequence analysis showed that strain IMCC34852T belonged to the genus Flavobacterium and that the strain was most closely related to F. cheonhonense ARSA-15 T (97.6%), F. buctense T7T (96.7%), F. silvisoli RD-2-33 T (96.1%), and F. paronense KNUS1T (96.1%). The whole-genome sequence of strain IMCC34852T was 3.2 Mbp in size, with a DNA G + C content 37.3%. The average nucleotide identities (ANI) and digital DNA-DNA hybridization (dDDH) values between strain IMCC34852T and its related species were all below 79.8% and 22.7%, respectively, which are significantly lower than the thresholds of 95% for ANI and 70% for DDH for species delineation. The major respiratory quinone of strain IMCC34852T was menaquinone-6 (MK-6) and the predominant cellular fatty acids were iso-C15:0 (32.6%), iso-C16:0 (11.7%), iso-C15:1 G (10.3%), and iso-C14:0 (6.7%). The major polar lipids of the strain were phosphatidylethanolamine, two unidentified aminolipids and six unidentified lipids. Based on these results, it was concluded that strain IMCC34852T represents a novel species in the genus Flavobacterium, for which the name Flavobacterium rivulicola sp. nov is proposed. The type strain of the proposed novel species is IMCC34852T (= KACC 23133 T = KCTC 82066 T = NBRC 114419 T).

Fecal metagenomic and metabolomic analyses reveal non-invasive biomarkers of Flavobacterium psychrophilum infection in ayu (Plecoglossus altivelis).

With the rapid growth of inland aquaculture worldwide, side effects such as the discharge of nutrients and antibiotics pose a threat to the global environments. A sustainable future for aquaculture requires an effective management system, including the early detection of disease through the monitoring of specific biomarkers in aquaculture tanks. To this end, we investigated whether fish feces in aquaculture tanks could be used for non-invasive health monitoring using ayu (Plecoglossus altivelis) infected with Flavobacterium psychrophilum, which causes bacterial cold-water disease worldwide. Feces that were subsequently produced in the tanks were used for metagenomic and metabolomic analyses. The relative abundances of the genera Cypionkella (0.6% ± 1.0%, 0.1% ± 0.2%), Klebsiella (11.2% ± 10.0%, 6.2% ± 5.9%), and F. psychrophilum (0.5% ± 1.0%, 0.0% ± 0.0%) were significantly higher in the feces of the infection challenge test tanks than in those of the control tanks. The abundances of cortisol, glucose, and acetate in the feces of the infection challenge test tanks were 2.4, 2.4, and 1.3 times higher, respectively, than those of the control tanks. Metagenome analysis suggested that acetate was produced by microbes such as Cypionkella. The abundances of indicated microbes or metabolites increased after day 4 of infection at the earliest, and were thus considered possible biomarkers. Our results suggest that feces produced in aquaculture tanks can potentially be used for non-invasive and holistic monitoring of fish diseases in aquaculture systems. IMPORTANCE: The aquaculture industry is rapidly growing, yet sustainability remains a challenge. One crucial task is to reduce losses due to diseases. Monitoring fish health and detecting diseases early are key to establishing sustainable aquaculture. Using metagenomic and metabolomic analyses, we found that feces of ayu infected with Flavobacterium psychrophilum contain various specific biomarkers that increased 4 days post-challenge, at the earliest. Our findings are the first step in establishing a novel, non-invasive, and holistic monitoring method for fish diseases in aquaculture systems, especially in ayu, which is an important freshwater fish species in Asia, promoting a sustainable future.

Flavobacterium poyangense sp. nov., an ammonifying bacterium isolated from a freshwater lake.

A Gram-stain-negative, aerobic, non-spore-forming, nonmotile, rod-shaped, and yellow-pigmented bacterium, designated strain JXAS1T, was isolated from a freshwater sample collected from Poyang Lake in China. Phylogenetic analysis based on 16S rRNA gene sequence revealed that the isolate belonged to the genus Flavobacterium, being closest to Flavobacterium pectinovorum DSM 6368T (98.61 %). The genome size of strain JXAS1T was 4.66 Mb with DNA G+C content 35.7 mol%. The average nucleotide identity and in silico DNA-DNA hybridization values between strain JXAS1T and its closest relatives were below the threshold values of 95 and 70 %, respectively. The strain contained menaquinone 6 (MK-6) as the predominant menaquinone and the major polar lipids were phosphatidylethanolamine, one unidentified glycolipid, and one unidentified polar lipid. The major fatty acids (>5 %) were iso-C15 : 0, summed feature 3 (C16 : 1 ω7c and/or C16 : 1 ω6c), C15 : 0, iso-C17 : 0 3OH, iso-C15 : 0 3OH, and summed feature 9 (iso-C17 : 1 ω9c and/or 10-methyl C16 : 0). Based on phylogenetic, genotypic, and phenotypic evidence, the isolated strain represents a new species in the genus Flavobacterium, and the name Flavobacterium poyangense is proposed. The type strain is JXAS1T (=GDMCC 1.1378T=KCTC 62719T).

Flavobacterium acetivorans sp. nov. and Flavobacterium galactosidilyticum sp. nov., psychrotolerant flavobacteria isolated from cultivated fish.

Two yellow-coloured strains, F-29T and F-340T, were isolated from fish farms in Antalya and Mugla in 2015 and 2017 during surveillance studies. The 16S rRNA gene sequence analysis showed that both strains belong to the genus Flavobacterium. A polyphasic approach involving a comprehensive genome analysis was employed to ascertain the taxonomic provenance of the strains. The overall genome-relatedness indices of digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) between the strains and the other members of the genus Flavobacterium were found to be well below the established thresholds of 70 and 95 %, respectively. The whole-genome-based phylogenetic analysis revealed that strain F-29T is closely related to Flavobacterium granuli (dDDH 39.3 % and ANI 89.4 %), while strain F-340T has a close relationship with the type strain of Flavobacterium pygoscelis (dDDH 25.6 % and ANI 81.5 %). Both strains were psychrotolerant with an optimum growth temperature of 25 °C. The chemotaxonomic characteristics of the strains were typical of the genus Flavobacterium. Both strains had phosphatidylethanolamine, aminolipids and unidentified lipids in their polar lipid profile and MK-6 as the isoprenoid quinone. The major fatty acids were iso-C15 : 0 and anteiso-C15 : 0. The genome size of the strains was 3.5 Mb, while G+C contents were 35.3 mol% for strain F-29T and 33.4 mol% for strain F-340T. Overall, the characterizations confirmed that both strains are representatives of two novel species within the genus Flavobacterium, for which the names Flavobacterium acetivorans sp. nov. and Flavobacterium galactosidilyticum sp. nov. are proposed, with F-29T (JCM 34193T=KCTC 82253T) and F-340T (JCM 34203T=KCTC 82263T) as the type strains, respectively.

Comprehensive genome analysis of five novel flavobacteria: Flavobacterium piscisymbiosum sp. nov., Flavobacterium pisciphilum sp. nov., Flavobacterium flavipigmentatum sp. nov., Flavobacterium lipolyticum sp. nov. and Flavobacterium cupriresistens sp. nov.

Eight isolates were obtained through a study on culture-dependent bacteria from fish farms and identified as members of the genus Flavobacterium based on pairwise analysis of the 16S rRNA gene sequences. The highest pairwise identity values were calculated as 98.8 % for strain F-30 T and Flavobacterium bizetiae, 99.0 % for strain F-65 T and Flavobacterium branchiarum, 98.7 % for strain F-126 T and Flavobacterium tructae, 98.2 % for strain F-323 T and Flavobacterium cupreum while 99.7 % identity level was detected for strain F-70 T and Flavobacterium geliluteum. In addition, strains F-33, Fl-77, and F-70 T shared 100 % identical 16S rRNA genes, while strains F-323 T and Fl-318 showed 99.9 % identity. A polyphasic approach including comparative analysis of whole-genome data was employed to ascertain the taxonomic provenance of the strains. In addition to the morphological, physiological, biochemical and chemotaxonomic characteristics of the strains, the overall genome-relatedness indices of dDDH and ANI below the established thresholds confirmed the classification of the strains as five novel species within the genus Flavobacterium. The comprehensive genome analyses of the strains were also conducted to determine the biosynthetic gene clusters, virulence features and ecological distribution patterns. Based on the polyphasic characterisations, including comparative genome analyses, it is concluded that strains F-30 T, F-65 T, F-70 T, F126T and F-323 T represent five novel species within the genus Flavobacterium for which Flavobacterium piscisymbiosum sp. nov. F-30 T (=JCM 34194 T = KCTC 82254 T), Flavobacterium pisciphilum sp. nov. F-65 T (=JCM 34197 T = KCTC 82257 T), Flavobacterium flavipigmentatum sp. nov. F-70 T (Fl-33 = Fl-77 = JCM 34198 T = KCTC 82258 T), Flavobacterium lipolyticum sp. nov. F-126 T (JCM 34199 T = KCTC 82259 T) and Flavobacterium cupriresistens sp. nov. F-323 T (Fl-318 = JCM 34200 T = KCTC 82260 T), are proposed.

Analysis of microbial dynamics in the soybean root-associated environments from community to single-cell levels.

Plant root-associated environments such as the rhizosphere, rhizoplane, and endosphere, are notably different from non-root-associated soil environments. However, the microbial dynamics in these spatially divided compartments remain unexplored. In this study, we propose a combinational analysis of single-cell genomics with 16S rRNA gene sequencing. This method enabled us to understand the entire soil microbiome and individual root-associated microorganisms. We applied this method to soybean microbiomes and revealed that their composition was different between the rhizoplane and rhizosphere in the early growth stages, but became more similar as growth progressed. In addition, a total of 610 medium- to high-quality single-amplified genomes (SAGs) were acquired, including plant growth-promoting rhizobacteria (PGPR) candidates while genomes with high GC content tended to be missed by SAGs. The whole-genome analyses of the SAGs suggested that rhizoplane-enriched Flavobacterium solubilizes organophosphate actively and Bacillus colonizes roots more efficiently. Single-cell genomics, together with 16S rRNA gene sequencing, enabled us to connect microbial taxonomy and function, and assess microorganisms at a strain resolution even in the complex soil microbiome.

Flavobacterium endoglycinae sp. nov., an endophytic bacterium isolated from soybean (Glycine max L. cv. Gwangan) stems.

A Gram-stain-negative, facultatively anaerobic, motile by gliding, rod-shaped, oxidase- and catalase-positive bacterial strain, designated BB8T, was isolated from the stems of a Korean soybean cultivar (Glycine max L. cv. Gwangan). The strain produced a yellow pigment on tryptic soy agar. Growth of strain BB8T occurred at pH 5.0-8.0 (optimum, pH 7.0), at 10-35 °C (optimum, 25-30 °C) and in the presence of 0-1 % (w/v) NaCl (optimum, 0.5%). Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain BB8T formed a lineage within the genus Flavobacterium and was most closely related to Flavobacterium artemisiae SYP-B1015T (96.9 % 16S rRNA gene sequence similarity) and Flavobacterium ustbae T13T (96.8%). The complete genome sequence of strain BB8T was 5 513 159 bp long with a G+C content of 34.1 mol%. The major fatty acids (>10 %) of strain BB8T were iso-C15 : 0 (21 %), summed feature 3 (comprising C16 : 1 ω7c and/or C16 : 1 ω6c, 20.3%) and iso-C16 : 0 3-OH (13.7%). The predominant polar lipids were phosphatidylethanolamine and unidentified aminolipids, and the major respiratory quinone was menaquinone-6. Based on these phenotypic, genotypic and chemotaxonomic characteristics, strain BB8T is considered to represent a novel species of the genus Flavobacterium, for which the name Flavobacterium endoglycinae sp. nov. is proposed. The type strain is BB8T (=KCTC 82167T=CCTCC AB 2020070T).

Flavobacterium coralii sp. nov., a marine bacterium isolated from coral culture seawater.

A Gram-stain-negative, non-motile, strictly aerobic, rod-shaped bacterium, with one polar flagellum and named D11R37T, was isolated from coral culture seawater of Acropora digitifera. Strain D11R37T grew with 0-6 % (w/v) NaCl (optimum, 0.5%), at 10-41 °C (optimum, 28 °C) and at pH 6.0-7.0 (optimum, 7.0). Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain D11R37T formed a lineage within the genus Flavobacterium, and it was distinct from the most closely related species Flavobacterium suzhouense XIN-1T and Flavobacterium suaedae G16-7T with 16S rRNA gene sequences similarities of 95.97% and 95.48 %. The major respiratory quinone was menaquinone-6. The polar lipids comprised one phosphatidylethanolamine, two aminolipids and one unknown polar lipid. The predominant fatty acids (more than 10 % of total fatty acids) were iso-C15 : 0 (18.0%), iso-C17 : 0 3-OH (11.9 %) and summed feature 3 (10.9 %). The DNA G+C content was 41.3 mol%. Based on polyphasic taxonomic data, strain D11R37T is considered to represent a novel species within the genus Flavobacterium, for which the name Flavobacterium coralii sp. nov. is proposed. The type strain is D11R37T (=KCTC 82968T=MCCC 1K06440T).

Flavobacterium hydrocarbonoxydans sp. nov., isolated from polluted soil.

This paper presents a polyphasic taxonomic study of a Gram-stain-negative bacterium designated GA093T, a soil isolate capable of benzo(α)pyrene degradation. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain GA093T is a member of the genus Flavobacterium, and formed an independent phylogenetic line while clustering with the type strains of Flavobacterium hibernum, Flavobacterium branchiarum and Flavobacterium hydatis. Strain GA093T was facultatively anaerobic, and could grow at 4-33 °C (optimum, 30 °C), at pH 6-11 (optimum, pH 7) and in the presence of 0-2 % (w/v) NaCl (optimum, 0 %). Strain GA093T was capable of producing acid from various carbon sources, which was comparable to other related species of Flavobacterium. The strain contained MK-6 as the only isoprenoid quinone, iso-C15 : 0 as the major cellular fatty acid, phosphatidylethanolamine and phosphatidylinositol as diagnostic polar lipids, and sym-homospermidine as the major polyamine. The chemotaxonomic properties of strain GA093T were consistent with the general properties of Flavobacterium except the presence of phosphatidylinositol, which distinguished it from other related species. The total stretch of the obtained genome of GA093T was 5.05 Mbp, and the DNA G+C content was 34.79 mol%. The genome contained genes potentially related to the degradation of aromatic hydrocarbons. On the basis of the present polyphasic analysis, strain GA093T was found to have properties that distunguished it as representing a novel species of the genus Flavobacterium, for which the name Flavobacterium hydrocarbonoxydans sp. nov. is proposed. The type strain is GA093T (=KCTC 72594T=LMG 31760T).

Flavobacterium dauae sp. nov., isolated from rhizosphere soil of a tomato plant.

A bacterial strain, designated TCH3-2T, was isolated from the rhizosphere of tomato plant grown at Dong-A University Agricultural Experiment Station, Republic of Korea. The strain was Gram-stain-negative, obligate aerobic, orange yellow-coloured, motile by gliding and short rod-shaped. Strain TCH3-2 T only grew on 1/2 tryptic soy agar and Luria-Bertani agar among the media tested, with optimum growth at 28 °C and pH 7. Salt of 1 % NaCl was necessary to support the growth of TCH3-2T. Strain TCH3-2T produced flexirubin-type pigments. The predominant cellular fatty acids were iso-C15 : 0 (55.6 %), iso-C17 : 0 3-OH (17.9 %), summed feature 9 (comprising C16 : 0 10-methyl and/or iso-C17 : 1 ω9c; 10.5 %), iso-C15 : 0 3-OH (4.8 %) and anteiso-C15 : 0 (2.3 %). The major menaquinone was menaquinone-6 and the major polar lipids were phosphatidylethanolamine, five unknown aminolipids and three unknown lipids. Phylogenetic analysis based on 16S rRNA sequences indicated that TCH3-2T was closely related to Flavobacterium ummariense DS-12T (95.16 %), Flavobacterium marinum SW105T (95.14 %) and Flavobacterium viscosus YIM 102796T (94.54 %). The draft genome of TCH3-2T comprised ca. 2.8 Mb with a G+C content of 34.61 mol%. The average nucleotide identity and digital DNA-DNA hybridization values between TCH3-2T and closely related Flavobacterium species showed that it belongs to a distinct species. Furthermore, the results of morphological, physiological and biochemical tests allowed further phenotypic differentiation of TCH3-2T from its closest relatives. Thus, chemotaxonomic characteristics together with phylogenetic affiliation illustrate that TCH3-2T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium dauae sp. nov. (type strain TCH3-2T=KACC 19054T=JCM 34025T) is proposed.

Flavobacterium solisilvae sp. nov. and Flavobacterium silvaticum sp. nov., isolated from forest soil.

Two Gram-stain-negative, yellow-pigmented and strictly aerobic bacteria, designated strains SE-s27T and SE-s28T, were isolated from forest soil. Both strains were non-motile rods that were catalase-positive and oxidase-negative and grew optimally at 25-30 °C, pH 8.0 and with 0 % (w/v) NaCl. Strain SE-s28T produced flexirubin-type pigments, but strain SE-s27T did not produce them. Both strains contained menaquinone-6 as the sole respiratory quinone and phosphatidylethanolamine as a major polar lipid. As the major cellular fatty acids (>10 %), SE-s27T contained iso-C15 : 1 and iso-C15 : 1G, whereas SE-s28T contained iso-C15 : 0 and summed feature 3 comprising C16 : 1ω7c and/or C16 : 1ω6c and/or iso-C15 : 0 2-OH. The DNA G+C contents of strains SE-s27T and SE-s28T were 33.1 and 44.3 mol%, respectively. The results of phylogenetic analysis based on 16S rRNA gene sequences revealed that SE-s27T and SE-s28T formed respective distinct phylogenetic lineages within the genus Flavobacterium. Strains SE-s27T and SE-s28T were most closely related to Flavobacterium macrobrachii an-8T and Flavobacterium piscinae ICH-30T with 98.0 and 94.5 % 16S rRNA gene sequence similarities, respectively. In conclusion, strains SE-s27T and SE-s28T represent novel species of the genus Flavobacterium, for which the names Flavobacterium solisilvae sp. nov. and Flavobacterium silvaticum sp. nov. are proposed. The type strains of F. solisilvae and F. silvaticum are SE-s27T (=KACC 18802T=JCM 31544T) and SE-s28T (=KACC 18803T=JCM 31545T), respectively.

Flavobacterium franklandianum sp. nov. and Flavobacterium gawalongense sp. nov., isolated from glaciers on the Tibetan Plateau.

The genus Flavobacterium (family Flavobacteriaceae) can be found in diverse environments. In this study, seven novel strains were isolated from glaciers in PR China and subjected to taxonomic research. Phylogenetic analyses based on the 16S rRNA gene revealed that the strains belonged to the genus Flavobacterium. None of the seven strains grew at temperatures above 22 °C, indicating that they are psychrophilic. Furthermore, the average nucleotide identity (ANI) values of the seven strains were calculated and indicated that they represented two novel species in Flavobacterium. Strain LB3P56T was most closely related to Flavobacterium soyangense IMCC26223T (97.70 %) and strain GSP16T was most closely related to Flavobacterium sinopsychrotolerans 0533T (98.03 %). The ANI values between the two Flavobacterium strains and their closest relatives were less than 83.47 %, which was much lower than the threshold for species delineation of 95-96 %. Therefore, we propose two novel species, Flavobacterium franklandianum sp. nov. (LB3P56T=CGMCC 1.11934T=NBRC 113651T) and Flavobacterium gawalongense sp. nov. (GSP16T=CGMCC 1.24642T=NBRC 113664T).

Flavobacterium muglaense sp. nov. isolated from internal organs of apparently healthy rainbow trout.

Two yellow-pigmented isolates, F-60T and F-392, were isolated from the internal organs of an apparently healthy rainbow trout (Oncorhynchus mykiss). The strains were identified as members of the genus Flavobacterium based on the results of 16S rRNA gene sequence analysis. Strains F-60T and F-392 had the highest 16S rRNA gene sequence identity level of 97.4 % to the type strain of Flavobacterium crassostreae LPB0076T. A polyphasic taxonomic approach including phenotypic, chemotaxonomic and genomic characterization was employed to ascertain the taxonomic position of the strains within the genus Flavobacterium. Digital DNA-DNA hybridization (dDDH) and average nucleotide identity based on blast (ANIb) values for strains F-60T and F-392 were calculated as 100 %. However, dDDH and ANI analyses between the strains and their close neighbours confirmed that both strains represent a novel species in the genus Flavobacterium. The strains shared the highest dDDH and ANIb levels of 23.3 and 77.9%, respectively, with the type strain of Flavobacterium frigidarium DSM 17623T while those values for F. crassostreae LPB0076T were obtained as 21.4-21.5 % and 76.3 %. The DNA G+C content of the strains was 34.5 mol%. Chemotaxonomic and phylogenomic analyses of these isolates confirmed that both strains are representatives of a novel species for which the name Flavobacterium muglaense sp. nov. is proposed, with F-60T as the type strain (=JCM 34196T=KCTC 82256T).

Flavobacterium difficile sp. nov., isolated from a freshwater waterfall.

A bacterial strain designated KDG-16 T is isolated from a freshwater waterfall in Taiwan and characterized to determine its taxonomic affiliation. Cells of strain KDG-16 T are Gram-stain-negative, strictly aerobic, motile by gliding, rod-shaped and form light yellow colonies. Optimal growth occurs at 20-25 °C, pH 6-7, and with 0% NaCl. Phylogenetic analyses based on 16S rRNA gene sequences and an up-to-date bacterial core gene set reveal that strain KDG-16 T is affiliated with species in the genus Flavobacterium. Analysis of 16S rRNA gene sequences shows that strain KDG-16 T shares the highest similarity with Flavobacterium terrigena DSM 17934 T (97.7%). The average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values between strain KDG-16 T and the closely related Flavobacterium species are below the cut-off values of 95-96, 90 and 70%, respectively, used for species demarcation. Strain KDG-16 T contains iso-C15:0, iso-C15:1 G and iso-C17:0 3-OH as the predominant fatty acids. The polar lipid profile consists of phosphatidylethanolamine, one uncharacterized aminophospholipid, one uncharacterized phospholipid, two uncharacterized aminolipids and two uncharacterized lipids. The major polyamine is homospermidine. The major isoprenoid quinone is MK-6. Genomic DNA G + C content of strain KDG-16 T is 31.6%. Based on the polyphasic taxonomic data obtained, strain KDG-16 T is considered to represent a novel species in the genus Flavobacterium, for which the name Flavobacterium difficile sp. nov. is proposed. The type strain is KDG-16 T (= BCRC 81194 T = LMG 31332 T).

Flavobacterium agrisoli sp. nov., a novel bacterium isolated from soil.

A Gram-stain negative, rod shaped, motile by gliding, yellow-pigmented, aerobic bacterium, designated SE-1-eT, was isolated from a soil sample collected on Dokdo Island, South Korea. The isolate was characterized taxonomically using a polyphasic approach based on the phenotypic and genomic analyses. The phylogenetic analysis based on 16S rRNA gene sequence revealed that strain SE-1-eT belonged to the genus Flavobacterium in the family Flavobacteriaceae and had the highest sequence similarity with Flavobacterium cheongpyeongense IMCC34759T (97.5%), Flavobacterium arsenitoxidans S2-3HT (97.4%), Flavobacterium resistens BD-b365T (97.4%), and Flavobacterium chungangense CJ7T (97.4%). The predominant respiratory quinone of the isolate was found to be MK-6; the main polar lipid was phosphatidylethanolamine; and the major fatty acids were identified as summed feature 3 (C16:1 ω7c/C16:1 ω6c), C15:0 iso, and C16:0. The draft genome of strain SE-1-eT had a length of 3,715,609 bp and a DNA G + C content of 34.8 mol%. The nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values between the novel isolate and F. cheongpyeongense IMCC34759T, F. resistens BD-b365T, and Flavobacterium chungangense CJ7T ranged from 74.9 to 75.3% and from 20.2 to 20.9%, respectively. On the basis of its phenotypic properties, genotypic distinctiveness, and chemotaxonomic features, strain SE-1-eT represents a novel species in the genus Flavobacterium, for which the name Flavobacterium agrisoli sp. nov. is proposed. The type strain is SE-1-eT (= KCTC 82352 T = JCM 34302 T).

Flavobacterium inviolabile sp. nov. isolated from stream water.

A Gram-stain-negative, rod-shaped, aerobic and non-motile bacterium, designated P2-65T, was isolated from Moonsan stream water in the Republic of Korea. The temperature, NaCl concentration and pH ranges for growth of strain P2-65T were 10-37 °C, 0.0-3.0% (w/v) and 6.5-8.5 with optimum growth at 25-30 °C, 0.0-1.0% and 7.0-7.5, respectively. Comparison of 16S rRNA gene sequence showed that strain P2-65T was closely related to Flavobacterium cauense (95.4%) and Flavobacterium cheniae (95.3%). The major fatty acids were iso-C15:0, iso C17:0 3-OH, summed feature 3 (C16:1ω7c and/or C16:1ω6c), summed feature 9 (iso-C17:1 ω9c and/or 10-methyl C16:0) and iso-C15:0 3-OH. The predominant respiratory quinone was menaquinone-6 (MK-6). The major polar lipids detected in the strain were phosphatidylethanolamine, one aminophospholipid, one unidentified aminolipid and one unidentified polar lipid. The G + C content of the genomic DNA was 39.7%. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) values for strain P2-65T with closely related Flavobacterium species were below 74.8% and 20%, respectively. Based on polyphasic features, strain P2-65T is considered to represent a novel species of the genus Flavobacterium, for which the name Flavobacterium inviolabile sp. nov. is proposed. The type strain is P2-65T (= KCTC 62055T = NBRC 112953T).

Flavobacterium turcicum sp. nov. and Flavobacterium kayseriense sp. nov. isolated from farmed rainbow trout in Turkey.

During a study on culturable microorganisms from fish farms, four yellow-pigmented gram negative, rod shaped isolates, F-47T, F-339T, F-380 and F-400, were recovered from rainbow trout samples exhibiting clinical signs. Based on 16S rRNA gene sequence analysis, the strains were identified as members of the genus Flavobacterium. Strains F-47T and F-380 shared the highest 16S rRNA gene sequence identity level of 97.6% with the type strain of Flavobacterium frigoris DSM 15719T while strains F-339T and F-400 shared the highest identity level of 97.6% with the type strain of F. caseinilyticum AT-3-2T. A polyphasic taxonomic approach including phenotypic and genomic characterization as well as whole-cell MALDI-TOF mass spectrometry analyses was employed to ascertain the taxonomic position of the strains within the genus Flavobacterium. Digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) analyses between strains F-47T, F-339T and their close neighbours F. frigoris DSM 15719T and F. caseinilyticum AT-3-2T, respectively, confirmed that both strains represent novel species in the genus Flavobacterium. The DNA G+C contents of the strains F-47T and F-339T are 34.3% and 35.3%, respectively. It can be concluded on the basis of polyphasic characterization as well as pairwise genome comparisons that the strains F-47T and F-339T represent two novel species within the genus Flavobacterium, for which Flavobacterium kayseriense sp. nov. F-47T (=JCM 34195T=KCTC 82255T) and Flavobacterium turcicum sp. nov. F-339T (=JCM 34202T=KCTC 82262T) are proposed, respectively.