IJSEM Papers in Press. Published May 15, 2015 as doi:10.1099/ijs.0.000334

International Journal of Systematic and Evolutionary Microbiology Winogradskyella eckloniae sp. nov., a marine bacterium isolated from a brown alga Ecklonia cava --Manuscript Draft-Manuscript Number:

IJSEM-D-14-00411R1

Full Title:

Winogradskyella eckloniae sp. nov., a marine bacterium isolated from a brown alga Ecklonia cava

Short Title:

Winogradskyella eckloniae sp. nov., a marine bacterium isolated from a brown alga Ecklonia cava

Article Type:

Note

Section/Category:

New taxa - Bacteroidetes

Corresponding Author:

Duck-Chul Oh, Ph.D Jeju National Universtiy Jeju, KOREA, REPUBLIC OF

First Author:

Ji-Young Kim

Order of Authors:

Ji-Young Kim So-Hyun Park Ga-Young Seo Young-Ju Kim Duck-Chul Oh, Ph.D

Manuscript Region of Origin:

KOREA, REPUBLIC OF

Abstract:

A novel bacterial strain, designated EC29T, was isolated from the brown alga Ecklonia cava collected on Jeju Island, Republic of Korea. Cells of strain EC29T were Gramnegative, aerobic, rod-shaped and motile by gliding. Growth was observed at 10-30 °C (optimum, 20-25°C), at pH 6.0-9.5 (optimum, pH 7.5) and in the presence of 1-5 % (w/v) NaCl. Phylogenetic analyses based on the 16S rRNA gene sequence revealed that the strain belonged to the genus Winogradskyella. Strain EC29T exhibited the highest 16S rRNA gene sequence similarities of 96.5-97.8 % to the type strains of Winogradskyella pulchriflava EM106T, Winogradskyella echinorum KMM 6211T and Winogradskyella ulvae KMM6390 T. Strain EC29T exhibited < 27% DNA-DNA relatedness with Winogradskyella pulchriflava EM106T and Winogradskyella echinorum KMM 6211T. The predominant fatty acids of strain EC29T were iso-C15:0, iso-C15:1 G, C15:0, iso-C17:0 3-OH, iso-C15:0 3-OH and anteiso C15:0. The DNA G+C content was 31.1 mol% and the major respiratory quinone was menaquinone-6 (MK-6). Based on a polyphasic study, strain EC29T is considered to represent a novel species of the genus Winogradskyella, for which the name Winogradskyella eckloniae sp. nov. is proposed. The type strain is EC29T (=KCTC 32172 T = JCM 18703 T).

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Winogradskyella eckloniae sp. nov., a marine bacterium isolated from

2

a brown alga Ecklonia cava

3 4

Ji-Young Kim, 1, 2 So-Hyun Park,3 Ga-Young Seo,1 Young-Ju Kim,2* and Duck-Chul Oh 1*

5 6

1

Department of Biology, Jeju National University, Jeju 690-756, Republic of Korea

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2

Jeju Biological Resource Co., Ltd, CTC Business Incubator, Jeju Tourism College, Jeju

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690-756, Republic of Korea

9

3

10

Department of Aquatic Life Medicine, College of Ocean Science, Jeju 690-756, Republic of

Korea

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Author for correspondence: Duck-Chul Oh; E-mail: [email protected]; Tel : +82-64-754-

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3524; Fax : +82-64-756-3541. Young-Ju Kim; E-mail: [email protected] jejunu.ac.kr

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Running title: Winogradskyella eckloniae sp. nov.

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Subject category: New taxa (Bacteroidetes)

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The GenBank accession number for the 16S rRNA gene sequence of strain EC29T (=KCTC

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32172 T = JCM 18703 T) is JF820845.

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Abstract

22 23

A novel bacterial strain, designated EC29T, was isolated from the brown alga Ecklonia

24

cava collected on Jeju Island, Republic of Korea. Cells of strain EC29T were Gram-negative,

25

aerobic, rod-shaped and motile by gliding. Growth was observed at 10-30 °C (optimum, 20-

26

25°C), at pH 6.0-9.5 (optimum, pH 7.5) and in the presence of 1-5 % (w/v) NaCl.

27

Phylogenetic analyses based on the 16S rRNA gene sequence revealed that the strain

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belonged to the genus Winogradskyella. Strain EC29T exhibited the highest 16S rRNA gene

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sequence similarities of 96.5-97.8 % to the type strains of Winogradskyella pulchriflava

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EM106T, Winogradskyella echinorum KMM 6211T and Winogradskyella ulvae KMM6390 T.

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Strain EC29T exhibited < 27% DNA-DNA relatedness with Winogradskyella pulchriflava

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EM106T and Winogradskyella echinorum KMM 6211T. The predominant fatty acids of strain

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EC29T were iso-C15:0, iso-C15:1 G, C15:0, iso-C17:0 3-OH, iso-C15:0 3-OH and anteiso C15:0. The

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DNA G+C content was 31.1 mol% and the major respiratory quinone was menaquinone-6

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(MK-6). Based on a polyphasic study, strain EC29T is considered to represent a novel species

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of the genus Winogradskyella, for which the name Winogradskyella eckloniae sp. nov. is

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proposed. The type strain is EC29T (=KCTC 32172 T = JCM 18703 T).

38 39

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The genus Winogradskyella was first reported by Nedashkovskaya et al. (2005) and belongs

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to the family Flavobacteriaceae (Bernardet et al., 2002; Bernardet, 2011) within the phylum

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Bacteroidetes. At the time of writing, the genus comprised 20 species with validly published

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names (http://www.bacterio.cict.fr/). Members of the genus Winogradskyella have been

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isolated from various marine environments, such as marine animals (Lau et al., 2005;

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Nedashkovskaya et al., 2009; Ivanova et al., 2010), marine sediments (Romanenko et al.,

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2009; Begum et al., 2013; Kang et al., 2013; Kim et al., 2013; Park et al., 2014), seawater

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(Pinhassi et al., 2009; Kim & Nedashkovskaya, 2010; Yoon et al., 2011; Yoon & Lee, 2012;

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Lee et al., 2012, 2013) and seaweeds (Nedashkovskaya et al., 2005; Nedashkovskaya et al.,

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2012; Park & Yoon, 2013).

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During our investigation of the biodiversity of marine bacteria from seaweed, strain EC29T

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was isolated from brown alga and its taxonomic position was investigated using a polyphasic

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approach. The brown alga sample of Ecklonia cava was collected from the coast of Jeju

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Island, Republic of Korea. The seaweed sample was homogenized with sterile seawater, and

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the homogenate was spread on marine agar 2216 (MA; Difco) using a dilution-plating

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technique. After incubation for 7 days at 25 °C, the purified colonies were maintained in a

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glycerol solution (20 %, w/v) at -70 °C. For phenotypic comparison and DNA-DNA

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hydridization experiments, Winogradskyella pulchriflava KCTC 23858T, Winogradskyella

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echinorum KCTC 22026T and Winogradskyella ulvae KCTC 23626T were used as reference

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strains in parallel tests.

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The 16S rRNA gene was amplified from genomic DNA by PCR using the universal

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primers of bacterial 27F and 1522R (Weisburg et al., 1991). The resultant products were

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cloned using the TOPO cloning kit (Invitrogen) and the sequencing of the cloned 16S rRNA

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gene was subject to a service by Genotech (Korea). A full sequence of the 16S rRNA gene

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was generated using SeqMan software (DNASTAR) and compared with the corresponding

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sequences

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(http://blast.ncbi.nlm,nih.gov/Blast.cgi)

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e.ezbioclould.net/; Kim et al., 2012). Multiple alignments were performed using CLUSTAL

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X version 1.83 program (Thompson et al., 1997) and gaps (at 5`- and 3`ends) were excluded

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by using the BioEdit program (Hall, 1999). Phylogenetic trees were reconstructed using

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MEGA program (version 6.0; Tamura et al., 2013) including the neighbor-joining (Saitou &

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Nei, 1987), maximum likelihood (Felsenstein, 1981) and maximum parsimony (Kluge and

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Farris, 1969) algorithms. The reliability of tree topology was evaluated by bootstrap analysis

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(Felsenstein, 1985) based on 1,000 replicates. DNA–DNA hybridization was tested to

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determine the relationship between strains EC29T and the type strain W. pulchriflava KCTC

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23858T and W. echinorum KCTC 22026T according to the measured photobiotin-labelled

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DNA probes using the microplate hybridization method (Ezaki et al., 1989; Christensen et al.,

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2000).

of

related

taxa

obtained and

from EzTaxon-e

the

GenBank server

database

(http://eztaxon-

80 81

The cell morphology of strain EC29T was observed by light microscopy (ECLIPS 80I; Nikon)

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and scanning electron microscopy (SIE-3000M; SEC), with cells grown on MA for 3 days at

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25 °C. Cell motility was investigated by the development of turbidity throughout a tube of

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semi-solid media (Bowman, 2000). The Gram reaction was determined using the Gram stain

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kit (BD) according to the manufacturer’s instructions. The temperature (4, 10, 15, 20, 25, 30,

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35, 37, 40, and 45 °C) and pH (4.0 - 10.0, at intervals of 0.5 pH unit) range for growth were

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examined on MA. The requirement and tolerance of various NaCl concentrations were

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adjusted to 0-10 % (w/v, at interval of 1.0 %) using Zobell's marine agar medium (Zobell,

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1941; 5.0 g Bacto peptone, 1.0 g yeast extract, 0.1 g ferric citrate, 5.94 g MgSO4·7H2O, 4.53g

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MgCl2·6H2O, 0.64g KCl, 1.3g CaCl2 and 15.0 g Bacto agar in 1 liter distilled water, pH 7.0).

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Anaerobic growth was investigated after incubation for 1 month on MA in the AnaeroPack

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system (Oxoid).

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Hydrolysis of casein, carboxymethylcellulose, starch and Tween 20, 40, 60, and 80 were

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carried out using MA, supplemented with each substance at a concentration as described

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previously (Smibert & Krieg, 1994). Hydrolysis of DNA was tested using DNase test agar

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(Difco) prepared with artificial seawater. Catalase and oxidase activities were examined in a

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3 % solution (v/v) of hydrogen peroxide and 1 % (w/v) tetramethyl p-phenylenediamine,

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respectively. Other physiological and biochemical properties were determined with the API

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20E, API 20NE and API ZYM (bioMérieux) according to the manufacturer`s instructions.

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Susceptibility to antibiotic was investigated using filter-paper discs (BBL) as described by

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Kim & Oh (2012).

102 103

For the analysis of cellular fatty acids, the cell mass of strain EC29T and reference strains

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were harvested on MA after incubation for 3 days at 25 °C. The identification of fatty acid

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methyl esters was performed using the Sherlock Microbial Identification System (MIDI;

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version 6.1) and the TSBA6 database (Sasser, 1990). The DNA G+C content of strain EC29T

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was determined by extracting genomic DNA according to the methods described by Marmur

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(1961) and the thermal denaturation method (Tm) with Escherichia coli K-12 as a reference

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strain (Marmur & Doty, 1962). Analysis of the respiratory quinones was performed using a

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service of the Korean Culture Center of Microorganisms (KCCM). Polar lipids were

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separated by two-dimensional thin-layer chromatography (TLC) and analyzed according to

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the methods described by Minnikin et al. (1984). For the detection of polar lipids,

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molybdophosphoric acid, ninhydrin, Zinzade and α-naphthol reagents were applied and

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through co-migration with authentic standards (Minnikin et al., 1984; Komagata and Suzuki,

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1987).

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The nearly complete 16S rRNA gene sequence of strain EC29T was a continuous stretch of

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1,442 bp and was GenBank accession number JF820845. Phylogenetic analysis established

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that strain EC29T belonged to the genus Winogradskyella, forming a well separated branch

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with type strains of W. pulchriflava EM106T (97.8 %), W. echinorum KMM 6211T (97.5 %),

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W. ulvae KMM 6390T (96.5 %), W. eximia KMM 3994T (96.2%) and W. lutea A73T (96.1 %)

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in the neighbor-joining phylogenetic tree (Fig. 1). Sequence similarities of < 93.8 % was

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observed with the other species of the genus Winogradskyella. Although the bootstrap value

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for the node between the isolate and closely related type strains was relatively low (56 %),

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the topologies of the phylogenetic trees built using the maximum-likelihood and maximum-

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parsimony algorithms supported the finding that strain EC29T formed lineage separated from

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members of the genus Winogradskyella (Fig S1). The levels of DNA–DNA relatedness

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between strain EC29T and the type strains W. pulchriflava KCTC 23858T and W. echinorum

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KCTC 22026T was 26.8 ± 0.7 and 24.9 ± 1.3  %, respectively. These values confirmed the

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assignment of the novel isolate EC29T to a separate species according to the cut-off value of

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70 % proposed Wayne et al. (1987) for the definition of members of a species.

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Cells of strain EC29T are Gram-negative, aerobic, motile by gliding and rod-shaped, able to

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grow at temperatures of 10-30 °C (optimum 25 °C), at pH 6.0-9.5 (optimum pH 7.5) and in

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the presence of 1-5 % NaCl. Colonies are circular, have a rough surface and are yellow-

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colored when grown on MA at 25 °C. Other phenotypic and biochemical characteristics of

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the strain EC29T are presented in Table 1 and the related Winogradskyella species description.

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The predominant cellular fatty acids of the strain EC29T (> 5 % of the total fatty acids) were

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iso-C15:0 (24.9 %), iso-C15:1 G (23.5 %), C15:0 (9.1 %), iso-C17:0 3-OH (8.8 %), iso-C15:0 3-OH

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(7.1 %), and anteiso C15:0 (5.6 %). This fatty acid profile was similar to those of closely related

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members within the genus Winogradskyella, with difference in the respective proportions of

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some fatty acids (Table 2). The predominant respiratory quinone found in the strain EC29T

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was menaquinone-6 (MK-6), which is a characteristic quinone system of the family

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Flavobacteriaceae (Bernardet and Nakagawa, 2006).

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was 31.1 mol%, this supports their affiliation with the genus Wingradskyella, members of

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which have DNA G+C content between 30.1 and 36.4 mol%. The polar lipid profiles of strain

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EC29T were phosphatidylethanolamine (PE), unknown aminolipids (AL1-AL2), and

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unknown lipids (L1-L5) (Fig. S2). The presence of phosphatidylethanolamine and aminolipid

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was consistent with results of analyses of the genus Winogradskyella (Nedashkovskaya et al.,

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2012; Kim et al., 2012; Kim et al., 2013; Park et al., 2014).

The G+C content of the strain EC29T

151 152

The genetic distinctiveness and differential phenotypic characteristics of strain EC29T are

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sufficient to show that this strain is separate from species of the genus Winogradskyella. Based on

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data from the present polyphasic taxonomic study, we considered that strain EC29T represent a

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novel species of the genus Winogradskyella, for which the name Winogradskyella eckloniae sp.

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nov. is proposed.

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Description of Winogradskyella eckloniae sp. nov.

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Winogradskyella eckloniae (eck.lo.ni`ae. N.L. fem. n. Ecklonia scientific genus name of a

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marine alga; N.L. gen. n. eckloniae of Ecklonia, referring to the isolation of the type strain

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from Ecklonia kurome).

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Cells are Gram-negative, aerobic, motile by gliding and rod-shaped (0.2–0.3 µm wide and

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0.7–1.2 µm long). After incubation for 3 days at 25 °C, colonies on MA are circular to rough

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surface, yellow-colored and 1-1.5 mm in diameter. Growth is observed at 10-30 °C (optimum,

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20-25 °C) and pH 6.0-9.5 (optimum, pH 7.5). Growth occurs in the presence of 1-5 % NaCl,

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but not below 1 % or above 5 % NaCl. Anaerobic growth is not observed after 4 weeks at

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25 °C on MA. Catalase- and oxidase- positive. Hydrolyses aesculin, gelatin, starch and Tween

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40, but does not hydrolyse casein, cellulose, Tweens 20, 60, and 80. Nitrate is not reduced.

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Acetoin, indole and H2S are not produced. In the API ZYM system, positive for alkaline

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phosphatase, esterase (C4), esterase lipase (C8), cystine-, valine-, and leucine arylamidases,

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α-chymotrypsin, trypsin, acid phosphatase, and naphthol-AS-BI-phosphohydrolase, and

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negative for lipase (C14), α–galactosidase, β–galactosidase, α-glucosidase, β-glucosidase, β-

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glucuronidase, N-acetyl-β-glucosaminidase, α-mannosidase and α-fucosidase. Susceptible to

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ampicillin (1µg), cephalothin (30µg), chloramphenicol (30µg), erythromycin (15µg),

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licomycin (2µg), novobiocin (30µg), penicillin G (10IU), and tetracycline (30µg), but

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resistant to gentamycin (10µg), kanamycin (30µg), neomycin (30µg), polymyxin B (300IU),

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and streptomycin (10µg). The predominant cellular fatty acids are iso-C15:0, iso-C15:1 G, C15:0,

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iso-C17:0 3-OH, iso-C15:0 3-OH and anteiso C15:0. The major respiratory quinone is

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menaquinone-6. The major polar lipids are phosphatidylethanolamine, two unknown

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aminolipids, and five unknown lipids. The DNA G+C content of the type strain is 31.1 mol%.

182 183

The type strain is EC29T (= KCTC 23835T = JCM 18703T), which was isolated from brown

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alga Ecklonia cava collected along the coast of Jeju Island, Republic of Korea.

185

186

References

187 188

Begum, Z., Srinivas, T. N. R., Manasa, P., Sailaja, B., Sunil, B., Prasad, S. & Shivaji, S.

189

(2013). Winogradskyella psychrotolerans sp. nov., a marine bacterium of the family

190

Flavobacteriaceae isolated from Arctic sediment. Int J Syst Evol Microbiol 63, 1646–1652.

191 192

Bernardet, J.-F. (2011). Family I. Flavobacteriaceae Reichenbach 1992. In Bergey's

193

Manual of Systematic Bacteriology, 2nd ed.,vol. 4, pp. 106-111. Edited by N. R. Krieg, W.

194

Ludwig, W. B. Whitman, B. P. Hedlund, B. J. Paster, J. T. Staley, N. Ward, D. Brown & A.

195

Parte. New York: Springer.

196 197

Bernardet, J.-F., Nakagawa, Y. & Holmes, B. (2002). Proposal minimal standards for

198

describing new taxa of the family Flavobacteriaceae and emended description of the family.

199

Int J Syst Evol Microbiol 52, 1049-1070.

200 201

Bernardet, J.-F. & Nakagawa, Y. (2006). An introduction to the family Flavobacteriaceae.

202

In The Prokaryotes. A Handbook on the Biology of Bacteria, 3rd edn, vol. 7, pp. 455–480.

203

Edited by M. Dworkin, S. Falkow, E. Rosenberg, K. H. Schleifer & E. Stackebrandt. New

204

York: Springer.

205 206

Bowman, J. P. (2000). Description of Cellulophaga algicola sp. nov., isolated from the

207

surfaces of Antarctic algae, and reclassification of Cytophaga uliginosa (ZoBell and Upham

208

1944) Reichenbach 1989 as Cellulophaga uliginosa comb. nov. Int J Syst Evol Microbiol 50,

209

1861–1868.

210

211

Christensen H., Angen O., Mutters R., Olsen J. E. & Bisgaard M. (2000). DNA-DNA

212

hybridization determined in micro-wells using covalent attachment of DNA. Int J Syst Evol

213

Microbiol 50, 1095–1102.

214 215

Ezaki, T., Hashimoto, Y. & Yabuuchi, E. (1989). Fluorometric deoxyribonucleic acid-

216

deoxyribonucleic acid hybridization in microdilution wells as an alternative to membrane

217

filter hybridization in which radioisotopes are used to determine genetic relatedness among

218

bacterial strains. Int J Syst Bacteriol 39, 224–229.

219 220

Felsenstein, J. (1981). Evolutionary trees from DNA sequences: a maximum likelihood

221

approach. J Mol Evol 17, 368-376.

222 223

Felsenstein, J. (1985). Confidence limits on phylogenies: an approach using the bootstrap.

224

Evolution 39, 783-791.

225 226

Hall, T. A. (1999). BioEdit: a user-friendly biological sequence alignment editor and analysis

227

program for Windows 95/98/NT. Nucleic Acids Symp Ser 41, 95-98.

228 229

Ivanova, E. P., Christen, R., Gorshkova, N. M., Zhukova, N. V., Kurilenko, V. V.,

230

Crawford, R. J. & Mikhailov, V. V. (2010). Winogradskyella exilis sp. nov., isolated from

231

the starfish Stellaster equestris, and emended description of the genus Winogradskyella. Int J

232

Syst Evol Microbiol 60, 1577-1580.

233 234

Kang, C.-H., Lee, S.-Y. & Yoon, J.-H. (2013). Winogradskyella litorisediminis sp. nov.,

235

isolated from coastal sediment. Int J Syst Evol Microbiol 63, 1793–1799.

236 237

Kim, O. S., Cho, Y. J., Lee, K., Yoon, S. H., Kim, M., Na, H., Park, S. C., Jeon, Y. S., Lee,

238

J. H., Yi, H., Won, S. & Chun, J. (2012). Introducing EzTaxon: a prokaryotic 16S rRNA

239

Gene sequence database with phylotypes that represent uncultured species. Int J Syst Evol

240

Microbiol 62, 716–721.

241

242

Kim, J-Y. & Oh, D-C. (2012). Winogradskyella jejuensis sp. nov., a marine bacterium

243

isolated from a brown alga Carpopeltis affinis. J Microbiol 50, 888-892.

244 245

Kim, S. B. & Nedashkovskaya, O. I. (2010). Winogradskyella pacifica sp. nov., a marine

246

bacterium of the family Flavobacteriaceae. Int J Syst Evol Microbiol 60, 1948-1951.

247 248

Kim, S.-J., Choi, Y.-R., Park, S.-J., Kim, J.-G., Shin, K.-S., Roh, D.-H. & Rhee, S.-K.

249

(2013). Winogradskyella pulchriflava sp. nov., isolated from marine sediment. Int J Syst Evol

250

Microbiol 63, 3062-3068.

251 252

Kluge, A. G. & Farris, F. S. (1969). Quantitative phyletics and the evolution of anurans. Syst

253

Zool 18, 1–32.

254 255

Komagata, K. & Suzuki. K. (1987). Lipids and cell-wall analysis in bacterial systematics.

256

Methods Microbiol. 19, 161-207.

257 258

Lau, S. C. K., Tsoi, M. M. Y., Li, X., Plakhotnikova, I., Dobretsov, S., Lau, K. W. K.,

259

Wu, M., Wong, P.-K., Pawlik, J. R. & Qian, P.-Y. (2005). Winogradskyella poriferorum sp

260

nov., a novel member of the family Flavobacteriaceae isolated from a sponge in the Bahamas.

261

Int J Syst Evol Microbiol 55, 1589-1592.

262 263

Lee, D.-H., Cho, S. J., Kim, S. M. & Lee, S. B. (2013). Winogradskyella damuponensis sp.

264

nov., isolated from seawater. Int J Syst Evol Microbiol 63, 321–326.

265 266

Lee, S.-Y., Park, S., Oh, T.-K. & Yoon, J.-H. (2012). Winogradskyella aquimaris sp. nov.,

267

isolated from seawater. Int J Syst Evol Microbiol 62, 1814-1818.

268 269

Marmur, J. (1961). A procedure for the isolation of deoxyribonucleic acid from micro-

270

organisms. J Mol Biol 3, 208-218.

271 272

Marmur, J. & Doty, P. (1962). Determination of the base composition of deoxyribonucleic

273

acid from its thermal denaturation temperature. J Mol Biol 5, 109-118.

274 275

Minnikin, D. E., O’Donnell, A. G., Goodfellow, M., Alderson, G., Athalye, M., Schaal, A.

276

& Parlett, J. H. (1984). An integrated procedure for the extraction of bacterial isoprenoid

277

quinones and polar lipids. J Microbiol Methods 2, 233–241.

278 279

Nedashkovskaya, O. I., Kim, S. B., Han, S. K., Snauwaert, C., Vancanneyt, M., Swings,

280

J., Kim, K. O., Lysenko, A. M., Rohde, M., Frolova, G. M., Mikhailov, V. V. & Bae, K.

281

S. (2005). Winogradskyella thalassocola gen. nov., sp. nov., Winogradskyella epiphytica sp.

282

nov. and Winogradskyella eximia sp. nov., marine bacteria of the family Flavobacteriaceae.

283

Int J Syst Evol Microbio. 55, 49-55.

284

285

Nedashkovskaya, O. I., Vancanneyt, M., Kim, S. B. & Zhukova, N. V. (2009).

286

Winogradskyella echinorum sp. nov., a marine bacterium of the family Flavobacteriaceae

287

isolated from the sea urchin Strongylocentrotus intermedius. Int J Syst Evol Microbiol 59,

288

1465-1468.

289 290

Nedashkovskaya, O. I., Kukhlevskiy, A. D. & Zhukova, N. V. (2012). Winogradskyella

291

ulvae sp. nov., an epiphyte of a Pacific seaweed, and emended descriptions of the genus

292

Winogradskyella

293

Winogradskyella exilis and Winogradskyella eximia. Int J Syst Evol Microbiol 62: 1450-1456.

and

Winogradskyella

thalassocola,

Winogradskyella

echinorum,

294 295

Park, S. & Yoon, J.-H. (2013). Winogradskyella undariae sp. nov., a member of the family

296

Flavobacteriaceae isolated from a brown algae reservoir. Antonie Van Leeuwenhoek 104,

297

619-626.

298 299

Park, S., Park, J.-M., Won, S.-M, Bae, K. S. & Yoon, J-H. (2014). Winogradskyella

300

wandonensis sp. nov., isolated from a tidal flat. Int J Syst Evol Microbiol 64, 1520-1525.

301 302

Pinhassi, J., Nedashkovskaya, O. I., Hagström, A. & Vancanneyt, M. (2009).

303

Winogradskyella rapida sp. nov., isolated from protein-enriched seawater. Int J Syst Evol

304

Microbiol 59, 2180-2184.

305 306

Romanenko, L. A., Tanaka, N., Frolova, G. M. & Mikhailov, V. V. (2009).

307

Winogradskyella arenosi sp. nov., a member of the family Flavobacteriaceae isolated from

308

marine sediments from the Sea of Japan. Int J Syst Evol Microbiol 59, 1443-1446.

309

310

Saitou, N. & Nei, M. (1987). The neighbor-joining method: a new method for reconstructing

311

phylogenetic trees. Mol Biol Evol 4, 406-425.

312 313

Sasser, M. (1990). Identification of bacteria by gas chromatography of cellular fatty acids,

314

MIDI Technical Note 101. Newark, DE: MIDI Inc.

315 316

Smibert, R. M. & Krieg, N. R. (1994). Phenotypic characterization. In Methods for General

317

and Molecular Bacteriology, pp. 607–654. Edited by P. Gerhardt. Washington, D.C., USA.

318

American Society for Microbiology.

319 320

Tamura, K., Stecher, G., Peterson, D., Peterson, Filipski, A. & Kumar, S. (2013).

321

MEGA6: Molecular evolutionary genetics analysis version 6.0. Mol Biol Evol 30, 2725–2729.

322 323

Thompson, J. D., Gibson, T. J., Plewniak, F., Jeanmougin, F. & Higgins, D. G. (1997).

324

The CLUSTAL_X windows interface: flexible strategies for multiple sequence alignment

325

aided by quality analysis tools. Nucleic Acids Res 25, 4876–4882.

326 327

Wayne, L. G., Brenner, D. J., Colwell, R. R., Grimont, P. A. D., Kandler, O.,

328

Krichevsky, M. I., Moore, L. H., Moore, W. E. C., Murray, R. G. E. & other authors

329

(1987). International Committee on Systematic Bacteriology. Report of the ad hoc committee

330

on reconciliation of approaches to bacterial systematics. Int J Syst Bacteriol 37, 463–464.

331 332

Weisburg, W. G., Barns, S. M., Pelletier, D. A., and Lane, D. J. (1991). 16S ribosomal

333

DNA amplification for phylogenetic study. J Bacteriol 173, 697-703.

334

335

Yoon, B.-J., Byun, H.-D., Kim, J.-Y., Lee, D.-H., Kahng, H.-Y. & Oh, D.-C. (2011).

336

Winogradskyella lutea sp. nov., isolated from seawater, and emended description of the genus

337

Winogradskyella. Int J Syst Evol Microbiol 61, 1539-1543.

338 339

Yoon, J.-H. & Lee, S.-Y. (2012). Winogradskyella multivorans sp. nov., a polysaccharide-

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degrading bacterium isolated from seawater of an oyster farm. Antonie van Leeuwenhoek 102,

341

231–238.

342 343

ZoBell, C. E. (1941). Studies on marine bacteria. I. The cultural requirements of

344

heterotrophic aerobes. J Mar Res 4, 42-75.

345

346

Table 1. Differential phenotypic characteristics of strain EC29T and the closely related

347

type strains of genus Winogradskyella

348

Strains: 1, W. eckloniae EC29T; 2, W. pulchriflava KCTC 23858 T; 3, W. echinorum KCTC

349

22026T; 4. W. ulvae KCTC 23626 T. All data are from this study. +, Positive; w, weakly

350

positive; -, negative. All strains are positive for the following: catalase and oxidase activities;

351

hydrolysis of aesculin and gelatin; and activities of acid phosphatase, alkaline phosphatase,

352

esterase (C4), esterase lipase (C8), leucine arylamidase, valine arylamidase, and cystein

353

arylamidase; susceptibility of cephalothin, chloramphenicol, erythromycin, lincomycin and

354

novobiosin. All strains are negative for the following: hydrolysis of cellulose, urea and

355

Tween 20; activities of lipase (C14), α-galactosidase, β-galactosidase, β-glucuronidase, α-

356

glucosidase, β-glucosidase, N-acetyl- β-glucosaminidase, α-mannosidase and α-fucosidase;

357

susceptibility of gentamicin, neomycin, polymyxin B and streptomycin. Characteristic

1

2

3

4

Anarobic

-

-

-

+

37 °C

-

-

+

+

1-5

0.5-5.0

1.0-6.0

0.5-4.0

-

-

-

+

DNA

-

+

+

+

Starch

+

-

-

+

Casein

-

+

+

+

Tween 40

+

+

-

+

Tween 80

-

+

-

+

D-Glucose

-

-†

-†

+

D-mannose

-

-†

-†

+

w

-

+

-

Growth at/with

NaCl (%) Fermentation of Glucose Hydrolysis of:

Utilization of:

Enzyme activities (API ZYM): Trypsin

α-chymotrypsin

+

+

+

-

Naphtol-AS-BI-phosphohydrolase

-

+

+

+

Ampicillin

+

+

-

+

Kanamycin

-

+

-

-

Penicillin

+

+

-

+

Tetracyline

+

+

-

+

31.1

33.3 a

33.6 b

34.2 c

Susceptibility to:

DNA G+C content (mol%)* 358

* DNA G+C content data from: a, Kim et al.,(2013); b, Nedashkovskaya et al., (2009); c,

359

Nedashkovskaya et al., (2012).

360

† Different results were reported by Kim et al., (2013) and Nedashkovskaya et al., (2009).

361

362

Table 2. Cellular fatty acid profiles of strain EC29T and the closely related type strains

363

of genus Winogradskyella

364

Strains: 1, W. eckloniae EC29T; 2, W. pulchriflava KCTC 23858 T; 3, W. echinorum KCTC

365

22026T; 4, W. ulvae KCTC 23626 T. All data were obtained in this study from cultures on MA

366

at 25°C for 3 days. Fatty acids founds in amounts

Winogradskyella eckloniae sp. nov., a marine bacterium isolated from the brown alga Ecklonia cava.

A novel bacterial strain, designated EC29(T), was isolated from the brown alga Ecklonia cava collected on Jeju Island, Republic of Korea. Cells of str...
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