Planktomarina temperata gen. nov., sp. nov., belonging to the globally distributed RCA cluster of the marine Roseobacter clade, isolated from the German Wadden Sea.

Four heterotrophic bacterial strains belonging to the globally distributed marine RCA (Roseobacter clade-affiliated) cluster (family Rhodobacteraceae, class Alphaproteobacteria) were obtained from coastal seawater samples. Strain RCA23(T) was isolated from a 10(-7) dilution culture inoculated with seawater from the German Wadden Sea (southern North Sea), reflecting the high abundance of RCA bacteria in this habitat. Strains IMCC1909, IMCC1923 and IMCC1933 were isolated from diluted seawater (10(-3)) of the Yellow Sea, South Korea. Based on 16S rRNA gene sequence comparison, Octadecabacter antarcticus 307(T) is the closest described relative of the RCA strains, with 95.4-95.5 % sequence similarity. Cells of RCA23(T), IMCC1909, IMCC1923 and IMCC1933 are small motile rods requiring sodium ions. Optimal growth of RCA23(T) occurs at 25 °C and within a very narrow pH range (pH 7-8, optimum pH 7.5). The DNA G+C base content of RCA23(T) is 53.67 mol%. The major respiratory lipoquinone is ubiquinone-10 (Q-10) and the dominant fatty acids (>1 %) are 12 : 1 3-OH, 16 : 1ω7c, 16 : 0, 18 : 1ω7c, 18 : 0 and 11-methyl 18 : 1ω7c. The polar lipid pattern indicated the presence of phosphatidylglycerol, two unidentified aminolipids and two unidentified phospholipids. On marine agar, RCA23(T) forms non-pigmented, transparent to light beige, small (<1 mm), circular, convex colonies. Strain RCA23(T) harbours all genes for the production of bacteriochlorophyll a (BChl a). Genes encoding the light-harvesting reaction centre of BChl a (pufM) were identified in all RCA strains. No visible pigmentation was observed for any of the strains under laboratory conditions, but spectrophotometric analysis revealed weak production of BChl a by RCA23(T). Morphological, physiological and genotypic features of strain RCA23(T) suggest that it represents a novel species of a new genus within the Rhodobacteraceae, for which we propose the name Planktomarina temperata gen. nov., sp. nov., described previously by Giebel et al. [ISME J 5 (2011), 8-19] as 'Candidatus Planktomarina temperata'. The type strain of Planktomarina temperata is RCA23(T) ( = DSM 22400(T) = JCM 18269(T)).

Phenotypic landscape of a bacterial cell.

The explosion of sequence information in bacteria makes developing high-throughput, cost-effective approaches to matching genes with phenotypes imperative. Using E. coli as proof of principle, we show that combining large-scale chemical genomics with quantitative fitness measurements provides a high-quality data set rich in discovery. Probing growth profiles of a mutant library in hundreds of conditions in parallel yielded > 10,000 phenotypes that allowed us to study gene essentiality, discover leads for gene function and drug action, and understand higher-order organization of the bacterial chromosome. We highlight new information derived from the study, including insights into a gene involved in multiple antibiotic resistance and the synergy between a broadly used combinatory antibiotic therapy, trimethoprim and sulfonamides. This data set, publicly available at http://ecoliwiki.net/tools/chemgen/, is a valuable resource for both the microbiological and bioinformatic communities, as it provides high-confidence associations between hundreds of annotated and uncharacterized genes as well as inferences about the mode of action of several poorly understood drugs.

Perlucidibaca piscinae gen. nov., sp. nov., a freshwater bacterium belonging to the family Moraxellaceae.

A freshwater bacterium, designated IMCC1704(T), was isolated from a eutrophic pond. The strain was Gram-negative, oxidase-positive, catalase-negative, chemoheterotrophic and facultatively aerobic with cells that were motile rods with a single polar flagellum. Based on 16S rRNA gene sequence similarity analyses, the novel strain was most closely related to the genera Alkanindiges (91.7%), Acinetobacter (89.0-91.2%), Moraxella (87.9-90.1%), Psychrobacter (87.2-89.5%) and Enhydrobacter (87.8%). Phylogenetic trees generated using 16S rRNA gene sequences showed that the novel isolate belonged to the family Moraxellaceae of the class Gammaproteobacteria and formed a distinct phyletic lineage within the family. The DNA G+C content of the strain was 63.1 mol% and the predominant constituents of the cellular fatty acids were C(16:1)omega7c and/or iso-C(15:0) 2-OH (21.2%), C(18:1)omega7c (12.8%) and C(12:0) 3-OH (12.3%). These chemotaxonomic properties, together with several phenotypic characteristics, differentiated the novel strain from other members of the family Moraxellaceae. From the taxonomic data, which revealed the distant relationship of the new strain to the related genera, the strain should be classified as a novel genus and species in the family Moraxellaceae, for which the name Perlucidibaca piscinae gen. nov., sp. nov. is proposed. The type strain of Perlucidibaca piscinae sp. nov. is IMCC1704(T) (=KCCM 42363(T)=NBRC 102354(T)).

Lutimonas vermicola gen. nov., sp. nov., a member of the family Flavobacteriaceae isolated from the marine polychaete Periserrula leucophryna.

A Gram-negative, yellow-coloured, non-motile, chemoheterotrophic, strictly aerobic, rod-shaped bacterium, designated IMCC1616(T), was isolated from the marine polychaete Periserrula leucophryna inhabiting tidal flat sediment of the Yellow Sea, and characterized by a polyphasic approach. The temperature, pH and NaCl ranges for growth were 3-37 degrees C, pH 5.0-11.0 and 0.5-7.5 %. Based on 16S rRNA gene sequence similarity analyses, the strain was most closely related to members of the genera Lutibacter (90.7 %), Tenacibaculum (89.2-90.4 %) and Polaribacter (88.4-90.2 %). Phylogenetic analysis using three treeing algorithms based on 16S rRNA gene sequences indicated that the strain formed a distinct lineage within the family Flavobacteriaceae. The DNA G+C content of the strain was 40.1 mol% and the predominant cellular fatty acids were iso-C(15 : 0) (16.5 %), anteiso-C(15 : 0) (10.9 %), iso-C(17 : 0) 3-OH (8.8 %) and iso-C(17 : 1) omega 9c (8.2 %). The DNA G+C content, large amount of iso-C(17 : 1) omega 9c and several phenotypic characteristics, including growth temperature and catalase activity, differentiated the strain from other related genera in the family. Therefore, from the taxonomic evidence collected in this study, it is proposed that strain IMCC1616(T) represents a new genus and species named Lutimonas vermicola gen. nov., sp. nov. The type strain of Lutimonas vermicola is strain IMCC1616(T) (=KCCM 42379(T) =NBRC 102041(T)).

Litoricolaceae fam. nov., to include Litoricola lipolytica gen. nov., sp. nov., a marine bacterium belonging to the order Oceanospirillales.

A Gram-negative, non-motile, chemoheterotrophic, facultatively aerobic, short-rod-shaped bacterium, designated IMCC1097(T), was isolated from coastal seawater (10 m depth) of the East Sea, Korea. The temperature, pH and NaCl ranges for growth were 15-30 degrees C, pH 5.0-10.0 and 1.5-10 % NaCl. The colonies of the strain were very small, having a mean diameter of 0.05 mm. 16S rRNA gene sequence data indicated that the strain was most closely related to genera within the class Gammaproteobacteria. Members of the most closely related genera showed less than 90 % sequence similarity and included Saccharospirillum (89.3 %), Oleiphilus (88.7 %), Reinekea (88.2 %), Alcanivorax (86.4-87.6 %) and Zooshikella (87.6 %), which represent five different families of the order Oceanospirillales. Phylogenetic analyses showed that this marine strain represented a distinct phylogenetic lineage in the order Oceanospirillales and could not be assigned to any of the defined families in the order. The predominant fatty acids were C(16 : 1) omega 7c and/or iso-C(15 : 0) 2-OH, C(18 : 1) omega 7c and C(10 : 0) 3-OH, and the DNA G+C content was 57.9 mol%. These chemotaxonomic properties, together with phenotypic characteristics, served to differentiate the strain from phylogenetically closely related genera. The very low sequence similarities (<90 %) and distant relationships between IMCC1097(T) and members of the order Oceanospirillales suggested that the strain merited classification within a novel genus within a novel family in the order. On the basis of taxonomic evidence collected in this study, a novel genus and species are proposed, Litoricola lipolytica gen. nov., sp. nov., within a new family Litoricolaceae fam. nov. Strain IMCC1097(T) (=KCCM 42360(T) =NBRC 102074(T)) is the type strain of Litoricola lipolytica.

Ruegeria pelagia sp. nov., isolated from the Sargasso Sea, Atlantic Ocean.

Gram-negative, facultatively aerobic, chemoheterotrophic, short rod-shaped marine bacterial strains HTCC2662(T) and HTCC2663, isolated from the Sargasso Sea by using a dilution-to-extinction culturing method, were investigated to determine their taxonomic position. Characterization of the two strains by phenotypic and phylogenetic analyses revealed that they belonged to the same species. The DNA G+C content of strain HTCC2662(T) was 58.4 mol% and the predominant cellular fatty acids were C(18 : 1) omega 7c (52.5 %), C(16 : 0) 2-OH (13.5 %) and C(18 : 1) 11-methyl omega 7c (12.2 %). Phylogenetic analysis of the 16S rRNA gene sequences showed that the strains represented a distinct line of descent within the genus Ruegeria, with highest sequence similarities to Ruegeria atlantica DSM 5823(T) (97.2 %), Ruegeria lacuscaerulensis DSM 11314(T) (96.5 %) and Ruegeria pomeroyi DSM 15171(T) (95.6 %). Several phenotypic characteristics, including facultatively requiring NaCl and oxygen for growth, together with the cellular fatty acid composition, differentiated strain HTCC2662(T) from other members of the genus Ruegeria. Based on phenotypic, chemotaxonomic and phylogenetic traits, it is suggested that strains HTCC2662(T) and HTCC2663 represent a novel species of the genus Ruegeria, for which the name Ruegeria pelagia sp. nov. is proposed. The type strain is HTCC2662(T) (=KCCM 42378(T)=NBRC 102038(T)).

Maritimibacter alkaliphilus gen. nov., sp. nov., a genome-sequenced marine bacterium of the Roseobacter clade in the order Rhodobacterales.

A Gram-negative, chemoheterotrophic, strictly aerobic, alkaliphilic, rod-shaped marine bacterium, designated HTCC2654(T), was isolated from the western Sargasso Sea by using a dilution-to-extinction culturing method. Phylogenetic analyses based on 16S rRNA gene sequences showed that strain HTCC2654(T) belonged to the Roseobacter clade of the order Rhodobacterales. The 16S rRNA gene sequence similarity of the strain with respect to other members of the Roseobacter clade ranged from 90.4 to 95.1 %. In the phylogenetic analyses, the strain formed an independent phyletic line and could not be assigned to any other known genera of the Rhodobacterales. The DNA G+C content of strain HTCC2654(T) was 61.7 mol% by HPLC and 64.1 mol% from genome sequences. The predominant constituents of the cellular fatty acids were C(16 : 0) 2-OH (27.3 %), 11-methyl C(18 : 1) omega 7c (19.6 %) and C(18 : 1) omega 7c (17.3 %), and the major polar lipids were phosphatidylethanolamine, phosphatidylglycerol and phosphatidylcholine, which served to differentiate the strain from other members of the Roseobacter clade. On the basis of the taxonomic data obtained in this study, strain HTCC2654(T) represents a novel genus and species, for which the name Maritimibacter alkaliphilus gen. nov., sp. nov. is proposed. The type strain is HTCC2654(T) (=KCCM 42376(T)=NBRC 102057(T)).

Marinobacterium litorale sp. nov. in the order Oceanospirillales.

A bacterial strain named IMCC1877(T) was obtained from surface seawater collected near the coast of Deokjeok island (Yellow Sea), using a standard dilution-plating method. The strain was Gram-negative, chemoheterotrophic and facultatively anaerobic, requiring NaCl, and cells were motile rods with a single polar flagellum. Colonies on marine agar were very small (average diameter 0.1 mm). Based on 16S rRNA gene sequences, the most closely related species to strain IMCC1877(T) was Marinobacterium stanieri (93.7 % sequence similarity to the type strain). Phylogenetic analyses based on 16S rRNA gene sequences showed that this marine isolate belonged to the order Oceanospirillales and formed an independent phyletic line within the clade forming the genus Marinobacterium. The DNA G+C content of the strain was 60.7 mol% and the predominant constituents of the cellular fatty acids were C(18 : 1) omega 7c (36.6 %), C(16 : 1) omega 7c and/or iso-C(15 : 0) 2-OH (26.7 %) and C(16 : 0) (24.3 %). Based on the taxonomic data, only a distant relationship could be established between strain IMCC1877(T) and other Marinobacterium species; the strain therefore represents a novel species of the genus Marinobacterium, for which the name Marinobacterium litorale sp. nov. is proposed. The type strain is IMCC1877(T) (=KCTC 12756(T)=LMG 23872(T)).

Puniceicoccus vermicola gen. nov., sp. nov., a novel marine bacterium, and description of Puniceicoccaceae fam. nov., Puniceicoccales ord. nov., Opitutaceae fam. nov., Opitutales ord. nov. and Opitutae classis nov. in the phylum 'Verrucomicrobia'.

A Gram-negative, chemoheterotrophic, facultatively anaerobic coccus, designated IMCC1545(T), was isolated from the digestive tract of a marine clamworm, Periserrula leucophryna, inhabiting a tidal flat of the Yellow Sea. Cells of strain IMCC1545(T) are non-motile, dividing by binary fission. The predominant fatty acids are anteiso-C(15 : 0) and C(18 : 0). The respiratory quinone is menaquinone-7 and the DNA G+C content is 52.1 mol%. Phylogenetic analyses based on 16S rRNA gene sequences using three treeing algorithms revealed that the strain formed a novel genus-level lineage within the phylum 'Verrucomicrobia'. The most closely related named organisms to strain IMCC1545(T) are 'Fucophilus fucoidanolyticus' SI-1234 (86.5 % 16S rRNA gene sequence similarity), Alterococcus agarolyticus ADT3(T) (81.8 %) and Opitutus terrae PB90-1(T) (80.3 %), which belong to subdivision 4 of the 'Verrucomicrobia'. Subdivision 4 of the 'Verrucomicrobia' (here named Opitutae classis nov.) was divided into two clades, a clade containing strain IMCC1545(T) and a clade containing Opitutus terrae. From the taxonomic data obtained in this study, it is proposed that the new marine isolate be placed into a novel genus and species named Puniceicoccus vermicola gen. nov., sp. nov. (the type strain of Puniceicoccus vermicola is IMCC1545(T)=KCCM 42343(T)=NBRC 101964(T)) within Puniceicoccaceae fam. nov and Puniceicoccales ord. nov in the class Opitutae. The family Opitutaceae fam. nov. and order Opitutales ord. nov. are also formally proposed.

Detection of human adenoviruses and enteroviruses in Korean oysters using cell culture, integrated cell culture-PCR, and direct PCR.

Oysters are known to be carriers of food-born diseases, but research on viruses in Korean oysters is scarce despite its importance for public health. We therefore tested oysters cultivated in Goheung, Seosan, Chungmu, and Tongyeong, for viral contamination using cell culture and integrated cell culture PCR (ICC-PCR) with Buffalo green monkey kidney (BGMK) and human lung epithelial (A549) cells. Additional screens via PCR, amplifying viral nucleic acids extracted from oysters supplemented our analysis. Our methods found 23.6%, 50.9%, and 89.1% of all oysters to be positive for adenoviruses when cell culture, ICC-PCR, and direct PCR, respectively, was used to conduct the screen. The same methodology identified enteroviruses in 5.45%, 30.9%, and 10.9% of all cases. Most of the detected enteroviruses (81.3%) were similar to poliovirus type 1; the remainder resembled coxsackievirus type A1. A homology search with the adenoviral sequences revealed similarities to adenovirus subgenera C (type 2, 5, and 6), D (type 44), and F (enteric type 40 and 41). Adenovirus-positive samples were more abundant in A549 cells (47.3%) than in BGMK cells (18.2%), while the reverse was true for enteroviruses (21.8% vs. 14.5%). Our data demonstrate that Korean oysters are heavily contaminated with enteric viruses, which is readily detectable via ICC-PCR using a combination of A549 and BGMK cells.