Arthrobacter nitroguajacolicus sp. nov., a novel 4-nitroguaiacol-degrading actinobacterium Ludmila Kotoucˇ kova´, 1 Peter Schumann, 2 Eva Durnova´, 3 Cathrin Spro¨ er, 2 Ivo Sedla´cˇek, 4 Jirˇı ´ Necˇa, 5 Zbyneˇ k Zdra´ hal 6,7 and Miroslav Neˇ mec 1 Correspondence Ludmila Kotoucˇkova´ lida@sci.muni.cz 1 Department of Microbiology, Faculty of Science, Masaryk University Brno, Tvrde´ho 14, 602 00 Brno, Czech Republic 2 DSMZ – German Collection of Microorganisms and Cell Cultures, Braunschweig, Germany 3 Regional Institute of Public Health, Ostrava, Czech Republic 4 CCM – Czech Collection of Microorganisms, Masaryk University Brno, Czech Republic 5 Veterinary Research Institute, Brno, Czech Republic 6 Institute of Analytical Chemistry, Brno, Czech Republic 7 Laboratory of Functional Genomics and Proteomics, Faculty of Science, Masaryk University Brno, Czech Republic Three bacterial isolates from soil, capable of degradation or transformation of nitroaromatic compounds and displaying a rod–coccus growth cycle, were studied by a polyphasic approach. On the basis of 16S rRNA sequence analysis and of chemotaxonomic characteristics, such as type A3a peptidoglycan with an interpeptide bridge Ala–Thr–Ala, the major menaquinone MK-9(H 2 ) and fatty acid composition, the isolates were assigned to the genus Arthrobacter. DNA–DNA hybridization, riboprinting and phenotypic studies revealed that the three strains constitute a single species, distinct from phylogenetically neighbouring Arthrobacter aurescens and Arthrobacter ilicis. A novel species, Arthrobacter nitroguajacolicus sp. nov., with the type strain G2-1 T (=CCM 4924 T =DSM 15232 T ) is proposed. The genus Arthrobacter (Conn & Dimmick, 1947; Koch et al., 1995) embraces more than 30 species of bacteria isolated mostly from soil. Arthrobacters are metabolically versatile, and some species of this genus are characterized by the ability to metabolize xenobiotics (Westerberg et al., 2000). Strain 8/3 (=CCM 4925=DSM 15233), identified at first as Corynebacterium sp., was isolated from con- taminated soil (Kotoucˇkova´ et al., 1997) and showed the ability to degrade nitroaromatic compounds. Two addi- tional strains, G2-1 T and P1P (=CCM 7049=DSM 15234), capable of degrading 4-nitroguaiacol (4-NG), were later isolated from forest soil. For isolation, approximately 1 g soil was transferred into 100 ml mineral medium (MM) supplemented with 0?1 mM 4-nitrophenol (4-NP), and incubated at 28 u C on a rotary shaker. The composition of MM (g l 21 ) was as follows: CaCl 2 , 0?0275; MgSO 4 .7H 2 O, 0?0225; FeCl 3 .6H 2 O, 0?00025; (NH 4 ) 2 SO 4 ,0?05; KH 2 PO 4 ,0?17; K 2 HPO 4 ,0?42; Na 2 HPO 4 .12H 2 O, 0?88. The enrichment products (10 ml) were transferred to 100 ml fresh MM supplemented with 4-NP (strain P1P), or to MM supplemented with 4-NG (strain G2-1 T ). After degradation of the nitroaromatic compounds, indicated by decolorization of the medium, the procedure was repeated three times. Subsequent streaking on solid MM supplemented with 4-NP or 4-NG resulted in the formation of single colonies. Degradation of nitroaro- matic compounds was confirmed by cultivation of the strains in liquid MM supplemented with 4-NP (0?05 g l 21 ) or 4-NG (0?004 g l 21 ), and by HPLC analysis. A Waters HPLC system, with a reverse-phase Nova-Pak C18 column and acetonitrile/0?1 % acetic acid (15 : 85) as the mobile phase, was used for HPLC analysis. Strain 8/3 was cultivated in MM with 0?2 % (w/v) peptone and 0?05 g 4-NP l 21 . Identification of 4-NG as a product of 4-NP transforma- tion by strain 8/3 was performed with a liquid HP 1100 chromatograph (Hewlett-Packard) coupled with an ion trap mass spectrometer (Bruker Daltonik). Nutrient agar (CM2, Oxoid) was used for the characteri- zation of colonies and cell morphology. Cell size was Abbreviations: 4-NG, 4-nitroguaiacol; 4-NP, 4-nitrophenol. Published online ahead of print on 28 November 2003 as DOI 10.1099/ijs.0.02923-0. The GenBank/EMBL/DDBJ accession number for the 16S rDNA sequence of strain G2-1 T is AJ512504. Tables showing the differentiating physiological reactions and cellular fatty acid composition of Arthrobacter nitroguajacolicus sp. nov. strains are available as supplementary material in IJSEM Online. 02923 G 2004 IUMS Printed in Great Britain 773 International Journal of Systematic and Evolutionary Microbiology (2004), 54, 773–777 DOI 10.1099/ijs.0.02923-0