- Volume 66, Issue 3, 2016

A Gram-stain-negative, non-flagellated, non-spore-forming bacterial strain motile by gliding, designated RSS1-6T, was isolated from a golden sea squirt Halocynthia aurantium and its taxonomic position was investigated by using a polyphasic approach. Strain RSS1-6T grew optimally at 30–37 °C and in the presence of 1.0–4.0 % (w/v) NaCl. Phylogenetic trees based on 16S rRNA gene sequences showed that strain RSS1-6T fell within the clade comprising species of the genus Tenacibaculum, clustering with the type strains of Tenacibaculum discolor, Tenacibaculum litoreum and Tenacibaculum gallaicum with which it exhibited 16S rRNA gene sequence similarity values of 98.5–99.5 %. Strain RSS1-6T contained MK-6 as the predominant menaquinone and iso-C15 : 0, iso-C17 : 0 3-OH and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) as the major fatty acids. The major polar lipids of strain RSS1-6T were phosphatidylethanolamine, two unidentified lipids, one unidentified aminophospholipid and one unidentified glycolipid. The DNA G+C content was 32.5 mol% and the mean DNA–DNA relatedness values with the type strains of T. discolor, T. litoreum and T. gallaicum were 17.3–25.2 %. The differential phenotypic properties, together with the phylogenetic and genetic distinctiveness, revealed that strain RSS1-6T is separated from other recognized species of the genus Tenacibaculum. On the basis of the data presented, strain RSS1-6T is considered to represent a novel species of the genus Tenacibaculum, for which the name Tenacibaculum ascidiaceicola sp. nov. is proposed. The type strain is RSS1-6T ( = KCTC 42702T = NBRC 111225T).

A Gram-staining-negative bacterial strain (termed CZ-AZ5T) was isolated from a biological filter in a marine recirculating aquaculture system in Tianjin, China. Its taxonomic status was determined using a polyphasic approach. CZ-AZ5T cells were non-spore-forming, non-motile rods, 0.6–0.7 μm wide and 3.0–3.7 μm long. CZ-AZ5T was strictly heterotrophic, aerobic, oxidase-negative and catalase-positive. Growth occurred in the temperature range 20–40 °C (optimal: 30 °C), pH range 6.0–8.5 (optimal: pH 7.5) and salinity range 0–5 % (w/v) NaCl (optimal: 1 %). In phylogenetic analyses based on 16S rRNA gene sequences, CZ-AZ5T was assigned to the family Saprospiraceae (phylum Bacteroidetes) and was clustered with the genera Saprospira and Aureispira within this family. It showed highest sequence similarity to ‘Candidatus Haliscomenobacter calcifugiens’ (86.2 %), followed by Saprospira grandis ATCC 23119T (85.7 %) and Lewinella persica T-3T (85.6 %). DNA G+C content was 40.1 mol%, the major menaquinone was MK-7, and the major cellular fatty acids (>10 %) were C16 : 1ω7c and iso-C15 : 0. Our phenotypic, chemotaxonomic and phylogenetic observations, taken together, led us to conclude that strain CZ-AZ5T represents a novel species and genus of the family Saprospiraceae, for which the name Membranicola marinus gen. nov., sp. nov. is proposed. The type strain is CZ-AZ5T ( = CGMCC 1.13179T = JCM 18886T).

A novel Gram-stain-negative, rod-shaped, non-spore-forming, non-flagellated, strictly aerobic strain, designated RZW2-1T, was isolated from coastal seawater of the Yellow Sea in China (35.475° N 119.613° E). The organism grew optimally at 24 °C, at pH 7.0 and in the presence of 3.0 % (w/v) NaCl. The strain requires seawater or artificial seawater for growth and NaCl alone does not support growth. Strain RZW2-1T contained MK-6 as the only respiratory quinone and iso-C15 : 0, iso-C15 : 1 G and 10-methyl C16 : 0 and/or iso-C17 : 1ω9c as the dominant fatty acids. The polar lipids of strain RZW2-1T were four unidentified phospholipids (PL1–PL4), two unknown lipids (L1, L2) and one unidentified aminolipid (AL1). The DNA G+C content of strain RZW2-1T was 32 mol%. Phylogenetic analysis based on 16S rRNA gene sequences showed that the novel strain was most closely related to the type strain of the only described species of genus Pseudofulvibacter, Pseudofulvibacter geojedonensis YCS-9T, with 95.1 % 16S rRNA gene sequence similarity. On the basis of polyphasic analyses, strain RZW2-1T represents a novel species of the genus Pseudofulvibacter, for which the name Pseudofulvibacter marinus sp. nov. is proposed. The type strain is RZW2-1T ( = JCM 30826T = MCCC 1K00695T).

Honey bees and bumble bees harbour a small, defined set of gut bacterial associates. Strains matching sequences from 16S rRNA gene surveys of bee gut microbiotas were isolated from two honey bee species from East Asia. These isolates were mesophlic, non-pigmented, catalase-positive and oxidase-negative. The major fatty acids were iso-C15 : 0, iso-C17 : 0 3-OH, C16 : 0 and C16 : 0 3-OH. The DNA G+C content was 29–31 mol%. They had ∼87 % 16S rRNA gene sequence identity to the closest relatives described. Phylogenetic reconstruction using 20 protein-coding genes showed that these bee-derived strains formed a highly supported monophyletic clade, sister to the clade containing species of the genera Chryseobacterium and Elizabethkingia within the family Flavobacteriaceae of the phylum Bacteroidetes. On the basis of phenotypic and genotypic characteristics, we propose placing these strains in a novel genus and species: Apibacter adventoris gen. nov., sp. nov. The type strain of Apibacter adventoris is wkB301T ( = NRRL B-65307T = NCIMB 14986T).

A novel heterotrophic, aerobic, Gram-stain-negative, rod-shaped and yellow bacterium, designated strain G18T, was isolated from a water sample collected from the deep South China Sea. Strain G18T grew at 4–40 °C (optimum 28–32 °C), at pH 6.0–8.0 (optimum pH 6.5–7.5) and with 0–12 % (w/v) NaCl (optimum 3–4 %). The organism was mesophilic and piezotolerant, its optimal growth pressure was 0.1 MPa, which was lower than that at the depth from which it was isolated. Its optimal growth temperature was higher than that at the depth of its isolation. The predominant cellular fatty acids were C15 : 0iso, C17 : 0iso 3-OH and C15 : 1iso. The major polar lipids were composed of phosphatidylethanolamine, one unknown aminolipid and one unknown polar lipid. The major respiratory quinone was menaquinone 6. The G+C content of the genomic DNA was 35 mol%. Phylogenetic analysis, based on 16S rRNA gene sequences, revealed that strain G18T clustered with species of the genus Leeuwenhoekiella with validly published names within the family Flavobacteriaceae with 95.9–98.2 % sequence similarity. DNA–DNA reassociation values ranged from 9 to 42 %. Differential phenotypic properties, together with the phylogenetic distinctiveness, suggest that strain G18T differs from species of the genus Leeuwenhoekiella with validly published names. On the basis of the polyphasic evidence, strain G18T represents a novel species, isolated from deep-sea, of the genus Leeuwenhoekiella for which the name Leeuwenhoekiella nanhaiensis sp. nov. is proposed. The type strain is G18T ( = CCTCC AB 2015204T = KCTC 42729T).

A Gram-stain-negative, strictly aerobic, yellow, rod-shaped bacterium, designated strain HQQT, was isolated from a municipal wastewater treatment plant in Hebei Province, PR China. Comparative 16S rRNA gene sequence analyses showed that strain HQQT is a member of the genus Flavobacterium and is closely related to ‘Flavobacterium shanxiense’ CCTCC AB 2014079T (94.8 %) and Flavobacterium macrobrachii DSM 22219T (94.7 %). Phylogenetic analysis showed that strain HQQT clustered with Flavobacterium fontis JCM 18212T and Flavobacterium squillarum KCTC 23915T. The polar lipid profile of strain HQQT revealed the presence of phosphatidylethanolamine, six unknown aminolipids, one unknown glycolipid and one unknown lipid and the only isoprenoid quinone was MK-6. The dominant fatty acids of strain HQQT were iso-C15 : 0, C15 : 0 and C16 : 1ω7c. The DNA G+C content of strain HQQT is 32 mol%. On the basis of the phylogenetic and phenotypic data, strain HQQT represents a novel species of the genus Flavobacterium, for which the name Flavobacterium lutivivi sp. nov. is proposed. The type strain is HQQT ( = CGMCC 1.15347T = KCTC 42935T).

Six strains of anaerobic bacteria, C13EG70T, C13EG118, C13EG186T, C13GAMG5, C13GAMG28 and C13GAMG40, were isolated from the caecum of a healthy chicken bred in Bogor, Indonesia. Phylogenetic analysis showed the isolates were separated into two groups. Group I (C13EG70T and C13EG118) showed nearly identical 16S rRNA gene sequences (99.9 % sequence similarity). Group II (C13EG186T, C13GAMG5, C13GAMG28 and C13GAMG40) showed nearly identical 16S rRNA gene sequences (>99.4 % sequence similarity). The isolates showed low 16S rRNA gene sequence similarities to recognized species of the genus Bacteroides. High gene sequence similarities were found between type strains (C13EG70T and C13EG186T) and Bacteroides salanitronis JCM 13657T (87.9, 91.5 %, respectively). Physiological, biochemical and genotypic characteristics demonstrated that these strains could be separated from the type strain of B. salanitronis. It is concluded that Group I and Group II represent novel species. Two novel species of the genus Bacteroides are proposed as Bacteroides caecicola sp. nov. (type strain C13EG70T = LIPI12-4-Ck732T = JSAT12-4-Ck732T = InaCC B449T = NBRC 110958T) and Bacteroides gallinaceum sp. nov. (type strain C13EG186T = LIPI12-4-Ck844T = JSAT12-4-Ck884T = InaCC B451T = NBRC 110963T).

Three novel, facultatively anaerobic bacteria of the family Porphyromonadaceae (phylum Bacteroidetes) were isolated from mesophilic laboratory-scale biogas reactors. The strains were Gram-negative rods. Optimal growth occurred between 35 and 45 °C and at pH 7.1–7.8. The main fermentation products were acetic and propionic acids. The predominant fatty acid in all strains was anteiso-C15 : 0, and the only respiratory quinone detected was menaquinone MK-8. 16S rRNA gene sequence comparison indicated that strains M3/6T and ING2-E5BT were most closely related to the type strain of Proteiniphilum acetatigenes, with sequence similarities of 97.3 and 94.5 %. Strain ING2-E5AT showed the closest affiliation to the type strain of Petrimonas sulfuriphila, with 97 % sequence identity. DNA–DNA hybridization of strain M3/6T and ING2-E5AT with the most closely related type strains showed 43.3–45.6 and 23.8–25.7 % relatedness, respectively, which supports the conclusion that both isolates represent novel species. Phylogenetic analysis and comparison of cellular fatty acid patterns indicated that strain ING2-E5BT cannot be classified as a member of any previously described genus. Therefore, because of the physiological, genotypic and chemotaxonomic differences, it is proposed to designate novel species within the genera Proteiniphilum and Petrimonas, Proteiniphilum saccharofermentans sp. nov. (type strain M3/6T = DSM 28694T = CECT 8610T = LMG 28299T) and Petrimonas mucosa sp. nov. (type strain ING2-E5AT = DSM 28695T = CECT 8611T), and a novel species of a new genus, Fermentimonas caenicola gen. nov., sp. nov. (type strain of Fermentimonas caenicola is ING2-E5BT = DSM 28696T = CECT 8609T = LMG 28429T). In addition, an emended description of the genus Proteiniphilum is provided.

A bacterial strain, designated KBP-30T, was isolated from a water sample taken from the Banping Lake Wetland Park in Taiwan and characterized taxonomically using a polyphasic approach. Cells of strain KBP-30T were Gram-stain-negative, aerobic, motile by gliding rods that were covered by large capsules and formed red colonies. Growth occurred at 10–37 °C (optimum 20 °C), at pH 6–8 (optimum pH 6) and with 0–1 % NaCl (optimum 0 %). Phylogenetic analyses based on 16S rRNA gene sequences showed that strain KBP-30T belonged to the genus Hymenobacter and was most closely related to Hymenobacter ocellatus Myx 2105T with a sequence similarity of 97.7 %; 16S rRNA gene sequence similarities were less than 95.1 % with other members of the genus. Strain KBP-30T contained iso-C15 : 0, summed feature 4 (iso-C17 : 1 I and/or anteiso-C17 : 1 B), anteiso-C15 : 0, iso-C17 : 0 3-OH and summed feature 3 (C16 : 1ω7c and/or C16 : 1ω6c) as the predominant fatty acids. The major isoprenoid quinone was MK-7. The polar lipid profile consisted of phosphatidylethanolamine, three unidentified aminophospholipids, an unidentified aminolipid, an unidentified glycolipid and eight unidentified lipids. The major polyamine was homospermidine. The DNA G+C content of the genomic DNA was 60.3 mol%. The DNA–DNA relatedness of strain KBP-30T with respect to Hymenobacter ocellatus Myx 2105T was less than 42 %. On the basis of the phylogenetic inference and phenotypic data, strain KBP-30T represents a novel species of the genus Hymenobacter, for which the name Hymenobacter paludis sp. nov. is proposed. The type strain is KBP-30T ( = LMG 27293T = KCTC 32237T).

A new phytoplasma was identified in naturally infected blackberry plants exhibiting witches’ broom symptoms in Portugal. The 16S rRNA gene sequence revealed that it is related to ‘Candidatus Phytoplasma rubi’ (16SrV-E ribosomal subgroup) and RFLP analysis revealed a unique profile following MseI endonuclease digestion of R16F2n/R2 amplicons that distinguished it from the strains belonging to previously established 16SrV phytoplasma subgroups. The in silico restriction analyses confirmed that the phytoplasma strain from blackberry is different from all the other strains reported in group 16SrV. Phylogeny of the 16S rRNA gene sequences, sequence analyses of 16S–23S, tuf, rplV-rpsC, rplF-rplR, rplO-SecY-map and uvrB-degV genetic loci, as well as the variability of unique oligonucleotide sequences defined for ‘Candidatus Phytoplasma rubi’ confirmed the uniqueness of this phytoplasma strain from Portugal for which a novel ribosomal subgroup, 16SrV-I, is proposed. The representative of this new subgroup was named blackPort phytoplasma (Portuguese blackberry phytoplasma).

Two species of the genus Borrelia, Borrelia bissettiae sp. nov. and Borrelia californiensis sp. nov., were first described by Postic and co-workers on the basis of genetic analyses of several loci. Multilocus sequence analysis of eight housekeeping loci confirmed that these two Borrelia genomospecies are distinct members of the Borrelia burgdorferi sensu lato complex. B. bissettiae sp. nov. was initially described in transmission cycles involving Neotoma fuscipes wood rats and Ixodes pacificus ticks in California, and Neotoma mexicana and Ixodes spinipalpis in Colorado. The preferred host of B. californiensis sp. nov. appears to be the California kangaroo rat, Dipodomys californicus; Ixodes jellisoni, I. spinipalipis and I. pacificus ticks are naturally infected with it. Thus, the ecological associations of the two genomospecies and their genetic distance from all other known Borrelia genomospecies species justify their description as separate genomospecies: B. bissettiae sp. nov. (type strain DN127T = DSM 17990T =  CIP 109136T) and B. californiensis (type strain CA446T = DSM 17989T = ATCC BAA-2689T).

Taking into account their 16S rRNA gene sequences, it appears that Acetomicrobium flavidum and the three species of the genus Anaerobaculum described so far belong to the same phylogenetic clade with high levels (>95 %) of similarity. In this respect, these three Anaerobaculum species should be reclassified within the genus Acetomicrobium, which has priority over the genus Anaerobaculum, which was validated since the genus Acetomicrobium. The DNA G+C content of Acetomicrobium flavidum is 47.1 mol%, which is of the same order as that of the three Anaerobaculum species. All these bacteria have in common iso-C15 : 0 as their main fatty acid. Based on further phylogenetic, genetic and chemotaxonomic studies, we propose that Anaerobaculum mobile ( = DSM 13181T = JCM 12221T), Anaerobaculum thermoterrenum ( = DSM 13490T = ACM 5076T) and Anaerobaculum hydrogeniformans ( = DSM 22491T = ATCC BAA-1850T) be reclassified as Acetomicrobium mobile comb. nov., Acetomicrobium thermoterrenum comb. nov. and Acetomicrobium hydrogeniformans comb. nov., respectively. The four bacterial species belong to the phylum Synergistetes.

Ninety-six yeast isolates associated with dung beetles (Heliocopris bucephalus Fabricius) were examined based on a culture-dependent method. A comparison of the colony morphology and PCR-fingerprints obtained by (GTG)5 microsatellite-primed PCR indicated that 84 of these isolates belonged to one group. Five strains (DD1-1T, DD2-33, DD4-11, DD5-15 and DD6-1) were selected as the representatives of this main group, where each of the five selected strains had been derived from a different dung beetle collected in northern Thailand. A comparison of the D1/D2 domain sequence of the large subunit rRNA gene (LSU D1/D2) and the internal transcribed spacer (ITS) sequences revealed that these five strains were the same and were related to the genus Trichosporon. Phylogenetic analysis based on the LSU D1/D2 plus ITS sequences placed this group within the Trichosporon brassicae clade, but it was clearly separated from any known species. In addition, physiological tests showed that this group had the unusual property of the inability to hydrolyse urea, which was distinctly different from the related taxon. Therefore a novel yeast species named Trichosporon heliocopridis sp. nov. (ex-type strain DD1-1T = TISTR 5946T = JCM 30786T = CBS 14168T) is proposed. The MycoBank number is MB812098.

Four strains of a novel ascomycetous yeast species were isolated from flowers in Iran and China. Phylogenetic analysis of the sequences of the ITS region (including 5.8S rRNA gene) and the LSU rRNA gene D1/D2 domains indicated that these strains belong to the Starmerella clade and show divergence from previously described species in this clade. Growth reactions on carbon and nitrogen sources were similar to those observed in related species of the Starmerella clade. Sexual reproduction was not observed after mating tests on different sporulation media. Based on physiological characteristics and phylogeny of rRNA gene sequences, the novel species is most closely related to Candida (iter. nom. Starmerella) powellii and Candida (iter. nom. Starmerella) floricola. It is therefore assigned to the genus Starmerella and described as Starmerella orientalis f.a., sp. nov. The type strain is SAM09T ( = IBRC-M 30204T = CBS 14142T). The MycoBank accession number is MB 814379.

Species of the genus Prototheca are achlorophyllous algae and ubiquitous in nature, and so far, six species have been listed in this genus: Prototheca wickerhamii, Prototheca zopfii, Prototheca blaschkeae, Prototheca cutis, Prototheca stagnora and Prototheca ulmea. A strain of the genus Prototheca, IFM 53848T, was isolated in Japan from a patient with systemic protothecosis and had been designated P. wickerhamii. Our previous study, by using PCR analysis, revealed that its SSU rRNA gene (rDNA) was distinctively larger than that of P. wickerhamii and other species of the genus Prototheca. In this study, molecular analysis showed that the exceptionally large SSU rDNA of IFM 53848T contains four group I introns. The morphology of IFM 53848T was indistinguishable from those of P. wickerhamii or P. cutis, and phylogenetic analyses, based on the sequences of the SSU rDNA exons and the D1/D2 region of the large subunit rDNA, indicated that IFM 53848T was closely related to P. cutis. On the other hand, unlike P. cutis, IFM 53848T failed to assimilate fructose or lysine and grew well at higher temperatures of up to 42 °C. In addition, the nucleotide sequence of the ribosomal internal transcribed spacer and the matrix assisted laser desorption ionization time-of-flight mass spectrometry profile of IFM 53848T were clearly distinct from those of P. cutis. The results strongly suggest that IFM 53848T represents a novel species, and so the seventh member of the genus Prototheca, which we have named Prototheca miyajii sp. nov. The unique characteristics of the strain may provide useful insights into the systematic taxonomy of the genus Prototheca.