IJSEM Papers in Press. Published May 26, 2015 as doi:10.1099/ijs.0.000352
International Journal of Systematic and Evolutionary Microbiology Pontivivens insulae gen. nov., sp. nov., isolated from seawater --Manuscript Draft-Manuscript Number:
IJS-D-15-00096R1
Full Title:
Pontivivens insulae gen. nov., sp. nov., isolated from seawater
Short Title:
Pontivivens insulae gen. nov., sp. nov.
Article Type:
Note
Section/Category:
New taxa - Proteobacteria
Corresponding Author:
Jung-Hoon Yoon Sungkyunkwan University Suwon, KOREA, REPUBLIC OF
First Author:
Sooyeon Park
Order of Authors:
Sooyeon Park Sung-Min Won Ji-MIn Park Yong-Taek Jung Jung-Hoon Yoon
Manuscript Region of Origin:
KOREA, REPUBLIC OF
Abstract:
A Gram-stain-negative, aerobic, non-motile and coccoid, ovoid or rod-shaped bacterial strain, designated GYSW-23T, was isolated from seawater off Geoje island on the South Sea, South Korea. Strain GYSW-23T grew optimally at 25 C, at pH 7.0-8.0 and in the presence of approximately 2.0-3.0 % (w/v) NaCl. Phylogenetic trees based on 16S rRNA gene sequences revealed that strain GYSW-23T forms a distinct evolutionary lineage independent of other taxa of the family Rhodobacteraceae. It exhibited 16S rRNA gene sequence similarity values of 94.0, 93.6, 93.5, 93.4 and 93.4 % to the type strains of Roseovarius aestuarii, 'Actinobacterium atlanticum', Ruegeria marina, Roseovarius pacificus and Oceanicola litoreus, respectively. Strain GYSW-23T contained Q-10 as the predominant ubiquinone and C18:1 ω7c as the major fatty acid. The major polar lipids of strain GYSW-23T were phosphatidylcholine, phosphatidylglycerol and one unidentified aminolipid. The fatty acid and polar lipid profiles of strain GYSW-23T were distinguished from those of the phylogenetically related taxa. The DNA G+C content of strain GYSW-23T was 60.6 mol%. On the basis of the phylogenetic, chemotaxonomic and other phenotypic properties, strain GYSW23T is considered to represent a new genus and species within the family Rhodobacteraceae, for which the name Pontivivens insulae gen. nov., sp. nov. is proposed. The type strain of Pontivivens insulae is GYSW-23T (= KCTC 42458T = CECT 8812T).
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1
Pontivivens insulae gen. nov., sp. nov., isolated from seawater
2 3
Sooyeon Park,1† Sung-Min Won,1† Ji-Min Park,1 Yong-Taek Jung1,2 and Jung-Hoon Yoon1
4 5
1
6
Suwon, South Korea
7
2
8
Korea
Department of Food Science and Biotechnology, Sungkyunkwan University, Jangan-gu,
University of Science and Technology (UST), 113 Gwahak-ro, Yuseong-gu, Daejeon, South
9 10 11
Running title: Pontivivens insulae gen. nov., sp. nov.
12 13
Subject category: New taxa - Proteobacteria
14 15 16 17 18 19 20
Author for correspondence: Prof. Jung-Hoon Yoon Department of Food Science and Biotechnology, Sungkyunkwan University, Jangan-gu, Suwon, South Korea Tel : +82-31-290-7800 Fax : +82-31-290-7882 e-mail :
[email protected] 21 22
†
These authors contributed equally to this work.
23 24 25 26
The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene sequence of strain GYSW-23T is KP662553. Four supplementary figures are available with the online version of this paper.
1
27
(Abstract)
28
A Gram-stain-negative, aerobic, non-motile and coccoid, ovoid or rod-shaped bacterial strain,
29
designated GYSW-23T, was isolated from seawater off Geoje island on the South Sea, South
30
Korea. Strain GYSW-23T grew optimally at 25 C, at pH 7.0-8.0 and in the presence of
31
approximately 2.0-3.0 % (w/v) NaCl. Phylogenetic trees based on 16S rRNA gene sequences
32
revealed that strain GYSW-23T forms a distinct evolutionary lineage independent of other taxa
33
of the family Rhodobacteraceae. It exhibited 16S rRNA gene sequence similarity values of 94.0,
34
93.6, 93.5, 93.4 and 93.4 % to the type strains of Roseovarius aestuarii, ‘Actinobacterium
35
atlanticum’, Ruegeria marina, Roseovarius pacificus and Oceanicola litoreus, respectively.
36
Strain GYSW-23T contained Q-10 as the predominant ubiquinone and C18:1 ω7c as the major
37
fatty acid. The major polar lipids of strain GYSW-23T were phosphatidylcholine,
38
phosphatidylglycerol and one unidentified aminolipid. The fatty acid and polar lipid profiles of
39
strain GYSW-23T were distinguished from those of the phylogenetically related taxa. The DNA
40
G+C content of strain GYSW-23T was 60.6 mol%. On the basis of the phylogenetic,
41
chemotaxonomic and other phenotypic properties, strain GYSW-23T is considered to represent a
42
new genus and species within the family Rhodobacteraceae, for which the name Pontivivens
43
insulae gen. nov., sp. nov. is proposed. The type strain of Pontivivens insulae is GYSW-23T (=
44
KCTC 42458T = CECT 8812T).
45 2
46
The family Rhodobacteraceae, belonging to the order Rhodobacterales of the class
47
Alphaproteobacteria (Garrity et al., 2005, 2006), has been known to be one of the most
48
abundant groups in global marine environments (Rappé et al., 2000; Buchan et al., 2005;
49
Brinkhoff et al., 2008). From our studies to screen novel bacteria from a variety of seawaters off
50
Geoje island on the South Sea of the Korean Peninsula, many novel bacterial strains belonging
51
to the Roseobacter clade have recently been isolated and characterized taxonomically (Park et
52
al., 2014a, d). One of these isolates, designated GYSW-23T, is described in this study.
53
Comparative 16S rRNA gene sequence analysis showed that this novel strain is
54
phylogenetically affiliated to members of the family Rhodobacteraceae but its exact taxonomic
55
status is not clear due to equidistant 16S rRNA gene sequence similarities with representatives
56
of several genera. The aim of the present work was to determine the exact taxonomic position of
57
strain GYSW-23T by using a polyphasic characterization which included the determination of
58
chemotaxonomic and other phenotypic properties and detailed phylogenetic investigations
59
based on 16S rRNA gene sequences.
60 61 62
A seawater sample was collected from Geoje island on the South Sea, South Korea, and used as
63
the source for the isolation of bacterial strains. Strain GYSW-23T was isolated by the standard
64
dilution plating technique at 25 °C on marine agar 2216 (MA; Becton, Dickinson and Company)
3
65
and cultivated routinely under the same culture conditions. Ruegeria marina JCM 16262T,
66
Roseovarius pacificus LMG 24575T and ‘Actibacterium atlanticum’ LMG 27158T, and
67
Roseovarius aestuarii SMK-122T and Oceanicola litoreus M-M22T, which were used as
68
reference strains for phenotypic characterization and the analyses of fatty acids and polar lipids,
69
were obtained from the Japan Collection of Microorganisms (JCM; Japan), the Laboratorium
70
voor Microbiologie Universiteit Gent (LMG; Belgium) and our previous studies (Yoon et al.,
71
2008; Park et al., 2013), respectively. The cell morphology, Gram reaction, pH range for growth
72
and anaerobic growth were determined as described by Park et al. (2014b). The presence of
73
poly- -hydroxybutyrate granules was investigated by epifluorescence microscopy (BX51;
74
Olympus) after staining with Nile blue A as described by Ostle & Holt (1982). Growth at 4, 10,
75
20, 25, 28, 30, 35, 37 and 40 °C was measured on MA to determine the optimal temperature and
76
temperature range for growth. Growth at various concentrations of NaCl (0, 0.5 and 1.0-10.0 %,
77
at increments of 1.0 %) was investigated by supplementing with appropriate concentrations of
78
NaCl in marine broth 2216 (MB) prepared according to the formula of the Becton, Dickinson
79
and Company medium except that NaCl was excluded. The requirement for Mg2+ ions was
80
investigated by using MB, prepared according to the formula of the Becton, Dickinson and
81
Company medium, that comprised all of the constituents except MgCl2 and MgSO4. Catalase
82
and oxidase activities were determined as described by Lányí (1987). Hydrolysis of aesculin,
83
casein, starch, hypoxanthine, Tween 80, L-tyrosine and xanthine was tested on MA using the
4
84
substrate concentrations described by Barrow & Feltham (1993). Nitrate reduction was
85
investigated as described previously (Lányí, 1987) with the modification that artificial seawater
86
was used for the preparation of media. Hydrolysis of gelatin and urea were investigated by
87
using nutrient gelatin and urea agar base media (Becton, Dickinson and Company), respectively,
88
with the modification that artificial seawater was used for the preparation of media. The
89
artificial seawater contained (l-1 distilled water) 23.6 g NaCl, 0.64 g KCl, 4.53 g MgCl2·6H2O,
90
5.94 g MgSO4·7H2O and 1.3 g CaCl2·2H2O (Bruns et al., 2001). Utilization of various
91
substrates for growth was tested according to Baumann & Baumann (1981), using
92
supplementation with 1 % (v/v) vitamin solution (Staley, 1968) and 2 % (v/v) Hutner’s mineral
93
salts (Cohen-Bazire et al., 1957). Susceptibility to antibiotics was tested on MA plates using
94
antibiotic discs (Advantec) containing the following (g per disc unless otherwise stated):
95
ampicillin (10), carbenicillin (100), cephalothin (30), chloramphenicol (100), gentamicin (30),
96
kanamycin (30), lincomycin (15), neomycin (30), novobiocin (5), oleandomycin (15), penicillin
97
G (20 IU), polymyxin B (100 IU), streptomycin (50) and tetracycline (30). Enzyme activities
98
were determined, after incubation for 8 h at 25 ºC, by using the API ZYM system (bioMérieux);
99
the strip was inoculated with cells suspended in artificial seawater from which CaCl 2 was
100
excluded to avoid the formation of precipitates. For spectral analysis of in vivo pigment
101
absorption, strain GYSW-22T was cultivated aerobically in the dark at 25 °C in MB. The culture
102
was washed twice using centrifugation with a MOPS buffer (MOPS/NaOH, 0.01 M; KCl, 0.1
5
103
M; MgCl2, 0.001 M; pH 7.5) and disrupted by means of sonication (VC505; Sonics & Materials,
104
Inc.). After removal of cell debris by centrifugation, the absorption spectrum of the supernatant
105
was examined on the Eon Microplate spectrophotometer (Biotek).
106 107
Cell biomass of strain GYSW-23T for DNA extraction and for the analyses of isoprenoid
108
quinones and polar lipids was obtained from cultures grown for 3 days in MB at 25 °C, and cell
109
biomass of R. aestuarii SMK-122T, R. pacificus LMG 24575T, ‘A. atlanticum’ LMG 27158T and
110
R.marina JCM 16262T for polar lipid analysis was obtained from the same culture conditions.
111
Chromosomal DNA was extracted and purified according to Yoon et al. (1996), with the
112
modification that RNase T1 was used in combination with RNase A to minimize contamination
113
of RNA. The 16S rRNA gene was amplified by PCR as described previously (Yoon et al.,
114
1998)
115
ACGGTTACCTTGTTACGACTT-3’. Sequencing of the amplified 16S rRNA gene and
116
phylogenetic analysis were performed as described by Yoon et al. (2003).
using
two
universal
primers,
5’-GAGTTTGATCCTGGCTCAG-3’
and
5’-
117 118
Isoprenoid quinones were extracted and analysed as described by Komagata & Suzuki (1987),
119
using reversed-phase HPLC and a YMC ODS-A (250×4.6 mm) column. The isoprenoid
120
quinones were eluted by a mixture of methanol/isopropanol (2:1, v/v) using a flow rate of 1 ml
121
min-1 at room temperature and detected by UV absorbance at 275 nm. For cellular fatty acid
6
122
analysis, cell mass of strain GYSW-23T was harvested from MA plates after cultivation for 3, 5
123
and 7 days at 25 °C, and cell mass of R. aestuarii SMK-122T, R. pacificus LMG 24575T, ‘A.
124
atlanticum’ LMG 27158T, R. marina JCM 16262T and O. litoreus M-M22T was harvested from
125
MA plates after cultivation for 5 days at 25 °C. Fatty acids were saponified, methylated and
126
extracted using the standard MIDI protocol (Sherlock Microbial Identification System, version
127
6.2B). The fatty acids were analysed by GC (Hewlett Packard 6890) and identified using the
128
TSBA6 database of the Microbial Identification System (Sasser, 1990). Polar lipids were
129
extracted according to the procedures described by Minnikin et al. (1984), and separated by
130
two-dimensional TLC using chloroform/methanol/water (65:25:3.8, by vol.) for the first
131
dimension and chloroform/methanol/acetic acid/water (40:7.5:6:1.8, by vol.) for the second
132
dimension as described by Embley & Wait (1994). Individual polar lipids were identified by
133
spraying the plates with 10 % ethanolic molybdophosphoric acid, molybdenum blue, ninhydrin
134
and -naphthol reagents (Minnikin et al., 1984; Komagata & Suzuki, 1987) and with
135
Dragendorff’s reagent (Sigma). The DNA G+C content was determined by the method of
136
Tamaoka & Komagata (1984) with the modification that DNA was hydrolysed and the resultant
137
nucleotides were analysed by reversed-phase HPLC equipped with a YMC ODS-A (2504.6
138
mm) column. The nucleotides were eluted by a mixture of 0.55 M NH4H2PO4 (pH 4.0) and
139
acetonitrile (40:1, v/v), using a flow rate of 1 ml min-1 at room temperature and detected by UV
140
absorbance at 270 nm.
7
141 142 143
Morphological, cultural, physiological and biochemical characteristics of strain GYSW-23T are
144
given in the genus and species descriptions (see below) or in Table 1 or Fig. S1 (available in the
145
online Supplementary Material). The almost-complete 16S rRNA gene sequence of strain
146
GYSW-23T determined in this study comprised 1385 nucleotides, representing approximately
147
95 % of the E. coli 16S rRNA gene sequence. Strain GYSW-23T exhibited the highest 16S
148
rRNA gene sequence similarity value (94.0 %) to the type strain of Roseovarius aestuarii. It
149
exhibited 16S rRNA gene sequence similarity values of 93.6, 93.5, 93.4 and 93.4 % to
150
‘Actibacterium atlanticum’ 22II-S11-z10T, Ruegeria marina ZH17T, Roseovarius pacificus 81-
151
2T and Oceanicola litoreus M-M22T, respectively. In the neighbour-joining, maximum-
152
likelihood and maximum-parsimony phylogenetic trees based on 16S rRNA gene sequences,
153
strain GYSW-23T was found to form a distinct evolutionary lineage independent of other taxa of
154
the family Rhodobacteraceae (Fig. 1; Fig. S2 & S3).
155 156
The predominant isoprenoid quinone detected in strain GYSW-23T was ubiquinone-10 (Q-10),
157
which is typical of the vast majority of the class Alphaproteobacteria. In Table 2, the cellular
158
fatty acid profile of strain GYSW-23T is compared with those of the type strains of some
159
phylogenetically related species (Table 2). The major fatty acid (> 10 % of total fatty acids)
8
160
detected in strain GYSW-23T was C18:1 ω7c. The fatty acid profile of strain GYSW-23T was
161
similar to those of the type strains of R. aestuarii, R. pacificus, ‘A. atlanticum’, R. marina and O.
162
litoreus in that C18:1 ω7c is major fatty acid, but distinguishable from the reference strains by
163
differences in proportions of some fatty acids, particularly C16:0, C18:0 and 11-methyl C18:1 ω7c
164
(Table 2). The major polar lipids detected in strain GYSW-23T were phosphatidylcholine,
165
phosphatidylglycerol and one unidentified aminolipid; minor amounts of one unidentified lipid
166
and one unidentified phospholipid were also present (Fig. S4). The polar lipid profile of strain
167
GYSW-23T was distinguished from those of the type strains of R. aestuarii, R. pacificus, ‘A.
168
atlanticum’, R. marina and O. litoreus by the absence of phosphatidylethanolamine,
169
sulfoquinovosyldiacylglycerol and other unidentified lipid(s) as major components (Fig. S4;
170
Park et al., 2013). The polar lipid profile of strain GYSW-23T was also distinguished from those
171
of other members of the genera Roseovarius, Actibacterium, Ruegeria and Oceanicola by the
172
absence of phosphatidylethanolamine and/or unidentified lipid(s) as major components (Kim et
173
al., 2012a, b; Lucena et al., 2012; Choi et al., 2013; Kämpfer et al., 2013; Park et al., 2013,
174
2014c; Rajasabapathy et al., 2014). The DNA G+C content of strain GYSW-23T was 60.6
175
mol%, a value in the range reported for members of the phylogenetically related taxa (Table 1).
176 177
From the results of the phylogenetic analyses, it does not seem appropriate to assign strain
178
GYSW-23T to any of the recognized genera of the family Rhodobacteraceae (Fig. 1; Fig. S2 &
9
179
S3). The differences in fatty acid and polar lipid profiles make it reasonable to differentiate
180
strain GYSW-23T from the phylogenetically related taxa (Table 2; Fig. S4). Strain GYSW-23T
181
could be distinguished from the type strains of some phylogenetically related species by
182
differences in several phenotypic characteristics, including motility, growth at 40 C, nitrate
183
reduction, hydrolysis and utilization of some substrates, activity of some enzymes and
184
susceptibility to some antibiotics (Table 1). Accordingly, on the basis of the phylogenetic and
185
chemotaxonomic distinctiveness and other differential phenotypic properties, strain GYSW-23T
186
is considered to represent a novel genus and species within the family Rhodobacteraceae, for
187
which the name Pontivivens insulae gen. nov., sp. nov., is proposed.
188 189 190
Description of Pontivivens gen. nov.
191 192
Pontivivens (Pon.ti.vi’vens. L. n. pontus the sea; L. part. adj. vivens living; N.L. part. adj.
193
Pontivivens living in the sea).
194 195
Cells are Gram-stain-negative, aerobic, non-motile and coccoid, ovoid or rod-shaped. Catalase-
196
and oxidase-positive. Nitrate reduction is positive. The predominant ubiquinone is Q-10. The
197
major fatty acid is C18:1 ω7c. The major polar lipids are phosphatidylcholine,
10
198
phosphatidylglycerol and one unidentified aminolipid. The DNA G+C content is 60.6 mol%.
199
The type species is Pontivivens insulae. A member of the family Rhodobacteraceae, the class
200
Alphaproteobacteria, according to 16S rRNA gene sequence analysis.
201 202 203
Description of Pontivivens insulae sp. nov.
204 205
Pontivivens insulae (in’su.lae. L. gen. n. insulae of an island, referring to the source of
206
isolation of the type strain).
207 208
Cells are Gram-stain-negative, non-flagellated and coccoid, ovoid or rod-shaped, approximately
209
0.2-0.7 μm in diameter and 0.3-6.0 μm in length. Colonies on MA are circular, slightly convex,
210
smooth, glistening, greyish yellow in colour and 0.5-1.0 mm in diameter after incubation for 5
211
days at 25 °C. Optimal temperature for growth is 25 °C; growth occurs at 15 and 35 °C, but not
212
at 10 and 37 °C. Optimal pH for growth is 7.0-8.0; growth occurs at pH 6.0, but not at pH 5.5.
213
Optimal growth occurs in the presence of approximately 2.0-3.0 % (w/v) NaCl; growth occurs
214
in the presence of 1.0-8.0 % (w/v) NaCl. Mg2+ ions are required for growth. Growth does not
215
occur under anaerobic conditions on MA and on MA supplemented with nitrate.
216
Bacterichlorophyll a is not produced. Catalase- and oxidase-positive. Nitrate is reduced to
11
217
nitrite. Aesculin and L-tyrosine are hydrolysed, but casein, gelatin, hypoxanthine, starch, Tween
218
80, urea and xanthine are not. D-Cellobiose, D-galactose, D-glucose, maltose, sucrose, D-xylose,
219
L-malate, pyruvate, succinate and salicin are utilized as sole carbon and energy sources, but L-
220
arabinose, D-fructose, D-mannose, D-trehalose, acetate, benzoate, citrate, formate and L-
221
glutamate are not. Susceptible to chloramphenicol, kanamycin, neomycin, novobiocin,
222
oleandomycin, polymyxin B, streptomycin and tetracycline, but not to ampicillin, carbenicillin,
223
cephalothin, gentamicin, lincomycin and penicillin G. Activity of alkaline phosphatase, esterase
224
(C 4) and esterase lipase (C 8) is present and activity of leucine arylamidase is weakly present,
225
but activity of lipase (C 14), valine arylamidase, cystine arylamidase, trypsin, α-chymotrypsin,
226
acid phosphatase, naphthol-AS-BI-phosphohydrolase, α-galactosidase, β-galactosidase, β-
227
glucuronidase, α-glucosidase, β-glucosidase, N-acetyl-β-glucosaminidase, α-mannosidase and
228
α-fucosidase is absent. The predominant ubiquinone is Q-10. The major fatty acid (> 10 % of
229
total fatty acids) is C18:1 ω7c. The major polar lipids are phosphatidylcholine,
230
phosphatidylglycerol and one unidentified aminolipid. The DNA G+C content of the type strain
231
is 60.6 mol%.
232
The type strain, GYSW-23T (= KCTC 42458T = CECT 8812T), was isolated from seawater off
233
Geoje island on the South Sea, South Korea.
234 235 12
236
Acknowledgements
237 238
This work was supported by the project on survey of indigenous species of Korea of the
239
National Institute of Biological Resources (NIBR) under the Ministry of Environment (MOE)
240
and the Programs for Collection, Management and Utilization of Biological Resources (grant
241
NRF-2013M3A9A5075953) from the Ministry of Science, ICT & Future Planning (MSIP) of
242
the Republic of Korea.
243 244 245
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375 376 377 378 379 380 381 382 383 384 385 386 387 20
388 389 390 391 392 393 394 395 396 397 398 399 400 401 402
Table 1. Differential characteristics of strain GYSW-23T and the type strains of some phylogenetically related species. Strain: 1, GYSW-23T; 2, Roseovarius aestuarii SMK-122T, data from Yoon et al. (2008); 3, Roseovarius pacificus LMG 24575T, data from Jung et al. (2012) and Kim et al. (2012) unless indicated otherwise; 4, ‘Actibacterium atlanticum’ LMG 27158T, data from this study unless indicated otherwise; 5, Ruegeria marina JCM 16262T, data from this study unless indicated otherwise; 6, Oceanicola litoreus M-M22T, data from Park et al. (2013). +, positive reaction; , negative reaction; w, weakly positive reaction; ND, not determined. All strains are positive for activity of catalase and oxidase; utilization of L-malate (weak for Ruegeria marina JCM 16262T) and succinate (weak for Ruegeria marina JCM 16262T and Oceanicola litoreus M-M22T); activity of alkaline phosphatase (weak for Roseovarius aestuarii SMK-122T), esterase (C 4) and esterase lipase (C 8); and susceptibility to chloramphenicol. All strains are negative for Gram-staining; hydrolysis of starch; utilization of D-trehalose, formate and L-glutamate; activity of lipase (C 14), cystine arylamidase, trypsin, -chymotrypsin, -galactosidase, -galactosidase, glucuronidase, -glucosidase, -glucosidase, N-acetyl--glucosaminidase, -mannosidase and -fucosidase; and susceptibility to lincomycin.
Characteristic Motility Growth at 4 C 40 C Nitrate reduction to nitrite Hydrolysis of Aesculin Casein Gelatin Tween 80 Utilization of L-Arabinose D-Cellobiose D-Fructose D-Galactose, sucrose, salicin D-Glucose Maltose D-Mannose D-Xylose Acetate Benzoate Citrate Pyruvate Enzyme activity (by API ZYM) Leucine arylamidase Valine arylamidase Acid phosphatase Naphthol-AS-BIphosphohydrolase Susceptibility to antibiotics Ampicillin Carbenicillin, cephalothin Gentamicin Kanamycin Neomycin Novobiocin Oleandomycin Penicillin G
1
2
3 *
4 *
5 *
6
W* ND *
* * *
* * *
* * * *
* * * *
w
w
w
w w
w
21
Polymyxin B Streptomycin Tetracycline DNA G+C content (mol%)
403 404
60.6
58.6
62.3*
59.0*
63.5*
w 67.6
*Data of R. pacificus LMG 24575T, A. atlanticum LMG 27158T and R. marina JCM 16262T taken from Wang et al. (2009), Li et al. (2014) and Huo et al. (2011), respectively.
405 406 407 408 409 410 411 412 413 414 415 416 417 418 419 420 421 22
422 423 424 425 426 427 428
Table 2. Cellular fatty acid compositions (%) of strain GYSW-23T and the type strains of some phylogenetically related species. Strains: 1, GYSW-23T (3 days); 2, GYSW-23T (5 days); 3, GYSW-23T (7 days); 4, Roseovarius aestuarii SMK-122T; 5, Roseovarius pacificus LMG 24575T; 6, ‘Actibacterium atlanticum’ LMG 27158T; 7, Ruegeria marina JCM 16262T; 8, Oceanicola litoreus M-M22T. Fatty acids that represented < 0.5 % in all columns were omitted. TR, Traces (< 0.5 %); –, Not detected; ECL, Equivalent chain length.
Fatty acid Straight-chain C10:0 C12:0 C16:0 C17:0 C18:0 Branched iso-C15:1 F anteiso-C15:1 A Unsaturated C17:1 ω8c C18:1 ω7c anteiso-C17:1 ω9c Hydroxy C10:0 3-OH C12:0 3-OH C12:1 3-OH C16:0 2-OH C18:1 2-OH iso-C13:0 3-OH 11-methyl C18:1 ω7c Cyclo C19:0 ω8c Summed features* 1 3 7
429 430 431
1
2
3
4
5
6
7
8
TR – 1.3 1.3 3.2
– – 1.1 1.7 3.3
– – 1.2 1.0 3.2
0.6 4.7 14.6 TR 1.2
0.6 3.9 3.4 TR 0.6
– – 1.2 – 4.0
2.0 1.7 5.8 TR 1.1
TR – 4.7 1.3 8.3
– –
TR –
2.1 0.9
– –
– –
– –
TR –
– –
1.0 84.1 –
1.3 81.4 –
1.4 73.4 1.2
– 69.1 –
– 66.5 –
– 75.6 –
– 72.8 –
– 79.3 –
0.8 – – – – – 2.4 –
1.1 – – – – TR 2.1 –
1.1 – – – 0.8 2.4 3.1 –
TR 5.4 – – – – 0.8 1.3
TR 2.9 2.2 2.8 – – 10.0 5.1
5.1 1.8 – – – – 10.3 1.4
TR 4.5 TR 2.3 TR TR 6.9 –
3.3 – – – – – TR 0.8
– 0.8 4.4
– 1.2 5.3
0.9 1.3 4.3
– TR –
– 0.7 –
– – –
TR TR –
– 1.5 –
*Summed feature 1 contains iso-C15:1 H and/or C13:0 3-OH; summed feature 3 contains C16:1 ω7c and/or C16:1 ω6c; summed feature 7 contains unknown fatty acid 18.846(ECL), C19:1 ω6c and/or cyclo C19:0 ω10c.
432 433 434 435 23
436 437
Legend to Figure
438 439
Fig. 1. Neighbour-joining phylogenetic tree based on 16S rRNA gene sequences showing the
440
positions of Pontivivens insulae GYSW-23T and representatives of some other related taxa.
441
Only bootstrap values (expressed as percentages of 1000 replications) greater than 50 % are
442
shown at branching points. Filled circles indicate that the corresponding nodes were also
443
recovered in the trees generated with the maximum-likelihood and maximum-parsimony
444
algorithms. Stappia stellulata IAM 12621T (GenBank accession number, D88525) was used as
445
an outgroup. Scale bar, 0.01 substitutions per nucleotide position.
24
Figure 1 Click here to download Figure: Fig. 1.ppt
Sulfitobacter dubius KMM 3554T (AY180102) Sulfitobacter pontiacus ChLG 10T (Y13155) Roseobacter litoralis ATCC 49566T (X78312) Aestuariihabitans beolgyonensis BB-MW15T (KC577450) Pelagicola litoralis CL-ES2T (EF192392) Pacificibacter maritimus KMM 9031T (AB558927) 99.6 Leisingera aquimarina LMG 24366T (AM900415) 55.9 Leisingera methylohalidivorans MB2T (AY005463) 66.5 Phaeobacter caeruleus LMG 24369T (AM943630) Phaeobacter gallaeciensis BS107T (Y13244) Marinovum algicola ATCC 51440T (X78315) 98.4 Ruegeria lacuscaerulensis ITI-1157T (U77644) 58.0 Ruegeria atlantica IAM 14463T (D88526) Ruegeria pomeroyi DSS-3T (AF098491) Ruegeria marina ZH17T (FJ872535) Ruegeria scottomollicae LMG 24367T (AM905330) 100 Marivita litorea CL-JM1T (EU512918) 84.2 Marivita cryptomonadis CL-SK44T (EU512919) Thalassobius maritimus GSW-M6T (HM748766) 50.6 Thalassococcus lentus YCS-24T (JX090308) Thalassococcus halodurans UST050418-052T (DQ397336) 95.1 69.4 Roseovarius pacificus 81-2T (DQ120726) 100 Roseovarius halotolerans HJ50T (EU431217) Roseovarius litoreus GSW-M15T (JQ390520) Donghicola eburneus SW-277T (DQ667965) 51.9 Citreimonas salinaria CL-SP20T (AY962295) Donghicola xiamenensis Y-2T (DQ120728) 56.2 Cribrihabitans marinus CZ-M5T (JX306766) 88.3 Roseovarius aestuarii SMK-122T (EU156066) Roseovarius nubinhibens ISMT (AF098495) 89.1 Roseovarius tolerans EL-172T (Y11551) Pontivivens insulae GYSW-23T (KP662553) 87.5 Rhodovulum kholense JA297T (AM748927) 99.7 Rhodovulum sulfidophilum DSM 1374T (D16423) Rhodovulum marinum JA128T (AJ891122) Rhodobacter capsulatus ATCC 11166T (D16428) Rhodobacter sphaeroides 2.4.1T (X53853) 99.7 ‘Confluentimicrobium lipolyticum’ SSK1-4T (KJ889015) 95.3 Paracoccus aminophilus ATCC 49673T (AY014176) Paracoccus denitrificans ATCC 17741T (Y16927) 100 Paracoccus saliphilus YIM 90738T (DQ923133) ‘Actibacterium atlanticum’ 22II-S11-z10T (KJ159064) Actibacterium mucosum R46T (HE590855) 99.7 69.1 Pseudoruegeria aquimaris SW-255T (DQ675021) Pseudoruegeria lutimaris HD-43T (FJ374173) Celeribacter neptunius H 14T (FJ535354) Celeribacter baekdonensis L-6T (HM997022) 99.1 99.6 Tropicimonas sediminicola M97T (JF748735) Tropicimonas isoalkanivorans B51T (AB302379) Oceanicola litoreus M-M22T (JX291104) Oceanicola granulosus HTCC2516T (AY424896) Roseisalinus antarcticus EL-88T (AJ605747) 90.1 Wenxinia marina HY34T (DQ640643) Rubellimicrobium thermophilum C-lvk-R2A-2T (AJ844281) Stappia stellulata IAM 12621T (D88525) 71.5 99.2
70.9
0.01
Fig. 1
Supplementary Materials (PDF) Click here to download Supplementary Material Files: Supplementary materials.pdf