Download Two novel species of marine phototrophic Gammaproteobacteria

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Roseomonas oryzae sp. nov., isolated from paddy rhizosphere
soil
Ramaprasad, E.V.V1., Sasikala, Ch.1*and Ramana, Ch.V.2
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Bacterial Discovery Laboratory, Centre for Environment, Institute of Science and
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Technology, J. N. T. University, Kukatpally, Hyderabad 500 085, INDIA.
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*Author for correspondence: Ch. Sasikala
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e-mail: [email protected]; [email protected]
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Subject category: New taxa - Proteobacteria
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Running title: Roseomonas oryzae sp. nov.
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Department of Plant Sciences, School of Life Sciences, University of Hyderabad,
P.O. Central University, Hyderabad 500 046, INDIA.
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The GenBank/EMBL/DDBJ accession numbers for the 16S rRNA gene sequences of
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strain JA288T is LN810637.
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A non-motile, cocci-shaped, pale-pink-pigmented bacterium, designated strain
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JC288T, was isolated from a paddy rhizosphere soil collected from Western Ghats,
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Kankumbi, Karnataka, India. Cells were found to be Gram-stain-negative, catalase and
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oxidase-positive, the major fatty acids were C16:0, C16:1ω7c/C16:1ω6c, C18:1ω7c/C18:16c
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and C18:12OH. The predominant respiratory quinone was Q-10 and genomic DNA G+C
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content
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phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, four unidentified
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aminolipids (AL1-4), three unidentified phospholipids (PL1,3,4), two unidentified lipids
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(L1-3), an aminophospholipid and a glycolipid. Hydroxyspirilloxanthin is the major
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carotenoid of strain JC288T. 16S rRNA gene sequence comparisons indicated that strain
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JC288T represents a member of the genus Roseomonas within the family
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Acetobacteraceae of the phylum Proteobacteria. Strain JC288T shared the highest 16S
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rRNA gene sequence similarity with Roseomonas rhizosphaerae YW11T (97.3 %),
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Roseomonas aestuarii JC17T (97.1%), R. cervicalis CIP104027T (95.9%) and other
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members of the genus Roseomonas (<95.5 %). The distinct genomic difference and
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morphological, physiological and chemotaxonomic differences from the previously
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described taxa support the classification of strain JC288T as a representative of a novel
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species of the genus Roseomonas, for which the name Roseomonas oryzae sp. nov. is
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proposed. The type strain is JC288T ( = KCTC 42542T = LMG 28711T).
was
67.5
mol%.
Strain
JC288T
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contained
diphosphatidylglycerol,
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Members of the genus Roseomonas are pink, Gram-stain-negative, coccobacilli
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and have oxidative metabolism. Though a few species are cultured from clinical
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specimens and are considered to be pathogenic for humans (Rihs et al., 1993; Sandoe et
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al., 1997; Bibashi et al., 2000; Subudhi et al., 2001; Han et al., 2003; McLean et al.,
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2006), large number of species were isolated from water samples (September et al., 2004;
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Gallego et al., 2006; Jiang et al., 2006; Furuhata et al., 2008; Baik et al., 2012), as
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associated with deep-water marine invertebrates (Sfanos et al., 2005), soil samples (Chen
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et al., 2014; Yoon et al., 2007) and also from air (Kim et al., 2013) indicating their
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ubiquitous nature of distribution. At the time of writing, 18 species with two subspecies
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names were validly published in the genus Roseomonas. Though “Roseomonas aceris”
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(Tonouchi & Tazawa, 2014), “Roseomonas musae” (Nutaratat et al., 2013), were
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effectively published, these species name still remains not validated. Phylogenetically,
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the genus Roseomonas (Rihs et al., 1993) is classified under the family Acetobacteraceae
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of the order Rhodospirillales in the class Alphaproteobacteria. In this communication, we
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propose a novel species of the genus Roseomonas.
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Strain JC288T was isolated from a paddy rhizosphere soil sample collected from
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Western Ghats, Kankumbi, Karnataka, India (GPS positioning of the sample collection
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site: N15.69487 E074.20980) on 1 November 2014. One gram dry soil was serially
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diluted [10- fold dilution in saline (0.6% NaCl)] and plated on a medium (pH 7.0)
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containing (g.l-1) yeast extract (0.5), peptone (0.5), casein enzyme hydrolysate (0.5), yeast
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extract (0.5), dextrose (0.5), water soluble starch (0.5), dipotassium phosphate (0.3),
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magnesium sulphate (0.05) and sodium pyruvate (0.3). Three distinct colony
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morphologies were observed from plates incubated at 30 ºC for 3 days. The pink
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pigmented colonies were purified by subsequent streaking on the same medium. The
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purified isolate was designated as strain JA288T, preserved as glycerol stocks and by
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lyophilization and stored at 4 ºC. The purified culture was grown in broth (conical flasks
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500 ml) with shaking (160 rpm) for chemoheterotrophic growth. For routine culturing
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and for physiological tests, strain JC288T was grown at pH 7.0 and at 30 ºC in the
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tryptone soya broth (TSB) medium.
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Genomic DNA was isolated and purified by the method described by Marmur
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(1961) and the G+C content (mol%) of the DNA as determined by HPLC (Mesbah et al.,
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1989) was 67.5 mol%. Cell material for 16S rRNA gene sequencing was taken from a
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well-isolated colony and the gene was PCR amplified and sequenced as described
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previously (Venkata Ramana et al., 2010). BLAST search analysis of the 16S rRNA gene
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sequence (1401 nt) was performed using the EzTaxon-e server (Kim et al., 2012). Strain
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JC288T showed the highest sequence similarity to Roseomonas rhizosphaerae YW11T
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(97.3 %), Roseomonas aestuarii JC17T (97.1%) and <95.9 % with other members of the
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genus Roseomonas. 16S rRNA gene sequences were aligned by MUSCLE program using
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UPGMA clustering method of MEGA version 6 (Tamura et al., 2013) software. For
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constructing NJ, ML and MP phylogenetic trees the following statistical methods were
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used. For NJ- Kimura 2-parameter model (Kimura, 1980), uniform rates, pairwise
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deletion was used. For ML- Kimura 2-parameter model, uniform rates. Heuristic search
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algorithm - Nearest-Neighbor-Interchange (NNI) with complete deletion was used. And
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For MP - Subtree-Pruning-Regrafting (SPR) as heuristic algorithm with complete
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deletion was used. The combined phylogenetic tree (NJ, ML, MP) confirmed the
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clustering of strain JC288T with members of the genus Roseomonas (Fig. 1).
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DNA–DNA reassociation between strains was determined using a membrane
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filter technique as described previously (Chakravarthy et al., 2012; Seldin & Dubnau,
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1985). When strain JC288T was labelled radioactively, the level of DNA–DNA
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reassociation with R. rhizosphaerae KACC 17225T (=YW11T) was 27±2% and 30±2%
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with R. aestuarii JC17T (Supplementary Fig. S1; near similar values were obtained when
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R. rhizosphaerae KACC17225T and R. aestuarii JC17T were used as probes for
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hybridization). Based on the hybridization results, strain JC288T represents a novel
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species according to recommendations for delineating a bacterial species (Stackebrandt &
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Goebel, 1994; Wayne et al., 1987). R. rhizosphaerae KACC 17225T, R. aestuarii JC17T
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and R. cervicalis CIP104027T(=E7107T) were used for comparative taxonomic analysis at
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authors’ laboratory.
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Under a light microscope (Olympus BH-2), cells of strain JC288T were cocci,
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0.6–0.8 µm (diameter; Supplementary Fig. S2). Cell division occurred by binary fission.
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Flagellar motility was not observed under the microscope and was confirmed by hanging
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drop method. Growth was measured turbidometrically at 540nm in a colorimeter
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(Systronics). In a buffered medium (K2HPO4–KH2PO4 buffer for pH 5–8 and NaHCO3–
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NaOH buffer for pH 9–11), growth of strain JC288T occurred between pH 6.0-7.5 and
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optimal pH was 7.0. Strain JC288T had no obligate requirement for salt (NaCl) for
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growth and could tolerate up to 6.0% (w/v) NaCl, while R. rhizosphaerae KACC17225T,
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R. aestuarii JC17T and R. cervicalis CIP104027T could tolerate 5%, 1% and 6,
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respectively. Temperature (4, 10, 15, 20, 25, 30, 35, 40, 45 and 50oC) ranges for growth
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were determined in TSB medium and growth was measured turbidometrically at 540 nm
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in a colorimeter (Systronics). Growth occurs for strain JC288T from 4-45 °C, while R.
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rhizosphaerae KACC17225T, R. aestuarii JC17T and R. cervicalis CIP104027T have
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growth range of 15-40 °C, 20-40 °C and 20-40 °C respectively, and growth optimum for
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strain JC288T, R. rhizosphaerae KACC17225T and R. aestuarii JC17T was 30 °C and
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35°C for R. cervicalis CIP104027T.
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Strain JC288T grew chemoorganoheterotrophically [aerobic with pyruvate (0.3%,
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w/v) as carbon source/electron donor]. Fermentative growth [anaerobic, dark with
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glucose/fructose/ pyruvate (0.3%, w/v)] could not be demonstrated. Organic carbon
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sources utilization was tested as described previously (Subhash et al., 2013). Strain
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JC288T utilized D-glucose, acetate, malate, succinate, D-arabinose, D-fructose, D-
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mannitol, D-ribose, D-xylose, 4-hydroxybenzoate, glycerol and tween 80. Citrate and
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lactose were not utilized by strain JC288T. Organic substrate utilization of strain JC288T
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differs from R. rhizosphaerae KACC17225T, R. aestuarii JC17T and R. cervicalis
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CIP104027T (Table 1).
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Vitamin (biotin, niacin, p-aminobenzoic acid and vitamin B12) requirements were
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tested by replacing yeast extract as described previously (Venkata Ramana et al., 2010)
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and all four strain had no additional requirement for vitamins for growth. Different
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biochemical tests such as hydrolysis of starch, casein and gelatin, indole production, and
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oxidase and catalase activity were carried out in the prescribed media as described
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previously (Venkata Ramana et al., 2010). All the strain did not hydrolyze casein, gelatin,
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starch and chitin. Urea was not hydrolyzed by strain JC288T, R. aestuarii JC17T and R.
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cervicalis CIP104027T. All the four strains were positive for catalase and oxidase
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activity; indole was produced from L-tryptophan by strain JC288T, R. rhizosphaerae
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KACC17225T and R. aestuarii JC17T. Sensitivity to different antibiotics was tested after
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spreading cells on tryptone soya agar plates (incubated for 15 days at 20 °C) using ready-
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made sensi-discs (HiMedia) with varying amount of antibiotics. Strain JC288T was
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sensitive to most of the antibiotics tested; nystatin, chloramphenicol, vancomycin,
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amikacin, gentamicin, nalidixic acid and tetracyclin. Strain JC288T was resistance to
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streptomycin and differs from R. rhizosphaerae KACC17225T and R. aestuarii JC17T.
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Strain JC288T was susceptible to nalidixic acid and tetracycline and differs from R.
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rhizosphaerae KACC17225T and R. aestuarii JC17T and R. cervicalis CIP104027T (Table
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1).
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Carotenoid composition of strain JC288T, R. rhizosphaerae KACC17225T, R.
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aestuarii JC17T and R. cervicalis CIP104027T as determined by C18-HPLC (Ramaprasad
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et al., 2013) indicated the presence of hydroxyspirilloxanthin (43.5%), spirilloxanthin
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(27%), ketospirilloxanthin (7.5%), rhodovibrin (7%) and two unidentified catotenoids
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(15%) (Supplementary Fig. S3).
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carotenoids, the carotenoid profile of all the three strains was similar. Cells grown in the
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TSB medium were harvested when growth of the cultures reached around 70% of the
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maximal optical density (exponential growth phase) and used for analysis of cellular
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fatty acids, polar lipids and quinones, which was done as described previously (Shalem
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Raj
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C16:1ω7c/C16:1ω6c, C18:1ω7c/C18:16c and C18:12OH with minor (>1% <%5) amounts of
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C18:0, C17:1ω7c and C16:03OH. The differences in fatty acid profile of strain JC288T with
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regard to R. rhizosphaerae KACC17225T, R. aestuarii JC17T and R. cervicalis
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CIP104027T is shown in Table 2.
Except for the relative abundance of difference
et al., 2013). Major (>5 % of total) fatty acids of strain JC288T were C16:0,
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Diphosphatidylglycerol,
phosphatidylglycerol,
unidentified aminolipids
phosphatidylethanolamine,
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phosphatidylcholine, four
(AL1-4), three
unidentified
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phospholipids (PL1,3,4), two unidentified lipids (L1-3), an aminophospholipid (APL)
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and a glycolipid (GL) are the polar lipid identified in strain JC288T (Supplementary Fig
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S3.) Strain JC288T differs from R. rhizosphaerae KACC17225T with regard to the
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presence ALP, AL3,4 and absence of unidentified lipid (L2) and unidentified
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phospholipid (PL2). The polar lipid profile of strain JC288T is also distinct from R.
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aestuarii JC17T in the presence of AL3,4, GL, L1,3, APL and absence of PL2
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(Supplementary Fig. S4). Q10 is the major (98%) quinone of the three strains.
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The genomic distinction of strain JC288T from its closest phylogenetic neighbors
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R. rhizosphaerae KACC17225T, R. aestuarii JC17T and R. cervicalis CIP104027T is well
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supported from the phenotypic traits which include cell size, organic carbon source
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utilization, antibiotic susceptibility, urea hydrolysis, fatty acid composition makes strain
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JC288T as a novel representative to the genus Roseomonas for which we propose the
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name Roseomonas oryzae sp. nov.
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Description of Roseomonas oryzae sp. nov.
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(o.ry’zae. L. gen. n. oryzae of rice, pertaining to the isolation of the type strain from rice
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paddy soil).
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Grows best on TSA, nutrient agar and R2A agar. Colonies on TSA are plane pink
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convex, circular, smooth and opaque with entire margins, approx. 2 mm in diameter after
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3-5 days at 30 °C (pH 7.0). Cells are Gram-stain-negative, non-motile cocci that occur
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singly and in pairs, strictly aerobic, non-motile and non-spore forming. Optimum growth
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occurs at 30 °C. Growth occurs from 4-45 °C. NaCl is not required for growth; tolerates
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up to 6% (w/v). Hydroxyspirilloxanthin, spirilloxanthin, ketospirilloxanthin, rhodovibrin
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and two unidentified catotenoids is the carotenoid composition. Bacteriochlorophyll-a is
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absent. Optimal growth occurs at pH 7.0 (range: pH 6.0–7.5). Catalase and oxidase are
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positive. Urea, starch, gelatin, Casein and chitin are not hydrolyzed. Indole is produced
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from tryptophan. Addition of vitamins is not required for growth. Good growth occurs
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with D-glucose, acetate, malate, citrate, succinate, D-arabinose, D-fructose, D-mannitol,
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D-ribose, D-xylose, 4-hydroxybenzoate and glycerol. Ammonium salts are used as a
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good nitrogen source. C16:0, C16:1ω7c/C16:1ω6c, C18:1ω7c/C18:16c, C18:12OH, C18:0,
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C17:1ω7c and C16:03OH is the fatty acid composition. Diphosphatidylglycerol,
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phosphatidylglycerol, phosphatidylethanolamine, phosphatidylcholine, four unidentified
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aminolipids (AL1-4), three unidentified phospholipids (PL1,3,4), two unidentified lipids
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(L1-3), an aminophospholipid (APL) and a glycolipid (GL) are the polar lipids. Q10 is
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the predominant quinone. DNA G+C content is 67.5 mol% (HPLC). Type strain JC288T
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(=KCTC 42542T = LMG 28711T), was isolated from a paddy rhizosphere soil sample
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collected from Western Ghats, Kankumbi, Karnataka, India.
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ACKNOWLEDGEMENTS
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We thank KACC, Korea for providing the type strain of Roseomonas rhizosphaerae
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YW11T = KACC 17225T which was received as gratis.
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REFERNCES
Baik, K.S., Park, S.C., Choe, H.N., Kim, S.N., Moon, J.-H. & Seong, C.N. (2012).
219
Roseomonas riguiloci sp. nov., isolated from wetland freshwater. Int J Syst Evol
220
Microbiol 62, 3024–3029.
9
221
Bibashi, E., Sofianou, D., Kontopoulou, K., Mitsopoulos, E. & Kokolina, E. (2000).
222
Peritonitis due to Roseomonas fauriae in a patient undergoing continuous
223
ambulatory peritoneal dialysis. J Clin Microbiol 38, 456–457.
224
Chakravarthy, S.K., Ramaprasad, E.V.V., Shobha, E., Sasikala, Ch. & Ramana,
225
Ch.V. (2012). Rhodoplanes piscinae sp. nov.. isolated from pond water. Int J Syst
226
Evol Microbiol 62, 2828–2834.
227
Chen, Q., Sun, L.N., Zhang, X.X., He, J., Kwon, S.W., Zhang, J., Li, S.P. & Gu, J.G.
228
(2014). Roseomonas rhizosphaerae sp. nov., a triazophos-degrading bacterium
229
isolated from soil. Int J Syst Evol Microbiol. 64, 1127-1133.
230
Furuhata, K., Miyamoto, H., Goto, K., Kato, Y., Hara, M. & Fukuyama, M. (2008).
231
Roseomonas stagni sp. nov., isolated from pond water in Japan. J Gen Appl
232
Microbiol 54, 167–171.
233
Gallego, V., Sánchez-Porro, C., García, M.T. & Ventosa, A. (2006). Roseomonas
234
aquatica sp. nov., isolated from drinking water. Int J Syst Evol Microbiol 56,
235
2291-2295.
236
Han, X.Y., Pham, A.S., Tarrand, J.J., Rolston, K.V., Helsel, L.O. & Levett, P.N.
237
(2003). Bacteriologic characterization of 36 strains of Roseomonas species and
238
proposal of Roseomonas mucosa sp. nov. and Roseomonas gilardii subsp rosea
239
subsp. nov.. Am J Clin Pathol 120, 256–264.
240
Kim, O.S., Cho, Y.J., Lee, K., Yoon, S. H., Kim, M., Na, H., Park, S.C., Jeon, Y.S.,
241
Lee, J.H. & other authors (2012). Introducing EzTaxon-e: a prokaryotic 16S
242
Rrna gene sequence database with phylotypes that represent uncultured species.
243
Int J Syst Evol Microbiol 62, 716–721.
10
244
Kim, S.-J., Weon, H.-Y., Ahn, J.-H., Hong, S.-B., Seok, S.-J., Whang, K.-S. & Kwon,
245
S.-W. (2013). Roseomonas aerophila sp. nov., isolated from air. Int J Syst Evol
246
Microbiol 63, 2334–2337.
247
Kimura, M. (1980). A simple method for estimating evolutionary rates of base
248
substitutions through comparative studies of nucleotide sequences. J Mol Evol 16,
249
111–120.
250
Jiang, C.-Y., Dai, X., Wang, B.-J., Zhou, Y.-G. & Liu, S.-J. (2006). Roseomonas lacus
251
sp. nov., isolated from freshwater lake sediment. Int J Syst Evol Microbiol 56, 25–
252
28.
253
254
Marmur, J. (1961). A procedure for the isolation of deoxyribonucleic acid from
microorganisms. J Mol Biol 3, 208–218.
255
Mesbah, M., Premachandran, U. & Whitman, W.B. (1989). Precise measurement of
256
the G+C content of deoxyribonucleic acid by high performance liquid
257
chromatography. Int J Syst Bacteriol 39, 159–167.
258
McLean, T. W., K. Rouster-Stevens, C. R. Woods. & A. K. Shetty. (2006). Catheter-
259
related bacteremia due to Roseomonas species in pediatric hematology/oncology
260
patients. Pediatr. Blood Cancer 46, 514-516.
261
Nutaratat P, Srisuk N, Duangmal K, Yurimoto H, Sakai Y, Muramatsu Y &
262
Nakagawa Y.(2013) Roseomonas musae sp. nov., a new bacterium isolated from
263
a banana phyllosphere. Antonie van Leeuwenhoek 103, 617-624.
264
Ramaprasad, E.V., Sasikala, Ch. & Ramana, Ch.V. (2013). Neurosporene is the major
265
carotenoid accumulated by Rhodobacter viridis JA737. Biotechnol Lett 35, 1093-
266
1097.
11
267
Rihs, J. D., Brenner, D. J., Weaver, R. E., Steigerwalt, A. G., Hollis, D. G. & Yu, V.
268
L. (1993) Roseomonas, a new genus associated with bacteremia and other human
269
infections. J Clin Microbiol 31, 3275–3283.
270
Sandoe, J. A., Malnick, H. & Loudon. K. W. (1997). A case of peritonitis caused by
271
Roseomonas gilardii in a patient undergoing continuous ambulatory peritoneal
272
dialysis. J Clin Microbiol 35, 2150-2152.
273
Seldin, L. & Dubnau, D. (1985). Deoxyribonucleic acid homology among Bacillus
274
polymyxa, Bacillus macerans, Bacillus azotofixans, and other nitrogen-fixing
275
Bacillus strains. Int J Syst Bacteriol 35, 151–154.
276
September, S. M., Brozel, V. S. & Venter, S. N. (2004). Diversity of non-tuberculoid
277
Mycobacterium species in biofilms of urban and semiurban drinking water
278
distribution systems. Appl Environ Microbiol 70, 7571–7573
279
Sfanos, K., Harmody, D., Dang, P., Ledger, A., Pomponi, S., McCarthy, P. & Lopez,
280
J. (2005). A molecular systematic survey of cultured microbial associates of
281
deep-water marine invertebrates. Syst Appl Microbiol 28, 242–264.
282
Shalem Raj, P., Ramaprasad, E.V.V., Vaseef, S., Sasikala, Ch. & Ramana, Ch.V.
283
(2012). Rhodobacter viridis sp. nov., a phototrophic bacterium isolated from
284
western ghats of India Int J Syst Evol Microbiol. 63, 181-186
285
Stackebrandt, E. & Goebel, B.M. (1994). Taxonomic note: a place for DNA-DNA
286
reassociation and 16S rRNA sequence analysis in the present species definition in
287
bacteriology. Int J Syst Evol Microbiol 44, 846–849.
12
288
Subhash, Y., Tushar, L., Sasikala Ch. & Ramana Ch.V (2013b). Erythrobacter
289
odishensis sp. nov. and Pontibacter odishensis sp. nov. isolated from a dry soil of
290
a solar saltern. Int J Syst Evol Microbiol.63, 4524-4532.
291
Subudhi, C.P.K., Adedeji, A., Kaufmann, M.E., Lucas, G.S. & Kerr, J.R. (2001).
292
Fatal Roseomonas gilardii bacteremia in a patient with refractory blast crisis of
293
chronic myeloid leukemia. Clin Microbiol Infect 7, 573–575.
294
Tamura, K., Stecher, G., Peterson, D., Filipski, A. & Kumar, S. (2013). MEGA6:
295
molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30, 2725–
296
2729.
297
Tonouchi, A. & Tazawa, D. (2014) Roseomonas aceris sp. nov. isolated from a mono
298
maple tree in the Shirakami Mountains in Japan. J Gen Appl Microbiol. 60: 38-43.
299
Venkata Ramana V, Sasikala Ch, Takaichi S. & Ramana ChV. (2010). Roseomonas
300
aestuarii sp. nov., a bacteriochlorophyll-a containing alphaproteobacterium
301
isolated from an estuarine habitat of India. Syst Appl Microbiol 33, 198-203.
302
Wayne, L. G., Brenner, D. J., Colwell, R. R., Grimont, P. A. D., Kandler, O.,
303
Krichevsky, M. I., Moore, L. H., Moore, W. E. C., Murray, R. G. E. & other
304
authors (1987). International Committee on Systematic Bacteriology. Report of
305
the ad hoc committee on reconciliation of approaches to bacterial systematic. Int J
306
Syst Bacteriol 37, 463–464.
307
308
Yoon, J. H., Kang, S. J., Oh, H. W. & Oh, T. K. (2007). Roseomonas terrae sp. nov.
Int J Evol Microbiol 57, 2485–2488.
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Table 1. Differentiating characteristics between strain JC288T, Roseomonas
rhizosphaerae KACC 17225T,Roseomonas aestuarii JC17T and R. cervicalis CIP104027T
Differentiating characteristics
Isolation source
Cell width (µm)
Cell length (µm)
Colony color
Temperature range for growth
(optimum) (ºC)
pH range for growth (optimum)
NaCl tolerance (%,w/v)
1
2
3
4
Paddy rhizosphere soil
0.6-0.8 (diameter)
Light Pink
4-45(30)
soil
0.5–1.0
1.0–1.5
Pink
15-40(30)
Estuary water
0.6–1.0
1.5–2.0
Dark pink
20-40(30)
Cervix
1.0-1.5
2.0-3.8
Pink
20-40(35)
6.0-7.5(7.0)
6
5.0-8.0(7.0)
5
5.0-8.0(7.0)
1
6.0-9.0 (8.0)
6
+
-
+
+
-
+
+
-
+
+
+
+
+
+
+
+
+
+
+
+
-
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
-
R
S
S
67.5
S
S
R
69.6
S
R
S
66.2
R
R
S
70.4
Urea hydrolysis
Indole from L-tryptophan
BChl-a
Carbon sources utilized for
growth
Sodium acetate
Malate
Citrate
Succinate
4-Hydroxybenzoate
D-Fructose
D-Glucose
D-Mannitol
D-Mannose
D-Ribose
D-Arabinose
Lactose
D-Xylose
Glycerol
Susceptibility to antibiotics(µg)
Streptomycin (10)
Nalidixic acid (30)
Tetracycline (10)
DNA G+C content (mol%)
316
317
Data pertain to analysis done at authors’ laboratory. 1, JA288T; 2, Roseomonas
318
rhizosphaerae KACC17225T; 3, Roseomonas aestuarii JC17T; 4. R. cervicalis
319
CIP104027T.
320
Hydroxyspirilloxanthin, spirilloxanthin, ketospirilloxanthin, rhodovibrin and two
321
unidentified catotenoids are the carotenoids present in all tested strains. Q10 is the
322
major quinone of all the strains. All strains are negative for starch, chitin and
All
the
strains
are
14
positive
for
catalase
and
oxidase.
323
gelatine, casein hydrolysis. All the strains utilize carbon D-fructose, maltose,
324
sodium acetate and pyruvic acid. All the strains are susceptible to
325
chloramphenicol and amikacin. +, Present/utilized; -, absent/not utilized. R,
326
resistant; S, sensitive; BChl-a, Bacteriochlorophyll-a.
327
328
329
330
331
332
333
334
335
336
337
338
339
340
341
342
343
344
345
15
346
Table. 2. Fatty acid compositions of strain JC288T and related type strains.
347
Fatty acid
C14:0
C16:0
C18:0
C16:1 5c
C16:1 7c/C16:1 6c
C17:1 7c
C17:1 6c
C18:1 7c/C18:1 6c
C16:03OH
C18:12OH
C19:0 cyclo 8c
1
7.6
1.2
6.8
4.5
55.8
1.6
18.1
-
2
13.4
1.0
1.2
3.0
1.0
61.5
0.9
11.6
-
3
10.7
4.4
12.3
52.6
1.8
8.7
3.8
4
1.0
16.6
4.8
3.8
56.0
1.4
6.9
1.4
348
349
Strains: 1, JA288T; 2, Roseomonas rhizosphaerae KACC17225T; 3, Roseomonas
350
aestuarii JC17T and 4. R. cervicalis CIP104027T. Data were obtained in this
351
study. Values are percentages of total fatty acids; -, not detected or present at less
352
than 1.0 %.
353
354
355
356
357
358
359
360
361
362
363
16
364
Fig. 1. Phylogenetic tree based on 16S rRNA gene sequences showing the relationship of strain
365
JC288T with the most closely related members of the genus Roseomonas. The tree was
366
constructed by the neighbor joining method using the MEGA6 software and rooted by using
367
Komagataeibacter xylinus NCIMB11664T (X75619) as the out-group. Numbers at nodes
368
represent bootstrap values (based on 1000 resamplings) and the bootstrap percentages refer
369
to NJ/ML/MP analysis. The GenBank accession numbers for 16S rRNA gene sequences are
370
shown in parentheses. Bar 1 nucleotide substitutions per 100 nucleotides.
371
372
17