Antonie van Leeuwenhoek (2012) 101:753–760
DOI 10.1007/s10482-011-9689-4
ORIGINAL PAPER
Pseudonocardia yuanmoensis sp. nov., a novel
actinobacterium isolated from soil in Yunnan, south-west
China
Guo-Xing Nie • Hong Ming • Da-Qiao Wei
En-Min Zhou • Xia Tang • Juan Cheng •
Shu-Kun Tang • Wen-Jun Li
•
Received: 18 September 2011 / Accepted: 13 December 2011 / Published online: 27 December 2011
Ó Springer Science+Business Media B.V. 2011
Abstract A novel Gram-stain positive, aerobic, nonmotile, spore-forming actinobacterium, designated
YIM 75926T, was isolated from a soil sample
collected at soil forest in Yuanmo county of Yunnan
province, south-west China. Its taxonomic position
was investigated by a polyphasic approach. Phylogenetic analyses based on 16S rRNA gene sequences
showed that the novel strain YIM 75926T belongs to
the genus Pseudonocardia and was closely related to
Guo-Xing Nie and Hong Ming contributed equally to this
work.
Electronic supplementary material The online version
of this article (doi:10.1007/s10482-011-9689-4) contains
supplementary material, which is available to authorized users.
G.-X. Nie
College of Life Sciences, Henan Normal University,
Xinxiang 453007, People’s Republic of China
H. Ming
Department of Life Sciences and Technology, Xinxiang
Medical University, Xinxiang 453003,
People’s Republic of China
Pseudonocardia halophobica DSM 43089T (98.1%
similarity). Strain YIM 75926T had MK-8 (H4) as the
predominant menaquinone. The whole organism
hydrolysates mainly consisted of meso-diaminopimelic acid, mannose, glucose, galactose and arabinose.
The major cellular fatty acids were iso-C16:0 (37.16%)
and C16:0 (12.43%). The DNA G?C content of strain
YIM 75926T was 70.6 mol%. The resultant phylogenetic trees further showed that strain YIM 75926T
belong to Pseudonocardia and had a distinct subclade
within the evolutionary radiation of the genus Pseudonocardia. On the basis of its comparative analysis of
phenotypic and genotypic characteristics, it is proposed that strain YIM 75926T represent a novel
species of the genus Pseudonocardia, named Pseudonocardia yuanmoensis sp. nov. The type strain is
S.-K. Tang
e-mail: [email protected]
D.-Q. Wei
Faculty of Life Science and Technology, Kunming
University of Science and Technology, Kunming 650224,
People’s Republic of China
H. Ming E.-M. Zhou X. Tang J. Cheng S.-K. Tang W.-J. Li (&)
Key Laboratory of Microbial Diversity in Southwest
China, Ministry of Education, and Laboratory for
Conservation and Utilization of Bio-resources, Yunnan
Institute of Microbiology, Yunnan University, Kunming
650091, People’s Republic of China
e-mail: [email protected]; [email protected]
123
754
Antonie van Leeuwenhoek (2012) 101:753–760
YIM 75926T (=CCTCC AA 2011017T = JCM
18055T).
the genus Pseudonocardia, for which the name
Pseudonocardia yuanmoensis sp. nov., is proposed.
Keywords Pseudonocardia yuanmoensi sp. nov. 16S rRNA Polyphasic taxonomy
Materials and methods
Strain and culture conditions
Introduction
The genus Pseudonocardia within the family Pseudonocardiaceae was originally described by Henssen
(1957), and since then the description of the genus has
been emended repeatedly (Warwick et al. 1994;
McVeigh et al. 1994; Reichert et al. 1998; Huang
et al. 2002; Park et al. 2008). So far, the genus
Pseudonocardia encompasses 44 species with validly
published names (http://www.bacterio.cict.fr/; Ara
et al. 2011; Kaewkla and Franco 2010, 2011; Li et al.
2010; Qin et al. 2010, 2011; Sakiyama et al. 2010;
Zhao et al. 2011a). Members of the genus Pseudonocardia have the characteristics as follow: have the type
IV cell wall, aerial and vegetative mycelium, display
spore chains which are produced by acropetal budding
or fragmentation, have MK-8 (H4) or MK-9 as their
major menaquinone, iso-branched hexadecanoic acid
as the predominant fatty acid. Mycolic acids are absent
and the phospholipids are type II, III or IV. The DNA
G?C contents are from 68 to 79 mol%. In recent
years, members of the genus Pseudonocardia have
been reported widely and have been isolated mostly
from plant samples (including stems, leaves, root
nodules, tree-bark composts and traditional medicinal
plants) (Gu et al. 2006; Chen et al. 2009; Duangmal
et al. 2009; Kaewkla and Franco 2010, 2011; Qin et al.
2010, 2011; Zhao et al. 2011b), some also have been
isolated from soil samples (including polluted soil
samples by industrial chemical) (Qin et al. 2008;
Mahendra and Cohen 2005), coastal sediment (Liu
et al. 2006) and so on.
In the course of our research on actinobacterial
sources, we obtained a lot of novel isolated strains
from soil forest samples in Yuanmo county of Yunnan
province, south-west China. In this study, the characteristics analysis of strain YIM 75926T, such as
phenotypic, chemotaxonomic and phylogenetic taxonomy were carried out to affiliate it to the genus
Pseudonocardia. The data we obtained also indicate
that isolate YIM 75926T represents a novel species of
123
Strain YIM 75926T was isolated from a soil forest
sample collected at Yuanmo, Yunnan province, southwest China. The soil sample was air-dried at room
temperature and then take 2 g soil sample in a flask
with 18 ml sterile water and several glass beads,
followed by shaking at 55°C and 200 rpm min-1 for
2 h. 1 ml mixture sample was taken into a tube with
9 ml sterilized water, and serial dilution of 1 ml soil
suspension to 10-2, and then 0.3 ml of the 10-2
dilution suspension was spread and incubated on
International Streptomyces Project (ISP) 5 medium
agar plate, supplemented with nalidixic acid
(25 mg l-1) and nystatin (50 mg l-1). The plate was
incubated at 28°C for 1 week. The purified strain YIM
75926T was obtained and routinely cultured on ISP 2
medium at 28°C and stored as a glycerol suspension
(20%, w/v) at -80°C.
Biomass for chemical and molecular studies was
obtained by cultivation in shaken flasks (about
200 rpm) using ISP 2 medium or Tryptic Soy Broth
(TSB, containing 15 g tryptone, 5 g soya peptone and
5 g NaCl per litre, pH 7.2) medium at 28°C for
1 week.
Morphological, cultural, physiological
and biochemical characteristics
Gram staining was carried out by using the standard
Gram reaction (Gram 1884). Cell motility was
confirmed by the development of turbidity throughout
a tube containing semisolid medium (Leifson 1960).
Cultured characteristics were tested on ISP media
(Shirling and Gottlieb 1966), Czapek’s agar, nutrient
agar and potato–glucose agar media. Aerial sporemass colour, substrate mycelium pigmentation and
coloration of the diffusible pigments were observed on
these media described as by Dong and Cai (2001).
Colours were determined by using the colour chips
from the ISCC–NBS colour charts (standard samples,
No. 2106) (Kelly 1964). The morphological characteristics of strain YIM 75926T were carried out by light
Antonie van Leeuwenhoek (2012) 101:753–760
microscopy (Philips XL30) and scanning electron
microscopy (ESEM-TMP), the cultures grown on ISP
2 medium at 28°C for 7–21 days and the spore-chain
morphology, spore size and surface ornamentation
were observed. Growth temperatures were tested at 4,
12, 28, 37, 45, 50 and 55°C on ISP 2 plates. The pH
range for growth at pH 4, 5, 6, 7, 8, 9 and 10 and NaCl
tolerance (0, 1, 3, 5, 7, 8, 9, 10, 15 and 20% w/v) were
tested at 28°C for 14–21 days by culturing the strains
in ISP 2 medium. Gelatinase, oxidase, urease and
catalase activities, nitrate reduction, starch and cellulose hydrolysis were observed as described by Smibert
and Krieg (1994). The strains grew in Pridham and
Gottlieb’s basal mineral salts medium with tested sole
carbon and energy sources. Acid production from
carbohydrates and other physiological and biochemical characteristics were assessed by using the media
and methods described by Gordon et al. (1974).
Chemotaxonomy
Menaquinones were extracted from lyophilized cell
described by Collins et al. (1977) and the extracts were
purified and analyzed by HPLC (Tamaoka et al. 1983;
Hu et al. 2001). Isomer of diaminopimelic acid and
sugar analysis of whole-cell hydrolysates were performed by using the procedures recorded by Hasegawa
et al. (1983), Lechevalier and Lechevalier (1970) and
Tang et al. (2009). The acyl type of the cell wall was
analysed according to the method of Uchida and Aida
(1984). Biomass for cellular fatty acids analysis were
cultured with TSB medium at 28°C for 3 days, the
cellular fatty acids were extracted, methylated and
analysed by using the Sherlock microbial identification system (MIDI) (Sherlock Version 6.1; MIDI
database: TSBA6). Phospholipids were extracted,
examined by two-dimensional thin-layer chromatography (TLC) and identified using the described
procedures (Minnikin et al. 1979; Collins and Jones
1980). Mycolic acids were extracted and analyzed by
one-dimensional TLC described by Minnikin et al.
(1980). The G?C content of the genomic DNA was
determined by using the HPLC method (Mesbah et al.
1989) with E. coli JM-109 as the reference strain.
Molecular analysis
Genomic DNA extraction, PCR amplification and the
16S rRNA gene sequences were performed as
755
described by Li et al. (2007). The 16S rRNA gene
sequence of strain YIM 75926T was compared against
cultured species via BLAST database (http://blast.
ncbi.nlm.nih.gov/Blast.cgi) and type strains via
Eztaxon
database
(http://147.47.212.35:8080/).
Throughout the two kinds of analysis, the most
sequences of recognized bacteria similar with YIM
75926T were retrieved. Multiple alignments were
performed by using the CLUSTAL_X software
package (Thompson et al. 1997). The Kimura twoparameter model (Kimura 1983) of theses sequences
was used to calculate evolutionary distance. And then,
the phylogenetic trees were constructed by neighbourjoining (Saitou and Nei 1987), maximum-parsimony
(Fitch 1971) and maximum-likelihood (Felsenstein
1981) tree-making algorithm by using the software
packages MEGA version 5.0 (Tamura et al. 2011) and
PHYML (Guindon and Gascuel 2003). The topologies
of the phylogenetic trees were evaluated by using the
bootstrap resampling method of Felsenstein (1985)
with 1000 replicates. The strain Kutzneria kofuensis
NRRL B-24061T (AF114801) was used as outgroup.
DNA–DNA relatedness was carried out according to
the fluorometric micro-well method (Ezaki et al. 1989;
Christensen et al. 2000; He et al. 2005), and the
hybridizations were performed with six replications.
Nucleotide sequence accession number
The 16S rRNA gene sequence of strain YIM 75926T
has been deposited in GenBank under the accession
number JN656709.
Results and discussion
The results showed that strain YIM 75926T had
morphological typical properties of the genus Pseudonocardia. The cells were Gram-positive, aerobic and
non-motile. Strain YIM 75926T grew well on ISP 2,
Czapek’s agar and potato-glucose agar media, grew
moderate on ISP 3 and ISP 4, and grew weak on ISP 5
nutrient agar medium. White aerial mycelium was
produced on all media except nutrient agar and ISP 5
medium. The substrate mycelium was coloured (yellow, orange-yellow and yellow-brown) when grown
on all media, and no diffusible pigment was observed
on each type of medium (Supplementary Table S1).
Morphological observation of a 14 day old culture of
123
756
strain YIM 75926T revealed that both aerial and
vegetative hyphae were abundant, well developed and
fragmented into rod-shaped elements (Supplementary
Fig. S1). Growth was observed at 10–45°C (optimum
28–37°C), pH 6.0–8.0 (optimum pH 7.0) and 0–5%
(w/v) NaCl (optimum 0–3%). Activities of urease and
catalase were positive and activities of gelatinase and
oxidase were negative. Tests of milk coagulation, milk
peptonization, reduction of nitrate, hydrolysis of
starch and Tweens 20, 40, 60, 80 were positive, and
hydrolysis of cellulose, H2S production and melanin
formation were negative. Detailed physiological characteristics differentiating strain YIM 75926T from its
closely related type strain are showed in Table 1.
The major respiratory menaquinone of strain YIM
75926T was MK-8 (H4), which is characteristic of
species of the genus Pseudonocardia. The cell wall
diamino acid of the strain YIM 75926T was mesodiaminopimelic acid (meso-DAP) in the peptidoglycan, and its whole organism hydrolysates were rich in
mannose, glucose, galactose and arabinose (Supplementary Fig. S2). The glycan moiety of the murein
was acetylated. Mycolic acids were absent. The major
fatty acids consisted of iso-C16:0 (37.16%) and C16:0
(12.43%), and the moderate amount fatty acids
included iso-C15:0 (7.50%), anteiso-C17:0 (7.81) and
iso-C17:0 (7.50%). Detailed fatty acid profiles of strain
YIM 75926T and the reference type strain of the
mostly related species Pseudonocardia halophobica
DSM 43089T are given in Supplementary Table S2.
The phospholipids of strain YIM 75926T consisted of
diphosphatidylglycerol, phosphatidylglycerol, phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylcholine, phosphatidylinositol,
phosphatidylinositol mannoside, two unknown glycolipid and two unknown polar lipids (Supplementary
Fig S3). The G?C content of strain YIM 75926T was
70.6 mol%, and it is in accordance with the level for
the genus Pseudonocardia (from 68 to 79 mol%).
The almost-complete 16S rRNA gene sequence
(1,518 nt) of strain YIM 75926T was aligned manually
with the corresponding 16S rRNA gene sequences of
all the members of the genus Pseudonocardia which
were retrieved from GenBank/EMBL/DDBJ. On the
basis of 16S rRNA gene sequences, phylogenetic
analyses showed that strain YIM 75926T displayed a
distinct subclade with the type strain of P. halophobica DSM 43089T, which shared a 16S rRNA gene
sequence similarity of 98.1%. The sequence
123
Antonie van Leeuwenhoek (2012) 101:753–760
Table 1 Different characteristics of strains YIM 75926T and
P. halophobica DSM 43089T
Characteristic
1
2
-
?
-
?
D-Trehalose
-
?
L-Rhamnose
-
?
Cellobiose
-
?
Succinic acid
-
?
Utilization as sole carbon source
D-Mannose
Glucose
Acid produced from
D-Mannose
-
?
Glucose
-
?
D-Sucrose
?
-
L-Arabinose
?
-
D-Galactose
-
?
-
?
Sodium acetate
?
?
-
Trisodium citrate
?
w
Succinic acid
-
?
L-Lysine
-
?
L-Histidine
±
?
-
?
pH 6.0
?
-
7% NaCl (w/v)
-
Glycerol
D-Trehalose
Utilization as sole nitrogen source
Hydrolysis of
Gelatinase
Growth at/on
?
T
Strains 1: P. yuanmoensis YIM 75926 , 2: P. halophobica
DSM 43089T
Data obtained during this study were carried out under
identical growth conditions
? Positive, utilized; - negative, not utilized; w weakly positive
similarities between strain YIM 75926T and other
members of the genus Pseudonocardia were below
98.0% (from 97.4 to 94.5%). An association which
was showed by neighbour-joining phylogenetic tree
supported by a bootstrap value of 93% (Fig. 1). From
the phylogenetic tree, the closest neighbour of strain
YIM 75926T is P. halophobica DSM 43089T, an
apparently distinct subclade was displayed between
them and the other phylogenetic trees such as Minimum evolution phylogenetic tree, Maximum parsimony phylogenetic tree and Maximum likelihood
phylogenetic tree supported this with the bootstrap
Antonie van Leeuwenhoek (2012) 101:753–760
757
P. kunmingensis YIM 63158T(FJ817377)
100*
P.sichuanensis KLBMP 1115T (HM153789)
95*
P.zijingensis JCM11117T (AF325725)
P. adelaidensis EUM 221T (FJ805427)
62*
P. serianimatus YIM 63233T (FJ817379)
P.petroleophila IMSNU 22072T (AJ252828)
99*
P.aurantiaca DSM 44773T (FR749916)
50
P.xinjiangensis CCTCC AA97020T (EU722520)
P.alaniniphila YIM 16303T (EU722519)
P.yunnanensis IFO 15681T (D85472)
100*
P. mongoliensis MN08-A0270 T (AB521671)
P.saturnea IMSNU 20052T (AJ252829)
P.chlorethenivorans SL-1T (AF454510)
P.asaccharolytica DSM 44247T (Y08536)
P.babensis VN05A0561T (AB514449)
P.artemisiae YIM 63587 T (GU227146)
P. bannaensis YIM63101T (FJ817375)
100
P.spinosispora LM 141T (AJ249206)
68*
P. spinosa JCM 3136T (AB547126)
58
P.acaciae GMKU095T (EU921261)
P. eucalypti EUM 374T (FJ805426)
54
P.compacta IMSNU 20111T (AJ252825)
100*
75
62*
P.autotrophica IMSNU 20050T(AJ252824)
P.kongjuensis LM 157T (AJ252833)
87
P.ammonioxydans AS 4.1877T (AY500143)
P.endophyticus YIM 56035T (DQ887489)
P.parietis 04-St-002T (FM863703)
95
P. tropica YIM 61452T GQ906587
89
72
P. nitrificans IFAM 379T (X55609)
P.antarctica DVS 5a1T (AJ576010)
71*
P. carboxydiovorans SWP-2006T (EF114314)
99
98*
P.alni IMSNU 20049T (AJ252823)
P.xishanensis YIM 63638 T (FJ817397)
85
P.oroxyli D10T (DQ343154)
90
P.ailaonensis YIM 45505 T (DQ344632)
77*
P.yuanmoensis YIM 75926T(JN656709)
P.halophobica IMSNU 21327T (AJ252827)
93*
P.tetrahydrofuranoxydans k1T (AJ249200)
100*
79
P.hydrocarbonoxydans IMSNU 22140T (AJ252826)
P.sulfidoxydans DSM 44248T (Y08537)
P.dioxanivorans CB1190T (AY340622)
100*
P.benzenivorans B5T (AJ556156)
P.thermophila IMSNU 20112T (AJ252830)
P. khuvsgulensis MN08-A0297 T (AB521672)
96*
100*
P. rhizophila YIM 67013T (GU322368)
Kutzneriakofuensis NRRL B-24061T (AF114801)
0.01
Fig. 1 Neighbour-joining phylogenetic tree showing the phylogenetic relationships of strain YIM 75926T and other closely
related Pseudonocardia species based on 16S rRNA gene
sequences. Asterisks indicate branches that were also recovered
using the maximum-parsimony and maximum-likelihood
methods. Bootstrap values (expressed as percentages of 1,000
replications) of above 50% are shown at branch points. Bar
0.005 substitutions per nucleotide position. Kutzneria kofuensis
NRRL B-24061T (AF114801) was used as an outgroup. 0.005
substitutions per nucleotide position
123
758
values of 92, 77 and 67%, respectively (Supplementary Figs. S4–6). DNA–DNA hybridization were
carried out between strain YIM 75926T and the most
phylogenetically closely related type strain P. halophobica DSM 43089T by applying the fluorometric
micro-well method at the optimal hybridization temperature (47°C). DNA–DNA relatedness values
between YIM 75926T and P. halophobica DSM
43089T was 43.5 ± 1.6%, which was significantly
lower than the threshold value (70%) for the recognition of genomic species (Stackebrandt and Goebel
1994), thus we suggest that strain YIM 75926T should
be considered as a representative of a different
genomic species to the genus Pseudonocardia.
Phylogenetic analysis, morphological and chemotaxonomic characteristics all supported strain YIM
75926T to be the member of the genus Pseudonocardia. Meanwhile, the differences in biochemical characteristics, 16S rRNA gene sequences, DNA G?C
content, DNA–DNA relatedness and fatty acids composition can be used to distinguish strain YIM 75926T
from any described species of the genus Pseudonocardia. Therefore, on the basis of these results, strain
YIM 75926T was recognized as a novel species of
genus Pseudonocardia, for which the name Pseudonocardia yuanmoensis sp. nov., is proposed.
Description of Pseudonocardia yuanmoensis sp.
nov.
Pseudonocardia
yuanmoensis
(yuan.mo.en’sis.
N.L.(fem). adj. yuanmoensis pertaining to Yuanmo
county of Yunnan Province of south-west China, the
source of the type strain isolated).
Cells are Gram-positive and aerobic, forms extensively branched substrate and aerial mycelia. Substrate
and aerial mycelia all fragments into rod-shaped
elements. Substrate mycelia are yellow, orange-yellow
and yellow-brown or white aerial mycelia are formed on
media tested. No diffusible pigment is produced.
Growth occurs between 10 and 45°C (optimum
28–37°C), pH 6.0–8.0 (optimum pH 7.0), and 0–5%
(w/v) NaCl (optimum 0–3%) on ISP 2 agar. Uses
L-arabinose, cellobiose, D-fructose, lactose, maltose,
D-mannitol, L-rhamnose, sodium acetate, sorbitol,
D-sucrose, trisodium citrate, D-xylitol and D-xylose as
the sole carbon and energy sources. Negative for use
D-glucose, glycerol, D-mannose, D-trehalose and succinic acid as the sole carbon and energy sources. Acid is
123
Antonie van Leeuwenhoek (2012) 101:753–760
produced from L-arabinose, D-fructose, D-sucrose and
D-xylose. Utilizes adenine, L-alanine, cystine, L-arginine, L-cysteine, L-asparagine, glycine, hypoxanthine,
L-ornithine, L-phenylalanine, L-proline, L-serine, L-threonine, L-tyrosine and xanthine as the sole nitrogen
sources, but does not utilize L-glutamic acid, L-histidine
and L-lysine as the sole nitrogen sources. Positive for
urease and catalase activities, milk peptonization and
coagulation, starch hydrolysis and nitrate reduction, but
negative for gelatin liquefaction, cellulose hydrolysis,
melanin formation and H2S production. The cell wall
belongs to type IV. The whole cell sugar profile consists
of mannose, glucose, galactose and arabinose. The
diagnostic diamino acid of the peptidoglycan is mesoDAP. The glycan moiety of the murein is acetylated.
Mycolic acids are absent. Predominant menaquinone is
MK-8(H4). The major fatty acids are iso-C16:0
(37.16%) and C16:0 (12.43%). The phospholipids compose of diphosphatidylglycerol, phosphatidylglycerol,
phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylcholine, phosphatidylinositol,
phosphatidylinositol mannoside, two unknown glycolipid and two unknown polar lipids. The DNA G?C
content is 70.6 mol%.
The type strain YIM 75926T(=CCTCC AA
2011017T = JCM 18055T) was isolated from a soil
forest collected from the county of Yuanmo, Yunnan
Province, south-west China.
Acknowledgments This research was supported by Science
and Technology Innovation Talents Program in Universities of
Henan Province (HASTIT, No. 2010HASTIT020) and Key
Technologies R & D Program of Henan Province of China
(112102310335).
References
Ara I, Tsetseg B, Daram D, Suto M, Ando K (2011) Pseudonocardia mongoliensis sp. nov., and Pseudonocardia
khuvsgulensis sp. nov., isolated from Mongolian soil. Int J
Syst Evol Microbiol 61:747–756
Chen HH, Qin S, Li J, Zhang YQ, Xu LH, Jiang CL, Kim CJ, Li
WJ (2009) Pseudonocardia endophytica sp. nov., isolated
from the pharmaceutical plant Lobelia clavata. Int J Syst
Evol Microbiol 59:559–563
Christensen H, Angen O, Mutters R, Olsen JE, Bisgaard M
(2000) DNA–DNA hybridization determined in microwells using covalent attachment of DNA. Int J Syst Evol
Microbiol 50:1095–1102
Collins MD, Jones D (1980) Lipids in the classification and
identification of coryneform bacteria containing
Antonie van Leeuwenhoek (2012) 101:753–760
peptidoglycan based on 2,4-diaminobutyric acid. J Appl
Bacteriol 48:459–470
Collins MD, Pirouz T, Goodfellow M, Minnikin DE (1977)
Distribution of menaquinones in actinomycetes and corynebacteria. J Gen Microbiol 100:221–230
Dong XZ, Cai MY (2001) Manual of systematics and identification of general bacteria. Science Press, Beijing
Duangmal K, Thamchaipenet A, Matsumoto A, Takahashi Y
(2009) Pseudonocardia acaciae sp. nov., isolated from
roots of Acacia auriculiformis A. Cunn. ex Benth. Int J Syst
Evol Microbiol 59:1487–1491
Ezaki T, Hashimoto Y, Yabuuchi E (1989) Fluorometric
deoxyribonucleic acid-deoxyribonucleic acid hybridization in microdilution wells as an alternative to membrane
filter hybridization in which radioisotopes are used to
determine genetic relatedness among bacterial strains. Int J
Syst Bacteriol 39:224–229
Felsenstein J (1981) Evolutionary trees from DNA sequences: a
maximum likelihood approach. J Mol Evol 17:368–376
Felsenstein J (1985) Confidence limits on phylogenies: an
approach using the bootstrap. Evolution 39:783–789
Fitch WM (1971) Toward defining the course of evolution:
minimum change for a specific tree topology. Syst Zool
20:406–416
Gordon RE, Barnett DA, Handerhan JE, Pang CHN (1974)
Nocardia coeliaca, Nocardia autotrophica, and the nocardin strain. Int J Syst Bacteriol 24:54–63
Gram HC (1884) Über die isolierte Färbung der Schizomyceten
in Schnitt- und Trockenpräparaten. Fortschritte der Medizin 2:185–189
Gu Q, Luo H, Zheng W, Liu Z, Huang Y (2006) Pseudonocardia
oroxyli sp. nov., a novel actinomycete isolated from surface-sterilized Oroxylum indicum root. Int J Syst Evol
Microbiol 56:2193–2197
Guindon S, Gascuel O (2003) A simple, fast, and accurate
algorithm to estimate large phylogenies by maximum
likelihood. Syst Biol 52(5):696–704
Hasegawa T, Takizawa M, Tanida S (1983) A rapid analysis for
chemical grouping of aerobic actinomycetes. J Gen
Microbiol 29:319–322
He L, Li W, Huang Y, Wang LM, Liu ZH, Lanoot BJ, Vancanneyt M, Swings J (2005) Streptomyces jietaisiensis sp.
nov., isolated from soil in northern China. Int J Syst Evol
Microbiol 55:1939–1944
Henssen A (1957) Beiträge zur Morphologie und Systematic der
thermophilen Actinomyceten. Arch Mikrobiol 26:377–414
Hu H, Lim B, Naohiro G, Koich FJ (2001) Analytical precision
and repeatability of respiratory quinones for quantitative
study of microbial community structure in environmental
samples. J Microbiol Methods 47:17–24
Huang Y, Wang L, Lu Z, Hong L, Liu Z, Tan GYA, Goodfellow
M (2002) Proposal to combine the genera Actinobispora
and Pseudonocardia in an emended genus Pseudonocardia, and description of Pseudonocardia zijingensis sp. nov.
Int J Syst Evol Microbiol 52:977–982
Kaewkla O, Franco CMM (2010) Pseudonocardia adelaidensis
sp. nov., an endophytic actinobacterium isolated from the
surface-sterilized stem of a Grey Box tree. Int J Syst Evol
Microbiol 60:2818–2822
Kaewkla O, Franco CMM (2011) Pseudonocardia eucalypti sp.
nov., an endophytic actinobacterium with a unique knobby
759
spore surface, isolated from the surface-sterilized root of a
native Australian eucalyptus tree. Int J Syst Evol Microbiol
61:742–746
Kelly KL (1964) Inter-Society Color Council–National Bureau
of Standards color name charts illustrated with centroid
colors. US Government Printing Office, Washington
Kimura M (1983) The neutral theory of molecular evolution.
Cambridge University Press, Cambridge
Lechevalier MP, Lechevalier H (1970) Chemical composition
as a criterion in the classification of aerobic actinomycetes.
Int J Syst Bacteriol 20:435–443
Leifson E (1960) Atlas of bacterial flagellation. Academic Press,
London
Li WJ, Xu P, Schumann P, Zhang YQ, Pukall R, Xu LH,
Stackebrandt E, Jiang CL (2007) Georgenia ruanii sp.
nov., a novel actinobacterium isolated from forest soil in
Yunnan (China) and emended description of the genus
Georgenia. Int J Syst Evol Microbiol 57:1424–1428
Li J, Zhao GZ, Huang HY, Zhu WY, Lee JC, Kim CJ, Xu LH,
Zhang LX, Li WJ (2010) Pseudonocardia rhizophila sp.
nov., a novel actinomycete isolated from a rhizosphere soil.
Antonie van Leeuwenhoek 98:77–83
Liu ZP, Wu JF, Liu ZH, Liu SJ (2006) Pseudonocardia ammonioxydans sp. nov., isolated from coastal sediment. Int J
Syst Evol Microbiol 56:555–558
Mahendra S, Cohen LA (2005) Pseudonocardia dioxanivorans
sp. nov., a novel actinomycete that grows on 1,4-dioxane.
Int J Syst Evol Microbiol 55:593–598
McVeigh HP, Munro J, Embley TM (1994) The phylogenetic
position of Pseudoamycolata halophobica (Akimov et al.
1989) and a proposal to reclassify it as Pseudonocardia
halophobica. Int J Syst Bacteriol 44:300–302
Mesbah M, Premachandran U, Whitman WB (1989) Precise
measurement of the G ? C content of deoxyribonucleic
acid by high-performance liquid chromatography. Int J
Syst Bacteriol 39:159–167
Minnikin DE, Collins MD, Goodfellow M (1979) Fatty acid and
polar lipid composition in the classification of Cellulomonas, Oerskovia and related taxa. J Appl Bacteriol
47:87–95
Minnikin DE, Hutchinson IG, Caldicott AB, Goodfellow M
(1980) Thin layer chromatography of methanolysates of
mycolic acid-containing bacteria. J Chromatogr
188:221–233
Park SW, Park ST, Lee JE, Kim YM (2008) Pseudonocardia
carboxydivorans sp. nov., a carbon monoxide-oxidizing
actinomycete, and an emended description of the genus
Pseudonocardia. Int J Syst Evol Microbiol 58:2475–2478
Qin S, Shu YY, Zhang YQ, Wang HB, Jiang CL, Xu LH, Li WJ
(2008) Pseudonocardia ailaonensis sp. nov., isolated from
soil in China. Int J Syst Evol Microbiol 58:2086–2089
Qin S, Zhu WY, Jiang JH, Klenk HP, Li J, Zhao GZ, Xu LH, Li
WJ (2010) Pseudonocardia tropica sp. nov., a novel
endophytic actinomycete isolated from the stem of Maytenus austroyunnanensis. Int J Syst Evol Microbiol
60:2524–2528
Qin S, Xing K, Fei SM, Lin Q, Chen XM, Cao CL, Sun Y, Wang
Y, Li WJ, Jiang JH (2011) Pseudonocardia sichuanensis
sp. nov., a novel endophytic actinomycete isolated from the
root of Jatropha curcas L. Antonie van Leeuwenhoek
99:395–401
123
760
Reichert K, Lipski A, Pradella S, Stackebrandt E, Altendorf K
(1998) Pseudonocardia asacccharolytica sp. nov. and
Pseudonocardia sulfidoxydans sp. nov., two new dimethyl
disulfide-degrading actinomycetes and emended description of the genus Pseudonocardia. Int J Syst Bacteriol
48:441–449
Saitou N, Nei M (1987) The neighbor-joining method: a new
method for reconstructing phylogenetic tree. Mol Biol Evol
4:406–425
Sakiyama Y, Thao NKN, Vinh HV, Giang NM, Miyadoh S, Hop
DV, Ando K (2010) Pseudonocardia babensis sp. nov.,
isolated from plant litter in Vietnam. Int J Syst Evol
Microbiol 60:2336–2340
Shirling EB, Gottlieb D (1966) Methods for characterization of
Streptomyces species. Int J Syst Bacteriol 16:313–340
Smibert RM, Krieg NR (1994) Phenotypic characterization. In:
Gerhardt P, Murray RGE, Wood WA, Krieg NR (eds)
Methods for general and molecular bacteriology. American
Society for Microbiology, Washington, pp 607–654
Stackebrandt E, Goebel BM (1994) Taxonomic note: a place for
DNA–DNA reassociation and 16S rRNA sequence analysis in the present species definition in bacteriology. Int J
Syst Bacteriol 44:846–849
Tamaoka J, Katayama-Fujimura Y, Kuraishi H (1983) Analysis
of bacterial menaquinone mixtures by high performance
liquid chromatography. J Appl Bacteriol 300:31–36
Tamura K, Peterson D, Peterson N, Stecher G, Nei M, Kumar S
(2011) MEGA5: molecular evolutionary genetics analysis
using maximum likelihood, evolutionary distance, and
123
Antonie van Leeuwenhoek (2012) 101:753–760
maximum parsimony methods. Mol Biol Evol
28:2731–2739
Tang SK, Wang Y, Chen Y, Lou K, Cao LL, Xu LH, Li WJ
(2009) Zhihengliuella alba sp. nov., and emended
description of the genus Zhihengliuella. Int J Syst Evol
Microbiol 59:2025–2032
Thompson JD, Gibson TJ, Plewniak F, Jeanmougin F, Higgins
DG (1997) The Clustal_X windows interface: flexible
strategies for multiple sequence alignment aided by quality
analysis tools. Nucleic Acids Res 25:4876–4882
Uchida K, Aida K (1984) An improved method for the glycolate
test for simple identification of acyl type of bacterial cell
walls. J Gen Appl Microbiol 30:131–134
Warwick S, Bowen T, McVeigh HP, Embley TM (1994) A
phylogenetic analysis of the family Pseudonocardiaceae
and the genera Actinokineospora and Saccharothrix with
16S rRNA sequences and a proposal to combine the genera
Amycolata and Pseudonocardia in an emended genus
Pseudonocardia. Int J Syst Bacteriol 44:293–299
Zhao GZ, Li J, Huang HY, Zhu WY, Park DJ, Kim CJ, Xu LH,
Li WJ (2011a) Pseudonocardia kunmingensis sp. nov., a
novel actinobacterium isolated from surface-sterilized
roots of Artemisia annua L. Int J Syst Evol Microbiol
61:2292–2297
Zhao GZ, Li J, Zhu WY, Li XP, Tian SZ, Zhao LX, Xu LH, Li
WJ (2011b) Pseudonocardia bannaensis sp. nov., a novel
actinomycete isolated from the surface-sterilized roots of
Artemisia annua L. Antonie van Leeuwenhoek 100:35–42