Arthrobactersiccitolerans sp. nov., a highly desiccation-tolerant, xeroprotectant-producing strain isolated from dry soil.

2013·0.19 MB·English

by  SantaCruz-Calvo, L.

InternationalJournalofSystematicandEvolutionaryMicrobiology(2013),63,4174–4180 DOI10.1099/ijs.0.052902-0
Arthrobacter siccitolerans sp. nov., a highly
desiccation-tolerant, xeroprotectant-producing
strain isolated from dry soil
L. SantaCruz-Calvo, J. Gonza´lez-Lo´pez and M. Manzanera
Correspondence Institutefor Water Research, and Department of Microbiology, University of Granada,Granada,
M.Manzanera Spain
[email protected]
Anovel desiccation-tolerant, xeroprotectant-producing bacterium, designatedstrain4J27T,was
isolatedfrom aNerium oleander rhizosphere subjected toseasonal droughtinGranada, Spain.
Phylogeneticanalysisbasedon16SrRNAgenesequencingplacedtheisolatewithinthegenus
Arthrobacter,its closestrelative beingArthrobacter phenanthrenivorans Shep3 DSM 18606T,
withwhich it showed 99.23% 16SrRNA gene sequencesimilarity. DNA–DNAhybridization
measurements showed less than25%relatedness betweenstrain 4J27TandArthrobacter
phenanthrenivorans DSM18606T.The DNA basecomposition of strain4J27Twas65.3 mol%.
The main fatty acidswere anteiso C ,anteisoC ,C andisoC andthe major
15:0 17:0 16:0 16:0
menaquinonewasMK-9(H2).ThepeptidoglycantypewasA3awithanL-Lys–L-Ser–L-Thr–L-Ala
interpeptide bridge. The bacterium testedpositive forcatalase activity andnegative foroxidase
activity.Phylogenetic, chemotaxonomic andphenotypic analysesindicated thatthe desiccation-
tolerantstrain4J27TrepresentsanovelspecieswithinthegenusArthrobacter,forwhichthename
Arthrobactersiccitoleransisproposed.Thetypestrainis4J27T(5CECT8257T5LMG27359T).
The genus Arthrobacter, first defined by Conn & Dimmick cellsagainstdifferentkindsofstress,suchasfreezing,drying
(1947), belongs to the class Actinobacteria and includes andheating(Brown,1976;Yanceyetal.,1982;Knappetal.,
Gram-stain-positive coryneform bacteria with aerobic 1999;Manzaneraetal.2002,Narva´ez-Reinaldoetal.,2010,
metabolism and little or no acid production from glucose. Julcaetal.,2012).Ourgrouphaspreviouslyreportedanew
Species of the genus Arthrobacter contain lysine in the method for the isolation of desiccation-tolerant micro-
peptidoglycanandhaveaDNAG+Ccontentrangingfrom organisms from dry soil using organic solvents as selective
59 mol%to66 mol%(Keddieetal.,1986;Jones&Keddie, agents (Manzanera et al., 2004a; Narva´ez-Reinaldo et al.,
1992). These bacteria typically take the shape of rods in 2010). Strain 4J27T displayed remarkably high tolerance to
younger cultures and cocci in older cultures (Keddie et al., desiccation and produced excellent xeroprotectants for the
1986), depending on their growth rate and nutritional dry stabilization of proteins (lipase enzymes) and whole
conditions(Germida&Cassida,1980).Thetransitiontothis prokaryotic cells (Escherichia coli MC4100) compared with
coccoid-like state has been shown to require manganese those when trehalose was used (Manzanera et al., 2004b;
(Germida&Cassida,1980).Thesmallcoccoid-likestatehas Narva´ez-Reinaldo et al., 2010). Among the 10 different
been described as being the most stable form. Due to their xeroprotectants tested, the best results were observed with
pleomorphic and heterogeneous appearance, strains of S4J27-D(composedoftrehalose,glutamineandglucose),a
species of the genus Arthrobacter were originally grouped synthetic mixture derived from strain 4J27T (Narva´ez-
Reinaldoetal.,2010).
withtheCorynebacteria(Keddieetal.,1986).
Here we describe the morphological, biochemical and
In response to changing extracellular osmolarity such as
phylogenetic characteristics of this desiccation-tolerant
desiccation or increased salinity some micro-organisms
strain(4J27T),isolatedfromdrysoilandwitharemarkable
accumulate small organic compounds (Brown, 1976;
potentialforthedrystabilizationofsomebiomaterials.On
Arakawa& Timasheff, 1982). These compatible solutes act
thebasisofthephylogeneticanalysisofthe16SrRNAgene
asprotectants, whichunderlaboratory conditions can also
sequence together with physiological, chemotaxonomic
stabilize enzymes, DNA, cell membranes and even whole
and DNA–DNA hybridization analyses we demonstrate
that strain 4J27T represents a novel species of the genus
Abbreviation: DSMZ, Deutsche Sammlung von Mikroorganismen und
Arthrobacter.
Zellkulturen.
Asupplementaryfigureandasupplementarytableareavailablewiththe Strain4J27Twasgrownat30 uC(±3 uC)intryptone soya
onlineversionofthispaper. agar(TSA)platesandintryptonesoyabroth(TSB)orM9
4174 052902G2013IUMS PrintedinGreatBritain
Arthrobactersiccitoleranssp.nov.
minimal medium (M6030; Sigma). Arthrobacter phenan- Arthrobacter phenanthrenivorans DSM 18606T. Phylo-
threnivorans DSM 18606T was included in the study as genetic analysis showed that strain 4J27T clearly belongs
reference. to the genus Arthrobacter, and its closest relative was
Arthrobacter phenanthrenivorans DSM 18606T.
Strain 4J27T, the object of this study, had already been
assigned to the genus Arthrobacter by partial analysis of its DNA–DNA hybridization was carried out at the Deutsche
16S rRNA gene sequence (GenBank accession number Sammlung von Mikroorganismen und Zellkulturen
GU815139; Narva´ez-Reinaldo et al., 2010), which was (DSMZ; Braunschweig, Germany). Cells of Artrhobacter
compared with those in the EzTaxon-e server (http:// phenanthrenivorans DSM 18606T and strain 4J27T were
eztaxon-e.ezbiocloud.net/, Kim et al., 2012). The nearly disrupted by using a French pressure cell (Thermo
complete sequence of the 16S rRNA gene of strain 4J27T Spectronic)andtheDNAofeachstraininthecrudelysate
(approximately 1500 bp) was aligned with the sequences were purified by chromatography on hydroxyapatite as
of closely related species of the genus Arthrobacter by described by Cashion et al. (1977). DNA–DNA hybridiza-
using the CLUSTAL X 2 program (Larkin et al., 2007). A tion was conducted as described by De Ley et al. (1970)
phylogenetictreewasinferredusingtheneighbour-joining with the modifications described by Huss et al. (1983)
(Saitou & Nei, 1987) and maximum-likelihood (Guindon using a model Cary 100 Bio UV/VIS-spectrophotometer
& Gascuel, 2003) methods with the MEGA 5.0 software equipped with a Peltier-thermostat-regulated 666 multi-
package(Tamuraetal.,2011).Bootstrapanalysiswasbased
cell charger and a temperature controller with in situ
on 1000 resamplings (Felsenstein, 1985). The distances
temperature probe (Varian). DNA–DNA hybridization of
were calculated according to Kimura’s two-parameter strain 4J27T with Arthrobacter phenanthrenivorans DSM
model (Kimura, 1980). The resulting neighbour-joining 18606T resulted in a DNA–DNA relatedness value of
tree obtained with Kimura’s two-parameter model is
22.3%(22.1%),thevalueinparenthesesbeingtheresultof
shown in Fig. 1 and the maximum-likelihood tree is
measurements in duplicate. On the basis of DNA–DNA
shown in Fig. S1, available in IJSEM Online. reciprocalhybridization,strain4J27Tdidnotbelongtothe
The sequence corresponding to the 16S rRNA gene of species Arthrobacter phenanthrenivorans according to the
strain 4J27T showed 99.23% similarity to that of recommendationsofathresholdvalueof70%DNA–DNA
Arthrobacter chlorophenolicus A6T (CP001341)
73 Arthrobacter equi IMMIB L-1606T (FN673551)
0.01 Arthrobacter defluvii 4C1-aT (AM409361)
100 Arthrobacter niigatensis LC4T (AB248526)
50
Arthrobacter phenanthrenivorans Sphe3T (CP002379)
59 Arthrobacter siccitolerans 4J27T (GU815139)
Arthrobacter polychromogenes DSM 20136T (X80741)
99 Arthrobacter oxydans DSM 20119T (X83408)
Arthrobacter scleromae YH-2001T (AF330692)
63 Arthrobacter roseus CMS 90rT (AJ278870)
Arthrobacter sulfonivorans ALLT AF23509 (AF235091)
Arthrobacter russicus GTC 863T (AB071950)
Arthrobacter livingstonensis LI2T (GQ406811)
98 Arthrobacter stackebrandtii CCM 2783T (AJ640198)
95 Arthrobacter cryoconiti Cr6-08T (GU784867)
Arthrobacter alpinus S6-3T (GQ227413)
Arthrobacter ramosus CCM 1646T (AM039435)
88 Arthrobacter nitroguajacolicus G2-1T (AJ512504)
52 97 Arthrobacter aurescens DSM 20116T (X83405)
Arthrobacter ilicis DSM 20138T (X83407)
Arthrobacter ureafaciens DSM 20126T (X80744)
74 Arthrobacter nicotinovorans DSM 420T (X80743)
99 Arthrobacter histidinolovorans DSM 20115T (X83406)
Arthrobacter agilis DSM 20550T (X80748)
Arthrobacter monumenti LMG 19502T (AJ315070)
Streptomyces albus AS 4.164T (AJ621602)
Fig.1.Neighbour-joiningphylogenetictreebasedon16SrRNAsequencecomparisonsofstrain4J27Tandits24closestrelatives.
Streptomyces albus AS 4.164T was used as the outgroup. The numbers at bifurcations indicate how many times each species
coincidedinthispositionaspercentagesandonlyvalues.50%areshown.Bar,0.01changespernucleotideposition.
http://ijs.sgmjournals.org 4175
L.SantaCruz-Calvo,J.Gonza´lez-Lo´pezandM.Manzanera
relatedness for the definition of bacterial species (Wayne to lowest, anteiso-C , 41.20%; anteiso-C , 30.86%;
15:0 17:0
et al., 1987). Therefore strain 4J27T probably represents a C , 10.21%; iso-C , 6.61%; iso-C , 4.40%; C ,
16:0 16:0 15:0 18:0
novel species of the genus Arthrobacter. 2.38%; iso-C , 1.79%; iso-C , 0.83%; C , 0.75%;
17:0 14:0 14:0
anteiso-C , 0.61% and iso-C , 0.36%. The fatty-acid
The G+C (mol%) content of the genomic DNA of strain 19:0 18:0
compositionofstrain4J27Twasconsistentwiththatofthe
4J27TwasanalysedattheDSMZ.ThedGanddTratiowas
genus Arthrobacter, with branched-chain fatty acid, ante-
calculatedaccordingtothemethodofMesbahetal.(1989).
sio-pentadecanoic acid (anteiso-C ) predominating
Species of the genus Arthrobacter have previously been 15:0
(Westerberg et al., 2000).
described as Gram-stain-positive actinobacteria with high
GC content (Keddie et al., 1986; Jones & Keddie ,1992), RespiratoryquinoneswereanalysedasdescribedbyTindall
which typically have a DNA G+C content in the range of (1990a; b), using TLC and UV mass spectroscopy, and
59–66 mol%(Keddieetal.,1986).TheDNAG+Ccontent found menaquinone to be the sole quinone component.
ofstrain4J27Twas65.3 mol%,whichwaswithintherange Analyses of the electron-transport system (isoprenoid
shown by all members of the genus Arthrobacter and quinones) for strain 4J27T resulted in detection of MK9
consideredtohaveahighGCcontent(Keddieetal.,1986). (II-H ) 68%; MK9 21% and MK8 (II-H ) 11%.
2 2
Chemotaxonomic analyses were carried out by the To analyse the whole cell sugars of strain 4J27T, cells were
Identification Service of the DSMZ. Peptidoglycans were hydrolysedin0.5 MH SO for2 hat100 uC.Sulfuricacid
2 4
isolated from strain 4J27T and their structures analysed was removed by 20% N,N-dioctylmethylamine in chlo-
(Schleifer & Kandler, 1972). After derivatization according roform according to the method of Whiton et al. (1985).
tothemethodofMacKenzie(1987)theapproximatemolar Sugars in the hydrolysate were analysed by TLC on
amino-acid ratio was determined by gas chromatography. cellulose plates according to the methods of Staneck &
Freeaminogroupswithinthepeptidoglycanweredetected Roberts(1974).Thewhole-cellsugarsoftheisolatedstrain
by labelling with 1-fluoro-2,4-dinitrobenzene (Schleifer, were galactose, glucose, mannose, ribose and rhamnose.
1985). The peptidoglycan of strain 4J27T was composed
Mobility was tested by stab-inoculating mannitol-mobility
of Ala, Ser, Thr, Glu and Lys at a molar ratio of
semi-solid agar (413782; Ultimed). This semi-solid agar
2.8:1.2:1.0:1.0:1.5. Two-dimensional TLC of the partial
mediumenabledustoanalysethenitratereductaseactivity
hydrolysate (4 M HCl, 100 uC, 45 min) of the peptidogly-
(capacity to reduce nitrate to nitrite) and catabolism of
can revealed the presence of the peptides L-Ala–D-Glu,
mannitol by using Griess–Ilosvay A and B reagents.
L-Lys–D-Ala, L-Lys–L-Ser, L-Lys–L-Ser–L-Thr, D-Ala–L-Lys–
Oxidaseactivitywasdeterminedusing1%w/vN,N,N9,N9-
L-Ser–L-Thr, L-Ser–L-Thr and L-Ala–D-Ala. On the basis of
tetramethyl-p-phenylenediamine and catalase activity was
these results it was concluded that strain 4J27T contains a
determined by the production of bubbles from 3% v/v.
type A3a peptidoglycan (Schleifer & Kandler, 1972) with
H O . Cells of strain 4J27T were identified as catalase-
an L-Lys–L-Ser–L-Thr–L-Ala interpeptide bridge (A11.23 2 2
positive, oxidase-negative, nitrate reductase-negative and
DSMZ-Catalogue of strains, 7th edition, 2001), which is
mannitol-positive. Arthrobacter phenanthrenivorans DSM
found in the more closely related members of the genus
18606T.
Arthrobacter, according to the neighbour-joining phylo-
genetic tree, such as Arthrobacter chlorophenolicus, Arthro- To characterize the growth of strain 4J27T at different
bacter oxydans, Arthrobacter polychromogenes, Arthrobacter temperatures, pH values and salinities, cultures were
sulfonivorans, Arthrobacter equi, Arthrobacter niigatensis, incubatedat150 r.p.m.inLuria–Bertani(LB)richmedium
Arthrobacterphenanthrenivorans,Arthrobacterdefluvii,Arthro- (L3152; Sigma). Cell growth was monitored at different
bacterroseusandArthrobacterscleromae(Borodinaetal.,2002; temperatures (5, 10, 15, 20, 25, 30, 35, 40, 45 and 50 uC),
Kodama et al., 1992; Westerberg et al., 2000; Reddy et al., pH(3,5,7,9,12and13)andNaClconcentrations(0,0.2,
2002; Huang et al., 2005; Kim et al., 2008; Ding et al., 2009; 0.4, 0.6, 0.8, 1 and 1.2 M) by measuring the OD in
600
Yassinetal.,2011).StrainscontainingatypeA3apeptidogly- triplicate at 0, 12 and 24 h using a UV-160A spectropho-
canmakeuparatheruniformgroup,althoughtheydoshowa tometer(Shimadzu).Strain4J27Tgrewbestat30 uCinLB
considerablenumberofdifferenttypesofinterpeptidebridge. medium.Itwasabletogrowat37 uCand15 uCbutnotat
MostofthesestrainsbelongtothegenusArthrobacterandare 40 uC or 10 uC. The pH range for growth was between 5
distinguished by strictly aerobic growth and a complete life and9withoptimum growth atpH 7.Strain4J27Tgrewin
cycle(Conn&Dimmick,1947;Schleifer&Kandler,1972). NaClconcentrationsrangingfrom0to0.8 Mbutgrewbest
at0.2 M.Thisdifferedclearlyfromthemostcloselyrelated
Fatty-acidmethylesterswereobtainedfrom40 mgcellsof
species, Arthrobacter phenanthrenivorans DSM 18606T,
strain 4J27T scraped from Petri dishes by saponification,
which was able to grow at 4 uC but not at pH 5.
methylation and extraction using the methods of Miller
(1982)andKuykendalletal.,(1988)withminormodifica- The following API kits were used for testing, API Coryne,
tions. The fatty-acid methyl-ester mixtures were separated API 20NE and API 20E (bioMe´rieux,). Each test was
using the Sherlock Microbial Identification System (MIS) interpreted according to the manufacturer’s instructions.
(MIDI, Microbial ID). The main cellular fatty acids of the Biolog tests were performed to investigate which com-
highly desiccation-tolerant strain 4J27T were, from highest poundsthestrainsinquestion coulduseforrespiration. A
4176 InternationalJournalofSystematicandEvolutionaryMicrobiology63
Arthrobactersiccitoleranssp.nov.
GP2 MicroPlate (Cat. No 1014; Biolog), containing 95 tolerance, in contrast to the novel strain, which is
differentcarboncompounds,wasusedtotestforsubstrate considered to be a desiccation-tolerant strain.
oxidation. The chemistry of these plates is based on
On thebasisofphylogenetic analysis ofits16SrRNA gene
tetrazolium reduction, in response to metabolic processes
sequence, together with physiological, chemotaxonomic
suchasfermentationandoxidation.Tetrazoliumreduction
and DNA–DNA hybridization analyses, strain 4J27T is
produced formazan in a variety of colours from dark blue
considered to represent a novel species of the genus
to deep red to orange, depending upon the original
Arthrobacter, for which the name Arthrobacter siccitolerans
tetrazolium salt used as the substrate for the reaction.
is proposed.
MicroPlates were inoculated and interpreted according to
the manufacturer’s instructions. The results were recorded
DescriptionofArthrobactersiccitoleranssp.nov.
after 12 h based on A . Antibiotic susceptibility testing
585
was performed using the disc-diffusion method in which Arthrobactersiccitolerans(sic.ci.to9le.rans.L.adj.siccusdry,
the antibiotic diffuses away from the disc in two L. part. adj. tolerans tolerating; N.L. part. adj. siccitolerans
dimensions,formingaconcentrationgradientthatinhibits dry-tolerating).
the growth of bacteria and causes an inhibition zone
Cells are non-motile, non-spore-forming, Gram-positive,
(Piddock, 1990).The results were interpretedaccordingto
aerobic and rod-to-coccus-shaped. Colonies on TSA are
the criteria established for staphylococci in 1997 by the
convex, circular, cream, opaque and usually 1–2 mm in
National Committee for Clinical Laboratory Standards
diameterwithin2 daysat30 uC.Catalase-positive,oxidase-
(2000). At the concentrations assayed, the inhibition zone
negative and nitrate-reductase-negative (no capacity to
caused by streptomycin was 157 mm, rifampicin 347 mm,
reducenitratetonitrite).Growsattemperaturesfrom15to
chloramphenicol 340 mm, kanamycin 150 mm and tetra-
35 uC,pH 5–9andwith0–0.8 MNaClinLBmedium.The
cycline157 mmandthusitcouldbeconcludedthatstrain
peptidoglycantypeisA3a(Schleifer&Kandler,1972),with
4J27T was susceptible to all the antibiotics tested. The
phenotypic differences between strain 4J27T and closely anL-Lys–L-Ser–L-Thr–L-Alainterpeptidebridge.Themajor
cellular fatty acids are anteiso C , anteiso C , C
related species are summarized in Table 1 and the 15:0 17:0 16:0
and iso C . The major menaquinone is MK9-(II-H ).
physiological differences between strain 4J27T and its 16:0 2
The whole-cell sugars of the strain are galactose, glucose,
closest relative species Arthrobacter phenanthrenivorans
mannose, ribose and rhamnose. It reduces nitrites to
DSM 18606T are summarized in Table S1.
nitrogen. Indole and acetoin (Voges–Proskauer) produc-
The degree of tolerance to desiccation shown by strain tion are positive. According to the results from the API
4J27T was compared with that of the previously described CORYNE, API 20NE and API 20E strips, the following
desiccation-tolerant bacteria Acinetobacter calcoaceticus enzyme activities are detected: pirazinamidase, b-glucur-
PADD68 (Narva´ez-Reinaldo et al., 2010), the desiccation- onidase,b-galactosidase, a-glucosidase, b-glucosidase(aes-
sensitive strain Pseudomonas putida KT2440 (Manzanera culin), b-galactosidase (p-nitrophenyl-b-D-galactopyrano-
et al., 2002) and the closely related Arthrobacter sidase). Assimilation of glucose, arabinose, mannose,
phenanthrenivorans DSM 18606T. A colony of a pure mannitol, N-acetyl-glucosamine, maltose, potassium glu-
culturegrownfor48 hofeachstrain,containing107to109 conate, malate, trisodium citrate, inositol, sorbitol, rham-
cells, was suspended in 1 ml M9 minimal medium. nose, sucrose, melibiose, amygdalin and arabinose are
Aliquots (100 ml) were placed on sterile Petri dishes and positive. The following enzyme activities are not present:
dried under a current of sterileair for 24 h. The cells were b-galactosidase(o-nitro-phenyl-b-D-galactopyranoside),argi-
then suspended in 1 ml sterile saline buffer, and serial nine dihydrolase, lysine decarboxylase, ornithine decarbox-
dilutions of the cell suspension were plated on TSA plates ylase,urease,tryptophandesaminase,gelatinase,pyrrolidonyl
before and after drying. All such procedures were arylamidase,alkalinephosphataseandN-acetyl-b-glucosami-
conducted at room temperature. The survival rate was nidase.ProductionofH Sisnegativeanddoesnotusecitrate.
2
calculated in terms of c.f.u. ml21 after drying compared In the Biolog GP2 MicroPlates the following substrates
with c.f.u. ml21 before drying, expressed as a percentage. were used for respiration: dextrin, inulin, L-arabinose, N-
The assays were performed in triplicate accordingly to the acetyl-D-glucosamine, N-acetyl-D-mannosamine, D-arbutin,
protocolofManzaneraetal.,2002.Strain4J27Tshowedthe cellobiose, D-fructose, D-galactose, D-galacturonic acid, a-D-
highest values of desiccation tolerance (31.58%±6.9%), glucose, gentiobiose, lactamide, L-lactic acid, lactulose,
whichweresignificantlydifferentfromthoseofthepositive maltose, maltotriose, D-mannitol, D-mannose, melezitose,
control, Acinetobacter calcoaceticus PADD68 (3.23%± melibiose, 3-methyl glucose, a-methyl D-mannoside, palati-
0.2%) and more so from the closely related strain, nose, D-psicose, D-rafinose, L-rhamnose, D-ribose, salicin, D-
Arthrobacter phenanthrenivorans DSM 18606T (1.5%± sorbitol,sucrose, trehalose, turanose, xylitol, D-xylose, acetic
0.41%). As expected, the desiccation tolerance of the acid,a-hydroxybutyricacid,b-hydroxybutyricacid,p-hydro-
negative control, P. putida KT2440T, was below detectable xyphenylacetic acid, a-ketovaleric acid, L-malicacid, pyruvic
levels. Therefore the closely related species Arthrobacter acid, L-alaninamide, L-alanyl glycine, glycyl-L-glutamic
phenanthrenivorans DSM 18606T is considered to be acid, putrescine, glycerol, adenosine, 29-deoxy adenosine,
desiccation-sensitive, due to its low degree of desiccation inosine, thymidine, uridine, thymidine-59 monophosphate,
http://ijs.sgmjournals.org 4177
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7 S
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Table1.Differential characteristics betweenstrain 4J27Tandthetype strainsofthe mostclosely related species ofthe genusArthrobacter za´
le
z
Strains:1,4J27T;2,ArthrobacterphenanthrenivoransDSM18606T;3,ArthrobacterniigatensisIAM15382T;4,Arthrobacter.defluviiDSM18782T;5.ArthrobacterequiDSM23395T;6.Arthrobacter -L
o´
chlorophenolicusDSM12829T;7.ArthrobacterpolychromogenesDSM20136T;8.ArthrobacteroxydansDSM20119T;9.ArthrobacterscleromaeJCM12642T.Dataofthereferencespeciesweretaken pe
z
fromKallimanisetal.(2009),Dingetal.(2009),Kimetal.,(2008),Yassinetal.(2011),Westerbergetal.(2000),Schippers-Lammertseetal.(2009),Sguros(1955),Huangetal.(2005)andthe a
n
presentstudy.+,Positive;2,negative;ND,notdetermined;CFA,cellularfattyacid. d
M
.
M
Trait 1 2 3 4 5 6 7 8 9 a
n
z
Motility 2 2 2 2 2 + 2 2 2 an
e
Colonycolour Cream Yellowish Grey/yellow Creamywhite Cream Grey Blue-green Grey-white White ra
Temperature 15–35 4–37 5–40 5–37 10–35 3–37 10–37 20–35 15–37
pH 5–9 6.5–8.5 6–11 6–10 6–9 5–9 6–11 5–9 6–9
Reductionofnitrate + + + + 2 2 + + +
Hydrolysisofgelatin 2 2 + 2 + + + + +
Utilizationof:
Inte Maltose + 2 2 2 + + + + +
rn D-Ribose + 2 + + 2 + + + 2
a
tio Sucrose + + + 2 + + +* + +
na Trehalose + 2* + 2 + +* 2* 2* +
l
Jo D-Xylose + + 2 2 + 2 + + +
u
rn D-Alanine 2 +* ND 2 + + +* 2* ND
a
lo Glucose-1-phosphate + 2* + ND ND 2* 2* 2* ND
fS Inulin + 2* ND 2 ND 2 2* 2 +
y
s MajorCFAs antesio-C antesio-C iso- antesio-C antesio- anteiso-C anteiso-C anteiso-C anteiso-C anteiso-C anteiso-C
te 15:0 15:0 15:0 15:0 15:0 15:0 15:0 15:0 15:0
m antesio-C C C iso-C iso-C iso-C antesio-C anteiso-C iso-C
a 17:0 15:0 17:0 16:0 15:0 16:0 17:0 15:0 15:0
tic Majormenaquinone MK-9(H2) MK-8(H2) MK-9(H2) MK-9(H2) MK-9(H2) MK-9(H2) MK-9(H2) MK-9(H2) MK-8(H2)
an DNAG+Ccontent 65.3 67.5 70.8 63.5–64.4 67.0 65±1 62.9 63.1 64.7
d
(mol%)
E
v
o
lu
tio *Datatakenfromthepresentstudy.
n
a
ry
M
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io
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y
6
3
Arthrobactersiccitoleranssp.nov.
glucose-1-phosphate and D-L-a-glycerol phosphate. The Felsenstein,J.(1985).Confidencelimitsonphylogenies:anapproach
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methyl-D-glucoside, palatinose, propionic acid, L-alanine, L- algorithmtoestimatelargephylogeniesbymaximumlikelihood.Syst
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