L. Zamora

Chryseobacterium oncorhynchi sp. nov., isolated from rainbow trout (Oncorhynchus mykiss)

Genotypic and phenotypic analyses were performed on five Gram-negative, catalase and oxidase-positive, rod-shaped bacteria isolated from the gill and liver of four rainbow trout. Studies based on comparative 16S rRNA gene sequence analysis showed that the five new isolates shared 99.8-100% sequence similarity and that they belong to the genus Chryseobacterium. The nearest phylogenetic neighbours of the strain 701B-08(T) were Chryseobacterium ureilyticum F-Fue-04IIIaaaa(T) (99.1% 16S rRNA gene sequence similarity) and Chryseobacterium joosteii LMG 18212(T) (98.6%). DNA-DNA hybridization values between the five isolates were 91-99% and ranged from 2 to 53% between strain 701B-08(T) and the type strains of phylogenetically closely related species of Chryseobacterium. Strain 701B-08(T) had a DNA G+C content of 36.3 mol%, the major fatty acids were iso-C(15:0), iso-C(17:1)ω9c, C(16:1)ω6c and iso-C(17:0) 3-OH and the predominant respiratory quinone was MK-6. The novel isolates were distinguished from related Chryseobacterium species by physiological and biochemical tests. The genotypic and phenotypic properties of the isolates from rainbow trout suggest their classification as representatives of a novel species of the genus Chryseobacterium, for which the name Chryseobacterium oncorhynchi sp. nov. is proposed. The type strain is 701B-08(T) (=CECT 7794(T)=CCUG 60105(T)).

Flavobacterium oncorhynchi sp. nov., a new species isolated from rainbow trout (Oncorhynchus mykiss).

Eighteen isolates of a Gram-negative, catalase and oxidase-positive, rod-shaped bacterium, recovered from diseased rainbow trout (Oncorhynchus mykiss), were characterized, using a polyphasic taxonomic approach. Studies based on comparative 16S rRNA gene sequence analysis showed that that the eighteen new isolates shared 99.2-100% sequence similarities. Phylogenetic analysis revealed that isolates from trout belonged to the genus Flavobacterium, showing the highest sequence similarities to F. chungangense (98.6%), F. frigidimaris (98.1%), F. hercynium (97.9%) and F. aquidurense (97.8%). DNA-DNA reassociation values between the trout isolates (exemplified by strain 631-08(T)) and five type strains of the most closely related Flavobacterium species exhibited less than 27% similarity. The G+C content of the genomic DNA was 33.0 mol%. The major respiratory quinone was observed to be menaquinone 6 (MK-6) and iso-C(15:0), C(15:0) and C(16:1) ω7c the predominant fatty acids. The polar lipid profile of strain 631-08(T) consisted of phosphatidylethanolamine, unknown aminolipids AL1 and AL3, lipids L1, L2, L3 and L4 and phospholipid PL1. The novel isolates were differentiated from related Flavobacterium species by physiological and biochemical tests. On the basis of the evidence from this polyphasic study, it is proposed that the isolates from rainbow trout be classified as a new species of the genus Flavobacterium, Flavobacterium oncorhynchi sp. nov. The type strain is 631-08(T) (= CECT 7678(T) = CCUG 59446(T)).

Flavobacterium tructae sp. nov. and Flavobacterium piscis sp. nov., isolated from farmed rainbow trout (Oncorhynchus mykiss).

Four Gram-staining-negative, catalase- and oxidase-positive, pale-orange pigmented bacterial strains (435-08(T), 47B-3-09, 412R-09(T) and 60B-3-09) were isolated from diseased rainbow trout. Analysis of their 16S rRNA gene sequences suggested their adscription to the genus Flavobacterium. Strains formed two phylogenetic groups represented by strains 435-08(T) and 47B-3-09 (group A), and strains 412R-09(T) and 60B-3-09 (group B) displaying 16S rRNA sequence similarities greater than 99.8-99.9% within their respective groups. Strain 435-08(T) exhibited the highest levels of similarity with Flavobacterium aquidurense WB-1.1.56(T) (98.6% sequence similarity) and strain 412R-09(T) with Flavobacterium frigidimaris KUC-1(T) and Flavobacterium aquidurense WB-1.1.56(T) (98.9% and 98.6% sequence similarity, respectively). DNA-DNA hybridization studies showed low levels of relatedness between strain 435-08(T) and strain 412R-09(T) and between both strains and the most closely related species of the genus Flavobacterium. The genomic DNA G+C contents of strains 435-08(T) and 412R-09(T) were 36.2 and 34.3 mol%, respectively. The predominant respiratory quinone of both strains was MK-6 and the major fatty acids were iso-C(15 : 0), C(16 : 1)ω7c and C(15 : 0). The two groups of strains could be distinguished from each other and from related species of the genus Flavobacterium by a number of phenotypic properties. Phylogenetic, genotypic and phenotypic evidence indicated that strains of groups A and B represent two novel species of the genus Flavobacterium, for which the names Flavobacterium tructae sp. nov. (type strain 435-08(T) = CECT 7791(T) = CCUG 60100(T)) and Flavobacterium piscis sp. nov. (type strain 412R-09(T) = CECT 7911(T) = CCUG 60099(T)) are proposed.

Chryseobacterium viscerum sp. nov., isolated from diseased fish

A taxonomic study was carried out on five Gram-staining-negative, catalase- and oxidase-positive, rod-shaped bacteria isolated from the gills and livers of five diseased rainbow trout. The five novel isolates were designated strains 687B-08(T), 445-08, 452-08, 453B-08 and 967B-08. In phylogenetic analyses based on 16S rRNA gene sequences, the five novel strains appeared almost identical (99.0-100 % sequence similarity) and to belong to the genus Chryseobacterium. Strain 687B-08(T) (the strain selected to represent the five novel isolates) was found to be most closely related to Chryseobacterium oncorhynchi 701B-08(T) (98.9% sequence similarity), Chryseobacterium ureilyticum F-Fue-04IIIaaaa(T) (98.6%), Chryseobacterium indologenes ATCC 29897(T) (98.3%), Chryseobacterium jejuense JS17-8(T) (98.1%) and Chryseobacterium gleum ATCC 35910(T) (98.1%). In DNA-DNA hybridizations, DNA-DNA relatedness values of 99-100% were recorded between the five novel strains. Lower DNA-DNA relatedness values (21-57%) were recorded between strain 687B-08(T) and C. oncorhynchi 701B-08(T), C. ureilyticum F-Fue-04IIIaaaa(T) and the type strains of other closely related, established species of the genus Chryseobacterium. The predominant respiratory quinone of strain 687B-08(T) was MK-6 and the major cellular fatty acids were iso-C(15:0), iso-C(17:1)ω9c, iso-C(17:0) 3-OH and C(16:1)ω6c. The G+C content of the genomic DNA of strain 687B-08(T) was 38.6 mol%. Based on the phenotypic and genotypic evidence, the five novel strains isolated from rainbow trout represent a single, novel species of the genus Chryseobacterium, for which the name Chryseobacterium viscerum sp. nov. is proposed. The type strain is 687B-08(T) ( = CECT 7793(T)  = CCUG 60103(T)).

First isolation and characterization of Chryseobacterium shigense from rainbow trout

BackgroundThere have been an increasing number of infections in fish associated with different species of Chryseobacterium, being considered potentially emerging pathogens. Nevertheless the knowledge of the diversity of species associated with fish disease is partial due to the problems for a correct identification at the species level based exclusively on phenotypic laboratory methods.ResultsChryseobacterium shigense was isolated from the liver, kidney and gills of diseased rainbow trout in different disease episodes that occurred in a fish farm between May 2008 and June 2009. Identity of the isolates was confirmed by 16 S rRNA gene sequencing and phenotypic characterization. Isolates represented a single strain as determined by random amplified polymorphic DNA analysis.ConclusionsThis is the first description of the recovery of C. shigense from clinical specimens in trout, a very different habitat to fresh lactic acid beverage where it was initially isolated.

Chryseobacterium tructae sp. nov., isolated from rainbow trout (Oncorhynchus mykiss)

Three pale-orange bacteria (strains 1083-08, 1084-08(T) and 1095B-08) were isolated from diseased rainbow trout. The isolates were Gram-staining-negative, catalase- and oxidase-positive, rod-shaped cells. Analyses of their 16S rRNA gene sequences confirmed their adscription to the genus Chryseobacterium. The three isolates shared 100% 16S rRNA gene sequence similarity and 98.5% similarity with Chryseobacterium indologenes CCUG 14556(T), being the closest phylogenetically related species. Genomic DNA-DNA hybridization similarity values between the three isolates were 94-100% and 2-39% between strain 1084-08(T) and the type strains of other related Chryseobacterium species, confirming that the isolates represent a novel species within the genus Chryseobacterium. The DNA G+C content of the species was 33.6-36.1mol%. The predominant respiratory quinone of strain 1084-08(T) was MK-6 and the major fatty acids were iso-C(15:0), iso-C(17:1)ω9c, iso-C(17:0) 3-OH and C(16:1)ω6c. The isolates were distinguished from related Chryseobacterium species by a number of phenotypic properties. Based on the phenotypic, genotypic and phylogenetic findings, it is proposed that the new isolates from rainbow trout be classified as a new species of the genus Chryseobacterium, with the name of Chryseobacterium tructae sp. nov. The type strain is 1084-08(T) (=CECT 7798(T)=CCUG 60111(T)).

Flavobacterium plurextorum sp. nov. Isolated from Farmed Rainbow Trout (Oncorhynchus mykiss)

Five strains (1126-1H-08T, 51B-09, 986-08, 1084B-08 and 424-08) were isolated from diseased rainbow trout. Cells were Gram-negative rods, 0.7 µm wide and 3 µm long, non-endospore-forming, catalase and oxidase positive. Colonies were circular, yellow-pigmented, smooth and entire on TGE agar after 72 hours incubation at 25°C. They grew in a temperature range between 15°C to 30°C, but they did not grow at 37°Cor 42°C. Based on 16S rRNA gene sequence analysis, the isolates belonged to the genus Flavobacterium. Strain 1126-1H-08T exhibited the highest levels of similarity with Flavobacterium oncorhynchi CECT 7678T and Flavobacterium pectinovorum DSM 6368T (98.5% and 97.9% sequence similarity, respectively). DNA–DNA hybridization values were 87 to 99% among the five isolates and ranged from 21 to 48% between strain 1126-1H-08T, selected as a representative isolate, and the type strains of Flavobacterium oncorhynchi CECT 7678T and other phylogenetic related Flavobacterium species. The DNA G+C content of strain 1126-1H-08T was 33.2 mol%. The predominant respiratory quinone was MK-6 and the major fatty acids were iso-C15∶0 and C15∶0. These data were similar to those reported for Flavobacterium species. Several physiological and biochemical tests differentiated the novel bacterial strains from related Flavobacterium species. Phylogenetic, genetic and phenotypic data indicate that these strains represent a new species of the genus Flavobacterium, for which the name Flavobacterium plurextorum sp. nov. was proposed. The type strain is 1126-1H-08T ( = CECT 7844T = CCUG 60112T).

Streptococcus porci sp. nov., isolated from swine sources.

Two unidentified Gram-positive, catalase-negative, coccus-shaped organisms were recovered from pigs and subjected to a polyphasic taxonomic analysis. Based on cellular morphology and biochemical criteria, the isolates were tentatively assigned to the genus Streptococcus, although the organisms did not appear to correspond to any recognized species. Comparative 16S rRNA gene sequence studies confirmed this identification and showed that the nearest phylogenetic relatives of the unknown cocci were Streptococcus plurextorum 1956-02(T) and Streptococcus suis NCTC 10234(T) (97.9 and 96.0 % 16S rRNA gene sequence similarity, respectively). The new isolates were related most closely to S. suis CIP 103217(T) based on rpoB gene sequence analysis (<8 % sequence divergence). DNA-DNA pairing studies showed that one of the unidentified strains (2923-03(T)) displayed DNA relatedness values of 26.6 and 27.2 % with S. plurextorum CECT 7308(T) and S. suis NCTC 10234(T), respectively. On the basis of phenotypic and phylogenetic evidence, it is proposed that the unknown isolates from pigs be classified in the genus Streptococcus as members of Streptococcus porci sp. nov., with the type strain 2923-03(T) (=CECT 7374(T) =CCUG 55896(T)).

Streptococcus cuniculi sp. nov., isolated from the respiratory tract of wild rabbits

Biochemical and molecular genetic studies were performed on four unknown Gram-stain-positive, catalase-negative, coccus-shaped organisms isolated from tonsils (n = 3) and nasal samples (n = 1) of four wild rabbits. The micro-organism was identified as a streptococcal species based on its cellular morphological and biochemical tests. Comparative 16S rRNA gene sequencing confirmed its identification as a member of the genus Streptococcus, but the organism did not correspond to any recognized species of this genus. The closest phylogenetic relative of the unknown cocci from wild rabbits was Streptococcus acidominimus NCIMB 702025(T) (97.9% 16S rRNA gene sequence similarity). rpoB and sodA sequence analysis of the novel isolate showed interspecies divergence of 16.2% and 20.3%, respectively, from the type strain of its closest 16S rRNA gene phylogenetic relative, S. acidominimus. The novel bacterial isolate could be distinguished from the type strain of S. acidominimus by several biochemical characteristics, such as the production of esterase C4, acid phosphatase and naphthol-AS-BI-phosphohydrolase and acidification of different sugars. Based on both phenotypic and phylogenetic findings, it is proposed that the unknown bacterium be classified as a novel species of the genus Streptococcus, Streptococcus cuniculi sp. nov. The type strain is NED12-00049-6B(T) ( = CECT 8498(T) = CCUG 65085(T)).

Bergeyella porcorum sp. nov., isolated from pigs.

Four Gram-stain-negative, catalase- and oxidase-positive, bacillus-shaped bacterial isolates were recovered from the lungs and tonsils of four pigs. Based on cellular morphology and biochemical criteria the isolates were tentatively assigned to the genus Bergeyella, although the organisms did not appear to correspond with Bergeyella zoohelcum, the only validly named species of this genus. 16S rRNA gene sequencing demonstrated that isolates represented a distinct subline within the genus Bergeyella with <97%. 16S rRNA gene sequence similarity with B. zoohelcum ATCC 43767(T). The predominant cellular fatty acids of strain 1350-03(T) were iso-C15:0 and iso-C17:0 3-OH and the major quinone was MK-6. The DNA G+C content of strain 1350-03(T) was 37.7mol%. The novel isolates can be phenotypically distinguished from B. zoohelcum based on physiological traits. On the basis of both phenotypic and phylogenetic findings, we describe a new species of the genus Bergeyella for which we propose the name of Bergeyella porcorum sp. nov. (1350-03(T)=CCUG 67887(T)=CECT 9006(T)).