Sphingobium algorifonticola sp. nov., isolated from a cold spring.

Strain TLA-22T, isolated from a cold spring in Taiwan, was characterized using a polyphasic taxonomy approach. Cells were Gram-stain-negative, aerobic, poly-ß-hydroxybutyrate-accumulating, motile by means of a single polar flagellum, rod-shaped and formed bright yellow colonies. Optimal growth occurred at 20-25 °C, pH 6-6.5, and in the presence of 0.5 % NaCl. The major fatty acids of TLA-22T were C18 : 1 ω7 c and C17 : 1ω6c. The predominant hydroxy fatty acids were C15 : 0 2-OH and C14 : 0 2-OH. The polar lipid profile consisted of a mixture of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidyldimethylethanolamine, sphingoglycolipid, an unidentified aminophospholipid, an unidentified phospholipid and three unidentified lipids. TLA-22T contained spermidine as the major polyamine and putrescine as the minor component. The only isoprenoid quinone was Q-10. The genomic DNA G+C content of TLA-22T was 63.2 mol%. Phylogenetic analyses based on 16S rRNA gene sequences and coding sequences of 92 protein clusters indicated that TLA-22T was a mem,ber of a phylogenetic lineage including members of the genus Sphingobium. TLA-22T was most closely related to Sphingobium aromaticiconvertens RW16T, with a 97.4 % 16S rRNA gene sequence similarity. TLA-22T showed 74.8-75.7 % average nucleotide identity and 20.1-22.0 % digital DNA-DNA hybridization identity with the strains of other species of the genus Sphingobium. On the basis of phenotypic and genotypic properties and phylogenetic inference, strain TLA-22T should be classified as representing a novel species of the genus Sphingobium, for which the name Sphingobium algorifonticola sp. nov. is proposed. The type strain is TLA-22T (=BCRC 81097T =LMG 30309T=KCTC 62189T).

Flavobacterium ichthyis sp. nov., isolated from a fish pond.

Bacterial strain NST-5T, isolated from a fish pond in Taiwan, was characterized using a polyphasic taxonomy approach. Results of phylogenetic analyses based on 16S rRNA gene sequences and coding sequences of 92 protein clusters indicated that strain NST-5T formed a phylogenetic lineage in the genus Flavobacterium. Analysis of 16S rRNA gene sequences showed that strain NST-5T showed the highest similarity to Flavobacterium enshiense DK69T (94.9 %), Flavobacterium ahnfeltiae 10Alg 130T (94.8 %) and Flavobacterium vireti THG-SM1T (94.8 %). Strain NST-5T showed 68.9-72.5% average nucleotide identity and 19.1-23.7% digital DNA-DNA hybridization identity with the type strains of other close related Flavobacterium species. Cells of the strain were Gram-stain-negative, strictly aerobic, motile by gliding, rod-shaped and formed yellow colonies. Optimal growth occurred at 30 °C, pH 7 and with 0.5% NaCl. Strain NST-5T contained iso-C15:0, C15:0 and iso-C16:0 as the predominant fatty acids. The major hydroxyl fatty acids were iso-C16:0 3-OH and iso-C17:0 3-OH. The polar lipid profile consisted of phosphatidylethanolamine, three uncharacterized aminophospholipids, two uncharacterized phospholipids and one uncharacterized aminolipid. The major polyamine was homospermidine. The major isoprenoid quinone was MK-6. The DNA G+C content of the genomic DNA was 35.5 mol%. Differential phenotypic properties, together with the phylogenetic inference, demonstrate that strain NST-5T should be classified as a novel species of the genus Flavobacterium, for which the name Flavobacterium ichthyis sp. nov. is proposed. The type strain is NST-5T (=BCRC 81198T=LMG 31341T).

Tabrizicola oligotrophica sp. nov. and Rhodobacter tardus sp. nov., two new species of bacteria belonging to the family Rhodobacteraceae.

Two Gram-stain-negative, aerobic, non-motile bacteria, designated KMS-5T and CYK-10T, were isolated from freshwater environments. 16S rRNA gene sequence similarity results indicated that these two novel strains belong to the family Rhodobacteraceae. Strain KMS-5T is closely related to species within the genus Tabrizicola (96.1-96.8 % sequence similarity) and Cypionkella (96.5-97.0 %). Strain CYK-10T is closest to Rhodobacter thermarum YIM 73036T with 96.6 % sequence similarity. Phylogenetic analyses based on 16S rRNA gene sequences and an up-to-date bacterial core gene set showed that strain KMS-5T is affiliated with species in the genus Tabrizicola and strain CYK-10T is placed in a distinct clade with Rhodobacter blasticus ATCC 33485T, Rhodobacter thermarum YIM 73036T and Rhodobacter flagellatus SYSU G03088T. These two strains shared common chemotaxonomic features comprising Q-10 as the major quinone, phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine as the principal polar lipids, and C18 : 1 ω7c as the main fatty acid. The average nucleotide identity, average amino acid identity and digital DNA-DNA hybridization values between these two novel isolates and their closest relatives were below the cut-off values of 95-96, 90 and 70 %, respectively, used for species demarcation. The obtained polyphasic taxonomic data suggested that strain KMS-5T represents a novel species within the genus Tabrizicola, for which the name Tabrizicola oligotrophica sp. nov. is proposed with KMS-5T (=BCRC 81196T=LMG 31337T) as the type strain, and strain CYK-10T should represent a novel species of the genus Rhodobacter, for which the name Rhodobacter tardus sp. nov. is proposed with CYK-10T (=BCRC 81191T=LMG 31336T) as the type strain.

Amniculibacterium aquaticum gen. nov., sp. nov., a new member of the family Flavobacteriaceae isolated from a stream.

Strain KYPW7T, isolated from the Funglin Stream in Taiwan, was characterized using a polyphasic taxonomy approach. Cells of strain KYPW7T were Gram-stain-negative, aerobic, non-spore forming, non-motile rods and formed white colonies. Growth occurred at 15-30 °C (optimum 25 °C), at pH 6-8 (optimum pH 6.5) and with 0-1% NaCl (optimum 0%). Phylogenetic analyses based on 16S rRNA and coding sequences of 92 protein clusters showed that strain KYPW7T represents a novel genus in the family Flavobacteriaceae. The 16S rRNA gene sequence of strain KYPW7T was related to the species of the genera Chryseobacterium (91.8-96.0% sequence similarity), Bergeyella (95.1-95.8%), Cloacibacterium (94.5-95.7%), Daejeonia (95.6%) and Riemerella (94.0-95.0%). Strain KYPW7T showed less than 72% average nucleotide identity and less than 24% digital DNA-DNA hybridization identity compared to the type strains of related genera within the family Flavobacteriaceae. The predominant fatty acids were iso-C15:0 and iso-C17:0 3-OH. The major isoprenoid quinone was MK-6 and the DNA G + C content was 36.8 mol%. The polar lipids had phosphatidylethanolamine, three uncharacterized aminophospholipids and an uncharacterized phospholipid. The polyamines contained homospermidine, putrescine and spermidine. On the basis of the genotypic and phenotypic data, strain KYPW7T represents a novel species of a new genus in the family Flavobacteriaceae, for which the name Amniculibacterium aquaticum gen. nov., sp. nov. is proposed. The type strain is KYPW7T (= BCRC 81123T = LMG 30598T = KCTC 62512T).

Pseudomethylobacillus aquaticus gen. nov., sp. nov., a new member of the family Methylophilaceae isolated from an artificial reservoir.

A bacterial strain, designated H-5T, was isolated from an artificial reservoir in Taiwan and characterized using a polyphasic taxonomic approach. Cells of strain H-5T were Gram-stain-negative, aerobic, motile by means of a single polar flagellum, rod-shaped, covered by large capsules and formed white colonies. Growth occurred at 15-30 °C (optimum, 25 °C), at pH 6-8 (optimum, pH 7) and with 0-0.5 % NaCl (optimum, 0 %). Phylogenetic analyses based on the 16S rRNA gene, the methanol dehydrogenase gene and the coding sequences of 92 protein clusters indicated that strain H-5T was affiliated with genera in the family Methylophilaceae in the class Betaproteobacteria. Strain H-5T was most closely related to Methylobacillus methanolivorans ZT with a 95.0 % 16S rRNA gene sequence similarity. Strain H-5T showed less than 73.7 % average nucleotide identity and less than 23.6 % digital DNA-DNA hybridization identity compared to the strains of related genera within the family Methylophilaceae. The predominant fatty acids were summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and C16 : 0. The major isoprenoid quinone was Q-8 and the DNA G+C content was 58.3 mol%. The major polar lipids were phosphatidylethanolamine, phosphatidylglycerol, one uncharacterized aminophospholipid, one uncharacterized phospholipid and one uncharacterized lipid. On the basis of the genotypic and phenotypic data presented here, strain H-5T represents a novel species of a new genus in the family Methylophilaceae, for which the name Pseudomethylobacillus aquaticus gen. nov., sp. nov. is proposed. The type strain is H-5T (=BCRC 81154T=KCTC 62865T).

Sphingomonas crocodyli sp. nov., isolated from a crocodile pond.

Strain CCP-7T, isolated from a freshwater pond in Taiwan, was characterized using a polyphasic taxonomy approach. Phylogenetic analyses based on 16S rRNA gene sequences and coding sequences of 92 protein clusters indicated that strain CCP-7T formed a phylogenetic lineage in the genus Sphingomonas. Strain CCP-7T was most closely related to Sphingomonas starnbergensis 382T and Sphingomonas naphthae DKC-5-1T with 96.2 % 16S rRNA gene sequence similarity. Strain CCP-7T showed 65.5-76.7 % average nucleotide identity and 20.2-22.5 % digital DNA-DNA hybridization identity with the strains of other related Sphingomonas species. Cells were Gram-stain-negative, aerobic, motile, rod-shaped and formed light orange-coloured colonies. Optimal growth occurred at 30 °C, pH 6 and in the absence of NaCl. The major fatty acid of strain CCP-7T was C18 : 1ω7c. The polar lipid profile consisted of phosphatidylethanolamine, phosphatidylglycerol, diphosphatidylglycerol, phosphatidylmonomethylethanolamine, three uncharacterized sphingoglycolipids, two uncharacterized phospholipids and six uncharacterized lipids. The predominant polyamine was homospermidine. The only isoprenoid quinone was Q-10. Genomic DNA G+C content of strain CCP-7T was 64.5 %. On the basis of phenotypic and genotypic properties and phylogenetic inference, strain CCP-7T should be classified in a novel species of the genus Sphingomonas, for which the name Sphingomonas crocodyli sp. nov. is proposed. The type strain is CCP-7T (=BCRC 81096T=LMG 30311T=KCTC 62190T).

Flavobacterium stagni sp. nov., isolated from a freshwater reservoir.

Strain WWJ-16T was isolated from a freshwater reservoir in Taiwan. Phylogenetic analyses based on 16S rRNA gene sequences and an up-to-date bacterial core gene set (92 protein clusters) indicated that strain WWJ-16T is affiliated with the species in the genus Flavobacterium. Strain WWJ-16T was most closely related to Flavobacterium fontis MIC3010T (97.9 % 16S rRNA gene sequence identity) and Flavobacterium squillarum CMJ-5T (97.6 %). Strain WWJ-16T showed 77.4 % average nucleotide identity and 20.6 % digital DNA-DNA hybridization identity with F. fontis MIC3010T. Cells of strain WWJ-16T were Gram-stain-negative, strictly aerobic, non-motile, rod-shaped and formed yellow colonies. Optimal growth occurred at 25 °C, pH 7 and in 0.5-1 % NaCl (w/v). Strain WWJ-16T contained iso-C15 : 1 G and iso-C15 : 0 as predominant fatty acids. The major hydroxyl fatty acids were iso-C15 : 0 3-OH, iso-C17 : 0 3-OH and iso-C16 : 0 3-OH. The polar lipid profile consisted of phosphatidylethanolamine, four uncharacterized aminophospholipids and four uncharacterized phospholipids. The major polyamine was homospermidine. The major isoprenoid quinone was MK-6. The genomic DNA G+C content of strain WWJ-16T was 39.4 mol%, as determined by genome sequencing. The genotypic and phenotypic characteristics indicate that strain WWJ-16T represents a novel species of the genus Flavobacterium, for which the name Flavobacterium stagni sp. nov. is proposed. The type strain is WWJ-16T (=BCRC 81125T =LMG 30600T=KCTC 62515T).

Salsuginimonas clara gen. nov., sp. nov., a member of the family Pseudoalteromonadaceae isolated from a brackish river.

A bacterial strain designated LSN-49T was isolated from a brackish river in Taiwan and characterized using a polyphasic taxonomy approach. Cells of strain LSN-49T were Gram-staining-negative, aerobic, poly-ß-hydroxybutyrate accumulating, motile by means of a monopolar flagellum, non-spore forming, straight rods and formed shiny and translucent colonies. Growth occurred at 20-40 °C (optimum, 25-30 °C), at pH 6-10 (optimum, pH 7-8) and with 0-3 % (w/v) NaCl [optimum, 0-1 % (w/v)]. The predominant fatty acids were summed feature 3 (comprising C16 : 1ω7c and/or C16 : 1ω6c), C17 : 1ω8c and C16 : 0. The polar lipid profile consisted of a mixture of phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phosphatidylcholine, (PC), two uncharacterized aminophospholipids (APL1 and APL2), one uncharacterized glycolipid (GL1), four uncharacterized phospholipids (PL1-PL4) and four uncharacterized lipids (L1-L4). The major polyamine was putrescine. The major isoprenoid quinone was Q-8 and the DNA G+C content was 51.0 mol%. The results of phylogenetic analyses based on 16S rRNA gene sequences indicated that LSN-49T formed a distinct lineage with respect to closely related genera in the family Pseudoalteromonadaceae. LSN-49T was most closely related to Pseudoalteromonas, Algicola and Psychrosphaera and showed 89.3-92.1 % sequence similarity with members of the family Pseudoalteromonadaceae with validly published names. On the basis of the genotypic and phenotypic data, LSN-49T represents a novel genus and species of the family Pseudoalteromonadaceae, for which the name Salsuginimonas clara gen. nov., sp. nov. is proposed. The type strain is LSN-49T (=BCRC 81005T=LMG 29726T=KCTC 52439T).

Cellvibrio zantedeschiae sp. nov., isolated from the roots of Zantedeschia aethiopica.

A bacterial strain, designated TPY-10T, was isolated from calla lily roots in Taiwan and characterized by using a polyphasic taxonomy approach. Cells of strain TPY-10T were Gram-stain-negative, strictly aerobic, motile and creamy white rods. Growth occurred at 15-35 °C (optimum, 25-30 °C), at pH 6-7 (optimum, pH 6) and with 0-1 % NaCl (optimum, 0 %). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain TPY-10T belonged to the genus Cellvibrio and was most closely related to Cellvibriomixtus ACM 2601T with sequence similarity of 97.8 %. Strain TPY-10T contained C16 : 0, summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) and C18 : 1ω7c as the predominant fatty acids. The only isoprenoid quinone was Q-9. The major polar lipids were phosphatidylethanolamine and phosphatidylglycerol. The DNA G+C content of the genomic DNA was 49.8 mol%. The DNA-DNA hybridization value for strain TPY-10T with Cellvibriomixtus ACM 2601T was less than 21 %. On the basis of the phylogenetic inference and phenotypic data, strain TPY-10T should be classified as a novel species, for which the name Cellvibrio zantedeschiae sp. nov. is proposed. The type strain is TPY-10T (=BCRC 80525T=LMG 27291T=KCTC 32239T).

Analysis of antibiotic fungicide kasugamycin in irrigation water by high performance liquid chromatography.

A high performance liquid chromatographic (HPLC) analysis method with an ultraviolet (UV) detector and an Aqua C18 (250 x 4.6 mm, Phenomenex) column were applied to analyze the antibiotic fungicide kasugamycin in water. An aromatic sulfonic acid spe column (Backerbond, J. T. Backer) was used to remove the interfering materials from irrigation water. A good linear relation existed between the concentration of the fungicide and the peak area, and correlation coefficient of linearity from 0.1 to 10.2 microg/mL was 0.998. The accuracies expressed as the recoveries of kasugamycin from irrigation water ranged from 112.2 to 111.7 %. The precisions expressed as relative standard deviations (RSD) were found to be below 7.0 %. The quantitative detection limit (LOQ) of kasugamycin in irrigation water was set at 2.2 microg/mL which was 2-times higher than the method detection limit (MDL) 1.03 microg/mL. Electrospray ionization-mass (ESI-MS) and fast-atom bombardment-mass (FAB-MS) were applied to compare the ability of identifying the component of the eluent peak from HPLC, and the result indicated that electrospray ionization-mass (ESI-MS) was more sensitive than fast-atom bombardment-mass (FAB-MS) in the detection of kasugamycin. There was no kasugamycin residue detected in irrigation water samples collected from paddyfields at Wufong, indicated that the residues of kasugamycin in water were less than 2.2 microg/mL, and the risk of water contamination was very low.

Extraction of DNA from soil for analysis of bacterial diversity in transgenic and nontransgenic papaya sites.

The influence of transgenic crops on the soil diversity of microorganisms is one of the major risk assessments being conducted in Taiwan since 2007, and a reliable soil DNA extraction method for denaturing gradient gel electrophoresis (DGGE) is required. Six soils of different type, organic matter content, cation exchange capacity, and pH were tested, and four previously reported soil DNA extraction methods were applied to these soils. Soil DNA extracts by Zhou's CS method plus QIAquick gel was recommended in our laboratory for DGGE to monitor the microbial diversity in soil. There were some differences on the bacterial diversity based on DGGE patterns at the beginning of planting, and the difference decreased after six months. The results also indicated that clay content (10.8-25.0%) and pH (4.4-6.9) of different soil samples we tested did not affect the DNA extraction efficiencies, but positive correlations were found between the organic matter content (1.2-3.9%) of soils and the DNA yields in Widmer's GS method (r = 0.93, p = 0.005) and the MoBio UC method (r = 0.92, p = 0.007). Coefficient of determinations between organic matter content and DNA yield were higher than those between clay content, CEC, and pH, indicating that organic matter content was more correlated with DNA yield than that clay content, CEC, and pH in our soil samples tested.