gyrB Gene Sequences Research Articles

Rheinheimera salexigens sp. nov., isolated from a fishing hook, and emended description of the genus Rheinheimera.

A Gram-negative, rod-shaped bacterium, designated KH87T, was isolated from a fishing hook that had been baited and suspended in seawater off O'ahu, Hawai'i. Based on a comparison of 1524 nt of the 16S rRNA gene sequence of strain KH87T, its nearest neighbours were the GammaproteobacteriaRheinheimera nanhaiensis E407-8T (96.2 % identity), Rheinheimera chironomi K19414T (96.0 %), Rheinheimera pacifica KMM 1406T (95.8 %), Rheinheimera muenzenbergensis E49T (95.7 %), Alishewanella solinquinati KMK6T (94.9 %) and Arsukibacterium ikkense GCM72T (94.6 %). Cells of KH87T were motile by a single polar flagellum, strictly aerobic, and catalase- and oxidase-positive. Growth occurred between 4 and 39 °C, and in a circumneutral pH range. Major fatty acids in whole cells of strain KH87T were cis-9-hexadecenoic acid, hexadecanoic acid and cis-11-octadecenoic acid. The quinone system contained mostly menaquinone MK-7, and a minor amount of ubiquinone Q-8. The polar lipid profile contained the major lipids phosphatidylglycerol, phosphatidylserine, phosphatidylethanolamine, an unidentified aminolipid, and a lipid not containing phosphate, an amino group or a sugar moiety. Putrescine was the major polyamine. Physiological, biochemical and genomic data, including obligate halophily, absence of amylolytic activity, a quinone system dominated by MK-7 and DNA G+C content (42.0 mol%) distinguished KH87T from extant Rheinheimera species; strain KH87T was also distinguished by a multi-locus sequence analysis of aligned and concatenated 16S rRNA, gyrB, rpoB and rpoD gene sequences. Based on phenotypic and genotypic differences, the species Rheinheimera salexigens sp. nov. is proposed to accommodate KH87T as the type strain (=ATCC BAA-2715T=CIP 111115T). An emended description of the genus Rheinheimera is also proposed.

FIRST REPORT OF PSEUDOMONAS VIRIDIFLAVA CAUSING CABBAGE BACTERIAL LEAF SPOT IN TURKEY

In the autumn of 2016, white head cabbage (Brassica oleracea var. capitata f. alba) showing circular to irregular lesions 2–3 mm in size surrounded by a yellow halo and necrotic areas on the leaves, were observed in the Samsun province of Turkey. Extract from symptomatic tissue macerated in plastic bags (Bioreba AG, Switzerland), plated on King’s medium B (King et al., 1954) and incubated at 26°C for 48 h yielded colonies with the same LOPAT profile of Pseudomonas viridiflava strain M66 from tomato. They were levan-, oxidase-, arginine-negative, caused hypersensitive reactions on tobacco leaves (Nicotiana tabacum cv. Xanthi) and macerated potato slices. Molecular characterization using ERIC-PCR (Rademaker et al., 1998) yielded the same band pattern for strains BT5X and M66. Blastn analysis of a 1400 bp 16S rDNA and a 835 bp gyrB gene sequence (GenBank accession Nos. KY769809, MF314821) showed 99% and 100% similarity to the sequences of the type strain ATCC13223T =CFBP 2107 (NR_114482- HM190239) of Ps. viridiflava deposited in GenBank. Multilocus sequence analysis (MLSA) produced by maximum likelihood method using concatenated sequences of 16S rDNA and gyrB allowed clustering of our strain in the same clade together with the type strain ATCC13223T and other representive P. viridiflava strains retrieved from GenBank. Cabbage plants (cv. Beyaz Bursa) were spray-inoculated with a suspension of 108 CFU ml-1 of an overnight culture grown in Luria-Bertani broth reacted six days post inoculation with spots similar to those observed in the field. Reisolated colonies were fluorescent under UV light and had the same morphology on King’s medium B as the original culture and positive strain M66. To our knowledge, this is the first report of P. viridiflava causing a leaf spot disease of head cabbage in Turkey.

Nocardia heshunensis sp. nov., an actinomycete isolated from soil

A novel Gram-stain-positive, aerobic, non-motile and acid-fast actinomycete strain, designated CFH S0067T, was isolated from a soil sample collected from Heshun old town in Tengchong, Yunnan province, in south-west PR China. The taxonomic position of strain CFH S0067T was studied in detail using a polyphasic approach. Phylogenetic analysis, based on 16S rRNA gene sequences, revealed that strain CFH S0067T belongs to the genus Nocardia and is closely related to Nocardia concava JCM 12351T (99.3 % similarity), forming a separated branch with this type strain. However, the strain shared 96.0 % gyrB gene sequence similarity with N. concava JCM 12351T. Furthermore, DNA-DNA hybridization showed 56.5±0.6 % DNA relatedness between the novel strain and N. concava JCM 12351T. The whole-cell hydrolysates contained meso-diaminopimelic acid (type IV) and arabinose, galactose, fructose and mannose. The major polar lipids were diphosphatidylglycerol, phosphatidylethanolamine, phosphatidylinositol, phosphatidylinositol mannoside and one unidentified lipid. Strain CFH S0067T contained MK-8 (H4ω-cycl) as the predominant menaquinone. C16 : 0, summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c), C18 : 1ω9c and C18 : 0 10-methyl (TBSA) were the major cellular fatty acids. Mycolic acids were also detected. The G+C content of the genomic DNA was determined to be 66.9 mol%. A combination of the low DNA-DNA hybridization values and phenotypic properties demonstrated that strain CFH S0067Tis clearly distinguishable from its most closely related strain, N. concava JCM 12351T. On the basis of this polyphasic study, it is concluded that strain CFH S0067T should be considered to represent a novel species of the genus Nocardia, for which the name Nocardia heshunensis sp. nov. is proposed. The type strain is CFH S0067T (=DSM 46764T=JCM 30085T).

Dyella caseinilytica sp. nov., Dyella flava sp. nov. and Dyella mobilis sp. nov., isolated from forest soil.

Three strains, DHOB09T, DHOC52T and DHON07T, were isolated from the forest soil of Dinghushan Biosphere Reserve, Guangdong Province, PR China. They were all Gram-stain-negative, aerobic, rod-shaped cells. The ranges (optimum) for the temperature, pH and NaCl concentration for growth of DHOB09T, DHOC52T and DHON07T were 10-42 (25-28) °C, pH 5.5-9.0 (7.0-7.5) and 0-4.0 (0-0.5) % (w/v); 10-42 (28) °C, pH 4.0-7.0 (4.5-6.5) and 0-2.0 (0) % (w/v) and 10-37 (25-28) °C, pH 4.0-7.5 (5.5-6.0) and 0-2.5 (0) % (w/v), respectively. Phylogenetic analysis based on 16S rRNA gene sequences showed that DHOB09T, DHOC52T and DHON07T formed a phyletic cluster with seven species of the genus Dyella within the major clade of Dyella with sequence similarities ranged from 96.9 to 98.6 %. This indicated that the three strains may represent three novel species of the genus Dyella. This result was also strongly supported by the concatenated analysis of partial gyrB, lepA and recA gene sequences. DNA-DNA hybridization between strains DHON07T and DHOB09T, as well as DHON07T and Dyella koreensis BB4T was much lower than 70 %. The G+C content of strains DHOB09T, DHOC52T and DHON07T were 59.4, 60.7 and 59.5 %, respectively. The major fatty acids of the three strains were iso-C15 : 0, iso-C16 : 0 and iso-C17 : 0 and the predominant respiratory lipoquinone was ubiquinone-8. All of the physiological, phylogenetic and chemotaxonomic data showed that strains DHOB09T, DHOC52T and DHON07T are distinctive from each other and from all species of the genus Dyellawith validly published names. Therefore, we suggest that they represent three novel species of the genus, for which the names Dyella caseinilytica sp. nov. (type strain DHOB09T=CGMCC 1.15434T=LMG 29202T), Dyella flava sp. nov. (type strain DHOC52T=NBRC 111979T=KCTC 52128T) and Dyella mobilis sp. nov. (type strain DHON07T=CGMCC 1.15400T=NBRC 111475T) are proposed.

Symbiotic bacteria associated with gut symbiotic organs and female genital accessory organs of the leaf beetle Bromius obscurus (Coleoptera: Chrysomelidae)

Leaf beetles (Coleoptera: Chrysomelidae) constitute one of the most species-rich insect families, and live exclusively on leaves or other plant parts. Early histological works described the presence of symbiotic bacteria in gut-associated symbiotic organs of some chrysomelid species, but their microbiological nature has been poorly characterized except for those associated with reed beetles of the subfamily Donaciinae. Here we investigated symbiotic bacteria of the leaf beetle Bromius obscurus (L.) belonging to the subfamily Eumolpinae. Specific bacterial 16S ribosomal RNA and gyrB gene sequences were consistently obtained from the symbiotic organs, which radially surround the foregut-midgut junction, of all adult males and females examined. In adult females, the same sequences were also obtained from a pair of genital accessory organs, which are presumably for vertical symbiont transmission. Whole mount in situ hybridization specifically detected the symbiont in the gut symbiotic organs endocellularly and also in the female genital accessory organs extracellularly. In the gut symbiotic organs, the endocellular symbiont cells were small and rosette-like or aggregated and granule-like, whereas in the female genital organs the extracellular symbiont cells were of a condensed form. Molecular phylogenetic analysis showed that the symbiont of B. obscurus constitutes a distinct lineage in the Gammaproteobacteria. Molecular evolutionary analysis has identified significantly accelerated molecular evolution and a highly adenine–thymine-biased nucleotide composition of the symbiont genes, presumably reflecting reductive evolution of the symbiont genome. These results suggest an intimate and stable host-symbiont association in B. obscurus, in which the symbiont may play some important, though hitherto unknown, biological roles in its herbivorous insect host.

Arcobacter haliotis sp. nov., isolated from abalone species Haliotis gigantea.

A Gram-negative, aerobic, polar-flagellated and rod-shaped, sometimes slightly curved bacterium, designated MA5T, was isolated from the gut of an abalone of the species Haliotis gigantea collected in Japan. Phylogenetic analyses based on 16S rRNA, gyrB, hsp60 and rpoB gene sequences placed strain MA5T in the genus Arcobacter in an independent phylogenetic line. Comparison of the 16S rRNA gene sequence of this strain with those of the type strains of the established Arcobacter species revealed A. nitrofigilis (95.1 %) as nearest neighbour. Strain MA5T grew optimally at 25 °C, pH 6.0 to 9.0 and in the presence of 2 to 5 % (w/v) NaCl under both aerobic and microaerobic conditions. The predominant fatty acids found were summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1 ω7c), C12 : 0 3-OH and C18 : 1 ω7c. Menaquinone-6 (MK-6) and menaquinone-7 (MK-7) were found as the major respiratory quinones. The major polar lipids detected were phosphatidylethanolamine and phosphatidylglycerol. Strain MA5T could be differentiated phenotypically from the phylogenetic closest Arcobacter species by its ability to grow on 0.05 % safranin and 0.01 % 2,3,5-triphenyl tetrazolium chloride (TTC), but not on 0.5 % NaCl. The obtained DNA G+C content of strain MA5T was 27.9 mol%. Based on the phylogenetic, chemotaxonomic and phenotypic distinctiveness of MA5T, this strain is considered to represent a novel species of the genus Arcobacter, for which the name Arcobacter haliotis sp. nov. is proposed. The type strain is MA5T (=LMG 28652T=JCM 31147T).

Serratia oryzae sp. nov., isolated from rice stems.

A novel endophytic bacterium, strain J11-6T, was isolated from rice stems. Its taxonomic position was investigated using a polyphasic approach. The novel strain was Gram-staining-negative, facultatively anaerobic, motile and rod-shaped. Although the results of phylogenetic analysis based on 16S rRNA gene sequences indicated that J11-6T represented a member of the genus Rahnella, multilocus sequence analysis (MLSA) on the basis of concatenated partial atpD, gyrB, rpoB and infB gene sequences showed a clear distinction of J11-6T from the type strains of species of the genus Rahnella but indicated that it lay within the clade of the genus Serratia. The phylogenetically closest species were Serratia fonticola and Serratia aquatilis on the basis of the results of the MLSA phylogenetic analysis. The predominant cellular fatty acids were C16 : 1ω7c (38.7 %) and C16 : 0 (25.0 %). The DNA G+C content was 53.2 mol%. The DNA-DNA relatedness was 17.4 % between J11-6T and Rahnella aquatilis CIP 78.65T, and 29.2 % between J11-6T and S. fonticola LMG 7882T which indicates that this strain represents a novel species of the genus Serratia. Characterization by genotypic and phenotypic analysis indicated that J11-6T (=ACCC 19934T=KCTC 52529T) represents a novel species of the genus Serratia, for which the name Serratia oryzae sp. nov. is proposed.

Arenimonas soli sp. nov., isolated from saline-alkaline soil.

A Gram-staining-negative, non-motile, aerobic bacterial strain, designated Y3L17T, was isolated from the saline-alkaline soil of a farmland, Hangjin Banner, Inner Mongolia, northern China. Y3L17T could grow at 15-45 °C (optimum 35 °C), pH 6.0-10.0 (optimum pH 8.0) and with 0-4 % (w/v) NaCl (optimum 0 %). The results of phylogenetic analysis based on the 16S rRNA gene and gyrB gene sequences revealed that Y3L17T tightly clustered with strains of members of the genus Arenimonas, sharing the highest 16S rRNA gene similarities with Arenimonas aestuarii S2-21T (99.5 %) and Arenimonas donghaensis HO3-R19T (98.2 %), and lower similarities (<97 %) with all the other type strains of species of this genus. However, Y3L17T shared only 92.62 % gyrB gene similarities with A. aestuarii S2-21T. The DNA-DNA hybridization values of Y3L17T with A. aestuariiS2-21T and A. donghaensis HO3-R19T were 20.1±2.5 and 18.2±3.2 %, respectively. Y3L17T contained phosphatidylethanolamine, diphosphatidylglycerol, phosphatidylglycerol, five unknown phospholipids and one unknown lipid as the major polar lipids. Ubiquinone-8 (Q-8) was the predominant respiratory quinone, while iso-C15 : 0, iso-C17 : 0ω9c and iso-C11 : 0 3-OH were the major cellular fatty acids. Its genomic DNA G+C content was 65.4 mol%. On the basis of its phenotypic, phylogenetic and genotypic characteristics, Y3L17T represents a novel species within the genus Arenimonas, for which the name Arenimonas soli sp. nov. is proposed, the type strain is Y3L17T (=CGMCC 1.15905T =KCTC 52420T).

Molecular characterization and antimicrobial resistance profile of atypical Citrobacter gillenii and Citrobacter sp. isolated from diseased rainbow trout (Oncorhynchus mykiss).

Over the last decade, Citrobacter species have been responsible for infections in fish and many species and also new Citrobacter species have been identified. In this study, molecular identifications and the phenotypic and genotypic antimicrobial-resistance characteristics of atypical and typical Citrobacter species were determined. Seven Citrobacter isolates were investigated from rainbow trout of different lengths with signs of disease. Biochemical characteristics were determined using conventional tests and rapid test kits; moreover, molecular identifications were conducted with 16S rRNA and the gyrB housekeeping gene region. The sequencing results obtained from the gyrB gene region were deposited in the GenBank database and compared with isolates from different countries that were registered in the database. Resistances to florfenicol, sulfonamides, and tetracycline antimicrobials were determined using the broth micro dilution method, and molecular resistance genes against these antimicrobials were identified. All detected resistance genes were confirmed by sequence analyses. It was determined that three of the Citrobacter species with biochemical characteristics were atypical and showed oxidase-positive reactions. All the Citrobacter species were identified as Citrobacter sp. using the 16S rRNA gene; three isolates were identified as Citrobacter gillenii and four as Citrobacter sp. based on gyrB gene sequence analysis. Some isolates were found in the same group as other countries' isolates in the GenBank database, while isolates with high identities were found in different genogroups. All isolates were found to be phenotypically resistant to sulfamethoxazole and susceptible to tetracycline; these isolates' resistance genes included sulI, tetA, tetB, and tetD.

Pseudomonas aestus sp. nov., a plant growth-promoting bacterium isolated from mangrove sediments.

Strain CMAA 1215T, a Gram-reaction-negative, aerobic, catalase positive, polarly flagellated, motile, rod-shaped (0.5-0.8×1.3-1.9µm) bacterium, was isolated from mangrove sediments, Cananéia Island, Brazil. Analysis of the 16S rRNA gene sequences showed that strain CMAA 1215T forms a distinct phyletic line within the Pseudomonas putida subclade, being closely related to P. plecoglossicida ATCC 700383T, P. monteilii NBRC 103158T, and P. taiwanensis BCRC 17751T of sequence similarity of 98.86, 98.73, and 98.71%, respectively. Genomic comparisons of the strain CMAA 1215T with its closest phylogenetic type strains using average nucleotide index (ANI) and DNA:DNA relatedness approaches revealed 84.3-85.3% and 56.0-63.0%, respectively. A multilocus sequence analysis (MLSA) performed concatenating 16S rRNA, gyrB and rpoB gene sequences from the novel species was related with Pseudomonas putida subcluster and formed a new phylogenetic lineage. The phenotypic, physiological, biochemical, and genetic characteristics support the assignment of CMAA 1215T to the genus Pseudomonas, representing a novel species. The name Pseudomonas aestus sp.nov. is proposed, with CMAA 1215T (=NRRL B-653100T=CBMAI 1962T) as the type strain.

Identification and characterization of Burkholderia multivorans CCA53

ObjectiveA lignin-degrading bacterium, Burkholderia sp. CCA53, was previously isolated from leaf soil. The purpose of this study was to determine phenotypic and biochemical features of Burkholderia sp. CCA53.ResultsMultilocus sequence typing (MLST) analysis based on fragments of the atpD, gltD, gyrB, lepA, recA and trpB gene sequences was performed to identify Burkholderia sp. CCA53. The MLST analysis revealed that Burkholderia sp. CCA53 was tightly clustered with B. multivorans ATCC BAA-247T. The quinone and cellular fatty acid profiles, carbon source utilization, growth temperature and pH were consistent with the characteristics of B. multivorans species. Burkholderia sp. CCA53 was therefore identified as B. multivorans CCA53.

Impact of Mannanase-Producing Bacillus spp. on the Accuracy of the 3M Petrifilm Aerobic Count Method

Consistent deviations of the 3M Petrifilm aerobic counts (AC) from the standard pour plate aerobic plate count (APC) were observed with dehydrated onion and garlic products. A large study was designed to determine the relationship of these two methods and the root cause for the deviations. A total of 3,800 dehydrated onion and garlic samples were analyzed by both the Petrifilm AC and the standard pour plate APC method. Large spreader-like liquefied areas were observed on numerous Petrifilm plates. These liquefied areas made enumeration inaccurate. "Liquefier" microorganisms from Petrifilm plates were isolated and identified to species level by 16S rRNA and gyrB gene sequencing. Enzyme diffusion assay was performed to determine potential enzymatic degradation of guar gum, the gelling agent used in Petrifilm plates. The results indicated that the correlation between Petrifilm AC and standard APC is relatively low. Paired t test results suggested that the Petrifilm AC method produced significantly different results compared with standard APC. The discrepancies were attributable at least partly to a liquefier organism that hydrolyzed guar gum, leading to liquefaction. Liquefaction of Petrifilm plates seems to have two effects on accuracy: (i) liquefied areas may allow motile organisms to move and multiply in the liquefied area during the incubation period, yielding more than one colony from one cell and, as a result, leading to overestimation of the microbial load and (ii) the blurred areas obscure other colonies, leading to potential underestimation. The liquefier organism was identified as Bacillus amyloliquefaciens , a potent mannanase producer and heat-resistant spore former. Enzyme diffusion assay confirmed that mannanase contained in the cell-free supernatant of B. amyloliquefaciens can hydrolyze the 1,4-β-mannopyranosyl bond, the backbone of guar gum. This is the first report of the role of B. amyloliquefaciens in the liquefaction of Petrifilm plates and its negative impact on accuracy.

Bifidobacterium apri sp. nov., a thermophilic actinobacterium isolated from the digestive tract of wild pigs (Sus scrofa).

Fresh samples of intestinal contents of three wild pigs originating from the Central Bohemia region were examined for the presence of bifidobacterial strains. During the study, we isolated many fructose-6-phosphate phosphoketolase-positive, strictly anaerobic, irregular rod-shaped bacterial isolates. Three of them were preliminarily identified as representing a novel species of the genus Bifidobacterium because their 16S rRNA gene sequence similarity with the closest relatives of thermophilic bifidobacteria (Bifidobacterium boum DSM 20432T, Bifidobacterium thermophilum DSM 20210T, Bifidobacterium thermacidophilumsubsp. porcinum LMG 21689T, Bifidobacterium thermacidophilumsubsp. thermacidophilum DSM 15837T) was in the range of 97.9 - 98.4 %. All three bacterial isolates had identical 16S rRNA, dnaJ1, fusA, gyrB and rplB gene sequences. Isolate RP115T was chosen as a representative of the bacterial group and DNA G+C content (mol%) determination, biochemical tests and analyses of physiological and morphological characteristics, habitat and chemotaxonomic traits (peptidoglycan structure, cellular fatty acids and polar lipids profile) were performed. The DNA-DNA hybridization analyses of RP115T and species representing the group of thermophilic bifidobacteria revealed values in the range from 33 to 53 %. This fact, together with relatively low sequence similarities of particular phylogenetic markers among examined bacterial strains and the phenotyping and chemotaxonomy results obtained, indicated that the evaluated bacterial isolate should be classified as representing a separate taxon within the specific group of thermophilic bifidobacteria. The name Bifidobacterium apri (of boar) sp. nov. has been proposed for the representative strain RP115T (=CCM 8605T=DSM 100238T=LMG 28779T).

Sporolactobacillus shoreicorticis sp.nov., a lactic acid-producing bacterium isolated from tree bark.

A Gram-stain-positive, lactic acid-producing bacterium designed strain MK21-7T, was isolated from tree bark collected from the north east of Thailand. This strain was a facultatively anaerobic spore-forming rod that was catalase-negative. It contained meso-diaminopimelic acid in the cell wall peptidoglycan and had seven isoprene units (MK-7) as the predominant menaquinone. Major fatty acids of MK21-7T were anteiso-C17 : 0, iso-C16 : 0, anteiso-C15 : 0 and C18 : 1ω9c. Polar lipids were phosphatidglycerol, diphosphatidylglycerol, an unknown phospholipid, three unknown glycolipids and an unknown lipid. The results of 16S rRNA gene sequence analysis indicated that it represented a member of the genus Sporolactobacillus. MK21-7T showed the highest 16S rRNA gene sequence similarity to Sporolactobacillus terrae NBRC 101527T with 98.4 % similarity and exhibited 97.6 % similarity with Sporolactobacillus kofuensis NRIC 0334T, 97.5 % with Sporolactobacillus laevolacticus NRIC 0361T, 97.3 % with Sporolactobacillus nakayamaesubsp.nakayamae NRIC 0347T and 97.1 % with Sporolactobacillus nakayamaesubsp.racemicus NBRC 101524T. Analysis of the phylogenetic relationship based on 16S rRNA and gyrB gene sequencing revealed that the position of MK21-7T was clearly separated from all related species of the genus Sporolactobacillus. It had low DNA-DNA relatedness (22.8-57.2 %) with S. terrae NBRC 101527T and related type strains. The DNA G+C content was 43.1 mol%. On the basis of the results of the phenotypic, genotypic and chemotaxonomic studies, MK21-7T should be classified as representing a novel species of the genus Sporolactobacillus for which the name Sporolactobacillus shoreicorticis sp. nov. is proposed. The type strain is MK21-7T (=NBRC 111517T=LMG 29111T=TISTR 2466T).

Association of Pantoea ananatis and Pantoea agglomerans with leaf spot disease on ornamental plants of Araceae Family

During a survey in 2011–2012, three ornamental plants of Araceae namely Aglaonema nitidum, Syngonium podophyllum and Dieffenbachia amoena showing foliar disease symptoms were collected from central region of Iran. Infected plants exhibited spots on their leaves which appeared as yellow and water-soaked with chlorotic haloes and necrotic center. To investigate the etiology of this disorder, symptomatic leaves were collected from affected plants and six bacterial strains (B2Y, J3Y, SY, E60Y, E68Y and E5MM) were isolated and identified as Pantoea ananatis or P. agglomerans based on morphological, physiological, biochemical and molecular characters. The pathogenicity tests of the isolates demonstrated that they were not host specific. Furthermore, 16S rRNA gene sequencing revealed that the strains were phylogenetically closely related to genus Pantoea. Multilocus sequence analysis (MLSA) of concatenated partial atpD, gyrB and rpoB gene sequences of the six isolates showed a high similarity of B2Y, J3Y, and SY strains to P. ananatis and also of E60Y, E5MM and E68Y strains to P. agglomerans. These results were confirmed by phylogenetic analysis. To the best of our knowledge, this is the first report of leaf spot and necrosis of A. nitidum, S. podophyllum and D. amoena caused by the genus Pantoea.

Bacteremia due to Gordonia polyisoprenivorans: case report and review of literature

BackgroundGordonia polyisoprenivorans is a ubiquitous aerobic actinomycetes bacterium that rarely cause infections in humans. Here, we report a case of G. polyisoprenivorans catheter-related bacteremia in an AIDS patient.Case presentationA 37-year-old man with a past medical history of AIDS-related lymphoma suffered bacteremia caused by a Gram-positive corynebacterium. The strain was identified as a Gordonia species by matrix-assisted laser desorption ionization–time of flight mass spectrometry and confirmed to G. polyisoprenivorans by 16S rRNA combined with gyrB gene sequencing analyses. The patient was treated with imipenem and had a good outcome.ConclusionsThe findings from our case and previously reported cases indicate that malignant hematologic disease, immunosuppression, and indwelling catheter heighten the risk for G. polyisoprenivorans infection. Molecular methods should be employed for proper identification of G. polyisoprenivorans to the species level.

Sphingobacterium alkalisoli sp. nov., isolated from a saline-alkaline soil.

A Gram-staining-negative, non-motile, non-spore-forming bacterium designated Y3L14T was isolated from the saline-alkaline soil of a farmland, Inner Mongolia, northern China. Strain Y3L14T could grow at 10-40 °C (optimally at 30 °C), pH 6.0-10.0 (optimally at pH 8.0), and in the presence of 0-6.0 % (w/v) NaCl (optimally with 0-2.0 %). Phylogenetic analysis based on the 16S rRNA gene and DNA gyrase subunit B (gyrB) gene sequences revealed that strain Y3L14T clustered with strains belonging to the genus Sphingobacterium, sharing the highest 16S rRNA gene sequence similarity with Sphingobacterium lactis WCC 4512T (94.99 %). Its major cellular fatty acids contained iso-C15 : 0, C16 : 0, iso-C17 : 0 3-OH and summed feature 3 (iso-C15 : 0 2-OH and/or C16 : 1ω7c). Menaquinone-7 (MK-7) was the only isoprenoid quinone. Strain Y3L14T contained phosphatidylethanolamine, sphingophospholipid, two unknown phospholipids and three unknown lipids as the major polar lipids. The genomic DNA G+C content of strain Y3L14T was 36.0 mol%. Based on the phenotypic, phylogenetic and genotypic characteristics, strain Y3L14T represents a novel species within the genus Sphingobacterium, for which Sphingobacterium alkalisoli sp. nov. is proposed; the type strain is Y3L14T (=CGMCC 1.15782T=KCTC 52379T).

Acinetobacter defluvii sp. nov., recovered from hospital sewage.

A Gram-stain-negative, non-motile Acinetobacter strain, WCHA30T, was isolated from hospital sewage in West China Hospital of Sichuan University in Chengdu, south-western China. Strain WCHA30T was a non-spore-forming, catalase-positive, oxidase-negative, strictly aerobic coccobacillus. The DNA G+C content was 38 mol%. Phylogenetic analysis based on 16S rRNA, rpoB and gyrB gene sequences revealed that the strain was distinct from any previously described species of the genus Acinetobacter. Strain WCHA30T could be distinguished from all known Acinetobacter species by its ability to assimilate β-alanine but not l-glutamate. Genotypic and phenotypic characteristics from this study indicate that strain WCHA30T should be considered to represent a novel species of the genus Acinetobacter, for which the name Acinetobacter defluvii sp. nov. is proposed. The type strain is WCHA30T (=CCTCC AB 2016203T=GDMCC 1.1101T=KCTC 52503T).

Xenorhabdus thuongxuanensis sp. nov. and Xenorhabdus eapokensis sp. nov., isolated from Steinernema species.

Two slightly yellowish-pigmented, oxidase-negative, rod-shaped and Gram-stain-negative bacterial strains (30TX1T and DL20T), isolated from Steinernema sangi and Steinernema eapokense, respectively, during soil sampling in Vietnam were studied using a polyphasic taxonomic approach. Strain 30TX1T showed highest 16S rRNA gene sequence similarity to the type strain of Xenorhabdus ehlersii (98.9 %) and strain DL20T to that of Xenorhabdus ishibashii (98.7 %). Sequence similarities to all other Xenorhabdus species were lower (<98.4 %). The two strains shared 98 % 16S rRNA gene sequence similarity. Multilocus sequence analysis (MLSA) based on concatenated partial recA, dnaN, gltX, gyrB and infB gene sequences showed a clear distinction of strains 30TX1T and DL20T among each other and to the closest related type strains. DNA-DNA hybridizations between strain DL20T and the type strain of X. ishibashii resulted in a relatedness value of 53 %. Genome-to-genome-based comparisons gave average nucleotide identities of 93.6 % (reciprocal 93.5 %) for strain 30TX1T and X. ehlersii DSM 16337T, of 92.8 % (reciprocal 93 %) for strain DL20T and X. ishibashiiDSM 22670Tand of 93.0 % (reciprocal 93.2 %) for the two novel strains. The fatty acid profile of the strains consisted of the major fatty acids C14 : 0, C16 : 0, C17 : 0 cyclo, C16 : 1ω7c and/or iso-C15 : 0 2-OH, and C18 : 1ω7c. Genome-to-genome comparison and MLSA results together with the differential biochemical and chemotaxonomic properties showed that strains 30TX1T and DL20T represent novel Xenorhabdus species, for which the names Xenorhabdus thuongxuanensis sp. nov. (type strain 30TX1T=CCM 8727T=LMG 29916T) and Xenorhabdus eapokensis sp. nov. (type strain DL20T=CCM 8728T=LMG 29917T) are proposed, respectively.

Molecular characterization of Mycobacterium avium subsp. paratuberculosis (MAP) isolates

Objective: Johne’s disease, caused by Mycobacterium avium subsp. paratuberculosis (MAP), is highly prevalent in domestic ruminants. In India, the exact prevalence of MAP genotypes still remains unknown limited, and the systematic disease control programs are also limited. This study was undertaken to study the molecular characterization of MAP isolates.Methods: About 22 MAP isolates were from cattle, sheep, and goat under gone the molecular characterization by three different methods (1) IS1311 polymerase chain reaction (PCR) with restriction enzyme analysis (REA), (2) GyrA and GyrB PCR with sequencing, and (3) digital microfluidic chip (DMC)-PCR. The study demonstrated that a) IS1311 PCR with REA (based on point mutations) identified all 22 MAP isolates as “intermediate type” irrespective of a host of origin and also belong to Indian Bison type. Molecular typing based on the gyrA and gyrB genes partial amplification and sequencing revealed that the MAP isolates exhibited more lineages toward the reference Type III, Intermediate strain.Results: The MAP isolate of sheep origin showed more lineages toward the sheep type than the isolates of cattle and goats. This variation may be due to host-pathogen interactions and adaptation to different hosts and environmental conditions in the nature.Conclusions: The DMC-PCR, which is based on sequence difference at 5` end of IS900 of MAP, differentiated rapidly all the isolates as sheep type. The application of DMC-PCR to differentiate sheep and Intermediate types is limited as the Intermediate type (Type III) and sheep type (Type I) are very closely related to each other and all the MAP isolates were confirmed as Intermediate or Type III by three different methods which are commonly present in India, Spain, and Iceland.Keywords: Johne’s disease, Mycobacterium avium subsp. paratuberculosis, Polymerase chain reaction, Digital microfluidic chip-polymerase chainreaction.