Untitled

International Journal of Systematic and Evolutionary Microbiology (2006), 56, 2927–2930 Cyclobacterium lianum sp. nov., a marinebacterium isolated from sediment of an oilfield inthe South China Sea, and emended description ofthe genus Cyclobacterium Jiao-Yan Ying,1,2 Bao-Jun Wang,2 Su-Sheng Yang1and Shuang-Jiang Liu2 College of Biological Sciences, China Agricultural University, Beijing 100094, People’s 2State Key Laboratory of Microbial Resources, Institute of Microbiology, Chinese Academy of Sciences, ZhongGuanCun, Haidian, Beijing 100080, People’s Republic of China The marine bacterial strain HY9T was isolated from sediment from the South China Sea. StrainHY9T is aerobic, heterotrophic and rose-pigmented. The cells are non-motile and curved, i.e.
ring-like or horseshoe-shaped. The 16S rRNA gene sequence of strain HY9T was determined and BLAST searches revealed that it possessed significant sequence similarities with respect to Cyclobacterium species (92?8–93?6 %). Phylogenetic analysis confirmed that strain HY9T wastightly clustered with members of the genus Cyclobacterium. The cellular morphology andchemotaxonomic and phenotypic properties of strain HY9T showed that it should be classified as amember of the genus Cyclobacterium. Significant evolutionary distances and a range of phenotypicfeatures distinguished strain HY9T from previously described Cyclobacterium species. Hence,strain HY9T represents a novel species in the genus Cyclobacterium, for which the nameCyclobacterium lianum sp. nov. is proposed. The type strain is HY9T (=CGMCC 1.6102T=JCM14011T). On the basis of this study and previously described properties of Cyclobacteriumspecies, an emended description of the genus Cyclobacterium is proposed.
Many members of the phylum Bacteroidetes (or Cytophaga– designated strain HY9T and isolated from sediment of the Flavobacterium–Bacteroides) are involved in biogeochemical South China Sea. A polyphasic analysis of strain HY9T cycles in aquatic environments (Bowman et al., 2003; and indicated that the isolate represents a novel species of the references therein). For instance, members of the genus Cyclobacterium displaying the unique ring-like and horse-shoe-shaped morphology are common constituents of Strain HY9T was isolated from sediment of the Xijiang marine environments. At the time of writing, the genus oilfield in the South China Sea, near Fujian Province, China; Cyclobacterium (Raj & Maloy, 1990) contains two species sampling was performed at a depth of about 100 m. For with validly published names: Cyclobacterium marinum, isolation, serially diluted sediment samples were spread onto isolated from the deep sea in the Gulf of California (Raj low-organic marine agar 2216 plates [containing 0?5 g & Maloy, 1990; Euze´by, 1998), and Cyclobacterium peptone l21 and 0?1 g yeast extract l21; the salt composition amurskyense, isolated from water in the Sea of Japan and concentration were the same as in marine agar 2216 (Nedashkovskaya et al., 2005). Organisms sharing high (MA; Difco)] and incubated at 30 uC for 10 days. A colony levels of 16S rRNA gene sequence similarity with Cyclo- of HY9T was collected and subcultured on MA. The bacterium species have also been found in salt-marsh temperature and pH ranges for growth, the tolerance of sediment (GenBank accession no. AY259502) and a soda NaCl and the substrates used as sole carbon sources were lake (AF275712). In this study, we describe a rose-pigmented determined according to the methods described by Cho & bacterial strain with ring-like and horseshoe-shaped cells, Giovannoni (2003). Strain HY9T was tested on API ZYM(bioMe´rieux) and Biolog GN2 MicroPlate systems, accord-ing to the manufacturers’ instructions. All other biochem- The GenBank/EMBL/DDBJ accession number for the 16S rRNA genesequence of strain HY9T is DQ534063.
ical tests were performed according to Dong & Cai (2001).
Susceptibility to antibiotics was determined using filter- A transmission electron micrograph of strain HY9T, two phylogenetictrees and a table of fatty acid compositions are available as paper discs containing various antibiotics, as specified in the supplementary material in IJSEM Online.
species description. The morphology of cells grown on MA Table 1. Differential properties of strain HY9T and the type 27F and 1492R (Lane, 1991) and sequenced. Preliminary comparisons with sequences held in GenBank, performedusing BLASTN searches, revealed that the isolate was related Strains: 1, HY9T; 2, C. marinum LMG 13164T; 3, C. amurskyense to C. marinum. Further analysis of 16S rRNA gene sequences KMM 6143T. Data are from Nedashkovskaya et al. (2005) and was performed using MEGA, version 3.1 (Kumar et al., 2004).
Strain HY9T showed highest sequence similarity with C.
marinum DSM 745T (93?6 %) and C. amurskyense KMM6143T (92?8 %), followed by members of the genera (87?8–90?3 %), Chimaereicella (88?4 %) and Algoriphagus (88?3–89?5 %). Phylogenetic trees were constructed with MEGA, version 3.1, using the neighbour-joining method (Saitou & Nei, 1987), maximum parsimony (Fitch, 1971) and minimum evolution (Felsenstein, 1997). The resulting tree topology was evaluated using the Kimura two- parameter calculation model (Kumar et al., 2004) based on 1000 replicates. As shown in Fig. 1 and Supplementary Fig. S2 (available in IJSEM Online), strain HY9T formed a monophyletic clade with C. marinum and C. amurskyense.
The closest relatives of this clade were members of the genera Aquiflexum and Belliella (Brettar et al., 2004a, b).
The whole-cell fatty acid composition was determined from cells cultured at 30 uC on MA for 2 days. The cellular fatty acids were extracted, methylated and analysed by using the Sherlock Microbial Identification System (MIDI) according to the manufacturer’s instructions. The major cellular fattyacids were iso-C15 : 0 (28?3 %), summed feature 3 (iso-C15 : 02-OH and/or C for 2 days at 30 uC was studied using transmission electron 16 : 1v7c; 16?6 %), iso-C17 : 1v9c (10?3 %), 17 : 0 3-OH (8?0 %) and anteiso-C15 : 0 (6?4 %), similar to the profiles reported for C. marinum and C. amurskyense The novel isolate displayed the basic characteristics of (Nedashkovskaya et al., 2005). The complete fatty acid members of the genus Cyclobacterium, e.g. the colonies were composition of strain HY9T is given in Supplementary rose-pigmented and the cells were curved, ring-like or Table S1 (available in IJSEM Online). The G+C content of horseshoe-shaped (see Supplementary Fig. S1 available in the DNA was determined by thermal denaturation (Marmur IJSEM Online). Other phenotypic properties of strain HY9T & Doty, 1962; Seidler & Mandel, 1971) using DNA from are given in the species description and in Table 1.
Escherichia coli K-12 as a control. The G+C content ofstrain HY9T was 45?2 mol%, which is slightly higher than The almost-complete 16S rRNA gene (1485 bp) of strain those of the type strains of C. marinum (41?9 mol%) and C.
HY9T was amplified using the bacterial universal primers amurskyense (41?3 mol%) (Nedashkovskaya et al., 2005).
sequences of strain HY9T and related mem-bers Bootstrap values (expressed as percentagesof 1000 replicates) greater than 50 % areshown at branching points. Bar, 0?02 substi-tutions per nucleotide position.
International Journal of Systematic and Evolutionary Microbiology 56 On the basis of these results, we concluded that strain HY9T 1-phosphate are oxidized. Weak or variable results are represents a novel species within the genus Cyclobacterium, detected with glycogen, N-acetyl-D-galactosamine, i-ery- for which the name Cyclobacterium lianum sp. nov. is thritol, D-mannitol, D-psicose, L-rhamnose, D-sorbitol, proposed. The properties that serve to differentiate strain monomethyl succinate, D-gluconic acid, a-ketovaleric HY9T, C. marinum and C. amurskyense from each other are acid, succinic acid, L-alanyl glycine, L-asparagine, L-aspartic listed in Table 1. As some of the properties of C. lianum acid, glutamic acid, L-ornithine, L-proline, L-pyroglutamic (from this study) and C. amurskyense (Nedashkovskaya acid, DL-serine, L-threonine, c-aminobutyric acid, uridine, et al., 2005) were not included in or are not compatible with 2-aminoethanol and glucose 6-phosphate. Resistant to the the original description of the genus Cyclobacterium (Raj & following antibiotics (mg): gentamicin (10), neomycin (30), Maloy, 1990), an emended description of the genus is also polymyxin B (300), streptomycin (10) and tetracycline (30).
Sensitive to the following antibiotics (mg): ampicillin (10),carbenicillin (100), vancomycin (30), ciprofloxacin (5), Description of Cyclobacterium lianum sp. nov.
rifampicin (5), norfloxacin (10), chloramphenicol (30),benzyl penicillin (10), kanamycin (30) and erythromycin Cyclobacterium lianum (N.L. neut. adj. lianum pertaining to (15). The major cellular fatty acids (>5 %) are iso-C Li, named in honour of Professor Ji-Lun Li, who devotes himself to microbiological research and education in 16 : 1v7c; 16?6 %), iso-C17 : 1v9c (10?3 %), iso-C17 : 0 3-OH (8?0 %) and anteiso-C15 : 0 (6?4 %). The molar G+C content Cells are Gram-negative, aerobic and heterotrophic, non- motile, curved, ring-like or horseshoe-shaped, 0?4–0?5 mm The type strain, HY9T (=CGMCC 1.6102T=JCM 14011T), wide, and the outer diameter of rings is 1?5–1?8 mm.
was isolated from sediment from the Xijiang oilfield in the Colonies grown for 3 days on MA are circular (2–3 mm in diameter), light rose in colour and shiny. Growth occurs at15–42 uC (optimum 33 uC), at pH 6?5–9?0 (optimum pH 7?5–8?0) and with 0?1–12 % NaCl (optimum 1–4 %).
Positive for oxidase and catalase activities, but negative forarginine dihydrolase, urease and lecithinase activities.
Colonies on MA are pink-pigmented and shiny. Cells are Indole and H2S are not produced and nitrate is not reduced.
curved, ring-like or horseshoe-shaped. Neutrophilic and Aesculin and Tween 20 are hydrolysed. Tweens 40 and 80 are mesophilic. Optimal growth temperature range is 25–30 uC.
hydrolysed weakly; agar, casein, gelatin, starch, DNA and NaCl is required for growth. The major cellular fatty acids carboxymethyl-cellulose are not hydrolysed. Glucose, are iso-C15 : 0, summed feature 3 (iso-C15 : 0 2-OH and/or sucrose, D-melibiose, ribose, lactose, galactose, maltose, C16 : 1v7c), iso-C17 : 1v9c, iso-C17 : 0 3-OH and anteiso-C15 : 0.
melezitose, inulin, L-rhamnose, L-arabinose, D-raffinose, The DNA G+C content is 41–45 mol%.
trehalose, cellobiose, methyl a-D-glucoside and gluconateare utilized as sole carbon sources. Glycerol, mannitol, D-mannose, D-fructose, D-xylose, lactic acid, succinate, malate, pyruvate and L-glutamic acid are weakly utilized.
This work was supported by grants from the National Natural Science L-Fucose, L-sorbose, dulcitol, adonitol, myo-inositol, citrate, malonate, L-lysine, L-alanine, formic acid, butyric acid andcaprate are not utilized. Forms acid from glucose, ribose(weakly), sucrose, D-melibiose, lactose, galactose, maltose, melezitose, inulin, L-rhamnose, L-arabinose, D-raffinose, Bowman, J. P., Nichols, C. M. & Gibson, J. A. E. (2003). Algoriphagus trehalose, cellobiose, D-xylose, glycerol (weakly) and methyl ratkowskyi gen. nov., sp. nov., Brumimicrobium glaciale gen. nov., sp.
a-D-glucoside. Shows strong activity in the API ZYM nov., Cryomorpha ignava gen. nov., sp. nov. and Crocinitomix system for alkaline and acid phosphatases, leucine and catalasitica gen. nov., sp. nov., novel flavobacteria isolated from valine arylamidases, naphthol-AS-BI-phosphohydrolase, b- various polar habitats. Int J Syst Evol Microbiol 53, 1343–1355.
galactosidase, a- and b-glucosidases and N-acetyl-b-gluco- Brettar, I., Christen, R. & Ho¨fle, M. G. (2004a). Belliella baltica gen.
saminidase. Shows weak activity for esterases C4 and C8, nov., sp. nov., a novel marine bacterium of the Cytophaga– Flavobacterium–Bacteroides group isolated from surface water of the central Baltic Sea. Int J Syst Evol Microbiol 54, 65–70.
activity is shown for trypsin, a-chymotrypsin, b-glucur- Brettar, I., Christen, R. & Ho¨fle, M. G. (2004b). Aquiflexum balticum onidase, a-fucosidase or lipase (C14). In GN2 MicroPlates, gen. nov., sp. nov., a novel marine bacterium of the Cytophaga– dextrin, N-acetyl-D-glucosamine, L-arabinose, D-cellobiose, Flavobacterium–Bacteroides group isolated from surface water of the D-fructose, D-galactose, gentiobiose, a-D-glucose, a-D- central Baltic Sea. Int J Syst Evol Microbiol 54, 2335–2341.
Cho, J. C. & Giovannoni, S. J. (2003). Parvularcula bermudensis gen.
methyl b-D-glucoside, D-raffinose, sucrose, D-trehalose, nov., sp. nov., a marine bacterium that forms a deep branch in the a- turanose, D-galacturonic acid, DL-lactic acid, glucurona- Proteobacteria. Int J Syst Evol Microbiol 53, 1031–1036.
mide, L-alaninamide, L-alanine, DL-carnitine, 2,3-butane- Dong, X.-Z. & Cai, M.-Y. (2001). Determinative Manual for Routine Bacteriology. Beijing: Scientific Press.
Euze´by, J. P. (1998). Taxonomic note: necessary correction of Nedashkovskaya, O. L., Kim, S. B., Lee, M. S., Park, M. S., Lee, K. H., specific and subspecific epithets according to Rules 12c and 13b of Lysenko, A. M., Oh, H. W., Mikhailov, V. V. & Bae, K. S. (2005).
the International Code of Nomenclature of Bacteria (1990 Revision).
Cyclobacterium amurskyense sp. nov., a novel marine bacterium Int J Syst Bacteriol 48, 1073–1075.
isolated from sea water. Int J Syst Evol Microbiol 55, 2391–2394.
Felsenstein, J. (1997). An alternative least-squares approach to Raj, H. D. & Maloy, S. R. (1990). Proposal of Cyclobacterium inferring phylogenies from pairwise distances. Syst Biol 46, 101–111.
marinum gen. nov., comb. nov. for a marine bacterium previously Fitch, W. M. (1971). Toward defining the course of evolution: assigned to the genus Flectobacillus. Int J Syst Bacteriol 40, 337–347.
minimum changes for a specific tree topology. Syst Zool 20, 406–416.
Saitou, N. & Nei, M. (1987). The neighbor-joining method: a new Kumar, S., Tamura, K. & Nei, M. (2004). MEGA3: integrated software method for reconstructing phylogenetic trees. Mol Biol Evol 4, 406–425.
for molecular evolutionary genetics analysis and sequence alignment.
Seidler, R. J. & Mandel, M. (1971). Quantitative aspects of deoxyribonucleic acid renaturation: base composition, state of Lane, D. J. (1991). 16S/23S rRNA sequencing. In Nucleic Acid chromosome replication, and polynucleotide homologies. J Bacteriol E. Stackebrandt & M. Goodfellow. Chichester: Wiley.
Zhang, D., Yang, H., Huang, Z., Zhang, W. & Liu, S.-J. (2002).
Marmur, J. & Doty, P. (1962). Determination of the base composition Rhodopseudomonas faecalis sp. nov., a phototrophic bacterium of deoxyribonucleic acid from thermal denaturation temperature.
isolated from an anaerobic reactor that digests chicken faeces. Int J Syst Evol Microbiol 52, 2055–2060.
International Journal of Systematic and Evolutionary Microbiology 56

Source: http://eco.ibcas.ac.cn/group/baiyf/pdf/jiaoyan_ying/Int_J_Syst_Evol_Microbiol_Ying_2006.pdf