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1 State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100080, P. R. China
2 College of Biological Sciences, China Agricultural University, Beijing 100094, P. R. China
Correspondence
Shuang-Jiang Liu
liusj{at}sun.im.ac.cn
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ABSTRACT |
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5 % of the total fatty acids) were iso-15 : 1 (5.0 %), iso-15 : 0 (6.8 %), anteiso-15 : 0 (6.4 %), 15 : 0 (10.4 %), iso-16 : 0 (13.5 %), summed feature 3 (comprising iso-15 : 0 2-OH and/or 16 : 1
7c; 6.3 %), iso-17 : 0 3-OH (5.2 %) and 17 : 0 2-OH (5.0 %). Cells contained menaquinone 6. Based on the phylogenetic and phenotypic analyses, strain HY1T should be classified as representing a novel species within the genus Salegentibacter, for which the name Salegentibacter catena sp. nov. is proposed. The type strain is HY1T (=CGMCC 1.6101T=JCM 14015T). Based on this study and on previously described Salegentibacter species, an emended description of the genus Salegentibacter is given.
Maximum-parsimony and minimum-evolution phylogenetic trees based on 16S rRNA gene sequences and electron micrographs of cells of strain HY1T are available as supplementary material in IJSEM Online.
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to accommodate strictly aerobic, non-motile, yellow-pigmented and moderately halophilic (optimal growth with about 5 % NaCl) bacteria. The type species Salegentibacter salegens was isolated from a hypersaline Antarctic lake (Dobson et al., 1993). Other members of this genus are as follows: Salegentibacter holothuriorum, isolated from the edible holothurian Apostichopus japonicus (Nedashkovskaya et al., 2004), Salegentibacter mishustinae from the sea urchin Strongylocentrotus intermedius (Nedashkovskaya et al., 2005), Salegentibacter agarivorans associated with sponge (Nedashkovskaya et al., 2006) and Salegentibacter flavus from sea sediment (Ivanova et al., 2006). Some of the recently described species showed characteristics that are not compatible with the original description of the genus Salegentibacter, e.g. S. agarivorans is motile by gliding (Nedashkovskaya et al., 2006) and most Salegentibacter species grow optimally with NaCl concentrations lower than 5 %. Besides Salegentibacter, many members of the family Flavobacteriaceae are found in various marine habitats, e.g. the genera Formosa (Ivanova et al., 2004), Gillisia (Van Trappen et al., 2004), Subsaximicrobium, Subsaxibacter and Lacinutrix (Bowman & Nichols, 2005). The abundance of marine Flavobacteriaceae suggests that members of the family potentially play important roles in marine ecosystems and in biogeochemical cycles in interaction with other marine organisms (Bowman & McCuaig, 2003).
During a study of microbial diversity of marine sediments in the South China Sea, a novel heterotrophic, aerobic, saffron yellow-pigmented, non-gliding, Gram-negative bacterial strain (HY1T) was isolated. Here we describe the taxonomic investigation of the new isolate following the minimal standards for describing new taxa of the family Flavobacteriaceae (Bernardet et al., 2002).
Strain HY1T was isolated from sediment of Xijiang oilfield in the South China Sea (114° E 21° N), from a depth of about 100 m. For isolation, serially diluted sediment samples were spread on low-organic marine (LOM) agar plates [containing (g l–1): 0.5 peptone, 0.1 yeast extract and 15 agar, with the same salts composition and concentrations as Bacto marine broth 2216 (Difco)]. Colonies were picked and repeatedly restreaked on fresh LOM agar plates, until purity was confirmed. The isolates were subsequently tested for growth on plain marine agar 2216 at 30 °C, and cultivation and routine maintenance were subsequently carried out in plain marine broth 2216.
The universal primers 27F and 1492R were used to amplify the 16S rRNA gene (Lane, 1991). The PCR product was ligated to T-vector and sequenced. BLASTN searches revealed that the 16S rRNA gene sequence of strain HY1T (1487 bp) showed high similarity with those of members of the genera Gillisia (91.7–93.8 %) and Salegentibacter (92.6–93.5 %). Alignments of the 16S rRNA gene sequences revealed that strain HY1T shared characteristic nucleotide(s) at positions 71 and 72, 79 and 80 and 117 with members of the genus Gillisia [numbering according to the 16S rRNA gene sequence of strain HY1T (DQ640642)] and characteristic nucleotide(s) at positions 125 and 936–942 with members of the genus Salegentibacter (Fig. 1). When phylogenetic trees were constructed using three different algorithms (neighbour-joining, maximum-parsimony and minimum-evolution) with MEGA version 3.1 (Kumar et al., 2004), strain HY1T clustered more frequently with Salegentibacter species than with Gillisia species, although bootstrap values were low. The neighbour-joining tree is shown in Fig. 2, while the maximum-parsimony and minimum-evolution trees are available as Supplementary Fig. S1 in IJSEM Online.
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; Seidler & Mandel, 1971), with DNA from Escherichia coli K-12 as a control. The DNA G+C content of strain HY1T was 44.4 mol%. Biomass for the analysis of isoprenoid quinones was grown in marine broth 2216 and quinones were determined according to Collins (1985) and Wu et al. (1989). The isoprenoid quinone of strain HY1T was menaquinone-6. The whole-cell fatty acid composition of strain HY1T was determined using the Sherlock Microbial Identification System, following the manufacturer's instructions. The fatty acid composition is given in the species description. Overall, it was similar to those of other Salegentibacter species, with the exception of Salegentibacter holothuriorum, which contains a lower amount of anteiso-15 : 0.
Physiological and biochemical characteristics of strain HY1T were determined according to the methods described by Cho & Giovannoni (2003) and Bernardet et al. (2002) and by using API ZYM kits (bioMérieux), according to the manufacturer's instructions. Susceptibility to antibiotics was determined using filter-paper discs containing various antibiotics on cultures incubated at 30 °C on marine agar 2216 for 2 days. Cells often occurred in chains and some cells of strain HY1T possessed appendages (Supplementary Fig. S2 in IJSEM Online) in older cultures. Other morphological, physiological and biochemical properties of strain HY1T are given in the species description. Characteristics that differentiate strain HY1T from recognized Salegentibacter species are given in Table 1.
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), an emended description of the genus is also given.
Emended description of the genus Salegentibacter McCammon and Bowman 2000
Cells are Gram-negative rods, occurring as single cells, pairs or chains. Non-motile or motile by gliding. Do not form spores or other resting stages. Colonies are yellow-pigmented. Flexirubin-type pigments are absent. Possess a strictly aerobic, chemo-organotrophic metabolism. Catalase-positive and positive for alkaline phosphatase (not determined for Salegentibacter flavus). Grow in 0–20 % NaCl (all grow in 1–8 % NaCl) and NaCl is required by most strains. Grow at 0–42 °C. Predominant fatty acids are iso-15 : 1, iso-15 : 0, anteiso-15 : 0, 15 : 0, iso-16 : 0, summed feature 3 (comprising iso-15 : 0 2-OH and/or 16 : 17c), iso-17 : 0 3-OH and 17 : 0 2-OH. The major respiratory quinone is menaquinone-6. The DNA G+C content is 36.8–44.4 mol%. Known habitats are seawater and marine sediments, marine plants and invertebrates and hypersaline lakes. The type species is Salegentibacter salegens.
Description of Salegentibacter catena sp. nov.
Salegentibacter catena (ca.te'na. L. n. catena chain, referring to the fact that cells frequently occur in chains).
Cells are Gram-negative rods, 0.5–0.8x2–6 µm, frequently occurring in chains and producing appendages in older cultures. Strictly aerobic. Devoid of flagellar and gliding motilities. Catalase-positive and oxidase-negative. Colonies on marine agar 2216 are saffron yellow, slimy, circular, convex and shiny, 3–5 mm in diameter after 5 days of incubation at 30 °C. Growth occurs at 15–42 °C (optimum, 33 °C), at pH 6.5–9.0 (optimum, 7.5–8.2) and in 0.1–10 % NaCl (optimum, 1–4 %). Arginine dihydrolase, urease and lecithinase are negative. H2S is produced, but indole is not. Nitrate is not reduced. Aesculin, casein, gelatin, starch and Tween 20 are hydrolysed, but agar, DNA and carboxymethylcellulose are not. Weak growth and acid formation occur with glucose, but not with sucrose, mannose, maltose, arabinose, lactose, rhamnose, melibiose, fucose, raffinose, xylose, adonitol, inositol, sorbitol, gluconate, malonate, galactose, malate, citrate, pyruvate, lactate, propionate, acetate, alanine or proline. In API ZYM kits, activities of alkaline and acid phosphatases, esterase lipase (C8), leucine and valine arylamidases, naphthol-AS-BI-phosphohydrolase and 
-glucosidase are strong and activities of esterase (C4), lipase (C14), cystine arylamidase and trypsin are weak. Negative for activities of -chymotrypsin, 
-galactosidase, -glucosidase, -glucuronidase, -fucosidase, -galactosidase, -mannosidase and N-acetyl--glucosaminidase. Resistant to gentamicin (10 µg), neomycin (30 µg), polymyxin B (300 µg) and streptomycin (10 µg), but sensitive to ampicillin (10 µg), carbenicillin (100 µg), tetracycline (30 µg), vancomycin (30 µg), ciprofloxacin (5 µg), rifampicin (5 µg), norfloxacin (10 µg), chloramphenicol (30 µg), benzylpenicillin (10 µg), kanamycin (30 µg) and erythromycin (15 µg). Cellular fatty acids (>1 % of the total fatty acids) are iso-14 : 0 (2.0 %), iso-15 : 1 (5.0 %), anteiso-15 : 1 (1.3 %), iso-15 : 0 (6.8 %), anteiso-15 : 0 (6.4 %), 15 : 16c (2.6 %), 15 : 0 (10.4 %), iso-16 : 1 (2.8 %), iso-16 : 0 (13.5 %), summed feature 3 (comprising iso-15 : 0 2-OH and/or 16 : 17c; 6.3 %), 16 : 0 (4.3 %), iso-15 : 0 3-OH (1.2 %), 15 : 0 2-OH (1.8 %), iso-17 : 19c (2.3 %), anteiso-17 : 19c (2.3 %), 17 : 18c (1.5 %), 17 : 16c (2.4 %), iso-16 : 0 3-OH (3.6 %), 18 : 17c (1.3 %), 18 : 0 (2.6 %), iso-17 : 0 3-OH (5.2 %) and 17 : 0 2-OH (5.0 %).
The G+C content of the DNA of the type strain is 44.4 mol%. The type strain is HY1T (=CGMCC 1.6101T=JCM 14015T), which was isolated from sediment of Xijiang oilfield in the South China Sea.
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ACKNOWLEDGEMENTS |
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