Downloaded from www.microbiologyresearch.org by IP: 54.90.167.105 On: Fri, 03 Jun 2016 03:42:22 International Journal of Systematic and Evolutionary Microbiology (2001), 51, 1773–1782 Printed in Great Britain Classification of metal-resistant bacteria from industrial biotopes as Ralstonia campinensis sp. nov., Ralstonia metallidurans sp. nov. and Ralstonia basilensis Steinle et al. 1998 emend. 1 Laboratorium voor Microbiologie, Universiteit Gent, K. L. Ledeganckstraat 35, B-9000 Gent, Belgium 2 BCCM/LMG Bacteria Collection, Laboratorium voor Microbiologie, Universiteit Gent, K. L. Ledeganckstraat 35, B-9000 Gent, Belgium 3,4 Environmental Technology, Vlaamse Instelling voor Technologisch Onderzoek (VITO) 3 and Laboratory for Microbiology, Radioactive Waste & Clean-up Division, SCK/CEN (Center of Studies for Nuclear Energy) 4 , Boeretang 200, B-2400 Mol, Belgium Johan Goris, 1 Paul De Vos, 1 Tom Coenye, 1 Bart Hoste, 2 Danielle Janssens, 2 Hassan Brim, 3 Ludo Diels, 3 Max Mergeay, 3,4 Karel Kersters 1,2 and Peter Vandamme 1 Author for correspondence : Johan Goris. Tel : 32 9 264 51 01. Fax: 32 9 264 53 46. e-mail : johan.gorisrug.ac.be Thirty-one heavy-metal-resistant bacteria isolated from industrial biotopes were subjected to polyphasic characterization, including 16S rDNA sequence analysis, DNA–DNA hybridizations, biochemical tests, whole-cell protein and fatty-acid analyses. All strains were shown to belong to the Ralstonia branch of the β-Proteobacteria. Whole-cell protein profiles and DNA–DNA hybridizations revealed two clearly distinct groups, showing low similarity to known Ralstonia species. These two groups, of 8 and 17 isolates, were assigned to two new species, for which the names Ralstonia campinensis sp. nov. and Ralstonia metallidurans sp. nov. are proposed. The type strains are WS2 T ( LMG 19282 T  CCUG 44526 T ) and CH34 T ( LMG 1195 T  DSM 2839 T ), respectively. Six isolates were allocated to Ralstonia basilensis, which presently contains only the type strain ; an emendation of the latter species description is therefore proposed. Keywords : Ralstonia, metal-resistance, taxonomy, industrial biotopes INTRODUCTION Metal resistance has been reported for a number of Gram-negative bacteria belonging to the Ralstonia lineage of the β-Proteobacteria (Diels & Mergeay, 1990 ; Mergeay et al., 1985 ; Schmidt et al., 1991; Schmidt & Schlegel, 1994). Resistance to Cd+ , Zn+ and Co+ has been shown to be located on a czc operon of the plasmid pMOL30 (240 kb), while Co+ combined with Ni+ resistance is located on the cnr operon of the plasmid pMOL28 (163 kb) (Liesegang et al., 1993 ; Mergeay et al., 1985; Nies et al., 1987; Siddiqui et al., 1989). Other plasmid-borne resistances to metals include resistance to copper (cop), lead ( pbr) and mercury (mer) on pMOL30 and resistance to chromate (chr) and mercury on pMOL28 (for a review see Mergeay, 2000). Hybridization with a czc gene ................................................................................................................................................. The EMBL accession numbers for the 16S rRNA gene sequences of R. campinensis WS2 T and R. basilensis ER121 and DSM 11853 T are AF312020, AF312021 and AF312022, respectively. probe showed the spread of the former resistance types over a variety of closely related taxa (Brim et al., 1999). The application of metal-resistant bacteria in bio- remediation processes offers attractive perspectives for biomonitoring (via biosensors), treatment of waste- water and the recycling of polluted soils (Collard et al., 1994 ; Diels et al., 1995). In the case of the Ralstonia strains mentioned above, knowledge of their exact taxonomic position is of the utmost importance. Indeed, in addition to organisms that can be used for bioremediation of polluted soils, the genus Ralstonia also contains an important plant pathogen (Ralstonia solanacearum ; Hayward, 1991) and several oppor- tunistic human pathogens such as Ralstonia pickettii (Ralston et al., 1973), Ralstonia mannitolytica (De Baere et al., 2001), Ralstonia gilardii (Coenye et al., 1999) and Ralstonia paucula (Vandamme et al., 1999). Ralstonia strains with potential large-scale appli- cations for bioremediation should obviously be dis- tinct from (opportunistic) pathogens of humans and plants. Recent taxonomic studies describing novel Ralstonia species highlight not only the unsuspected 01846  2001 IUMS 1773