Staphylococcus devriesei sp. nov., isolated from teat apices and milk of dairy cows.

Ten non-motile, Gram-stain-positive, coagulase-negative staphylococci were isolated from bovine milk and teat apices. All isolates were catalase-positive, with anteiso-C(15 : 0), iso-C(15 : 0), anteiso-C(17 : 0), iso-C(17 : 0) and C(18 : 0) as predominant fatty acids and diphosphatidylglycerol and phosphatidylglycerol as major polar lipids. The results of sequence analysis of the 16S rRNA gene and four housekeeping genes (rpoB, hsp60, tuf and dnaJ) in combination with tRNA-intergenic spacer length analysis showed that the isolates form a separate branch within the genus Staphylococcus. Based on 16S rRNA gene sequencing, the phylogenetically most closely related species are Staphylococcus haemolyticus, S. hominis and S. lugdunensis, with >98.7 % sequence similarity. The DNA G+C content varies from 33.3 to 33.7 mol%, and DNA-DNA hybridization with the nearest neighbours, based on 16S rRNA gene sequences, confirmed that the isolates represent a novel Staphylococcus species. All isolates induced a small zone of complete haemolysis on Columbia agar with 5 % sheep blood and exhibited a homogeneous biochemical fingerprint that is discriminative from the phylogenetically most closely related species. Based on these results, it is proposed to classify the ten isolates as Staphylococcus devriesei sp. nov., with strain KS-SP 60(T) (=LMG 25332(T) =CCUG 58238(T)) as the type strain.

Emended description of the genus Pantoea, description of four species from human clinical samples, Pantoea septica sp. nov., Pantoea eucrina sp. nov., Pantoea brenneri sp. nov. and Pantoea conspicua sp. nov., and transfer of Pectobacterium cypripedii (Hori 1911) Brenner et al. 1973 emend. Hauben et al. 1998 to the genus as Pantoea cypripedii comb. nov.

Bacterial strains belonging to DNA hybridization groups (HG) II, IV and V, in the Erwinia herbicola-Enterobacter agglomerans complex, of Brenner et al. [Int J Syst Bacteriol 34 (1984), 45-55] were suggested previously to belong to the genus Pantoea, but have never been formally described and classified. Additionally, it has been shown in several studies that Pectobacterium cypripedii is more closely related to species of Pantoea than to those of Pectobacterium. In this study, the phylogenetic positions of Brenner's DNA HG II, IV and V and Pectobacterium cypripedii were re-examined by both 16S rRNA gene sequencing and multilocus sequence analyses (MLSA) based on the gyrB, rpoB, atpD and infB genes. The analyses revealed that DNA HG II, IV and V and Pectobacterium cypripedii form five separate branches within the genus Pantoea (strains from HG V were split into two branches). DNA-DNA hybridization data further confirmed that DNA HG II, IV and V constitute four separate species. Pectobacterium cypripedii was shown to be a close phylogenetic relative of Pantoea dispersa and DNA HG IV by both 16S rRNA gene sequence and MLSA analyses. Biochemical analyses performed on strains from DNA HG II, IV and V and Pectobacterium cypripedii confirmed their taxonomic position within the genus Pantoea and revealed phenotypic characteristics that allow the differentiation of these species from each other and from their closest phylogenetic neighbours. It is proposed to emend the description of the genus Pantoea and to describe Pantoea septica sp. nov. for DNA HG II (type strain LMG 5345(T) =BD 874(T) =CDC 3123-70(T)), Pantoea eucrina sp. nov. for DNA HG IV (type strain LMG 2781(T) =BD 872(T) =CDC 1741-71(T) =LMG 5346(T)), Pantoea brenneri sp. nov. for strains of DNA HG V excluding LMG 24534 (type strain LMG 5343(T) =BD 873(T) =CDC 3482-71(T)) and Pantoea conspicua sp. nov. for the remaining strain of DNA HG V (type strain LMG 24534(T) =BD 805(T) =CDC 3527-71(T)) and to transfer Pectobacterium cypripedii to the genus as Pantoea cypripedii comb. nov. (type strain LMG 2657(T) =ATCC 29267(T) =DSM 3873(T) =LMG 2655(T)).

Pantoea vagans sp. nov., Pantoea eucalypti sp. nov., Pantoea deleyi sp. nov. and Pantoea anthophila sp. nov.

Bacteria isolated from eucalyptus leaves and shoots showing symptoms of blight and die-back collected in Uganda, Uruguay and Argentina and from maize displaying brown stalk rot symptoms in South Africa were tentatively placed in the genus Pantoea on the basis of phenotypic and biochemical tests. These isolates, together with two strains (LMG 2558 and LMG 2560) previously assigned to Pantoea agglomerans based on protein electrophoregrams but later excluded from this species, were further investigated using molecular techniques. 16S rRNA gene sequencing and multilocus sequence analyses (MLSA) revealed that the strains were phylogenetically closely related to Pantoea agglomerans, Pantoea stewartii and Pantoea ananatis. MLSA and amplified fragment length polymorphism analysis placed the strains into four separate clusters, not containing any of the type strains of species of the genus Pantoea. DNA-DNA hybridization confirmed the classification of the isolates into four novel species, for which the names Pantoea vagans sp. nov. (type strain R-21566T=LMG 24199T=BCC 105T=BD 765T), Pantoea eucalypti sp. nov. (type strain R-25678T=LMG 24197T=BCC 076T=BD 769T), Pantoea deleyi sp. nov. (type strain R-31523T=LMG 24200T=BCC 109T=BD 767T) and Pantoea anthophila sp. nov. (type strain LMG 2558T=BD 871T=NCPPB 1682T) are proposed.

Acetobacter fabarum sp. nov., an acetic acid bacterium from a Ghanaian cocoa bean heap fermentation.

Six acetic acid bacterial isolates, obtained during a study of the microbial diversity of spontaneous fermentations of Ghanaian cocoa beans, were subjected to a polyphasic taxonomic study. (GTG)(5)-PCR fingerprinting grouped the isolates together, but they could not be identified using this method. Phylogenetic analysis based on 16S rRNA gene sequences allocated the isolates to the genus Acetobacter and revealed Acetobacter lovaniensis, Acetobacter ghanensis and Acetobacter syzygii to be nearest neighbours. DNA-DNA hybridizations demonstrated that the isolates belonged to a single novel genospecies that could be differentiated from its phylogenetically nearest neighbours by the following phenotypic characteristics: no production of 2-keto-D-gluconic acid from D-glucose; growth on methanol and D-xylose, but not on maltose, as sole carbon sources; no growth on yeast extract with 30% D-glucose; and weak growth at 37 degrees C. The DNA G+C contents of four selected strains were 56.8-58.0 mol%. The results obtained prove that the isolates should be classified as representatives of a novel Acetobacter species, for which the name Acetobacter fabarum sp. nov. is proposed. The type strain is strain 985(T) (=R-36330(T) =LMG 24244(T) =DSM 19596(T)).

Photobacterium kishitanii sp. nov., a luminous marine bacterium symbiotic with deep-sea fishes.

Six representatives of a luminous bacterium commonly found in association with deep, cold-dwelling marine fishes were isolated from the light organs and skin of different fish species. These bacteria were Gram-negative, catalase-positive, and weakly oxidase-positive or oxidase-negative. Morphologically, cells of these strains were coccoid or coccoid-rods, occurring singly or in pairs, and motile by means of polar flagellation. After growth on seawater-based agar medium at 22 degrees C for 18 h, colonies were small, round and white, with an intense cerulean blue luminescence. Analysis of 16S rRNA gene sequence similarity placed these bacteria in the genus Photobacterium. Phylogenetic analysis based on seven housekeeping gene sequences (16S rRNA gene, gapA, gyrB, pyrH, recA, rpoA and rpoD), seven gene sequences of the lux operon (luxC, luxD, luxA, luxB, luxF, luxE and luxG) and four gene sequences of the rib operon (ribE, ribB, ribH and ribA), resolved the six strains as members of the genus Photobacterium and as a clade distinct from other species of Photobacterium. These strains were most closely related to Photobacterium phosphoreum and Photobacterium iliopiscarium. DNA-DNA hybridization values between the designated type strain, Photobacterium kishitanii pjapo.1.1(T), and P. phosphoreum LMG 4233(T), P. iliopiscarium LMG 19543(T) and Photobacterium indicum LMG 22857(T) were 51, 43 and 19 %, respectively. In AFLP analysis, the six strains clustered together, forming a group distinct from other analysed species. The fatty acid C(17 : 0) cyclo was present in these bacteria, but not in P. phosphoreum, P. iliopiscarium or P. indicum. A combination of biochemical tests (arginine dihydrolase and lysine decarboxylase) differentiates these strains from P. phosphoreum and P. indicum. The DNA G+C content of P. kishitanii pjapo.1.1(T) is 40.2 %, and the genome size is approximately 4.2 Mbp, in the form of two circular chromosomes. These strains represent a novel species, for which the name Photobacterium kishitanii sp. nov. is proposed. The type strain, pjapo.1.1(T) (=ATCC BAA-1194(T)=LMG 23890(T)), is a luminous symbiont isolated from the light organ of the deep-water fish Physiculus japonicus.

Acetobacter senegalensis sp. nov., a thermotolerant acetic acid bacterium isolated in Senegal (sub-Saharan Africa) from mango fruit (Mangifera indica L.).

A thermotolerant acetic acid bacterium, designated strain CWBI-B418(T), isolated in Senegal from mango fruit (Mangifera indica), was characterized in detail by means of genotypic and phenotypic methods. The novel strain was strictly aerobic and exhibited optimal growth on YGM medium at 35 degrees C. Cells were Gram-negative, motile and coccoid. The strain was assigned to the genus Acetobacter on the basis of 16S rRNA gene sequence analysis. DNA-DNA hybridization experiments with its phylogenetically closest relatives showed that strain CWBI-B418(T) represented a novel Acetobacter genospecies. The DNA G+C content of strain CWBI-B418(T) was 56.0 mol%. Phenotypic characteristics enabling the differentiation of strain CWBI-B418(T) from phylogenetically related Acetobacter species were: production of 2-keto-D-gluconic acid from D-glucose, but not 5-keto-D-gluconic acid, production of catalase but not oxidase, growth on yeast extract with 30 % d-glucose, growth with ammonium as sole nitrogen source with ethanol as carbon source, utilization of glycerol and ethanol but not maltose or methanol as carbon sources, and growth in the presence of 10 % ethanol. Based on the genotypic and phenotypic data presented, strain CWBI-B418(T) clearly represents a novel Acetobacter species, for which the name Acetobacter senegalensis sp. nov. is proposed. The type strain is CWBI-B418(T) (=LMG 23690(T)=DSM 18889(T)).

Acetobacter ghanensis sp. nov., a novel acetic acid bacterium isolated from traditional heap fermentations of Ghanaian cocoa beans.

Twenty-three acetic acid bacteria, isolated from traditional heap fermentations of Ghanaian cocoa beans, were subjected to a polyphasic taxonomic study. The isolates were catalase-positive, oxidase-negative, Gram-negative rods. They oxidized ethanol to acetic acid and were unable to produce 2-ketogluconic acid, 5-ketogluconic acid and 2,5-diketogluconic acid from glucose; therefore, they were tentatively identified as Acetobacter species. 16S rRNA gene sequencing and phylogenetic analysis confirmed their position in the genus Acetobacter, with Acetobacter syzygii and Acetobacter lovaniensis as their closest phylogenetic neighbours. (GTG)(5)-PCR fingerprinting grouped the strains in a cluster that did not contain any type strains of members of the genus Acetobacter. DNA-DNA hybridization with the type strains of all recognized Acetobacter species revealed DNA-DNA relatedness values below the species level. The DNA G+C contents of three selected strains were 56.9-57.3 mol%. The novel strains had phenotypic characteristics that enabled them to be differentiated from phylogenetically related Acetobacter species, i.e. they were motile, did not produce 2-ketogluconic acid or 5-ketogluconic acid from glucose, were catalase-positive and oxidase-negative, grew on yeast extract with 30 % glucose, grew on glycerol (although weakly) but not on maltose or methanol as carbon sources, and did not grow with ammonium as sole nitrogen source and ethanol as carbon source. Based on the genotypic and phenotypic data, the isolates represent a novel species of the genus Acetobacter for which the name Acetobacter ghanensis sp. nov. is proposed. The type strain is R-29337(T) (=430A(T)=LMG 23848(T)=DSM 18895(T)).

Reclassification of Lactobacillus brevis strains LMG 11494 and LMG 11984 as Lactobacillus parabrevis sp. nov.

A polyphasic study revealed taxonomic heterogeneity among reference strains of the species Lactobacillus brevis. Representative strains of L. brevis and related taxa were investigated by partial sequence analysis of the housekeeping gene encoding the alpha-subunit of phenylalanyl-tRNA synthase (pheS). Species-specific clusters were delineated for all taxa studied except for two L. brevis strains, LMG 11494 and LMG 11984, respectively isolated from cheese and wheat, which occupied a distinct position. Their phylogenetic affiliation was determined using 16S rRNA gene sequence analysis and it was found that both strains (with 99.9 % gene sequence similarity between them) belonged to the Lactobacillus buchneri group, with nearest neighbours Lactobacillus hammesii and L. brevis (gene sequence similarities of 99.2 and 98.1 %, respectively). Further genotypic and phenotypic studies, including fluorescent amplified fragment length polymorphism, DNA-DNA hybridization and DNA G+C content, clearly demonstrated that the two strains represent a single novel taxon for which the name Lactobacillus parabrevis sp. nov. is proposed (type strain LMG 11984(T)=ATCC 53295(T)).

Description of Gluconacetobacter swingsii sp. nov. and Gluconacetobacter rhaeticus sp. nov., isolated from Italian apple fruit.

Two Gram-negative, rod-shaped, non-spore-forming bacteria (DST GL01T and DST GL02T) were isolated from apple fruit juice in the region of the Italian Alps. On the basis of 16S rRNA gene sequence similarities, strains DST GL01T and DST GL02T were shown to belong to the alpha-subclass of the Proteobacteria, and, in particular, to the genus Gluconacetobacter, in the Gluconacetobacter xylinus branch (98.5-100 %). Chemotaxonomic data (major ubiquinone, Q10; predominant fatty acid, C(18 : 1omega7c), accounting for approximately 50 % of the fatty acid content) support the affiliation of both strains to the genus Gluconacetobacter. The results of DNA-DNA hybridizations, together with physiological and biochemical data, allowed genotypic and phenotypic differentiation between strains DST GL01T and DST GL02T and from the 11 validly published Gluconacetobacter species. They therefore represent two new species, for which the names Gluconacetobacter swingsii sp. nov. and Gluconacetobacter rhaeticus sp. nov. are proposed, with the type strains DST GL01T (=LMG 22125T=DSM 16373T) and DST GL02T (=LMG 22126T=DSM 16663T), respectively.

Reclassification of Lactobacillus ferintoshensis as a later heterotypic synonym of Lactobacillus parabuchneri.

Lactobacillus ferintoshensis has recently been described as a novel species, distinct from its close phylogenetic neighbours Lactobacillus buchneri, Lactobacillus kefiri and Lactobacillus hilgardii. Two highly related species with validly published names, Lactobacillus parakefiri and Lactobacillus parabuchneri, were not considered in the study due to the lack of 16S rRNA gene sequence data at that time. Since the publication of the study, the sequences have become available and have revealed that L. ferintoshensis and L. parabuchneri share 99.7% 16S rRNA gene sequence similarity. Further genomic and phenotypic data, derived from fluorescent amplified fragment length polymorphism, DNA-DNA hybridization and API 50 CHL analyses, have demonstrated that the species are synonymous.

Reclassification of [Cytophaga] marinoflava Reichenbach 1989 as Leeuwenhoekiella marinoflava gen. nov., comb. nov. and description of Leeuwenhoekiella aequorea sp. nov.

Five heterotrophic, aerobic, halotolerant and pigmented bacterial strains with gliding motility were isolated from Antarctic sea water; one other isolate was collected from the sea urchin Strongylocentrotus intermedius in the Gulf of Peter the Great in the Sea of Japan. 16S rRNA gene sequence analysis indicated that the strains are members of the family Flavobacteriaceae, the nearest neighbour (with 97.1 % sequence similarity) being the misclassified species [Cytophaga] marinoflava. DNA-DNA hybridization experiments and chemotaxonomic and phenotypic analyses demonstrated that the six novel isolates represent a single species distinct from [C.] marinoflava. On the basis of its separate phylogenetic lineage (the nearest neighbours show 92 % sequence similarity), [C.] marinoflava is reclassified as Leeuwenhoekiella marinoflava gen. nov., comb. nov. A second species of this new genus, Leeuwenhoekiella aequorea sp. nov., is proposed for the six novel isolates, with strain LMG 22550(T) (=CCUG 50091(T)) as the type strain.

Reclassification of Staphylococcus pulvereri Zakrzewska-Czerwinska et al. 1995 as a later synonym of Staphylococcus vitulinus Webster et al. 1994.

A polyphasic taxonomic approach was applied to strains of the species Staphylococcus vitulinus and Staphylococcus pulvereri in order to clarify their taxonomic relatedness. Four reference strains, representing both species, and seven strains isolated from human clinical material were characterized by biotyping, ribotyping and SDS-PAGE analysis of whole-cell proteins, and none of the screening approaches allowed the two taxa to be distinguished. DNA-DNA hybridization experiments between four selected representative strains, including the type strains, confirmed that Staphylococcus pulvereri is a later synonym of Staphylococcus vitulinus.

Vibrio trachuri Iwamoto et al. 1995 is a junior synonym of Vibrio harveyi (Johnson and Shunk 1936) Baumann et al. 1981.

The taxonomic position of Vibrio trachuri was examined through a polyphasic approach using 16S rDNA sequencing, fluorescent amplified fragment length polymorphisms (FAFLP), DNA-DNA hybridization experiments, G+C content of DNA and phenotypical tests. Phylogenetic analysis showed that Vibrio harveyi is the closest neighbour of V. trachuri, sharing about 98.8% similarity in the 16S rDNA gene. Moreover, numerical analysis of the FAFLP patterns revealed that both species have highly related genomes, sharing 55% pattern similarity. DNA-DNA hybridization experiments and G+C content measurements reinforced these results, since V. trachuri and V. harveyi had at least 74% DNA similarity and 44.5-45.2 mol % G+C. Phenotypical features of both species were also very similar, except that V. trachuri utilized itaconic acid, whereas V. harveyi did not. Therefore, it is proposed that the species V. trachuri should be reclassified as V. harveyi.