ISOLATION AND CHARACTERIZATION OF DEHALOBACTERIUM-FORMICOACETICUM GEN-NOV SP-NOV, A STRICTLY ANAEROBIC BACTERIUM UTILIZING DICHLOROMETHANE AS SOURCE OF CARBON AND ENERGY

A strictly anaerobic, dichloromethane-utilizing bacterium was isolated from a previously described dichloromethane-fermenting, two-component mixed culture. In a mineral medium with vitamins, the organism conver ted 5 mM dichloromethane within 7 days to formate plus acetate in a mo lar ratio of 2:1 and to biomass and traces of pyruvate. Of 50 potentia l substrates and combinations of substrates tested, only dichlorometha ne supported growth. The organism had a DNA G+C content of 42.7 mol%. From its phylogenetic position deduced from 16S rDNA analysis and from its unique substrate range, we conclude that the organism represents a new genus and a new species within the phylum of the gram-positive b acteria for which we propose the name Dehalobacterium formicoaceticum. Cell extracts were found to contain carbon monoxide dehydrogenase, me thylene tetrahydrofolate dehydrogenase, formyl tetrahydrofolate synthe tase, and hydrogenase activities, whereas activities of methenyl tetra hydrofolate cyclohydrolase and methylene tetrahydrofolate reductase we re not detectable. Activity for dehalogenation of dichloromethane was lost on preparation of cell extracts, but was maintained in cell suspe nsions. Oxygen and reagents that react with thiol groups caused irreve rsible inhibition, and propyl iodide caused reversible inhibition of d ehalogenation. Our observations suggest: 1) conversion of dichlorometh ane to methylene tetrahydrofolate, which gives rise to both formate an d the methyl group of acetate, or 2) conversion of two molecules of di chloromethane to methylene tetrahydrofolate (which is oxidized to form ate) and parallel reductive dehalogenation of one dichloromethane to t he methyl group of the corrinoid-protein involved in acetate formation .