ORIGINAL PAPER

Isolation and characterization of a novel violacein-likepigment producing psychrotrophic bacterial speciesJanthinobacterium svalbardensis sp. nov

Jerneja Ambrozic Avgustin •

Darja Zgur Bertok • Rok Kostanjsek •

Gorazd Avgustin

Received: 19 April 2012 / Accepted: 18 November 2012 / Published online: 29 November 2012

� Springer Science+Business Media Dordrecht 2012

Abstract A bacterial strain designated JA-1, related

to Janthinobacterium lividum, was isolated from glacier

ice samples from the island Spitsbergen in the Arctic.

The strain was tested for phenotypic traits and the most

prominent appeared to be the dark red brown to black

pigmentation different from the violet pigment of

Janthinobacterium, Chromobacterium and Iodobacter.

Phylogenetic analysis based on 16S rRNA gene

sequences and DNA–DNA hybridization tests showed

that strain JA-1 belongs to the genus Janthinobacterium

but represents a novel lineage distinct from the two

known species of this genus, J. lividum and Janthino-

bacterium agaricidamnosum. The DNA G ? C content

of strain JA-1 was determined to be 62.3 mol %. The

isolate is a psychrotrophic Gram negative bacterium,

rod-shaped with rounded ends, containing intracellular

inclusions and one polar flagellum. On the basis of the

presented results strain JA-1 is proposed as the type

strain of a novel species of the genus Janthinobacterium,

for which the name Janthinobacterium svalbardensis

sp. nov. is proposed (JA-1T = DSM 25734, ZIM B637).

Keywords Psychrotrophic bacterium �Janthinobacterium svalbardensis sp. nov.

Violacein-like pigment

Introduction

Bacterial strains identified as members of the beta-

proteobacterial genus Janthinobacterium have been

isolated from a number of different environments,

most commonly from soil and water ecosystems in

temperate and cold climates (Sneath 1984). Only two

species have been described so far, J. lividum (Eisen-

berg 1891; De Ley et al. 1978), a violet pigment

producing psychrotrophic, motile, Gram negative rod,

and J. agaricidamnosum, a non-pigmented soft rot

pathogen of a cultivated mushroom Agaricus bisporus

(Lincoln et al. 1999). ‘‘Atypical’’ strains of J. lividum

have also been described differing in several bio-

chemical test reactions (Moss and Ryall 1981; Logan

1989) or in the spectral properties of the produced

pigment (Schloss et al. 2010); however, no attempts

were made to taxonomically delineate these strains as

J. Ambrozic Avgustin � D. Zgur Bertok

Department of Biology, Chair for Molecular Genetics and

Biology of Microorganisms, University of Ljubljana,

Biotechnical Faculty, Vecna Pot 111, 1000 Ljubljana,

Slovenia

R. Kostanjsek

Department of Biology, Chair of Zoology, University of

Ljubljana, Biotechnical Faculty, Vecna Pot 111,

1000 Ljubljana, Slovenia

G. Avgustin (&)

Animal Science Department, Chair for Microbiology and

Microbial Biotechnology, University of Ljubljana,

Biotechnical Faculty, Groblje 3, 1230 Domzale, Slovenia

e-mail: gorazd.avgustin@bf.uni-lj.si

123

Antonie van Leeuwenhoek (2013) 103:763–769

DOI 10.1007/s10482-012-9858-0

novel taxa. In the present study we report the isolation

of a pigmented bacterial strain from glacier ice

samples from Spitsbergen Island at the Svalbard

archipelago on the very border of the Arctic Ocean.

Initial identification based on morphological charac-

teristics suggested that the strain is related to

J. lividum. Further investigation showed that the strain

represents a novel taxonomic lineage and a proposal of

a new species is thus described here.

Materials and methods

Isolation, bacterial strains and culture conditions

Glacier ice samples were taken from the Austre

Broggerbreen glacier near Ny Alesund, located on the

north-west coast of Spitsbergen island (78o 550 N, 11o

560 E) in the summer of 2003 by Nina Gunde-

Cimerman. The collecting strategy and transport to the

laboratory where the ice samples were molten were the

same as described elsewhere (Gunde-Cimerman et al.

2003). Samples of the molten glacier water were

subsequently plated onto different agar media (i.e.

Luria–Bertani (LB), Brain heart infusion (BHI) and

minimal medium (E) (Vogel and Bonner 1956) and

incubated at temperatures ranging from 2� C to 30 �C

for up to two months. After three week incubation at

18 �C, among others, dark reddish-brown to black

colonies appeared on minimal medium agar plates.

Colonies of this morphotype were re- streaked several

times on minimal and LB agar plates until pure

cultures of uniform colonies were obtained. One of

them was chosen for further investigations and

designated as strain JA-1. Other bacterial strains used

in this study were type strains of J. lividum (DSM

1522T) and J. agaricidamnosum (DSM 9628T). Addi-

tionally Chromobacterium violaceum CV026 mutant

strain was kindly provided by Dr. Paul Williams from

the University of Nothingham, UK. All strains were

grown in LB, BHI or E broths or agar plates at 20 �C

for 24–28 h.

Physiological and biochemical characterization

The temperature growth range of the strain JA-1 was

tested on LB and BHI agar plates at 2, 8, 10, 15, 20, 22,

24, 26, 30 and 37 �C. Resistance to antibiotics was

determined on Mueller–Hinton (MH) agar (CLSI, 2007)

using standard antibiotic discs (BBL, Oxoid) accord-

ing to the manufacturer’s instructions. Zones of

inhibition were examined after 48 h incubation at

20 �C.

Strains JA-1, J. lividum DSM 1522T and J.

agaricidamnosum DSM 9628 T were tested for

carbohydrate utilization capability with API 50 CH

kit (bioMerieux, Inc., France). All strains were grown

in LB broth at 20 �C for 30 h and resuspended in the

appropriate test media according to the manufacturer’s

instruction. The readings were done after 48 h incu-

bation at 20 �C. Catalase activity was detected with

BD Catalase Reagent droppers (BD, USA) according

to the manufacturer’s instructions.

Analysis of the cellular fatty acids composition was

carried out by the Identification Service of the DSMZ,

Braunschweig, Germany. Briefly, the fatty acid

methyl esters were obtained using minor modifications

of the method of Miller (1982) and Kuykendall et al.

(1988) and analyzed by gas chromatography accord-

ing to the standard protocol of the Sherlock Microbial

Identification System (MIDI, Microbial ID, Newark,

DE 19711 U.S.A.). The profiles of cellular fatty acids

were compared using the TSBA40 library database

version 4.10 (Microbial ID, Newark, DE, USA).

Analysis of respiratory quinones and polar lipids

were carried out by the Identification Service of the

DSMZ and Dr. B.J.Tindall, DSMZ, Braunschweig,

Germany. The TLC plates have been stained with 5 %

molybdophosphoric acid to show all lipids.

Pigment extraction and subsequent spectrophoto-

metric analysis were performed from cells of strains

JA-1 and J. lividum DSM 1522T, grown on LB agar

plates for 6 days at 22 �C. Spectrophotometric anal-

ysis of the pigment extracted from a loopful of bacteria

with 96 % ethanol (v/v) or 10 % H2SO4 (v/v) in 96 %

(v/v) ethanol, and filtered through 0.22 lm filter

(Millipore), was performed as described by (Logan

and Moss 1992). When necessary the pigment solution

was diluted in the same solvent. The absorption

spectrum between 200–800 nm was measured using a

UV-160 A (Shimadzu, Japan) spectrophotometer.

Strains JA-1 and J. lividum DSM 1522T were tested

for the ability to produce homoserine lactones which

are able to restore quorum sensing regulated violacein

synthesis in the white mutant strain Chromobacterium

violaceum CV026. The test was carried out on solid

LB, BHI and MG medium in a streak plate assay as

described by Steindler and Venturi (2007).

764 Antonie van Leeuwenhoek (2013) 103:763–769

123

Electron microscopy

For transmission electron microscopy freshly grown

pigmented colonies of strain JA-1 were cut out of LB

agar medium and fixed with 3.5 % glutaraldehyde in

0.1 M cacodylate buffer (pH 7.2) for 2 h at 4 �C. After

washing with 0.1 M cacodylate buffer for 30 min,

colonies were postfixed in 1 % OsO4 for 1 h at 4 �C.

After additional washing with the cacodylate buffer

the colonies were dehydrated in a graded alcohol

series at room temperature and embedded in ERL

4026 resin (SPI, USA). Ultrathin sections were stained

with uranyl aceate and Reynold’s lead citrate (Rey-

nolds 1963) and analyzed with the Philips CM 100

transmission electron microscope. For negative stain-

ing transmission electron microscopy, bacterial colo-

nies were resuspended in filtered water, transferred on

grids covered with Formvar support film (Ted Pella,

Inc. Redding, California) and negatively stained with

an equal volume of 0.5 % w/v aqueous uranyl acetate

(Bozzola and Russell 1999). Stained bacteria were air

dried at room temperature and examined with a Philips

CM 100 electron microscope. The images were

recorded by 792 Bioscan CCD camera (Gatan Inc.,

USA), using DigitalMicrograph software.

DNA isolation, 16S rRNA gene amplification,

sequencing and phylogenetic analysis

Genomic DNA was isolated as described by Ausubel

et al. (1992) from strain JA-1 grown in LB broth. The

16S rRNA genes were PCR amplified with oligonu-

cleotide primers fD1 (Weisburg et al. 1991) and 1495r

(Bandi et al. 1994). The reaction mixture containing

100 ng of the isolated DNA, 10 pg of each primer and

PCR Master mix from Fermentas (Lithuania) was first

denatured at 94 �C for 5 min and then subjected to 30

PCR cycles as follows: denaturation at 94 �C for 40 s,

primer annealing at 66 �C for 30 s, and elongation at

72 �C for 1 min 30 s. The final elongation step was

20 min at 72 �C. The PCR products were analyzed by

electrophoresis on 0.9 % (w/v) agarose gel and

subsequently purified with the QIA Gel Extraction

Kit (Qiagen). The purified PCR products were

sequenced on our request at the Microsynth GmbH

(Balgach, Switzerland) using the fD1 and 1495r

sequencing primers.

Phylogenetic analysis of the retrieved 16S rRNA

sequences was performed with the ARB phylogenetic

software package (Ludwig et al. 2004) using the all-

species living tree project database (release LTPs

106_SSU, August 2011) (Munoz et al. 2011). The final

tree was reconstructed after multiple analysis employ-

ing maximum parsimony, neighbor-joining, and max-

imum likelihood algorithms. Confidence in the tree

topology constructed by the Neighbor-joining method

was determined by the boostrap analysis employing

1.000 resamplings of the analyzed sequences.

DNA–DNA relatedness and G ? C content

determination

The G ? C content of DNA and the DNA–DNA

hybridization analysis were carried out by the identi-

fication Service of the DSMZ, Braunschweig, Ger-

many. Briefly, the G ? C content was determined

using the HPLC method after P1 nuclease hydrolysis

(Mesbah et al. 1989) of the DNA retrieved from cells

crushed with a French pressure cell and purified on

hydroxyapatite (Cashion et al. 1977). The DNA–DNA

hybridization analysis was performed from renatur-

ation rates as described by De Ley et al. (1970) and

with modifications described by Huss et al. (1983).

Nucleotide sequence accession number

The 16S rRNA gene sequence of strain JA-1 has been

deposited in GenBank under the accession number

DQ355146.2.

Results and discussion

Isolation and phenotypic characterization

of the strain JA-1

Strain JA-1 was isolated after three week incubation of

the molten glacier ice on E medium at 20 �C. JA-1 was

subsequently subcultivated on LB agar until a pure

culture of uniform, reddish-brown colonies containing

short Gram negative rods was obtained. Strain JA-1 is an

aerobic psyhrotrophic bacterium able to grow on various

growth media at temperatures between 2 and 25 �C.

Growth at lower temperatures was not tested. Whereas

visible colonies can be observed after 48 h incubation

at room temperature when grown on LB medium, a one

week incubation time is required when the strain is

grown on minimal growth medium or LB medium at

Antonie van Leeuwenhoek (2013) 103:763–769 765

123

2 �C. Cells of the strain JA-1 are rod-shaped with

rounded ends (2.5 ± 0.3 lm long and 0.7 ± 0.1 lm

wide), sometimes slightly curved, containing intracel-

lular inclusions (Fig. 1). JA-1 possesses one polar

flagellum (Fig. 2), whereas no subpolar or lateral

flagella, which are common in J. lividum, were

observed. The color of the colonies depends on the

medium, incubation time and temperature and varies

from reddish-brown to dark-brown or even black.

Similar applies for the shape of the colonies, changing

from tough and concave to soft and low-convex.

The biochemical characteristics of strain JA-1, in

comparison with related strains J. lividum DSM 1522T

and J. agaricidamnosum DSM 9628T, are shown in

Table 1. JA-1 can be easily differentiated by its ability

to utilize adonitol, N-acetylglucosamine and xylitol

using the API 50 CH kit. Additionally, only J. lividum

DSM 1522T can grow on LB medium supplemented

with 2 % NaCl (Table 1). Strain JA-1 is, like other

janthinobacteria, catalase positive.

The cellular fatty acids profile of the strain JA-1 is

very similar to the fatty acid profile of strain J. lividum

DSM 1522T. The major fatty acids of strain JA-1 consist

of C16:1 x7c/15 iso 2OH and C16:0, and the moderate amount

fatty acids include C18:1 x7c, C12:0, C10:0 3OH, and

C12:0 2OH. The fatty acids profile of strain J. lividum

DSM 1522T differs in the concentrations of the major

two fatty acids C16:1 x7c/15 iso 2OH (less) and C16:0 (more).

Analysis of the respiratory quinones indicated that

like in other janthinobacteria only ubiquinone Q8 is

present in strain JA-1. The polar lipid composition

consisted of phosphatidylethanolamine, phosphatidyl-

glycerol, an unknown phospholipid, an unknown

aminophospholipid and an additional unknown polar

lipid. The overall figure resembled the two-dimensional

TLC separation of polar lipids of J. lividum DSM

1522T and J. agaricidamnosum DSM 9628T (Lincoln

et al. 1999).

A violacein-like pigment is produced by strain JA-1

when grown on LB or MG agar plates or in the same

liquid media without shaking. Nevertheless, pigmen-

tation is distinct from the typical violacein pigmenta-

tion of C. violaceum, J. lividum or I. fluviatilis.

Colonies of strain JA-1 are reddish-brown rather than

violet. When grown under unfavorable conditions (e.g.

incubation on minimal growth medium or presence of

ampicillin) the pigment appears to accumulate and

colonies become dark brown or even black. After

prolonged incubation the pigment begins to diffuse

away from colonies into the surrounding media.

Spectrophotometric analysis of a pigment solution in

96 % (v/v) ethanol and in 10 % (v/v) H2SO4 in 96 %

(v/v) ethanol reveals clearly different absorption

maximums and minimums in tested strains (Table 2).

Fig. 1 Transmission electron micrograph of an ultrathin

section of strain J. svalbardensis JA-1 grown in LB medium.

Bar 1 lm

Fig. 2 Transmission electron micrograph of a negatively

contrasted strain J. svalbardensis JA-1 grown in LB medium.

Bar 2 lm

766 Antonie van Leeuwenhoek (2013) 103:763–769

123

Strains J. lividum DSM 1522T and JA-1 were also

tested for the production of homoserine lactones

which are able to restore quorum sensing regulated

violacein production in the white mutant strain

C. violaceum CV026 on solid LB, BHI and E medium

in a streak plate assay. Whereas J. lividum can restore

the violacein production in C. violaceum CVO26,

JA-1 cannot, even after prolonged incubation on

various media.

Resistance to a number of antibiotics was tested at

8, 15 and 22 �C. JA-1 was resistant to penicillin G (10

U disc), trimethoprim (5 lg disc), erythromycin (2 lg

disc), streptomycin and colistin (both 10 lg disc).

Phylogenetic analysis and DNA relatedness

Comparative sequence analysis was done using the

1403 bp long stretch of the 16S rRNA gene and

revealed that strain JA-1 undoubtedly belongs to the

Gram negative b-proteobacterial genus Janthinobac-

terium. The phylogenetic distance to the J. lividum

type strain DSM 1522T is not large, below 1 %, which

is similar as was found for the distances between

J. lividum and J. agaricidamnosum strains (Lincoln

et al. 1999). The distance to the J. agaricidamnosum

type strain DSM 9628T is slightly larger, 1.07 %. The

boostrap values of 96–98 % confirm the stability of

the nods within the genus Janthinobacterium (Fig 3.).

Due to the low DNA distances and thus limited

resolution power of the 16S rRNA phylogenetic

analysis, DNA–DNA hybridization tests were per-

formed. The % of DNA–DNA similarity was 15.9

(± 1.05) between JA-1 and J. lividum DSM 1522T and

8.85 (± 4.25) between JA-1 and J. agaricidamnosum

DSM 9628T, which is clearly below the recommended

threshold value of 70 % (Wayne et al. 1987) and

indicating that JA-1 does not belong to the J. lividum

nor to J. agaricidamnosum. The mol % G ? C value

of the DNA of the strain JA-1 was 62.3. We propose

that strain JA-1 is classified as representing a novel

species of the genus Janthinobacterium, designated as

J. svalbardensis.

Description of Janthinobacterium svalbardensis

sp. nov

Janthinobacterium svalbardensis (sval.bard.en’sis.

N.L. neut. adj. svalbardensis pertaining to Svalbard,

Table 1 Differential characteristics of strain J. svalbardensisJA-1T and related strains J. lividum DSM 1522T and J. aga-ricidamnosum DSM 9628T

Characteristics\strain JA-1Ta 1522Tb 9628Tc

Growth in LB with increased NaCl

Supplementation of NaCl 1 % ? ? nd

2 % – ? nd

3 % – – nd

Carbohydrate utilization in API 50CHd

D-arabinose – ? –

L-arabinose ? ? –

D-xylose – ? –

Adonitol ? – –

Galactose ? ? –

Sorbitol ? ? –

N-acetylglucosamine ? – –

Arbutine ? ? –

Salicine – ? –

Cellobiose – ? –

Maltose ? ? –

Trehalose ? – ?

Xylitol ? – –

b gentiobiose – – ?

D-lyxose ? ? –

L-fucose – ? –

2-ketogluconate – ? –

a J. svalbardensis JA-1T

b J. lividum DSM 1522T

c J. agaricidamnosum DSM 9628T

d Tests negative for all strains were: erythritol, L-xylose,

methyl b-xyloside, L-sorbose, rhamnose, dulcitol, methyl aD-

mannoside, methyl aD-glucoside, amygdalin, esculine, lactose,

melibiose, inuline, melezitose, D-raffinose, amidon, glycogen,

D-turanose, D-tagatose, D-fucose, L-arabitol, gluconate,

5-ketogluconate. Tests positive for all strains were: glycerol,

ribose, D-glucose, D-fructose, D-mannose, inositol, mannitol,

saccharose, D-arabitol

Table 2 Spectral properties of extracted pigments from strains

J. svalbardensis JA-1T and J. lividum DSM 1522T

Spectral properties/strain JA-1T 1522T

Absorption minimum of pigment extract in (in nm)

-96 % ethanol 330 430

-10 % H2SO4 in 96 % ethanol 330 495

Absorption maximum of pigment extract in (in nm)

-96 % ethanol 500 580

-10 % H2SO4 in 96 % ethanol 625 and 500 700

Antonie van Leeuwenhoek (2013) 103:763–769 767

123

the Norwegian archipelago in the Arctic, from where

the type strain was isolated).

J. svalbardensis displays the following properties:

Cells are Gram negative, rod-shaped with rounded ends,

sometimes slightly curved, motile by the means of single

polar flagellum. Upon growth on Luria–Bertani agar

medium cells contain abundant intracellular inclusions

visible under transmission electron microscopy. Col-

onies are small, tough, reddish-brown to almost black.

Growth occurs at temperatures between 2 and 25 �C,

but not higher. Visible colonies appear after 48 h

incubation at room temperature on LB medium,

whereas a one week incubation time is required when

grown on minimal growth medium or LB medium at

2 �C. Growth occurs in the presence of 1 % but not at

2 % NaCl (w/v). Catalase positive. In API 50CH test

strips, reactions are positive for glycerol, ribose, D-

glucose, D-fructose, D-mannose, inositol, mannitol,

saccharose, D-arabitol, L-arabinose, adonitol, galact-

ose, sorbitol, N-acetylglucosamine, arbutine, maltose,

trehalose, xylitol, and D-lyxose. A violacein-like

pigment is produced, different from the typical viola-

cein pigmentation of the C. violaceum, J. lividum or

I. fluviatilis, with absorption maximum and minimum

of 520 and 418 nm (extract in 96 % ET-OH).

J. svalbardensis does not produce homoserine lactones

which could restore the quorum sensing regulated

violacein production in the white mutant strain

C. violaceum CVO26. The DNA G ? C content of

the type strain JA-1T is 62.3 mol %. The major fatty

acids of strain JA-1 consist of C16:1 x7c/15 iso 2OH and

C16:0. The respiratory quinone of J. svalbardensis is Q8

and the major polar lipids are phosphatidylethanol-

amine and phosphatidylglycerol.

The type strain is J. svalbardensis JA-1T (= DSM-

25734; = ZIM B637), isolated from the ice sample of

the Austre Broggerbreen glacier located on the north-

west coast of Spitsbergen island of the Svalbard

archipelago, Norway.

The GenBank accession number of the 16S rRNA

sequence of the strain JA-1T is DQ355146.2.

Acknowledgments We would like to express our gratitude to

Prof. Dr. Nina Gunde Cimerman for the arctic glacier ice samples.

References

Ausubel FM, Brent R, Kingston RE, Moore DD, Seidmen JG,

Smith JA, Struhl K (eds) (1992) Current protocols in

molecular biology, vol 1. John Wiley and Sons, New York

Bandi C, Damiani G, Magrassi L, Grigolo A, Fani R, Sacchi L

(1994) Flavobacteria as intracellular symbionts in cock-

roaches. Proc R Soc Lond B 257:43–48

Fig. 3 Phylogenetic reconstruction based on a neighbour joining algorithm. The tree was constructed with the ARB phylogenetic

software package using the all-species living tree project database. Bar indicates 1.0 % of sequence divergence

768 Antonie van Leeuwenhoek (2013) 103:763–769

123

Bozzola JJ, Russell LD (1999) Electron Microscopy, 2nd edn.

Jones and Bartlett Publishers, Sudbury

Cashion P, Holder-Franklin MA, McCully J, Franklin M (1977)

A rapid method for the base ratio determination of bacterial

DNA. Anal Biochem 81:461–466

CLSI (2007) Performance standards for antimicrobial disk

susceptibility testing: seventeenth international supple-

ment M100–S17. CLSI, Wayne

De Ley J, Cattoir H, Reynaerts A (1970) The quantitative

measurement of DNA hybridization from renaturation

rates. Eur J Biochem 12:133–142

De Ley J, Segers P, Gillis M (1978) Intra and intergeneric

similarities of Chromobacterium and Janthinobacteriumribosomal ribonucleicacid cistrons. Int J Syst Bacteriol

25:154–168

Eisenberg J (1891) Bacteriologische Diagnostik Hilfstabellen

zum Gebrauche beim Praktischen Arbeiten 3 Aufl. Leopold

Voss, Hamburg

Gunde-Cimerman N, Sonjak S, Zalar P, Frisvad JC, Diderichsen

B, Plemenitas A (2003) Extremophilic fungi in arctic ice: a

relationship between adaptation to low temperature and

water activity. Phys Chem Earth 28:1273–1278

Huss VAR, Festl H, Schleifer KH (1983) Studies on the spec-

trophotometric determination of DNA hybridization from

renaturation rates. Syst Appl Microbiol 4:184–192

Kuykendall LD, Roy MA, O’Neill JJ, Devine TE (1988) Fatty

Acids, antibiotic resistance, and deoxyribonucleic acid

homology groups of bradyrhizobium japonicum. Int J Syst

Bacteriol 38:358–361

Lincoln SP, Fermor TR, Tindall BJ (1999) Janthinobacteriumagaricidamnosum sp. nov. a soft rot pathogen of Agaricusbisporus. Int J Syst Bacteriol 49:1577–1589

Logan NA (1989) Numerical taxonomy of violet-pigmented,

gram-negative bacteria and description of Iodobacter flu-viatile gen. nov. comb. nov. Int J Syst Bacteriol 39:450–456

Logan NA, Moss MO (1992) Identification of Chromobacte-rium, Janthinobacterium and Iodobacter species. In:

JonesBoard Skinner (ed) Identification methods in Applied

and Environmental Microbiology. The Society for Applied

Bacteriology Technical Series Blackwell Scientific Publi-

cations, Oxford

Ludwig W, Strunk O, Westram R, Richter L, Meier H, Yadh-

ukumar BA, Ali T, Steppi S, Jobb G, Forster W, Brettske I,

Gerber S, Ginhart AW, Gross O, Grumann S, Hermann S,

Jost R, Konig A, Liss T, Lußmann R, May M, Nonhoff B,

Reichel B, Strehlow R, Stamatakis A, Stuckmann N, Vilbig

A, Lenke M, Ludwig T, Bode A, Schleifer K-H (2004)

ARB: a software environment for sequence data. Nucleic

Acids Res 32:1363–1371

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

Miller LT (1982) A single derivatization method for bacterial

fatty acid methyl esters including hydroxy acids. J Clin

Microbiol 16:584–586

Moss MO, Ryall C (1981) The genus Chromobacterium. In:

Starr, Stolp, Truper, Balows and Schlegel (eds) The Pro-

karyotes, A Handbook on Habitats, Isolation, and Identi-

fication of Bacteria. Springer Verlag, Berlin

Munoz R, Yarza P, Ludwig W, Euzeby J, Amann R, Schleifer KH,

Glockner FO, Rossello-Mora R (2011) Release LTPs104 of

the all-species living tree. Syst Appl Microbiol 34:169–170

Reynolds ES (1963) The use of lead citrate at high pH as an

electron-opaque stain in electron microscopy. J Cell Biol

17:208–212

Schloss PD, Allen HK, Klimowicz AK, Mlot C, Gross JA,

Savengsuksa S, McEllin J, Clardy J, Ruess RW, Handels-

man J (2010) Psychrotrophic strain of Janthinobacteriumlividum from a cold alaskan soil produces prodigiosin.

DNA Cell Biol 29:533–541Sneath PHA (1984) Genus Janthinobacterium. In: Krieg NR,

Holt JG (eds) Bergey’s manual of systematic bacteriology.

The Williams & Wilkins Co., Baltimore, pp 376–377

Steindler L, Venturi V (2007) Detection of quorum-sensing

N-acyl homoserine lactone signal molecules by bacterial

sensors. FEMS Microbiol Lett 266:1–9

Vogel HJ, Bonner DM (1956) Acetylornithinase of Escherichiacoli: partial purification and some properties. J Biol Chem

218:97–106

Wayne LG, Brenner DJ, Colwell RR, Grimont PAD, Kandler O,

Krichevsky MI, Moore LH, Moore WEC, Murray RGE,

Stackebrandt E, Starr MP, Truper HG (1987) Report of the

Ad Hoc committee on reconciliation of approaches to

bacterial systematics. Int J Syst Bacteriol 37(4):463–464

Weisburg WG, Barms SM, Pelletier DA, Lane DJ (1991) 16S

ribosomal DNA amplification for phylogenetic study.

J Bacteriol 173(697–7):03

Antonie van Leeuwenhoek (2013) 103:763–769 769

123