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Background: The salmonid pathogen Flavobacterium psychrophilum poses a significant economic threat to global aquaculture, yet our understanding of its genetic and phenotypic diversity remains incomplete across much of its geographic range. In this study, we characterise the genetic and phenotypic diversity of 70 isolates collected from rainbow trout (Oncorhynchus mykiss) and brown trout (Salmo trutta m. fario) from fish farms in the Czech Republic between 2012 and 2019 to compare their genomic content with all draft or complete genomes present in the NCBI database (n = 187)., Results: The Czech isolates underwent comprehensive evaluation, including multiplex PCR-based serotyping, genetic analysis, antimicrobial resistance testing, and assessment of selected virulence factors. Multiplex PCR serotyping revealed 43 isolates as Type 1, 23 as Type 2, with sporadic cases of Types 3 and 4. Multi-locus sequence typing unveiled 12 sequence types (ST), including seven newly described ones. Notably, 24 isolates were identified as ST329, a novel sequence type, while 22 were classified as the globally-distributed ST2. Phylogenetic analysis demonstrated clonal distribution of ST329 in the Czech Republic, with these isolates lacking a phage sequence in their genomes. Antimicrobial susceptibility testing revealed a high proportion of isolates classified as non-wild type with reduced susceptibility to oxolinic acid, oxytetracycline, flumequine, and enrofloxacin, while most isolates were classified as wild type for florfenicol, sulfamethoxazole-trimethoprim, and erythromycin. However, 31 isolates classified as wild type for florfenicol exhibited minimum inhibitory concentrations at the susceptibility breakpoint., Conclusion: The prevalence of the Czech F. psychrophilum serotypes has evolved over time, likely influenced by the introduction of new isolates through international trade. Thus, it is crucial to monitor F. psychrophilum clones within and across countries using advanced methods such as MLST, serotyping, and genome sequencing. Given the open nature of the pan-genome, further sequencing of strains promises exciting discoveries in F. psychrophilum genomics., (© 2024. The Author(s).)

Four Gram-stain-positive and two Gram-stain-negative bacterial strains, designated as W4 T , FW7 T , TW48 T , UW52 T , PT-3 T , and RJY3 T , were isolated from soil samples collected from the Republic of Korea. The 16S rRNA gene sequence analysis showed that strains W4 T and FW7 T belonged to the genus Microbacterium , strains TW48 T and UW52 T were affiliated to the genus Paenibacillus, strain PT-3 T was related to the genus Flavobacterium , and strain RJY3 T was associated with the genus Aquabacterium . The closest phylogenetic taxa to W4 T , FW7 T , TW48 T , UW52 T , PT-3 T , and RJY3 T were Microbacterium bovistercoris NEAU-LLE T (97.7 %), Microbacterium protaetiae DFW100M-13 T (97.9 %), Paenibacillus auburnensis JJ-7 T (99.6 %), Paenibacillus allorhizosphaerae JJ-447 T (95.7 %), Flavobacterium buctense T7 T (97.1 %), and Aquabacterium terrae S2 T (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 W4 T , FW7 T TW48 T , and UW52 T was antiso-C 15 : 0 . Similarly, strain PT-3 T revealed iso-C 15 : 0 , iso-C 15 : 1 G, iso-C 17 : 0 3-OH, and iso-C 15 : 0 3-OH as its principal fatty acids. On the other hand, RJY3 T exhibited summed feature 3 (C 16 : 1  ω 7 c and/or C 16 : 1  ω 6 c ), C 16 : 0 , summed feature 8 (C 18 : 1  ω 7 c and/or C 18 : 1  ω 6 c ), and C 12 : 0 as its predominant fatty acids. Overall, the polyphasic taxonomic data indicated that strains W4 T , FW7 T , TW48 T , UW52 T , PT-3 T , and RJY3 T represent novel species within the genera Microbacterium , Paenibacillus , Flavobacterium , and Aquabacterium . Accordingly, we propose the names Microbacterium humicola sp. nov., with the type strain W4 T (=KCTC 49888 T =NBRC 116001 T ), Microbacterium terrisoli sp. nov., with the type strain FW7 T (=KCTC 49859 T =NBRC 116000 T ), Paenibacillus pedocola sp. nov., with the type strain TW48 T (=KCTC 43470 T =NBRC 116017 T ), Paenibacillus silviterrae sp. nov., with the type strain UW52 T (=KCTC 43477 T =NBRC 116018 T ), Flavobacterium terrisoli sp. nov., with the type strain PT-3 T (=KCTC 92106 T =NBRC 116012 T ), and Aquabacterium humicola sp. nov., with the type strain RJY3 T (=KCTC 92105 T =NBRC 115831 T ).

The generic term "Gill disease" refers to a wide range of disorders that affect the gills and severely impact salmonid aquaculture systems worldwide. In rainbow trout freshwater aquaculture, various etiological agents causing gill diseases have been described, particularly Flavobacterium and Amoeba species, but research studies suggest a more complex and multifactorial aetiology. Here, a cohort of rainbow trout affected by gill disease is monitored both through standard laboratory techniques and 16S rRNA Next-Generation Sequencing (NGS) analysis during a natural disease outbreak and subsequent antibiotic treatment with Oxytetracycline. NGS results show a clear clustering of the samples between pre- and post-treatment based on the microbial community of the gills. Interestingly, the three main pathogenic bacteria species in rainbow trout (Yersinia ruckeri, Flavobacterium psychrophilum, and Flavobacterium branchiophilum) appear to be weak descriptors of the diversity between pre-treatment and post-treatment groups. In this study, the dynamics of the gill microbiome during the outbreak and subsequent treatment are far more complex than previously reported in the literature, and environmental factors seem of the utmost importance in determining gill disease. These findings present a potential novel perspective on the diagnosis and management of gill diseases, showing the limitations of conventional laboratory methodologies in elucidating the complexity of this disease in rainbow trout. To the authors' knowledge, this work is the first to describe the microbiome of rainbow trout gills during a natural outbreak and subsequent antibiotic treatment. The results of this study suggest that NGS can play a critical role in the analysis and comprehension of gill pathology. Using NGS in future research is highly recommended to gain deeper insights into such diseases correlating gill's microbiome with other possible cofactors and establish strong prevention guidelines., (© 2024. The Author(s).)

A Gram-strain-negative, aerobic, yellow-colored, non-motile, and rod-shaped bacterial strain, designated IMCC34852 T , 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 IMCC34852 T belonged to the genus Flavobacterium and that the strain was most closely related to F. cheonhonense ARSA-15  T (97.6%), F. buctense T7 T (96.7%), F. silvisoli RD-2-33  T (96.1%), and F. paronense KNUS1 T (96.1%). The whole-genome sequence of strain IMCC34852 T 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 IMCC34852 T 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 IMCC34852 T was menaquinone-6 (MK-6) and the predominant cellular fatty acids were iso-C 15:0 (32.6%), iso-C 16:0 (11.7%), iso-C 15:1 G (10.3%), and iso-C 14:0 (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 IMCC34852 T 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 IMCC34852 T (= KACC 23133  T  = KCTC 82066  T  = NBRC 114419  T )., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

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., Competing Interests: The authors declare no conflict of interest.

Two strains, designated as SYSU M80004 T and SYSU M80005 T , were isolated from water sampled in the Pearl River Estuary, Guangzhou, Guangdong, PR China. The strains were Gram-stain-negative and aerobic. Strain SYSU M80004 T 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 M80005 T 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. C 16 : 0 and iso-C 15 : 0 were identified as the major fatty acids (>10 %) of strain SYSU M80004 T while strain SYSU M80005 T contained iso-C 15 : 0 and iso-C 17 : 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 M80004 T =CDMCC 1.4522 T =KCTC 102268 T ) and Flavobacterium maritimum sp. nov. (type strain=SYSU M80005 T =CGMCC 1.4523 T = KCTC 102269 T ) are proposed.

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 , F126 T 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., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier GmbH. All rights reserved.)

Strain T-12 T , 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-12 T belonged to the genus Flavobacterium , showing close relationships to Flavobacterium bernardetii F-372 T (99.48 %) and Flavobacterium terrigena DS-20 T (98.50 %). The genome size of strain T-12 T was 3.28 Mb, with a G+C content of 31.1 mol%. Genome comparisons aligned strain T-12 T with Flavobacterium bernardetii F-372 T (GCA&#95;011305415) and Flavobacterium terrigena DSM 17934 T (GCA&#95;900108955). The highest digital DNA-DNA hybridization (dDDH) values were 42.6 % with F. bernardetii F-372 T (GCA&#95;011305415) and 33.9 % with F. terrigena DSM 17934 T (GCA&#95;900108955). Pairwise average nucleotide identity (ANI) calculations were below the species cutoff, with the best results with F. bernardetii F-372 T being: ANIb, 90.33 %; ANIm, 91.85 %; and TETRA, 0.997 %. These dDDH and ANI results confirm that strain T-12 T represents a new species. The major fatty acids were iso-C 15 : 0 and C 15 : 1 ω6 с . 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-12 T (=CECT 30410 T =RGM 3222 T ) as representing a novel species of Flavobacterium , for which the name Flavobacterium facile sp. nov. is proposed.

A Gram-stain-negative, aerobic, non-spore-forming, nonmotile, rod-shaped, and yellow-pigmented bacterium, designated strain JXAS1 T , 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 6368 T (98.61 %). The genome size of strain JXAS1 T 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 JXAS1 T 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-C 15 : 0 , summed feature 3 (C 16 : 1  ω 7 c and/or C 16 : 1  ω 6 c ), C 15 : 0 , iso-C 17 : 0 3OH, iso-C 15 : 0 3OH, and summed feature 9 (iso-C 17 : 1  ω 9 c and/or 10-methyl C 16 : 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 JXAS1 T (=GDMCC 1.1378 T =KCTC 62719 T ).

Two yellow-coloured strains, F-29 T and F-340 T , 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-29 T is closely related to Flavobacterium granuli (dDDH 39.3 % and ANI 89.4 %), while strain F-340 T 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-C 15 : 0 and anteiso-C 15 : 0 . The genome size of the strains was 3.5 Mb, while G+C contents were 35.3 mol% for strain F-29 T and 33.4 mol% for strain F-340 T . 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-29 T (JCM 34193 T =KCTC 82253 T ) and F-340 T (JCM 34203 T =KCTC 82263 T ) as the type strains, respectively.

A Gram-stain-negative, facultatively anaerobic, motile by gliding, rod-shaped, oxidase- and catalase-positive bacterial strain, designated BB8 T , 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 BB8 T 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 BB8 T formed a lineage within the genus Flavobacterium and was most closely related to Flavobacterium artemisiae SYP-B1015 T (96.9 % 16S rRNA gene sequence similarity) and Flavobacterium ustbae T13 T (96.8%). The complete genome sequence of strain BB8 T was 5 513 159 bp long with a G+C content of 34.1 mol%. The major fatty acids (>10 %) of strain BB8 T were iso-C 15 : 0 (21 %), summed feature 3 (comprising C 16 : 1  ω7 c and/or C 16 : 1  ω6 c , 20.3%) and iso-C 16 : 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 BB8 T 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 BB8 T (=KCTC 82167 T =CCTCC AB 2020070 T ).

A Gram-stain-negative, non-motile, strictly aerobic, rod-shaped bacterium, with one polar flagellum and named D11R37 T , was isolated from coral culture seawater of Acropora digitifera . Strain D11R37 T 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 D11R37 T formed a lineage within the genus Flavobacterium , and it was distinct from the most closely related species Flavobacterium suzhouense XIN-1 T and Flavobacterium suaedae G16-7 T 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-C 15 : 0 (18.0%), iso-C 17 : 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 D11R37 T 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 D11R37 T (=KCTC 82968 T =MCCC 1K06440 T ).

Flavobacterium psychrophilum, the etiological agent of bacterial coldwater disease (BCWD) and rainbow trout fry syndrome, causes great economic losses in salmonid aquaculture worldwide. Recent molecular studies have uncovered important epidemiological and ecological aspects of this pathogen; however, such data are lacking for F. psychrophilum populations affecting aquaculture in China. Herein, F. psychrophilum phenotype, genotype, and virulence were characterized for isolates recovered from epizootics in multiple salmonid aquaculture facilities across China. Thirty-one F. psychrophilum isolates, originating from four provinces and three host fish species, were predominantly homogeneous biochemically but represented 5 sequence types (STs) according to multilocus sequence typing (MLST) that belonged to clonal complex CC-ST10 or 3 newly recognized singleton STs. PCR-based serotyping classified 19 and 12 F. psychrophilum isolates into molecular serotypes 1 and 0, respectively, showing an obvious relationship with host species. Antimicrobial susceptibility analysis via broth microdilution revealed reduced susceptibility to enrofloxacin, flumequine, and oxolinic acid, moderate susceptibility to gentamicin, erythromycin, and florfenicol, and variable susceptibility to ampicillin and oxytetracycline. In vivo challenge experiments confirmed the ability of two representative Chinese F. psychrophilum isolates to induce typical signs of BCWD and mortality in 1-year-old rainbow trout (Oncorhynchus mykiss). Findings collectively demonstrate (i) that BCWD outbreaks in China studied thus far are caused by F. psychrophilum lineages that are common on other continents (e.g., CC-ST10) and others that have not been reported elsewhere (e.g., ST355, ST356, ST357), (ii) that F. psychrophilum molecular serotypes distinguish isolates from different host fish species, even within STs, and (iii) reduced F. psychrophilum antimicrobial susceptibility against compounds used for BCWD control in China. IMPORTANCE Flavobacterium psychrophilum causes substantial economic losses in salmonid aquaculture worldwide. Although this bacterium is also believed to be a disease source in China, published reports of its presence do not yet exist. Herein, F. psychrophilum was linked to multiple disease outbreaks in several salmonid aquaculture facilities within four Chinese provinces, and polyphasic characterization revealed that most isolates were genetically distinct from strains recovered on other continents. Analyses further revealed the predominating molecular serotypes, antimicrobial susceptibility profiles, and pathogenic potential of two representative recovered isolates. Collectively, the results presented here provide important data on the epidemiology and disease ecology of F. psychrophilum in China and pave the way for targeted prevention and control methods to be pursued in the future.

This paper presents a polyphasic taxonomic study of a Gram-stain-negative bacterium designated GA093 T , a soil isolate capable of benzo(α)pyrene degradation. Phylogenetic analysis based on the 16S rRNA gene sequence indicated that strain GA093 T 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 GA093 T 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 GA093 T 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-C 15 : 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 GA093 T 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 GA093 T 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 GA093 T 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 GA093 T (=KCTC 72594 T =LMG 31760 T ).

A bacterial strain, designated TCH3-2 T , 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 TCH3-2 T . Strain TCH3-2 T produced flexirubin-type pigments. The predominant cellular fatty acids were iso-C 15 : 0 (55.6 %), iso-C 17 : 0 3-OH (17.9 %), summed feature 9 (comprising C 16 : 0 10-methyl and/or iso-C 17 : 1  ω 9 c ; 10.5 %), iso-C 15 : 0 3-OH (4.8 %) and anteiso-C 15 : 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 TCH3-2 T was closely related to Flavobacterium ummariense DS-12 T (95.16 %), Flavobacterium marinum SW105 T (95.14 %) and Flavobacterium viscosus YIM 102796 T (94.54 %). The draft genome of TCH3-2 T 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 TCH3-2 T 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 TCH3-2 T from its closest relatives. Thus, chemotaxonomic characteristics together with phylogenetic affiliation illustrate that TCH3-2 T represents a novel species of the genus Flavobacterium , for which the name Flavobacterium dauae sp. nov. (type strain TCH3-2 T =KACC 19054 T =JCM 34025 T ) is proposed.

Two yellow-pigmented isolates, F-60 T 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-60 T and F-392 had the highest 16S rRNA gene sequence identity level of 97.4 % to the type strain of Flavobacterium crassostreae LPB0076 T . 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-60 T 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 17623 T while those values for F. crassostreae LPB0076 T 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-60 T as the type strain (=JCM 34196 T =KCTC 82256 T ).

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 LB3P56 T was most closely related to Flavobacterium soyangense IMCC26223 T (97.70 %) and strain GSP16 T was most closely related to Flavobacterium sinopsychrotolerans 0533 T (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. (LB3P56 T =CGMCC 1.11934 T =NBRC 113651 T ) and Flavobacterium gawalongense sp. nov. (GSP16 T =CGMCC 1.24642 T =NBRC 113664 T ).

Two Gram-stain-negative, yellow-pigmented and strictly aerobic bacteria, designated strains SE-s27 T and SE-s28 T , 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-s28 T produced flexirubin-type pigments, but strain SE-s27 T 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-s27 T contained iso-C 15 : 1 and iso-C 15 : 1 G, whereas SE-s28 T contained iso-C 15 : 0 and summed feature 3 comprising C 16 : 1 ω 7 c and/or C 16 : 1 ω 6 c and/or iso-C 15 : 0 2-OH. The DNA G+C contents of strains SE-s27 T and SE-s28 T were 33.1 and 44.3 mol%, respectively. The results of phylogenetic analysis based on 16S rRNA gene sequences revealed that SE-s27 T and SE-s28 T formed respective distinct phylogenetic lineages within the genus Flavobacterium . Strains SE-s27 T and SE-s28 T were most closely related to Flavobacterium macrobrachii an-8 T and Flavobacterium piscinae ICH-30 T with 98.0 and 94.5 % 16S rRNA gene sequence similarities, respectively. In conclusion, strains SE-s27 T and SE-s28 T 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-s27 T (=KACC 18802 T =JCM 31544 T ) and SE-s28 T (=KACC 18803 T =JCM 31545 T ), respectively.

During a study on culturable microorganisms from fish farms, four yellow-pigmented gram negative, rod shaped isolates, F-47 T , F-339 T , 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-47 T and F-380 shared the highest 16S rRNA gene sequence identity level of 97.6% with the type strain of Flavobacterium frigoris DSM 15719 T while strains F-339 T and F-400 shared the highest identity level of 97.6% with the type strain of F. caseinilyticum AT-3-2 T . 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-47 T , F-339 T and their close neighbours F. frigoris DSM 15719 T and F. caseinilyticum AT-3-2 T , respectively, confirmed that both strains represent novel species in the genus Flavobacterium. The DNA G+C contents of the strains F-47 T and F-339 T 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-47 T and F-339 T represent two novel species within the genus Flavobacterium, for which Flavobacterium kayseriense sp. nov. F-47 T (=JCM 34195 T =KCTC 82255 T ) and Flavobacterium turcicum sp. nov. F-339 T (=JCM 34202 T =KCTC 82262 T ) are proposed, respectively., (Copyright © 2021 Elsevier GmbH. All rights reserved.)

Modern aquaculture systems are designed for intensive rearing of fish or other species. Both land-based and offshore systems typically contain high loads of biomass and the water quality in these systems is of paramount importance for fish health and production. Microorganisms play a crucial role in removal of organic matter and nitrogen-recycling, production of toxic hydrogen sulfide (H 2 S), and can affect fish health directly if pathogenic for fish or exerting probiotic properties. Methods currently used in aquaculture for monitoring certain bacteria species numbers still have typically low precision, specificity, sensitivity and are time-consuming. Here, we demonstrate the use of Digital PCR as a powerful tool for absolute quantification of sulfate-reducing bacteria (SRB) and major pathogens in salmon aquaculture, Moritella viscosa, Yersinia ruckeri and Flavobacterium psychrophilum. In addition, an assay for quantification of Listeria monocytogenes, which is a human pathogen bacterium and relevant target associated with salmonid cultivation in recirculating systems and salmon processing, has been assessed. Sudden mass mortality incidents caused by H 2 S produced by SRB have become of major concern in closed aquaculture systems. An ultra-sensitive assay for quantification of SRB has been established using Desulfovibrio desulfuricans as reference strain. The use of TaqMan® probe technology allowed for the development of multi-plex assays capable of simultaneous quantification of these aquaculture priority bacteria. In single-plex assays, limit of detection was found to be at around 20 fg DNA for M. viscosa, Y. ruckeri and F. psychrophilum, and as low as 2 fg DNA for L. monocytogenes and D. desulfuricans., (Copyright © 2021 The Author(s). Published by Elsevier B.V. All rights reserved.)

Flavobacterium psychrophilum affects salmonid health worldwide and causes economic losses. The genetic diversity of the pathogen must be considered to develop control methods. However, previous studies have reported both high and low levels of genetic diversity. The present longitudinal study aimed at assessing the genetic diversity of F. psychrophilum at a small temporal and geographic scale. Four farms located on the same watershed in France were studied. Rainbow trout ( Oncorhynchus mykiss ) batches were monitored, and apparently healthy individuals were sampled over 1 year. A total of 288 isolates were recovered from fish organs (gills and spleen) and eggs. Pulsed field gel electrophoresis revealed high genetic diversity. Multilocus sequence typing performed on a selection of 31 isolates provided congruent results, as follows: 18 sequence types (STs) were found, of which 13 were novel. The mean gene diversity ( H  = 0.8413) was much higher than that previously reported for this host species, although the sampling was restricted to a single watershed and 1 year. Seven isolates out of 31 were assigned to clonal complex ST10 (CC-ST10), which is the predominant clonal complex in the main salmonid production areas. A split decomposition tree reflected a panmictic population. This finding is important for aquaculture veterinarians in their diagnostic procedure, as the choice of adequate antibiotic treatment is conditioned by the correct identification of the causative agent. Furthermore, this study expands our knowledge on genetic diversity required for the development of an effective vaccine against F. psychrophilum IMPORTANCE The bacterium Flavobacterium psychrophilum is a serious pathogen in many fish species, especially salmonids, that is responsible for considerable economic losses worldwide. In order to treat infections and to develop vaccines, the genetic diversity of this bacterium needs to be known. We assessed the genetic diversity of F. psychrophilum isolates from apparently healthy rainbow trout raised in several fish farms in the same watershed in France. Two different genotyping methods revealed high diversity. The majority of isolates were unrelated to clonal complex sequence type 10 (CC-ST10), the clonal complex that is predominant worldwide and associated with disease in rainbow trout. In addition, we found 13 novel sequence types. These results suggest that a diverse subpopulation of F. psychrophilum may be harbored by rainbow trout., (Copyright © 2021 American Society for Microbiology.)

Genome sequence analysis of two strains collected in Canada at the end of the 1970s and deposited in 1998 at the Collection de l'Institut Pasteur has led to the taxonomic description of a novel fish-associated species in the genus Flavobacterium . Both strains, CIP 105534 T and CIP 105535, were yellow-pigmented, Gram-stain-negative, non-spore-forming rod-shaped bacteria that exhibited gliding motility. They grew aerobically in a temperature range from 5 to 30 °C with optimal growth at 25 °C on trypticase soy or Reasoner's 2A agar but they did not grow on marine agar. Their major fatty acid profiles were similar, consisting of iso-C 15 : 0 , C 16 : 1  ω7 c and/or iso-C 15 : 0 2-OH (shown as summed feature 3), C 16 : 0 3-OH, iso-C 17 : 0 3-OH and C 16 : 0 . The major polyamine was sym -homospermidine. Phosphatidylethanolamine and, most notably, ornithine-containing lipid OL2 and unidentified aminophospholipid APL1 were major polar lipids. A yellow pigment spot was visible after chromatographic analysis. The predominant respiratory quinone was MK-6. The G+C content of the two genomes was 34 mol% and their size was around 5.8 Mb. Comparison of the 16S rRNA gene sequences with those of the closely related type strains showed high levels of relatedness with Flavobacterium collinsii and Flavobacterium pectinovorum . All average nucleotide identity (ANI) and digital DNA-DNA hybridization values estimated against publicly available Flavobacterium genome assemblies were lower than 90 and 30 %, respectively. Phylogenetic, phenotypic and chemotaxonomic data indicated that the two strains represent a novel species of the genus Flavobacterium , for which the name Flavobacterium bizetiae sp. nov. is proposed. The type strain is CIP 105534 T (=LMG 1342 T ). The unique ability of F. bizetiae to use melibiose as a sole source of carbon could provide a simple phenotypic test to discriminate F. bizetiae from its closest relatives.

A Gram-staining-negative non endospore-forming strain, T13(2019) T was isolated from water samples from Atlantic salmon ( Salmo salar ) fry culture in Chile and studied in detail for its taxonomic position. The isolate shared highest 16S rRNA gene sequence similarities with the type strains of Flavobacterium chungangense (98.44 %) followed by Flavobacterium tructae and Flavobacterium spartansii (both 98.22 %). Menaquinone MK-6 was the predominant respiratory quinone in T13(2019) T . Major polar lipids were phosphatidylethanolamine, an ornithine lipid and the unidentified polar lipids L1, L3 and L4 lacking a functional group. The major polyamine was sym -homospermidine. The fatty acid profile contained major amounts of iso-C 15 : 0 , iso-C 15 : 0 3-OH, iso-C 17 : 0 3-OH, C 15 : 0 , summed feature 3 (C 16 : 1  ω7 c and/or iso-C 15 : 0 2-OH) and various hydroxylated fatty acids in smaller amounts, among them iso-C 16 : 0 3-OH, and C 15 : 0 3-OH, which supported the grouping of the isolate into the genus Flavobacterium . Physiological/biochemical characterisation and ANI calculations with the type strains of the most closely related species allowed a clear phenotypic and genotypic differentiation. In addition it became obvious, that the type strains of F. tructae and F. spartansii showed 100 % 16S rRNA gene sequence similarities and ANI values of 97.21%/ 97.59 % and DDH values of 80.40 % [77.5 and 83%]. These data indicate that F. tructae and F. spartansii belong to the same species and it is proposed that F. spartansii is a later heterotypic synonym of F. tructae . For strain T13(2019) T (=CIP 111411 T =LMG 30298 T =CCM 8798 T ) a new species with the name Flavobacterium salmonis sp. nov. is proposed.

Three bacterial strains, namely HYN0069 T , HYN0085 T and HYN0086 T , were isolated from freshwater samples taken from the Namhangan River system in Korea. 16S rRNA gene sequence similarities and phylogenetic tree topologies indicated that the three strains belonged to the genera Gemmobacter , Runella and Flavobacterium by showing the highest sequence similarities with Gemmobacter straminiformis (98.4 %), Runella aurantiaca (98.3 %) and Flavobacterium chungangense (98.1 %). No bacterial species with validly published names showed 98.7 % or higher sequence similarity with the novel isolates. The average nucleotide identities between the genome sequences of the three new isolates and the three closest neighbours were 80.2-92.0 %, all below the threshold for bacterial species delineation (95-96 %). Many biochemical and physiological features also discriminated the isolates from previously known species of the genera Gemmobacter , Runella and Flavobacterium . Based on the phylogenetic and phenotypic data presented in this study, we suggest three novel species with the following names: Gemmobacter aquarius sp. nov. (type strain HYN0069 T =KACC 19488 T =NBRC 113115 T ), Runella rosea sp. nov. (type strain HYN0085 T =KACC 19490 T =NBRC 113116 T ) and Flavobacterium fluviale sp. nov. (type strain HYN0086 T =KACC 19489 T =NBRC 113117 T ).

A Gram-staining-negative non endospore-forming strain, PXU-55 T , was isolated from the rhizosphere of the switchgrass Panicum virgatum and studied in detail to determine its taxonomic position. The results of 16S rRNA gene sequence analysis indicated that the isolate represented a member of the genus Flavobacterium . The isolate shared highest 16S rRNA gene sequence similarities with the type strains of Flavobacterium chungangense (98.78 %) and Flavobacterium chilense (98.64 %). The average nucleotide identity (ANI) and in silico DNA-DNA hybridization (isDDH) values between the PXU-55 T genome assembly and the ones of the most closely related type strains of species of the genus Flavobacterium were 87.3 and 31.9% ( Flavobacterium defluvii ), and 86.1 and 29.9% ( Flavobacterium johnsoniae ). Menaquinone MK-6 was the major respiratory quinone. As major polar lipids, phosphatidylethanolamine, an ornithine lipid and the unidentified polar lipids L2, L3 and L4 lacking a functional group were found. Moderate to minor amounts of another ornithine lipid, the unidentified lipid L1 and a glycolipid were present, as well. The major polyamine is sym -homospermidine. The fatty acid profiles contained major amounts of iso-C 15:0 , iso-C 15:0 3-OH, iso-C 17:0 3-OH, C 15:0 , summed feature 3 (C 16:1 ω7 c and/or iso-C 15:0 2-OH) and various hydroxylated fatty acids in smaller amounts, among them iso C 16:0 3-OH, C 16:0 3-OH and C 15:0 3-OH, which supported the classification of the isolate as a member of the genus Flavobacterium . Physiological and biochemical characterisation and ANI calculations with the type strains of the most closely related species allowed a clear phenotypic and genotypic differentiation of the strain. For this reason, we propose that strain PXU-55 T (=CIP 111646 T =CCM 8914 T ) represents a novel species with the name Flavobacterium panici sp. nov.

Nine plant-associated bacterial strains designated as L1I52 T , NRK F1, NRK F15, NRK F16, NRK F41, NRK F42, NRK F47, NRK F49, and NRK F50 originating from the roots and rhizosphere region of a coastal saline tolerant pokkali rice were taxonomically characterized in this study. Genomic fingerprinting using Enterobacterial Repetitive Intergenic Consensus (ERIC) primers discriminated the nine strains based on the DNA fingerprint patterns indicating that they were not clonal in origin. Phylogenetic analysis using 16S rRNA and other five housekeeping genes (gyrB, glyA, atpA, dnaK and murG) revealed that the novel strains constituted a single novel species within the genus Flavobacterium. In all tree construction methods, the novel strains formed a distinct phylogenetic branch, with Flavobacterium daejeonense GH1-10 T , F. sufflavum BBQ-12 T , and F. glycines Gm-149 T as their nearest phylogenetic neighbours. However, average nucleotide identity (ANI), average amino acid identity (AAI) and digital DNA-DNA hybridization (dDDH) comparison between the draft genomes of L1I52 T (representative isolate) and it's nearest phylogenetic neighbours were well below the proposed threshold values (<95 % and <70 %) used for species discrimination. Thus, based on the phenotypic, genotypic and chemotaxonomic data obtained in this study, we describe a novel Flavobacterium species for which we propose the name Flavobacterium pokkalii sp.nov., with strain L1I52 T (=MTCC 12454 T =KCTC 42429 T ) as the type strain. In addition, L1I52 T is a potential plant growth promoting rhizobacteria as they can promote pokkali rice growth and we identified several plant associated gene features in the genome of L1I52 T that are potentially involved in plant microbe interactions., (Copyright © 2020 Elsevier GmbH. All rights reserved.)

A novel bacterium designated SSM4.2 T was isolated from seaweed of Gouqi Island, which is the center of the Zhoushan fishing ground in the East China Sea. Strain SSM4.2 T was Gram-stain-negative, bright yellow-pigmented, short rod-shaped, non-flagellated, non-spore forming, aerobic and motile by gliding. Growth was observed at 4-37 °C (optimum 25-30 °C), pH 6.0-8.0 (optimum pH 7.0) and 0-2.0% (w/v) NaCl (optimum 0%) concentration. The strain was catalase- and oxidase-positive. Menaquinone-6 (MK-6) was found as the sole respiratory quinone and zeaxanthin as the main carotenoid pigment. The predominant fatty acids (≥ 10%) were iso-C 15:0 , iso-C 15:1 G, iso-C 17:0 3-OH and summed feature 3 (C 16:1 ω7c /C 16:1 ω6c). The major polar lipid was phosphatidylethanolamine (PE). The genome size was 5.7 Mbp. The DNA G + C content was 34.1 mol%. 16S rRNA gene sequence revealed that strain SSM4.2 T belongs to the genus Flavobacterium and shares high-sequence similarity with F. limi KACC 18851 T (98.1%), F. hydrophilum KACC 19591 T (97.6%), F. defluvii KCTC 12612 T (97.1%), F. cheongpyeongense KACC 19592 T (97.0%) and F. fluviatile KCTC 52446 T (96.9%). Strain SSM4.2 T had 73.2-84.6% average nucleotide identity and 19.1-29.4% digital DNA-DNA hybridization values with its closest type strains. Based on its phenotypic, chemotaxonomic, phylogenetic and genomic features, strain SSM4.2 T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium ajazii sp. nov. is proposed. The type strain is SSM4.2 T (= KCTC 72807 T  = MCCC 1K04370 T ).

We report the development of ddPCR assays for single and simultaneous detection of the bacterial pathogens Flavobacterium psychrophilum and Yersinia ruckeri in water from land-based recirculation aquaculture systems (RAS), producing Atlantic salmon (Salmo salar) smolt. The method was tested and verified for use in water analyses from RAS production sites, and proved to be specific and with sensitivity 0.0011 ng DNA for F. psychrophilum and 1.24 ng for Y. ruckeri. These bacteria are important fish pathogens that have caused reoccurring salmonid infection disease in RAS. Monitoring pathogen levels in water samples could be a useful alternative surveillance strategy to evaluate operational risk assessment connected to stress factors. Water quality is essential for fish health and growth in RAS production in general, and high or increasing levels of these pathogens in the RAS water may generate an early indication of unfavourable conditions in the RAS environment, and give directions to operational actions. This approach may reduce fish mortality, reduce production loss, and offer more effective and targeted preventive measures within RAS production., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 The Author(s). Published by Elsevier B.V. All rights reserved.)

Chile is currently the second largest producer of farmed salmon worldwide, but Flavobacterium psychrophilum, as one of the most detrimental pathogens, is responsible for major losses during the freshwater culturing step in salmonid fish farms. An antigenic study conducted 10 years ago reported four serological groups using 20 F. psychrophilum Chilean strains. To reduce disease outbreaks and to develop vaccine candidates, antigenic knowledge needs to be regularly updated using a significant number of additional recent F. psychrophilum isolates. The present study aimed at investigating the serological diversity of 118 F. psychrophilum isolates collected between 2006 and 2018 from farmed Atlantic salmon (Salmo salar), rainbow trout (Oncorhynchus mykiss) and coho salmon (Oncorhynchus kisutch). The current study supports an expansion of the known antigenic groups in Chile from 4 to 14. However, the use of the slide-agglutination technique for serotyping is costly, is labour-intensive and requires significant technical expertise. Addressing these points, the mPCR-based procedure was a very useful tool for serotyping the collected Chilean F. psychrophilum isolates. This technique revealed the presence of diverse mPCR serotypes (i.e. types 0, 1, 2 and 4). Therefore, mPCR should be employed to select the bacterial strain(s) for vaccine development and to conduct follow-up, selective breeding or epidemiological surveillance in Chilean fish farms. Given the presented findings, changes to Chilean fish-farming practices are vital for ensuring the continued productivity and well-being of farmed salmonids., (© 2020 John Wiley & Sons Ltd.)

Three strains, designated as LB1R34 T , LB3P52 T and ZT4R6 T , were isolated from glaciers located on the Tibetan Plateau, PR China. The strains were Gram-stain-negative, aerobic, rod-shaped, non-motile and yellow. Phylogenetic analysis based on the 16S rRNA gene and genomic sequences indicated that they were related to the members of the genus Flavobacterium . The 16S rRNA gene sequences similarities between the three strains were 92.31-96.93 %. The average nucleotide identity values and digital DNA-DNA hybridization values between these three strains and their closest relatives were 76.80-91.33 % and 21.3-44.2 %, respectively. LB1R34 T , LB3P52 T and ZT4R6 T contained MK-6 as the major menaquinone, summed feature 3 (comprising C 16 : 1 ω7 c and/or C 16 : 1 ω6 c ) and iso -C 15 : 0 as the major fatty acids. Phosphatidylethanolamine was present in their polar lipids profiles. On the basis of the phenotypic characteristics, he results of phylogenetic analysis and genotypic data, three novel species, Flavobacterium restrictum sp. nov. (type strain=LB1 R34 T =CGMCC 1.11493 T =NBRC 113650 T ), Flavobacterium rhamnosiphilum sp. nov. (type strain=LB3 P52 T =CGMCC 1.11446 T =NBRC 113776 T ) and Flavobacterium zepuense sp. nov. (type strain=ZT4 R6 T =CGMCC 1.11919 T =NBRC 113653 T ) are proposed.

Two yellow-pigmented, Gram-stain-negative, aerobic, rod-shaped bacteria were isolated from the water of the hypersaline Chaka Salt Lake (strain SaA2.12 T ) and sediment of Qinghai Lake (strain LaA7.5 T ), PR China. According to the 16S rRNA phylogeny, the isolates belong to the genus Flavobacterium , showing the highest 16S rRNA sequence similarities to Flavobacterium arcticum SM1502 T (97.6-97.7 %) and Flavobacterium suzhouense XIN-1 T (96.5-96.6 %). Moreover, strains SaA2.12 T and LaA7.5 T showed 99.73 % 16S rRNA sequence similarity to each other. Major fatty acids, respiratory quinones and polar lipids detected in these isolates were iso-C 15 : 0 , menaquinone-6 and phosphatidylethanolamine, respectively. Strains SaA2.12 T and LaA7.5 T showed significant unique characteristics between them as well as between the closest phylogenetic members. The highest digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values between SaA2.12 T and its closest neighbours were 25.3 and 82.8 %, respectively; whereas these values (highest) between LaA7.5 T and its closest members were 25.2 and 82.8 %, respectively. The dDDH and ANI values between strains SaA2.12 T and LaA7.5 T were calculated as 75.9 and 97.2 %, respectively. Therefore, based on polyphasic data, we propose that strain SaA2.12 T represents a novel species with the name Flavobacterium salilacus sp. nov., with the type strain SaA2.12 T (=KCTC 72220 T =MCCC 1K03618 T ) and strain LaA7.5 T as a subspecies within novel Flavobacterium salilacus with the name Flavobacterium salilacus subsp. altitudinum subsp. nov., with the type strain LaA7.5 T (=KCTC 72806 T =MCCC 1K04372 T ). These propositions automatically create Flavobacterium salilacus subsp. salilacus subsp. nov. with SaA2.12 T (=KCTC 72220 T =MCCC 1K03618 T ) as the type strain.

A bacterial strain CC-CTC003 T was isolated from a synthetic wooden board. Cells of strain CC-CTC003 T were Gram-stain-negative, rod-shaped, motile by gliding and formed yellow colonies. Optimal growth occurred at 25 °C, pH 7 and in the presence of 1 % NaCl. The phylogenetic analyses based on 16S rRNA genes revealed that strain CC-CTC003 T belonged to the genus Flavobacterium and was most closely related to Flavobacterium cerinum (95.3 % sequence identity), Flavobacterium maris (94.9 % sequence identity), Flavobacterium qiangtangense (94.8 %) and Flavobacterium subsaxonicum (94.7 %) and had less than 94.7 % sequence similarity to other members of the genus. Average nucleotide identity (ANI) values between strain CC-CTC003 T and the type strains of other closely related species were 70.1-74.1 %. The digital DNA-DNA hybridization (dDDH) with F. cerinum was 19.4 %. Strain CC-CTC003 T contained C 15 : 0 , iso -C 15 : 0 , iso -C 15 : 0 3-OH, iso -C 17 : 0 3-OH, summed feature 3 (C 16 : 1  ω 6c / C 16 : 1  ω 7c) and summed feature 9 (C 16 : 0 10-methyl / iso -C 17 : 1  ω 9c) as the predominant fatty acids. The polar lipid profile consisted of phosphatidylethanolamine, four uncharacterized aminophospholipids, two aminolipids and one unidentified glycolipid. The major polyamine was sym -homospermidine and contained MK-6 as major isoprenoid quinone. The DNA G+C content of the genomic DNA was 39.2 mol%. On the basis of the phylogenetic inference and phenotypic data, strain CC-CTC003 T should be classified as a novel species, for which the name Flavobacterium supellecticarium sp. nov. is proposed. The type strain is CC-CTC003 T (=BCRC 81146 T =JCM 32838 T ).

A Gram-negative, strictly aerobic, gliding motility, none-spore forming, yellow, rods bacterial strain, designated XS-5 T , was isolated from rhizosphere soil of Suaeda salsa , in Tumd Right Banner, Inner Mongolia, PR China. A phylogenetic tree based on the 16S rRNA gene sequences and the phylogenomic tree both showed that strain XS-5 T clustered with Flavobacterium beibuense F44-8 T (shared 97.2 % of 16S rRNA gene similarity) and Flavobacterium rakeshii FCS-5 T (97.6 %), and shared <96.0 % of 16S rRNA gene similarities with all other type strains. Strain XS-5 T contained MK-6 as the major respiratory quinone. Its major polar lipids were phosphatidylethanolamine, an unidentified aminolipid and an unidentified lipid; and the major fatty acids were iso-C 15 : 0 , iso-C 17 : 0 3-OH, C 16 : 0 , iso-C 15 : 0 3-OH, Summed feature 3 (iso-C 15 : 0 2-OH and/or C 16 : 1   ω 7c ), and Summed feature 9 (iso-C 17 : 1  ω 9 c and/or C 16 : 0 10-methyl). The genome consisted of a 3 985 855 bp circular chromosome, with a G+C content of 37.9 mol%, predicting 3616 coding sequences genes, 45 tRNA genes and three rRNA operons. The average nucleotide identity, amino acid identity and digital DNA-DNA hybridization values of strain XS-5 T to F. beibuense F44-8 T and F. rakeshii FCS-5 T were 79.2 and 79.2 %, 81.7 and 81.6 %, 22.3 and 22.2 %, respectively. The results of phylogenetic, physiological and biochemical tests allowed the discrimination of strain XS-5 T from its phylogenetic relatives. Flavobacterium alkalisoli sp. nov. is therefore proposed with strain XS-5 T (=CGMCC 1.17077 T =KCTC 72459 T ) as the type strain.

The Gram-stain-negative, rod-shaped, facultative anaerobic, motile bacterial strain, designated TP390 T , was isolated from a seamount near the Yap Trench in the tropical western Pacific. Phylogenetic analysis based on 16S rRNA gene sequence showed that strain TP390 T was related to the genus Flavobacterium and had highest 16S rRNA gene sequence identity with the type strain of Flavobacterium jejuense EC11 T (97.8 %). Sequence similarities to all other type strains of current species of the genus Flavobacterium were below 97 %. The predominant cellular fatty acids were iso-C 15 : 0 and iso-C 15 : 1 G. The quinone system for strain TP390 T comprised predominantly menaquinone MK-6 and the polar lipid profile contained phosphatidylethanolamine, four unknown aminolipids, one glycolipid and six unknown polar lipids. The genomic DNA G+C content of strain TP390 T was 31.2 mol%. In addition, the maximum values of in silico DNA-DNA hybridization ( is DDH) and average nucleotide identity (ANI) between strain TP390 T with F. jejuense KCTC 42149 T were 22.60 and 80.01% respectively. Combined data from phenotypic, phylogenetic, is DDH and ANI data demonstrated that the strain TP390 T is representative of a novel species of the genus Flavobacterium , for which we propose the name Flavobacterium profundi sp. nov. (type strain TP390 T =KACC 18559 T =CGMCC 1.15398 T ).

Flavobacterium columnare is the causative agent of columnaris disease in a variety of fish hosts. Using modifications to previously established protocols, a quantitative PCR (qPCR) assay was validated for the detection of 2 predominant F. columnare genomovars. The oligonucleotide primer and probe combination was designed to amplify a 203-bp region of the chondroitin AC lyase gene (GenBank AY912281) of F. columnare . There were no significant differences in amplification between genomovars. Comparable quantities of genomic DNA from 10 F. columnare strains, 5 representatives of each genomovar, produced similar results. Serial dilutions of purified PCR product demonstrated the limit of sensitivity for the assay was ~ 10 copies per reaction. The presence of gill and spleen tissue did not significantly affect the sensitivity of the assay. Comparably, bacterial DNA detected from the liver and kidney was less sensitive than pure bacterial DNA. However, detection from these tissues was within one order of magnitude of other tissues, indicating this reduction may have minimal analytic significance. This validated assay was used to approximate the minimum infectious dose for F. columnare isolate 94-081 in channel catfish and assess bacterial loads in gill and kidney tissues 48 h post-infection.

A Gram-negative, non-motile, rod-shaped bacterial strain, designated GS13 T , was isolated from sediments in a branch of the Nackdong River in Sangju, Korea. Optimal growth occurred at pH 7.0, 20 °C, and 0% NaCl. Phylogenetic analyses using 16S rRNA showed that strain GS13 T is a member of the genus Flavobacterium, with highest similarity to Flavobacterium soyangense IMCC26223 T (97.0%). The DNA G+C content of strain GS13 T was 36.2 mol%. The dominant fatty acids were summed feature 3 (C 16:1 ω7c and/or C 16:1 ω6c) and iso-C 15:0 . The major polar lipids were phosphatidylethanolamine, three unidentified aminolipids, three unidentified lipids, and one unidentified aminophospholipid. The predominant respiratory quinone was MK-6. Our data demonstrate that strain GS13 T can be distinguished from closely-related Flavobacterium species. Thus, strain GS13 T is a novel Flavobacterium species, and we propose the name Flavobacterium nackdongense sp. nov. The type strain is GS13 T (=KCTC 62569 T  = JCM 32765 T ).

This study presents taxonomic description of strains LB-D12 T , AT-3-2 T , AT-3-7, and TSA-D2 T isolated from Arctic soil. All strains were psychrophilic, Gram-stain-negative, aerobic, non-motile, and rod-shaped. Phylogenetic analysis showed that these strains belonged to the genus Flavobacterium . Strains LB-D12 T , AT-3-2 T and AT-3-7 were closest to Flavobacterium psychrolimnae LMG 22018 T (98.5-98.8% sequence similarity). Strain TSA-D2 T was closest to Flavobacterium degerlachei DSM 15718 T (98.3 % sequence similarity). These strains shared common chemotaxonomic features comprising MK-6 as a sole quinone, phosphatidylethanolamine as the principal polar lipid, and summed feature 3 (iso-C 15 : 0 2-OH and/or C 16 : 1 ω7 c ), iso-C 16 : 0 3-OH, C 15 : 1 ω6 c , iso-C 16 : 0 , and anteiso-C 15 : 0 as the main fatty acids. The ANI and dDDH values between these novel isolates and their closest relatives were below the cut-off values of 95 and 70 %, respectively used for species demarcation. The DNA G+C content of all strains ranged from 34.2 to 34.6 mol%. The obtained polyphasic taxonomic data suggested that the isolated strains represent novel species within the genus Flavobacterium , for which the names Flavobacterium sandaracinum sp. nov. (type strain LB-D12 T =KEMB 9005-737 T =KACC 21180 T =NBRC 113784 T ), Flavobacterium caseinilyticum sp. nov. (type strain AT-3-2 T =KEMB 9005-738 T =KACC 21176 T =NBRC 113785 T ), and Flavobacterium hiemivividum sp. nov. (type strain TSA-D2 T =KEMB 9005-741 T =KACC 21179 T =NBRC 113788 T ) are proposed.

A taxonomic study was carried out on strain SH27 T , which was isolated from seawater collected around Xiaoshi Island, PR China. Cells of strain SH27 T were Gram-stain-negative, non-motile, rod-shaped, orange-pigmented and grew at 15-37 °C (optimum, 28 °C), at pH 6.0-8.0 (pH 7.0) and in 1.0-7.0 % (w/v) NaCl (2.0-3.0 %). The isolate was positive for catalase, but negative for nitrate reduction, oxidase, indole production and urease. Carotenoid pigment was produced. Phylogenetic analysis based on the 16S rRNA gene placed strain SH27 T in the genus Dokdonia with the closest relative being Dokdonia donghaensis KCTC 12391 T , exhibiting 96.7 % 16S rRNA gene pairwise similarity. The results of genomic comparisons, including average nucleotide identity and digital DNA-DNA hybridization, showed 72.9 and 19.2 % identity to D. donghaensis KCTC 12391 T , respectively. The major cellular fatty acids were iso-C 15 : 0 , iso-C 15 : 1 G and iso-C 17 : 0 3-OH. The major polar lipids were phosphatidylethanolamine and two unidentified lipids. Menaquinone-6 was the only respiratory quinone. The G+C content of the genomic DNA was 32.9 mol%. On the basis of the phenotypic and phylogenetic data, strain SH27 T represents a novel species of the genus Dokdonia , for which the name Dokdonia sinensis sp. nov. is proposed, with the type strain SH27 T (MCCC 1H00358 T =CCTCC AB 2018323 T =KCTC 62962 T ).

One aerobic, Gram-stain-negative, rod-shaped bacterium, designated strain RB1R5 T , was isolated from Renlongba glacier in Tibet Autonomous Region, China. Growth was observed at 4-25 °C and pH 7.0-8.0. Phylogenetic analysis based on 16S rRNA gene sequences revealed that strain RB1R5 T belongs to the genus Flavobacterium , and its closest relatives are Flavobacterium luteum IMCC26026 T and Flavobacterium psychroterrae CCM 8827 T with 97.75 and 97.42% gene sequence similarities, respectively. The average nucleotide identity and DNA-DNA hybridization values between strain RB1R5 T and its closest relative Flavobacterium luteum IMCC26026 T were 78.02 and 21.8 %, respectively. The major cellular fatty acids were anteiso-C 15 : 0 and summed feature 3 (comprising C 16:1 ω7 c /C 16:1 ω6 c ). The sole menaquinone was MK-6. The major phospholipid was phosphatidylethanolamine. On the basis of phenotypic, phylogenetic and chemotaxonomic characterizations, strain RB1R5 T represents a novel species of the genus Flavobacterium , for which the name Flavobacterium psychrotolerans sp. nov. is proposed. The type strain is RB1R5 T (=CGMCC 1.23024 T =NBRC 113060 T ).

A Gram-stain-negative, yellow-pigmented, strictly aerobic bacterium, designated strain MaA-Y11 T , was isolated from a culture of Microcystis aeruginosa in the Republic of Korea. Cells were non-motile rods showing catalase- and oxidase-positive reactions. Growth of strain MaA-Y11 T was observed at 25-37 °C (optimum, 30 °C) and pH 6.0-9.0 (optimum, 7.0) and in the presence of 0-1.0 % (w/v) NaCl (optimum, 0 %). Strain MaA-Y11 T did not produce flexirubin-type pigments. Strain MaA-Y11 T contained iso-C 15 : 0 , iso-C 17 : 0 3-OH and summed feature 3 (C 16 : 1 ω7 c and/or C 16 : 1 ω6 c ) as major cellular fatty acids and menaquinone-6 as the sole isoprenoid quinone. Phosphatidylethanolamine, three unidentified aminolipids and four unidentified lipids were detected as the major polar lipids. The G+C content of the genomic DNA was 37.0 mol%. Phylogenetic analysis, based on 16S rRNA gene sequences, showed that strain MaA-Y11 T formed a phyletic lineage with Flavobacterium lindanitolerans IP-10 T within the genus Flavobacterium . Strain MaA-Y11 T was most closely related to Flavobacterium lindanitolerans IP-10 T , with a 98.85 % 16S rRNA gene sequence similarity, and shared less than 93.87 % sequence similarities with other type strains. Average nucleotide identity and in silico DNA-DNA hybridization values between strain MaA-Y11 T and the type strain of F. lindanitolerans were 87.0 and 32.3 %, respectively. Here, we conclude that strain MaA-Y11 T represents a new species of the genus Flavobacterium , for which the name Flavobacterium microcysteis sp. nov. is proposed. The type strain is MaA-Y11 T (=KACC 21225 T =JCM 33501 T ).

A novel psychrophilic, light-yellow-coloured, aerobic, Gram-stain-negative, rod-shaped, non-motile bacterium, designated strain AR-3-4 T was isolated from a sample of Arctic soil. Strain AR-3-4 T grew at 0-25 °C, pH 6.0-9.0 and 0-1.0 % (w/v) NaCl concentration. The 16S rRNA gene sequence analysis showed that strain AR-3-4 T belonged to the genus Flavobacterium , with nearest phylogenetic neighbour being Flavobacterium fluvii H7 T (97.5 % sequence similarity). The strain comprised phosphatidylethanolamine as the main polar lipid, MK-6 as predominant respiratory quinone, and summed feature 3 (C 16 : 1 ω7 c and/or C 16 : 1 ω6 c ), anteiso-C 15 : 0 and iso-C 15 : 0 as the major fatty acids. The average nucleotide identity and in silico DNA-DNA hybridization values between strain AR-3-4 T and closest members were below the threshold values of 95 % and 70 %, respectively. The DNA G+C content was 33.0 mol%. Based on the polyphasic taxonomic data, the novel species Flavobacterium cellulosilyticum sp. nov. is proposed with the type strain AR-3-4 T (=KEMB 9005-740 T =KACC 21171 T =NBRC 113787 T ).

A yellow, Gram-stain-negative, aerobic, non-gliding, non-spore-forming, rod-shaped strain, designated YIM 102600 T , was isolated from the faeces of Macaca mulatta dwelling in the Yunnan Wild Animal Park, Yunnan Province, South-West PR China. Phylogenetic analysis based on 16S rRNA gene sequences showed that strain YIM 102600 T was a member of the genus Flavobacterium , and closely related to Flavobacterium qiangtangense F3 T (96.9 % similarity) and Flavobacterium noncentrifugens R-HLS-17 T (96.0 % similarity). Phylogenetic trees showed that strain YIM 102600 T formed a clade with F. qiangtangense F3 T and F. noncentrifugens R-HLS-17 T . Growth occurred at 4-30 °C (optimum, 28 °C), pH 7.0-8.0 (pH 7.5) and NaCl concentration 0-2 % (w/v; 0-1 %, w/v). The major fatty acids were iso-C 15:0 and summed feature 3 (comprising C 16:1 ω 7 c and/or C 16:1 ω 6 c ). The predominant polar lipid was phosphatidylethanolamine and the sole respiratory quinone was menaquinone-6. The DNA G+C content was 36.4 mol%. The calculated digital DNA-DNA hybridization values between strain YIM 102600 T and other species of Flavobacterium ranged from 70.0 to 75.0 % and average nucleotide identity values were in a range between 13.7 to 23.5 %. Based above the consensus of phenotypic and phylogenetic analyses as well as whole genome comparisons, strain YIM 102600 T (=KCTC 52099 T =CCTCC AB 201632 T ) is proposed to represent type strain of a novel species, Flavobacterium macacae sp. nov.

Objective: Alkaline Carboxymethyl Cellulase (CMCase) is an attractive enzyme for the textile, laundry, pulp, and paper industries; however, commercial preparations with sufficient activity at alkaline conditions are scarce., Methods: High CMCase-producing bacterial isolate, SX9-4, was screened out from soil bacteria, which was identified as Flavobacterium sp. on the basis of 16S rDNA sequencing., Results: The optimum pH and temperature for CMCase reaction were 8.0 and 55°C, respectively. Alkaline CMCase was stable over wide pH (3.0-10.6) and temperature (25-55°C) ranges. Enzyme activity was significantly inhibited by the bivalent cations Mn 2+ and Cu 2+ , and was activated by Fe 2+ . To improve the alkaline CMCase production of SX9-4, fermentation parameters were selected through onefactor- at-a-time and further carried out by response surface methodologies based on a central composite design., Conclusion: High CMCase production (57.18 U/mL) was achieved under the optimal conditions: 10.53 g/L carboxymethylcellulose sodium, 7.74 g/L glucose, 13.71 g/L peptone, and 5.27 g/L ammonium oxalate., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

In the present paper, we describe a new species of the genus Flavobacterium , designated as strain 1E403 T , which was isolated from a soil sample collected from the Arctic. Strain 1E403 T was Gram-stain-negative, yellow-pigmented, rod-shaped, gliding and aerobic. Growth occurred at 4-37 °C (optimum, 28 °C), pH 6.0-9.0 (pH 7.0) and with 0-2 % (w/v) NaCl (0 %) on modified marine agar 2216. Phylogenetic analysis based on 16S rRNA gene sequencing revealed that strain 1E403 T was affiliated with the genus Flavobacterium and was more closely related to Flavobacterium subsaxonicum DSM 21790 T (96.6 %) than to other species. In silico genomic comparisons, including average nucleotide identity and the digital DNA-DNA hybridization values, showed 73.9 % and 18.8 % identity to the closest relative Flavobacterium subsaxonicum DSM 21790 T , respectively. The average amino acid identity value between strain 1E403 T and Flavobacterium aquatile DSM 1132 T was 63.8 %. The DNA G+C content of the strain was 36.8 %, while the sole respiratory quinone was menaquinone-6. Iso-C 15 : 0 and summed feature 3 (C 16 : 1 ω7 c and/or C 16 : 1 ω6 c ) were the major fatty acids (>10 %) of strain 1E403 T . The polar lipid profile of strain 1E403 T contained phosphatidylethanolamine, four unidentified aminolipids and two unidentified phospholipids. Based on the phenotypic characteristics, chemotaxonomic characteristics and phylogenetic inference, strain 1E403 T represents a novel species of the genus Flavobacterium , and we propose the name Flavobacterium cerinum sp. nov. The type strain of Flavobacterium cerinum sp. nov. is 1E403 T (=KCTC 62960 T =MCCC 1H00356 T ).

A yellow pigmented, Gram-stain-negative, strictly aerobic, rod-shaped, motile by means of gliding, catalase and oxidase positive bacterium, designated strain DS2-A T , was isolated from soil. Growth was observed at 4-32°C (optimum, 28°C), pH 6-9 (optimum, 7.0), and with 0-0.25% (w/v) NaCl (optimum, 0%). Phylogenetic analysis of 16S rRNA gene sequence revealed that strain DS2-A T belonged to the genus Flavobacterium and was most closely related to Flavobacterium aquatile LMG 4008 T (96.4%), Flavobacterium terrae DSM 18829 T (95.6%), Flavobacterium vireti THG-SM1 T (95.5%), Flavobacterium inkyongense IMCC27201 T (95.4%), Flavobacterium brevivitae TTM-43 T (95.2%), and Flavobacterium cucumis DSM 18830 T (95.2%). Strain DS2-A T produces flexirubin-type pigments. The major fatty acids were iso-C 15:0 , iso-C 17:0 3-OH, and iso-C 15:0 3-OH. The major respiratory quinone was identified as menaquinone-6. The major polar lipid was found to be phosphatidylethanolamine. The average nucleotide identity values between strain DS2-A T and selected taxa, F. aquatile LMG 4008 T , F. terrae DSM 18829 T , and F. cucumis DSM 18830 T , were 72, 72.7, and 71.6%, respectively. The draft genome of strain DS2-A T has a number of 14 contigs, scaffold N50 of 476,310 bp and a total size of 3,563,867 bp. Additionally, strain DS2-A T contains 3,127 of gene, 41 of tRNA, 6 of rRNA, and 3 of ncRNA. The DNA G + C content of stain DS2-A T was 40.7 mol%. Based on phylogenetic and phenotypic analyses, strain DS2-A T is considered as a novel species of the genus Flavobacterium, for which the name Flavobacterium humi sp. nov., (type strain DS2-A T = KACC 19715 T = JCM 32786 T ) has been proposed.

A yellow-pigmented bacterial strain, designated T13 T , was isolated from the rhizosphere soil of Alhagi sparsifolia collected from Xinjiang, PR China. Cells were rod-shaped, Gram-stain-negative, aerobic and gliding. Strain T13 T grew optimally at 25-30 °C and pH 7.0-8.0 with a NaCl tolerance of 0-2 % on Reasoner's 2A agar. Phylogenetic analysis based on the 16S rRNA gene sequence showed that strain T13 T belonged to the genus Flavobacterium within the family Flavobacteriaceae and was closely related to Flavobacterium nitrogenifigens KCTC 42884 T with a similarity value of 97.4 %. The major polar lipid was phosphatidylethanolamine; the only respiratory quinone was MK-6, and the polyamine profile contained sym -homospermidine as the major polyamine and a trace amount of spermidine. The major fatty acids were iso-C 15 : 0 , C16 :  1 ω7 c and summed feature 3 (comprising C 16 : 1 ω7 c or C 16 : 1 ω6 c ). The G+C content of the genomic DNA was 34.1 mol%. It is concluded from the phenotypic and genotypic data that strain T13 T represents a novel species of the genus Flavobacterium , for which the name Flavobacteriumustbae sp. nov. with the type strain T13 T (=KCTC 62874 T =ACCC 60126 T ) is proposed.

A yellow pigmented, Gram-stain-negative, aerobic bacterium designated A5.7 T was studied to evaluate the taxonomic position following the modern polyphasic approach. The strain was isolated from sediments near Zhairuo Island, which is situated in the East China Sea. Cells were non-spore forming rods without flagella but showed motility by gliding. Growth was observed at 15-35°C (optimum 28°C), pH 6.0-9.0 (optimum pH 6.5) and 0-2% (w/v) NaCl (optimum 0-0.5%) in LB broth. The major respiratory quinone of A5.7 T was menaquinone 6. The major polar lipid of A5.7 T was phosphatidylethanolamine and the predominant fatty acids (> 5%) were iso-C 15:0 , iso-C 17:0 3-OH, C 15:1 ω6c, iso-C 15:0 3-OH, iso-C 15:1 G, summed feature 3 (C 16:1 ω7c and/or C 16:1 ω6c) and summed feature 9 (iso-C 17:1 ω9c and/or C 16:0 10-methyl). Phylogenetic analysis based on 16S rRNA gene sequences showed that the isolate belongs to the genus Flavobacterium and shares the highest sequence similarities with Flavobacterium sharifuzzamanii A7.6 T (98.5%), Flavobacterium tistrianum GB 56.1 T (98.3%), Flavobacterium nitrogenifigens NXU-44 T (97.8%), Flavobacterium anhuiense D3 T (97.6%), Flavobacterium ginsenosidimutans THG 01 T (97.6%), and Flavobacterium foetidum CJ42 T (97.6%). Digital DNA-DNA hybridization and average nucleotide identity values between the strain and its closest phylogenetic neighbors showed the ranges from 19.6 to 34.1% and 73.7 to 87.9%, respectively. Therefore, based on polyphasic characteristics, strain A5.7 T represents a novel species of the genus Flavobacterium for which the name Flavobacterium zhairuonensis sp. nov. is proposed. The type strain is A5.7 T (= KCTC 62406 T = MCCC 1K03494 T ).

A Gram-stain-negative, aerobic, rod-shaped, devoid of flagella, orange-yellow-coloured bacterium (strain LB2P22 T ) was isolated from an ice sample collected from the Laigu glacier on the Tibetan Plateau, PR China. The results of 16S rRNA gene sequence analysis indicated that strain LB2P22 T belongs to the genus Flavobacterium with highest similarity to Flavobacterium laiguense LB2P30 T (98.14 %), Flavobacterium glaciei CGMCC 1.5380 T (98.02 %), Flavobacterium psychroterrae CCM 8827 T (97.94 %) and Flavobacterium granuli CGMCC 1.10125 T (97.91 %). Strain LB2P22 T had 77.99-84.99 % average nucleotide identity and 22.3-29.3 % digital DNA-DNA hybridization values with its closest relatives, indicating that it represents a novel species of the genus Flavobacterium . Strain LB2P22 T grew at 0-25 °C, pH 7.0-8.0 and up to 1.0 % (w/v) NaCl. The main cellular fatty acids were iso-C 15 : 0 , summed feature 3 (comprising C 16 : 1 ω7 c /C 16 : 1 ω6 c ) and anteiso-C 15 : 0 . The genomic DNA G+C content is 34.3 mol%. The predominant menaquinone was MK-6 and the polar lipids were phosphatidylethanolamine, one unidentified aminolipid and four unidentified lipids. Based on these data, a novel species, Flavobacteriumranwuense sp. nov., is proposed, with LB2P22 T (=CGMCC 1.11361 T =NBRC 113777 T ) as the type strain.

Recent research has identified four distinct genetic groups among isolates of Flavobacterium columnare through multilocus phylogenetic analyses; however, there are no quick methods to determine the genotype of an isolate. The objective of this research was to develop a multiplex PCR to rapidly genotype F. columnare to genetic group. Comparative bacterial genomics was used to identify regions in the genomes unique to each genetic group, and primers were designed to specifically amplify different sized amplicons for each genetic group. The optimized assay was demonstrated to be specific for each genetic group and F. columnare, and no specific amplicons were generated using gDNA from a panel of other Flavobacterium spp. and bacterial fish pathogens. The analytical sensitivity of the assay ranged from 209 to 883 genome equivalents depending on the genetic group. The multiplex PCR was evaluated by genotyping a panel of 22 unknown F. columnare isolates and performing DNA sequencing of the dnaK gene in parallel. The results demonstrated 100% accordance between multiplex PCR results and assignment to genetic group via phylogenetic analysis. The multiplex PCR provides a useful tool for assigning an unknown isolate to genetic group and may be used to determine which genetic groups of F. columnare are circulating and most predominant in different aquaculture industries., (Published 2019. This article is a U.S. Government work and is in the public domain in the USA.)

In late April 2015, the River Dee Trust informed Marine Scotland Science, Fish Health Inspectorate (FHI), that there had been observations of dead and moribund European eels on the River Dee. Later in May, the Spey Fishery Board also reported a number of moribund European eels in a rotary screw smolt trap on the River Spey. In total, 10 cases involving moribund eels were investigated in 2015 and one case in 2016. In addition, a health screen was conducted to investigate the potential presence of Flavobacterium psychrophilum in healthy eels and Atlantic salmon from the River Dee in 2015. Externally, the diseased eels demonstrated white patches in different locations of the body. In all cases, F. psychrophilum was detected by bacterial isolation and/or molecular methods. Three isolates were further characterized by whole-genome sequencing (WGS) as belonging to sequence type 15 (ST15). Histological examination of diseased European eels revealed lesions at the level of the integument. The pathogen screen for F. psychrophilum in wild healthy fish tested negative by PCR. Further investigation is required to understand the pathogenicity of this bacterium on the health of eels and the potential impact on the wild salmonid population., (© 2019 John Wiley & Sons Ltd.)