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Amantichitinum ursilacus gen. nov., sp. nov., a
chitin-degrading bacterium isolated from soil
Karin S. Moß,13 Stefan C. Hartmann,13 Isabell Mu ¨ller,2Christina Fritz,2
Sven Kru ¨gener,2Susanne Zibek,2Thomas Hirth1,2and Steffen Rupp2
1Institute for Interfacial Engineering, University of Stuttgart, Nobelstraße 12, 70569 Stuttgart,
2Fraunhofer Institute for Interfacial Engineering and Biotechnology, Nobelstraße 12,
70569 Stuttgart, Germany
A bacterial strain named IGB-41Twas isolated from a soil sample from an ant hill near Stuttgart,
Germany. The strain was Gram-negative, rod-shaped, motile and facultatively anaerobic.
Phylogenetic analysis based on 16S rRNA grouped the strain IGB-41Twithin the class
Betaproteobacteria into the family Neisseriaceae together with Silvimonas amylolytica NBRC
103189T, Silvimonas iriomotensis NBRC 103188Tand Silvimonas terrae KM-45Tas the closest
relatives with sequence similarities of 96.7, 96.6 and 96.1%, respectively. The G+C content of
the genomic DNA was determined to be 61.5 mol% and quinone analysis revealed Q-8 as the
only detectable quinone. Major cellular fatty acids were identified as C16:0, summed feature 3
(iso-C15:02-OH and/or C16:1v7c) and C18:1v7c . Strain IGB-41Twas unique in harbouring
phosphoaminolipids, aminolipids and glycoaminolipids when compared with Silvimonas
amylolytica NBRC 103189Tin polar lipid analysis. On the basis of the physiological, phenotypic
and genotypic characteristics of strain IGB-41T, we suggest that the novel strain should be
assigned to a new genus Amantichitinum and novel species Amantichitinum ursilacus. The type
species of the genus Amantichitinum is Amantichitinum ursilacus and the type strain is IGB-41T
(5DSM 23761T5CIP 110167T).
Chitin is the second most abundant polymer on earth,
synthesized for use as structural scaffold by a variety of
organisms including fungi, crustaceans and insects (Patil
et al., 2000). Due to its omnipresence, many organisms are
able to degrade chitin and use it as a carbon and energy
source in metabolic processes (Campbell & Williams,
1951). Habitats characterized by a natural enrichment of
chitinous polymer, such as ant hills and other agglomera-
tions of arthropoda, are more likely to harbour chitinolytic
micro-organisms than other habitats. Therefore, screening
for chitin-metabolizing micro-organisms in such habitats is
highly promising. During a research project focused on the
identification and characterization of new chitinolytic
bacteria for use in industrial processes, strain IGB-41T
was isolated from a soil sample taken near Stuttgart,
A soil sample with a temperature of 18 uC was taken from
an abandoned ant hill located near Lake Ba ¨rensee in
Stuttgart, Germany. Chitinolytic organisms were enriched
by sequential use of different media with chitin of rising
complexity. In a first and second step, acidic pretreated
chitin was used as carbon source, which in the third step
was supplemented with protein-free milled chitin. The
latter was the sole carbon source for the fourth culture
step. Three grams of the soil sample were transferred
into 100 ml screening medium (pH 4.2) containing
0.05 M potassium phosphate, 5 mg Ca(NO3)2.4H2O l21,
20 mg MgSO4.7H2O l21, 100 mg (NH4)2SO4l21, 1 mg
Fe(III)NH4-citrate l21, 100 mg ZnSO4.7H2O l21, 30 mg
MnCl2.4H2O l21,300 mgH3BO3l21,
6H2O l21, 10 mg CuCl2.2H2O l21, 20 mg NiCl2.6H2O l21,
30 mg Na2MoO4.2H2O l21, 25 mg tryptone l21, 12.5 mg
yeast extract l21, 25 mg NaCl l21and 0.5 g wet mass
colloidal chitin l21. The suspension was incubated at
20 uC. After 10 days, a 10 ml aliquot of the suspension was
added to 90 ml fresh culture medium. This procedure was
repeated twice with tryptone, yeast extract and NaCl omitted
and 1 g chitin (Senn Chemicals) added for the third step. In
a fourth step, colloidal chitin was also omitted.
The micro-organism suspension was diluted 1:106, spread
on Luria–Bertani (LB)-agar pH 4.2 and incubated for
5 days at 20 uC. Morphologically distinguishable colonies
3These authors contributed equally to this work.
The GenBank/EMBL/DDBJ accession number for the 16S rRNA gene
sequence of Amantichitinum ursilacus IGB-41Tis FN994890.
Two supplementary figures, a supplementary table and a sequence
alignment are available with the online version of this paper.
International Journal of Systematic and Evolutionary Microbiology (2013), 63, 98–103
98034447G2013 IUMSPrinted in Great Britain
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Cashion, P., Holder-Franklin, M. A., McCully, J. & Franklin, M. (1977).
A rapid method for the base ratio determination of bacterial DNA.
Anal Biochem 81, 461–466.
De Ley, J., Cattoir, H. & Reynaerts, A. (1970). The quantitative
measurement of DNA hybridization from renaturation rates. Eur J
Biochem 12, 133–142.
Huß, V. A. R., Festl, H. & Schleifer, K. H. (1983). Studies on the
spectrophotometric determination of DNA hybridization from
renaturation rates. Syst Appl Microbiol 4, 184–192.
Kuykendall, L. D., Roy, M. A., O’Neill, J. J. & Devine, T. E. (1988). Fatty
acids, antibiotic resistance, and deoxyribonucleic acid homology
groups of Bradyrhizobium japonicum. Int J Syst Bacteriol 38, 358–361.
Larkin, M. A., Blackshields, G., Brown, N. P., Chenna, R., McGettigan,
P. A., McWilliam, H., Valentin, F., Wallace, I. M., Wilm, A. & other
authors (2007). CLUSTAL W and CLUSTAL_X version 2.0. Bioinformatics
Ludwig, W., Strunk, O., Westram, R., Richter, L., Meier, H.,
Yadhukumar, Buchner, A., Lai, T., Steppi, S. & other authors
(2004). ARB: a software environment for sequence data. Nucleic Acids
Res 32, 1363–1371.
Mesbah, M., Premachandran, U. & Whitman, W. B. (1989). Precise
measurement of the G+C content of deoxyribonucleic acid by high-
performance liquid chromatography. Int J Syst Bacteriol 39, 159–167.
Miller, L. T. (1982). Single derivatization method for routine analysis
of bacterial whole-cell fatty acid methyl esters, including hydroxy
acids. J Clin Microbiol 16, 584–586.
Muramatsu, Y., Suzuki, K. & Nakagawa, Y. (2010). Silvimonas
iriomotensis sp. nov. and Silvimonas amylolytica sp. nov., new
members of the class Betaproteobacteria isolated from the subtropical
zone in Japan. Int J Syst Evol Microbiol 60, 174–178.
Patil, R. S., Ghormade, V. & Deshpande, M. V. (2000). Chitinolytic
enzymes: an exploration. Enzyme Microb Technol 26, 473–483.
Reddy, C. A., Beveridge, T. J., Breznak, J. A., Snyder, L., Schmidt,
T. M. & Marzluf, G. A. (2007). Methods for General and Molecular
Microbiology. Washington, DC: American Society for Microbiology.
Saitou, N. & Nei, M. (1987). The neighbor-joining method: a new
method forreconstructingphylogenetic trees. Mol Biol Evol 4, 406–425.
Sheu, S. Y., Chiu, T. F., Chou, J. H., Sheu, D. S., Arun, A. B., Young,
C. C., Chen, C. A., Wang, J. T. & Chen, W. M. (2009). Andreprevotia
lacus sp. nov., isolated from a fish-culture pond. Int J Syst Evol
Microbiol 59, 2482–2485.
Stackebrandt, E. & Goebel, B. M. (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–
Stamatakis, A. (2006). RAxML-VI-HPC: maximum likelihood-based
phylogenetic analyses with thousands of taxa and mixed models.
Bioinformatics 22, 2688–2690.
Tamaoka, J. & Komagata, K. (1984). Determination of DNA base
composition by reversed-phase high-performance liquid chromato-
graphy. FEMS Microbiol Lett 25, 125–128.
Tindall, B. J. (1990a). A comparative study of the lipid composition
of Halobacterium sacharovorum from various sources. Syst Appl
Microbiol 13, 128–130.
Tindall, B. J. (1990b). Lipid composition of Halobacterium lacu-
sprofundi. FEMS Microbiol Lett 66, 199–202.
Tindall, B. J., Sikorski, J., Smibert, R. M. & Kreig, N. R. (2007).
Phenotypic characterization and the principles of comparative
systematics. In Methods for General and Molecular Microbiology, 3rd
edn. pp. 330–393. Edited by C. A. Reddy, T. J. Beveridge, J. A. Breznak,
G. A. Marzluf, T. M. Schmidt & L. Snyder. Washington, DC, USA:
American Society for Microbiology.
Tindall,B. J.,Rossello ´-Mo ´ra,R., Busse,H. J.,Ludwig,W. & Ka ¨mpfer,P.
(2010). Notes on the characterization of prokaryote strains for
taxonomic purposes. Int J Syst Evol Microbiol 60, 249–266.
Krichevsky, M. I., Moore, L. H., Moore, W. E. C., Murray, R. G. E. & other
authors (1987). International Committee on Systematic Bacteriology.
Report of the ad hoc committee on reconciliation of approaches to
bacterial systematics. Int J Syst Bacteriol 37, 463–464.
Weon, H.-Y., Kim, B.-Y., Yoo, S.-H., Joa, J.-H., Kwon, S.-W. & Kim,
W.-G. (2007). Andreprevotia chitinilytica gen. nov., sp. nov., isolated
from forest soil from Halla Mountain, Jeju Island, Korea. Int J Syst
Evol Microbiol 57, 1572–1575.
Yang, H.-C., Im, W.-T., An, D.-S., Park, W. S., Kim, I. S. & Lee, S.-T.
(2005). Silvimonas terrae gen. nov., sp. nov., a novel chitin-degrading
facultative anaerobe belonging to the ‘Betaproteobacteria’. Int J Syst
Evol Microbiol 55, 2329–2332.
Yoon, J. H., Choi, J. H., Kang, S. J., Choi, N. S., Lee, J. S. & Song, J. J.
(2010). Jeongeupia naejangsanensis gen. nov., sp. nov., a cellulose-
degrading bacterium isolated from forest soil from Naejang Mountain
in Korea. Int J Syst Evol Microbiol 60, 615–619.
Amantichitinum ursilacus gen. nov., sp. nov.