Alteromonas flava sp. nov. and Alteromonas facilis sp. nov., two novel copper tolerating bacteria isolated from a sea cucumber culture pond in China

Two bacterial strains, P0211^T and P0213^T, were isolated from a sea cucumber culture pond in China. The strains were able to resist high copper levels. These two strains were characterized at the phenotypic, chemotaxonomic, and genomic level. They were completely different colors, but the 16S rRNA genes showed 99.30% similarity. Phylogenetic analysis based on the sequences of the 16S rRNA gene and five housekeeping genes (dnaK, sucC, rpoB, gyrB, and rpoD) supported the inclusion of these strains within the genus Alteromonas, and the two isolated strains formed a group separated from the closest species Alteromonas aestuariivivens KCTC 52655^T. Genomic analyses, including average nucleotide identity (ANIb and ANIm), DNA–DNA hybridization (DDH), and the percentage of conserved proteins (POCP), clearly separated strains P0211^T and P0213^T from the other species within the genus Alteromonas with values below the thresholds for species delineation. The chemotaxonomic features (including fatty acid and polar lipid analysis) of strains P0211^T and P0213^T also confirmed their differentiation from the related taxa.The results demonstrated that strains P0211^T and P0213^T represented two novel species in the genus Alteromonas, for which we propose the names Alteromonas flava sp. nov., type strain P0211^T (= KCTC 62078^T = MCCC 1H00242^T), and Alteromonas facilis sp. nov., type strain P0213^T (= KCTC 62079^T = MCCC 1H00243^T).     

Halomonas borealis sp. nov. and Halomonas niordiana sp. nov., two new species isolated from seawater

Two Gram-negative strains obtained from tank water in a scallop hatchery in Norway, were phenotypically and genotypically characterized in order to clarify their taxonomic position. On the basis of 16S rRNA gene sequence analysis, these isolates, ATF 5.2^T and ATF 5.4^T, were included in the genus Halomonas, being their closest relatives H. smyrnensis and H. taeanensis, with similarities of 98.9% and 97.7%, respectively. Sequence analysis of the housekeeping genes atpA, ftsZ, gyrA, gyrB, mreB, rpoB, rpoD, rpoE, rpoH, rpoN and rpoS clearly differentiated the isolates from the currently described Halomonas species, and the phylogenetic analysis using concatenated sequences of these genes located them in two robust and independent branches. DNA–DNA hybridization (eDDH) percentage, together with average nucleotide identity (ANI), were calculated using the complete genome sequences of the strains, and demonstrate that the isolates constitute two new species of Halomonas, for which the names of Halomonas borealis sp. nov. and Halomonas niordiana sp. nov. are proposed, with type strains ATF 5.2^T (=CECT 9780^T = LMG 31367^T) and ATF 5.4^T (=CECT 9779^T = LMG 31227^T), respectively.     

Photobacterium piscicola sp. nov., isolated from marine fish and spoiled packed cod

Five isolates from marine fish (W3^T, WM, W1S, S2 and S3) and three isolates misclassified as Photobacterium phosphoreum, originating from spoiled modified atmosphere packed stored cod (NCIMB 13482 and NCIMB 13483) and the intestine of skate (NCIMB 192), were subjected to a polyphasic taxonomic study. Phylogenetic analysis of 16S rRNA gene sequences showed that the isolates were members of the genus Photobacterium. Sequence analysis using the gapA, gyrB, pyrH, recA and rpoA loci showed that these isolates formed a distinct branch in the genus Photobacterium, and were most closely related to Photobacterium aquimaris, Photobacterium kishitanii, Photobacterium phosphoreum and Photobacterium iliopiscarium. The luxA gene was present in isolates W3^T, WM, W1S, S2 and S3 but not in NCIMB 13482, NCIMB 13483 and NCIMB 192. AFLP and (GTG)₅-PCR fingerprinting indicated that the eight isolates represented at least five distinct genotypes. DNA–DNA hybridizations revealed 89% relatedness between isolate W3^T and NCIMB 192, and values below 70% with the type strains of the phylogenetically closest species, P. iliopiscarium LMG 19543^T, P. kishitanii LMG 23890^T, P. aquimaris LMG 26951^T and P. phosphoreum LMG4233^T. The strains of this new taxon could also be distinguished from the latter species by phenotypic characteristics. Therefore, we propose to classify this new taxon as Photobacterium piscicola sp. nov., with W3^T (=NCCB 100098^T = LMG 27681^T) as the type strain.     

Caulobacter zeae sp. nov. and Caulobacter radicis sp. nov., novel endophytic bacteria isolated from maize root (Zea mays L.)

Four bacterial strains designated 410^T, 441, 695^T and 736 were isolated from maize root in Beijing, P. R. China. Based on 16S rRNA gene phylogeny, the four strains formed two clusters in the genus Caulobacter. Since strain 441 was a clonal variety of strain 410^T, only three strains were selected for further taxonomic studies. The whole genome average nucleotide identity (ANI) value between strains 410^T and 695^T was 94.65%, and both strains shared less than 92.10% ANI values with their close phylogenetic neighbors Caulobacter vibrioides DSM 9893^T, Caulobacter segnis ATCC 21756^T and Caulobacter flavus CGMCC 1.15093^T. Strains 410^T and 695^T contained Q-10 as the sole ubiquinone and their major fatty acids were C_{16:0, 11-methyl C18:1ω 0}, 11-methyl C_{18: 1}ω7c, summed feature 3 (C_{16:1ω7c and/or C16:1ω 1}ω7c and/or C_{16: 1}ω6c) and summed feature 8 (C_{18:1ω7c and/or C18:1ω 1}ω7c and/or C_{18: 1}ω6c). Their major polar lipids consisted of glycolipids and phosphatidylglycerol, and phenotypic tests differentiated them from their closest phylogenetic neighbors. Based on the results obtained, it is proposed that the three strains represent two novel species, for which the names Caulobacter zeae sp. nov. (type strain 410^T = CGMCC 1.15991 = DSM 104304) and Caulobacter radicis sp. nov. (type strain 695^T = CGMCC 1.16556 = DSM 106792) are proposed.     

Aeromonas piscicola sp. nov., isolated from diseased fish

Four Aeromonas strains (S1.2^T, EO-0505, TC1 and TI 1.1) isolated from moribund fish in Spain showed a restriction fragment length polymorphism (RFLP) pattern related to strains of Aeromonas salmonicida and Aeromonas bestiarum but their specific taxonomic position was unclear. Multilocus sequence analysis (MLSA) of housekeeping genes rpoD, gyrB, recA and dnaJ confirmed the allocation of these isolates to an unknown genetic lineage within the genus Aeromonas with A. salmonicida, A. bestiarum and Aeromonas popoffii as the phylogenetically nearest neighbours. Furthermore, a strain biochemically labelled as Aeromonas hydrophila (AH-3), showing a pattern of A. bestiarum based on 16S rDNA-RFLP, also clustered with the unknown genetic lineage. The genes rpoD and gyrB proved to be the best phylogenetic markers for differentiating these isolates from their neighbouring species. Useful phenotypic features for differentiating the novel species from other known Aeromonas species included their ability to hydrolyze elastin, produce acid from l-arabinose and salicin, and their inability to produce acid from lactose and use l-lactate as a sole carbon source. A polyphasic approach using phenotypic characterization, phylogenetic analysis of the 16S rRNA gene and of four housekeeping genes, as well as DNA–DNA hybridization studies and an analysis of the protein profiles by MALDI-TOF-MS, showed that these strains represented a novel species for which the name Aeromonas piscicola sp. nov. is proposed with isolate S1.2^T (=CECT 7443^T, =LMG 24783^T) as the type strain.     

Salinivibrio kushneri sp. nov., a moderately halophilic bacterium isolated from salterns

Ten Gram-strain-negative, facultatively anaerobic, moderately halophilic bacterial strains, designated AL184^T, IB560, IB563, IC202, IC317, MA421, ML277, ML318, ML328A and ML331, were isolated from water ponds of five salterns located in Spain. The cells were motile, curved rods and oxidase and catalase positive. All of them grew optimally at 37 °C, at pH 7.2–7.4 and in the presence of 7.5% (w/v) NaCl. Based on phylogenetic analyses of the 16S rRNA, the isolates were most closely related to Salinivibrio sharmensis BAG^T (99.6–98.2% 16S rRNA gene sequence similarity) and Salinivibrio costicola subsp. costicola ATCC 35508^T (99.0–98.1%). According to the MLSA analyses based on four (gyrB, recA, rpoA and rpoD) and eight (ftsZ, gapA, gyrB, mreB, pyrH, recA, rpoA and topA) concatenated gene sequences, the most closely relatives were S. siamensis JCM 14472^T (96.8–95.4% and 94.9–94.7%, respectively) and S. sharmensis DSM 18182^T (94.0–92.6% and 92.9–92.7%, respectively). In silico DNA–DNA hybridization (GGDC) and average nucleotide identity (ANI) showed values of 23.3–44.8% and 80.2–91.8%, respectively with the related species demonstrating that the ten isolates constituted a single novel species of the genus Salinivibrio. Its pangenome and core genome consist of 6041 and 1230 genes, respectively. The phylogeny based on the concatenated orthologous core genes revealed that the ten strains form a coherent phylogroup well separated from the rest of the species of the genus Salinivibrio. The major cellular fatty acids of strain AL184^T were C_16:0 and C_18:1. The DNA G + C content range was 51.9–52.5 mol% (T_m) and 50.2–50.9 mol% (genome). Based on the phylogenetic-phylogenomic, phenotypic and chemotaxonomic data, the ten isolates represent a novel species of the genus Salinivibrio, for which the name Salinivibrio kushneri sp. nov. is proposed. The type strain is AL184^T (= CECT 9177^T = LMG 29817^T).     

Flavobacterium circumlabens sp. nov. and Flavobacterium cupreum sp. nov., two psychrotrophic species isolated from Antarctic environmental samples

A taxonomic study of 24 Gram-stain-negative rod-shaped bacteria originating from the Antarctic environment is described. Phylogenetic analysis using 16S rRNA gene sequencing differentiated isolated strains into two groups belonging to the genus Flavobacterium. Group I (n = 20) was closest to Flavobacterium aquidurense WB 1.1-56^T (98.3% 16S rRNA gene sequence similarity) while group II (n = 4) showed Flavobacterium hydatis DSM 2063^T as its nearest neighbour (98.5–98.9% 16S rRNA gene sequence similarity). Despite high 16S rRNA gene sequence similarity, these two groups represented two distinct novel species as shown by phenotypic traits and low genomic relatedness assessed by rep-PCR fingerprinting, DNA-DNA hybridization and whole-genome sequencing. Common to representative strains of both groups were the presence of major menaquinone MK-6 and sym-homospermidine as the major polyamine. Common major fatty acids were C_15:0 iso, C_15:1 iso G, C_15:0 iso 3-OH, C_17:0 iso 3OH and Summed Feature 3 (C_16:1 ω7c/C_16:1 ω6c). Strain CCM 8828^T (group I) contained phosphatidylethanolamine, three unidentified lipids lacking a functional group, three unidentified aminolipids and single unidentified glycolipid in the polar lipid profile. Strain CCM 8825^T (group II) contained phosphatidylethanolamine, eight unidentified lipids lacking a functional group, three unidentified aminolipids and two unidentified glycolipids in the polar lipid profile. These characteristics corresponded to characteristics of the genus Flavobacterium. The obtained results showed that the analysed strains represent novel species of the genus Flavobacterium, for which the names Flavobacterium circumlabens sp. nov. (type strain CCM 8828^T = P5626^T = LMG 30617^T) and Flavobacterium cupreum sp. nov. (type strain CCM 8825^T = P2683^T = LMG 30614^T) are proposed.     

Photobacterium carnosum sp. nov., isolated from spoiled modified atmosphere packaged poultry meat

Analysis of spoilage-associated microbiota of modified-atmosphere packaged poultry meat revealed four different bacterial isolates that could not be assigned to known species. They showed a Gram-negative staining behavior, were facultatively aerobic, non-motile with variable cell morphology. Phylogenetic analysis of 16S rDNA and gyrB, rpoD and recA revealed a distinct lineage within the genus Photobacterium with Photobacterium (P.) iliopiscarium DSM 9896^T, P. phosphoreum DSM 15556^T, P. kishitanii DSM 19954^T, P. piscicola LMG 27681^T and P. aquimaris DSM 23343^T as closest relatives.The designated type strain TMW 2.2021^T is non-luminous and grew at 0–20 °C (optimum 10–15 °C), within pH 5.0–8.5 (optimum 6–8) and in the presence of 0.5–3% (w/v) NaCl (optimum 1%). Major cellular fatty acids of TMW 2.2021^T were summed feature 3 (C_16:1ω7c/iso-C₁₅ 3-OH), C_16:0, C_18:1ω7c and summed feature 2 (C_12:0 aldehyde and C_10.928 unknown). Quinone analysis revealed Q-8 as sole respiratory ubiquinone. The genome of TMW 2.2021^T has a size of 4.56 Mb and a G + C content of 38.49 mol%. The ANI value between TMW 2.2021^T and the type strain of closest relative P. iliopiscarium DSM 9896^T was 91.43%. Fingerprinting on the base of M13-RAPD-PCR band pattern and MALDI-TOF MS profiles allowed intraspecies differentiation between our isolates but also supported their distinct lineage to a novel species. Based on phylogenetic, genomic, phenotypic and chemotaxonomic data, strain TMW 2.2021^T and further strains represent a novel species of the genus Photobacterium, for which the name Photobacterium carnosum sp. nov. is proposed. The type strain is TMW 2.2021^T (=DSM 105454^T = CECT 9394^T).     

Description of Tropicibacter mediterraneus sp. nov. and Tropicibacter litoreus sp. nov.

Four strains (M15∅_3, M17^T, M49 and R37^T) were isolated from Mediterranean seawater at Malvarrosa beach, Valencia, Spain. Together with an older preserved isolate (strain 2OM6) from cultured oysters at Vinaroz, Castellón, Spain, the strains were thoroughly characterized in a polyphasic study and were placed phylogenetically within the Roseobacter clade in the family Rhodobacteraceae. Highest 16S rRNA sequence similarities of the five strains to the types of any established species corresponded to Tropicibacter multivorans (95.8–96.4%), Phaeobacter inhibens (95.9–96.3%) and Phaeobacter gallaeciensis (95.9–96.2%). On the other hand, whole genome (ANI) and protein fingerprinting (MALDI-TOF) data proved: (i) non clonality among the strains, and (ii) the existence of two genospecies, one consisting of strains M15∅_3, M17^T, M49 and 2OM6 and another one consisting of strain R37^T. Phenotypic traits determined allow differentiating both genospecies from each other and from closely related taxa. In view of all data collected we propose to accommodate these isolates in two species as members of the genus Tropicibacter, Tropicibacter mediterraneus sp. nov. (type strain M17^T = CECT 7615^T = KCTC 23058^T) and Tropicibacter litoreus sp. nov. (type strain R37^T = CECT 7639^T = KCTC 23353^T).     

Kocuria uropygioeca sp. nov. and Kocuria uropygialis sp. nov., isolated from the preen glands of Great Spotted Woodpeckers (Dendrocopos major)

Two new species of Gram-positive cocci were isolated from the uropygial glands of wild woodpeckers (Dendrocopos major) originating from different locations in Germany. A polyphasic approach confirmed the affiliation of the isolates to the genus Kocuria. Phylogenetic analysis based on the 16S rRNA gene showed high degree of similarity to Kocuria koreensis DSM 23367^T (99.0% for both isolates). However, low ANIb values of <80% unequivocally separated the new species from K. koreensis. This finding was further corroborated by DNA fingerprinting and analysis of polar lipid profiles. Furthermore, growth characteristics, biochemical tests, MALDI-TOF MS analysis, and G + C contents clearly differentiated the isolates from their known relatives. Besides, the woodpecker isolates significantly differed from each other in their whole-cell protein profiles, DNA fingerprints, and ANIb values. In conclusion, the isolated microorganisms constitute members of two new species, for which the names Kocuria uropygioeca sp. nov. and Kocuria uropygialis sp. nov. are proposed. The type strains are 36^T (DSM 101740^T = LMG 29265^T) and 257^T (=DSM 101741^T = LMG 29266^T) for K. uropygialis sp. nov. and K. uropygioeca sp. nov., respectively.     

Arcobacter cloacae sp. nov. and Arcobacter suis sp. nov., two new species isolated from food and sewage

Three strains recovered from mussels (F26), sewage (SW28-13^T) and pork meat (F41^T) were characterized as Arcobacter. They did not appear to resemble any known species on the basis of their 16S rDNA-RFLP patterns and the rpoB gene analyses. However, strains F26 and SW28-13^T appeared to be the same species. The 16S rRNA gene sequence similarity of strains SW28-13^T and F41^T to the type strains of all other Arcobacter species ranged from 94.1% to 99.6% and 93.4% to 98.8%, respectively. Phenotypic characteristics and the DNA–DNA hybridization (DDH) results showed that they belonged to 2 new Arcobacter species. A multilocus phylogenetic analysis (MLPA) with the concatenated sequences of 5 housekeeping genes (gyrA, atpA, rpoB, gyrB and hsp60) was used for the first time in the genus, showing concordance with the 16S rRNA gene phylogenetic analysis and DDH results. The MALDI-TOF mass spectra also discriminated these strains as two new species. The names proposed for them are Arcobacter cloacae with the type strain SW28-13^T (=CECT 7834^T = LMG 26153^T) and Arcobacter suis with the type strain F41^T (=CECT 7833^T = LMG 26152^T).     

Methyloligella halotolerans gen. nov., sp. nov. and Methyloligella solikamskensis sp. nov., two non-pigmented halotolerant obligately methylotrophic bacteria isolated from the Ural saline environments

Two newly isolated halotolerant obligately methylotrophic bacteria (strains C2^T and SK12^T) with the serine pathway of C₁ assimilation are described. The isolates are strictly aerobic, Gram negative, asporogenous, non-motile rods, forming rosettes, multiplying by binary fission. Mesophilic and neutrophilic, accumulate intracellularly compatible solute ectoine and poly-β-hydroxybutyrate. The novel strains are able to grow at 0 up to 16% NaCl (w/v), optimally at 3–5% NaCl. The major cellular fatty acids are C_18:1ω7c and C_19:0cyc and the prevailing quinone is Q-10. The predominant phospholipids are phosphatidylcholine, phosphatidylglycerol, phosphatidyldimethylethanolamine and phosphatidylethanolamine. Assimilate NH₄⁺ by glutamate dehydrogenase and via the glutamate cycle (glutamine synthetase and glutamate synthase). The DNA G + C contents of strains C2^T and SK12^T are 60.9 and 60.5 mol% (Tm), respectively. 16S rRNA gene sequence similarity between the two new isolates are 99% but below 94% with other members of the Alphaproteobacteria thus indicating that they can be assigned to a novel genus Methyloligella. Rather low level of DNA–DNA relatedness (53%) between the strains C2^T and SK12^T indicates that they represent two separate species of the new genus, for which the names Methyloligella halotolerans gen. nov., sp. nov. and Methyloligella solikamskensis sp. nov. are proposed. The type strain of Methyloligella halotolerans is C2^T (=VKM B-2706^T = CCUG 61687^T = DSM 25045^T) and the type strain of Methyloligella solikamskensis is SK12^T (=VKM B-2707^T = CCUG 61697^T = DSM 25212^T).     

Marinifilum flexuosum sp. nov., a new Bacteroidetes isolated from coastal Mediterranean Sea water and emended description of the genus Marinifilum Na et al., 2009

A facultatively anaerobe, moderately halophilic, Gram-negative, filamentous, non motile and unpigmented bacterium, designated M30^T, was isolated from coastal Mediterranean Sea water in Valencia, Spain. Phylogenetic analysis based on 16S rRNA sequences placed this strain in the phylum “Bacteroidetes” with Marinifilum fragile JC2469^T as its closest relative with 97% sequence similarity. Average nucleotide identity (ANI) values between both strains were far below the 95% threshold value for species delineation (about 89% using BLAST and about 90% using MUMmer). A comprehensive polyphasic study, including morphological, biochemical, physiological, chemotaxonomic and phylogenetic data, confirmed the independent species status of strain M30^T within the genus Marinifilum, for which the name Marinifilum flexuosum sp. nov. is proposed. The type strain of Marinifilum flexuosum is M30^T (=CECT 7448^T = DSM 21950^T).     

Photobacterium marinum sp. nov., a marine bacterium isolated from a sediment sample from Palk Bay,India

The novel, cream colored, Gram-staining-negative, rod-shaped, motile bacteria, designated strains AK15^T and AK18, were isolated from sediment samples collected from Palk Bay, India. Both strains were positive for arginine dihydrolase, lysine decarboxylase, oxidase, nitrate reduction and methyl red test. The major fatty acids were C_16:0, C_{18:1 ω7c}, C_{16:1 ω7c} and/or C_{16:1 ω6c} and/or iso-C_15:0 2-OH (summed feature 3). Polar lipids content of strains AK15^T and AK18 were found to bephosphatidylethanolamine (PE), two unidentified phospholipids (PL1 and PL2) and three unidentified lipids (L1-L3). The 16S rRNA gene sequence analysis indicated strains AK15^T and AK18 as the members of the genus Photobacterium and closely related to the type strain Photobacterium jeanii with pair-wise sequence similarity of 96.7%. DNA–DNA hybridization between strain AK15^T and AK18 showed a relatedness of 87%. Based on data from the current polyphasic study, strains AK15^T and AK18 are proposed as novel species of the genus Photobacterium, for which the name Photobacterium marinum sp. nov. is proposed. The type strain of Photobacterium marinum is AK15^T (=MTCC 11066^T = DSM 25368^T).     

Arcobacter molluscorum sp. nov., a new species isolated from shellfish

Nineteen bacteria isolates recovered from shellfish samples (mussels and oysters) showed a new and specific 16S rDNA-RFLP pattern with an Arcobacter identification method designed to recognize all species described up to 2008. These results suggested that they could belong to a new species. ERIC-PCR revealed that the 19 isolates belonged to 3 different strains. The sequence of the 16S rRNA gene of a representative strain (F98-3^T) showed 97.6% similarity with the closest species Arcobacter marinus followed by Arcobacter halophilus (95.6%) and Arcobacter mytili (94.7%). The phylogenetic analysis with the16S rRNA, rpoB, gyrB and hsp60 genes placed the shellfish strains within the same cluster as the three species mentioned (also isolated from saline habitats) but they formed an independent phylogenetic line. The DDH results between strain F98-3^T and A. marinus (54.8% ± 1.05), confirmed that it represents a new species. Several biochemical tests differentiated the shellfish isolates from all other Arcobacter species. Although the new species was different from A. mytili, they shared not only the same habitat (mussels) but also the characteristic of being so far the only Arcobacter species that are simultaneously negative for urea and indoxyl acetate hydrolysis. All results supported the classification of the shellfish strains as a new species, for which the name Arcobacter molluscorum sp. nov. with the type strain F98-3^T is proposed (=CECT 7696^T = LMG 25693^T).     

Pseudomonas caspiana sp. nov., a citrus pathogen in the Pseudomonas syringae phylogenetic group

In a screening by multilocus sequence analysis of Pseudomonas strains isolated from diverse origins, 4 phylogenetically closely related strains (FBF58, FBF102^T, FBF103, and FBF122) formed a well-defined cluster in the Pseudomonas syringae phylogenetic group. The strains were isolated from citrus orchards in northern Iran with disease symptoms in the leaves and stems and its pathogenicity against citrus plants was demonstrated. The whole genome of the type strain of the proposed new species (FBF102^T = CECT 9164^T = CCUG 69273^T) was sequenced and characterized. Comparative genomics with the 14 known Pseudomonas species type strains of the P. syringae phylogenetic group demonstrated that this strain belonged to a new genomic species, different from the species described thus far. Genome analysis detected genes predicted to be involved in pathogenesis, such as an atypical type 3 secretion system and two type 6 secretion systems, together with effectors and virulence factors. A polyphasic taxonomic characterization demonstrated that the 4 plant pathogenic strains represented a new species, for which the name Pseudomonas caspiana sp. nov. is proposed.     

Agrobacterium rosae sp. nov., isolated from galls on different agricultural crops

The plant tumorigenic strain NCPPB 1650^T isolated from Rosa × hybrida, and four nonpathogenic strains isolated from tumors on grapevine (strain 384), raspberry (strain 839) and blueberry (strains B20.3 and B25.3) were characterized by using polyphasic taxonomic methods. Based on 16S rRNA gene phylogeny, strains were clustered within the genus Agrobacterium. Furthermore, multilocus sequence analysis (MLSA) based on the partial sequences of atpD, recA and rpoB housekeeping genes indicated that five strains studied form a novel Agrobacterium species. Their closest relatives were Agrobacterium sp. R89-1, Agrobacterium rubi and Agrobacterium skierniewicense. Authenticity of the novel species was confirmed by average nucleotide identity (ANI) and in silico DNA–DNA hybridization (DDH) comparisons between strains NCPPB 1650^T and B20.3, and their closest relatives, since obtained values were considerably below the proposed thresholds for the species delineation. Whole-genome-based phylogeny further supported distinctiveness of the novel species, that forms together with A. rubi, A. skierniewicense and Agrobacterium sp. R89-1 a well-delineated sub-clade of Agrobacterium spp. named “rubi”. As for other species of the genus Agrobacterium, the major fatty acid of the strains studied was 18:1 w7c (73.42–78.12%). The five strains studied were phenotypically distinguishable from other species of the genus Agrobacterium. Overall, polyphasic characterization showed that the five strains studied represent a novel species of the genus Agrobacterium, for which the name Agrobacterium rosae sp. nov. is proposed. The type strain of A. rosae is NCPPB 1650^T (=DSM 30203^T = LMG 230^T = CFBP 4470^T = IAM 13558^T = JCM 20915^T).     

Pseudomonas gallaeciensis sp. nov., isolated from crude-oil-contaminated intertidal sand samples after the Prestige oil spill

Strains V113^T, V92 and V120 have been isolated from sand samples taken at the Atlantic intertidal shore in Galicia, Spain, after the Prestige oil spill. A preliminary analysis of the 16S rRNA and the partial rpoD gene sequences indicated that these strains belonged to the Pseudomonas genus, but they were distinct from any known Pseudomonas species. They were extensively characterized by a polyphasic taxonomic approach and phylogenetic data that confirmed that these strains belonged to the Pseudomonas pertucinogena group. Phylogenetic analysis of 16S rRNA, gyrB and rpoD gene sequences showed that the three strains were 99% similar and were closely related to members of the P. pertucinogena group, with less than 94% similarity to strains of established species; Pseudomonas pachastrellae was the closest relative. The Average Nucleotide Index based on blast values was 89.0% between V113^T and the P. pachastrellae type strain, below the accepted species level (95%). The predominant cellular fatty acid contents and whole cell protein profiles determined by MALDI-TOF mass spectrometry also differentiated the studied strains from known Pseudomonas species. We therefore conclude that strains V113^T, V92 and V120 represent a novel species of Pseudomonas, for which the name Pseudomonas gallaeciensis is proposed; the type strain is V113^T (= CCUG 67583^T = LMG 29038^T).     

Pseudocitrobacter gen. nov., a novel genus of the Enterobacteriaceae with two new species Pseudocitrobacter faecalis sp. nov., and Pseudocitrobacter anthropi sp. nov,isolated from fecal samples from hospitalized patients in Pakistan

Four isolates of Gram-negative facultatively anaerobic bacteria, three of them producing NDM-1 carbapenemase, were isolated from hospitalized patients and outpatients attending two military hospitals in Rawalpindi, Pakistan, and studied for their taxonomic position. Initially the strains were phenotypically identified as Citrobacter species. Comparative analysis of 16S rRNA gene sequences then showed that the four strains shared >97%, but in no case >98.3%, 16S rRNA gene sequence similarities to members of the genera Citrobacter, Kluyvera, Pantoea, Enterobacter and Raoultella, but always formed a separate cluster in respective phylogenetic trees. Based on multilocus sequence analysis (MLSA) including partial recN, rpoA, thdF and rpoB gene sequence and respective amino acid sequence analysis it turned out that the strains also here always formed separate clusters. Based on further comparative analyses including DNA–DNA hybridizations, genomic fingerprint analysis using rep- and RAPD-PCRs and physiological tests, it is proposed to classify these four strains into the novel genus Pseudocitrobacter gen. nov. with a new species Pseudocitrobacter faecalis sp. nov. with strain 25 CIT^T (= CCM 8479^T = LMG 27751^T) and Pseudocitrobacter anthropi sp. nov. with strain C138^T (= CCM 8478^T = LMG 27750^T), as the type strains, respectively.