Appl Microbiol Biotechnol (2002) 59:62–67 DOI 10.1007/s00253-002-0979-7 Abstract Thirty-five strains of soil bacteria were grown with biphenyl (BP) and tested for their capacity to cooxi- dize dibenzofuran (DBF). During metabolism of DBF, the culture medium of 17 strains changed from colorless to or- ange, indicating a meta-cleavage pathway of DBF degra- dation. The ring cleavage product of these isolates was shown to be 2-hydroxy-4-(3′-oxo-3′H-benzofuran-2′-yli- den)but-2-enoic acid (HOBB). The strain SBUG 271, studied in detail and identified as Rhodococcus erythropo- lis, degraded DBF via 1,2-dihydroxydibenzofuran. The ensuing meta-cleavage yielded HOBB and salicylic acid. In addition, the four monohydroxylated monomers of DBF and two metabolites, which were not further characterized, were detected. Thus, our results demonstrate that the met- abolic mechanism involves lateral dioxygenation of DBF followed by meta-cleavage and occurs in Gram-negative as well as in Gram-positive BP-degrading bacteria. Introduction Dibenzofuran (DBF) and the halogenated congeneres of DBF are reported to be environmental pollutants with potential risks for human health (Goerlitz et al. 1985; Ahlborg et al. 1992; van Birgelen et al. 1996). In recent years, several strains of bacteria have been isolated and characterized which are able to use DBF as a growth substrate. Most of these strains are members of the proteobacteria, namely Pseudomonas sp. NCIB 9816–4 (Resnick and Gibson 1996), Pseudomonas fluorescens TTC1 (Bianchi et al. 1997), Sphingomonas sp. HH69 (Fortnagel et al. 1990; Harms et al. 1995) and strain RW1 (Bünz et al. 1993; Wilkes et al. 1996). Additional- ly, Gram-positive bacteria like Brevibacterium sp. DPO1361 (Engesser et al. 1989), Terrabacter sp. DBF63 (Monna et al. 1993; Kasuga et al. 1997) and strain DPO360 (Schmid et al. 1997) seem to be able to oxidize DBF. The DBF degradation pathway of these bacteria starts with hydroxylation at carbon atoms 4 and 4a (an- gular dioxygenation), resulting in the cleavage of the ether bridge, and is followed by formation of 2,2′,3-tri- hydroxybiphenyl (Strubel et al. 1991). Subsequently, a second dioxygenase catalyzes the ring cleavage of this intermediate to 2-hydroxy-6-oxo-6-(2-hydroxyphenyl)- hexa-2,4-dienoic acid, a meta-cleavage product, which is transformed to 2-hydroxypenta-2,4-dienoic acid and sali- cylic acid (Fortnagel et al. 1990; Bertini et al. 1995). Ring cleavage of DBF after hydroxylation at carbon atoms 1 and 2 (lateral dioxygenation) by biphenyl (BP)- utilizing bacteria has been postulated by several authors, because of the yellow-orange coloration during incuba- tion with the substrate (Cerniglia et al. 1979). This as- sumption has been proven for Ralstonia sp. SBUG 290, which degraded DBF after cultivation with BP. Ring fis- sion of 1,2-dihydroxydibenzofuran (1,2-DiOH-DBF) re- sulted in production of 2-hydroxy-4-(3′-oxo-3′H-benzo- furan-2′-yliden)but-2-enoic acid (HOBB), which was de- graded via salicylic acid (Becher et al. 2000). The aim of our study was to investigate whether the lateral dioxygenation of DBF with subsequent meta- cleavage of the dihydroxylated intermediate can be car- ried out by other Gram-negative or Gram-positive bacte- ria growing with BP. The DBF metabolism of the Gram- positive isolate Rhodococcus erythropolis SBUG 271 has been characterized in more detail. Materials and methods Organisms and growth conditions The bacteria were isolated from sewage sludge and compost soil samples by enrichment cultivation performed in 500-ml flasks M.B. Stope ( ✉ ) Institute of Molecular Biology, Friedrich-Loeffler-Institutes, Federal Research Centre for Virus Diseases of Animals, Boddenblick 5a, 17498 Insel Riems, Germany e-mail: stope@rie.bfav.de Tel.: +49-38351-7267, Fax: +49-38351-7151 D. Becher · E. Hammer · F. Schauer Institute of Microbiology and Molecular Biology, Ernst-Moritz-Arndt-University of Greifswald, Friedrich-Ludwig-Jahnstrasse 15, 17487 Greifswald, Germany ORIGINAL PAPER M. B. Stope · D. Becher · E. Hammer · F. Schauer Cometabolic ring fission of dibenzofuran by Gram-negative and Gram-positive biphenyl-utilizing bacteria Received: 12 October 2001 / Revised: 12 February 2002 / Accepted: 15 February 2002 / Published online: 6 April 2002 © Springer-Verlag 2002