Development of a Rhodococcus recombinant strain for degradation of products from anaerobic dechlorination of PCBs

Abstract

The Gram-positive bacterium Rhodococcus sp. strain RHA1, naturally containing the biphenyl pathway, was electroporated with a broad host range plasmid containing the 4-chlorobenzoate (4-CBA) degradation operon (fcb) isolated from Arthrobacter globiformis strain KZT1. The recombinant strain grew in medium containing 4-CBA and 4-chlorobiphenyl (4-CB) as the only source of carbon, with stoichiometric release of chloride and a molar growth yield on 4-CB that suggested utilization of both biphenyl rings. In resting cell assays, similar rates of degradation were observed for wild-type and recombinant strains for the most common eight congeners from the anaerobic dechlorination of Aroclor 1242, but the recombinant strain accumulated lower amounts of chlorinated meta-cleavage products and no 4-CBA. Recombinant cells inoculated at 10(4) cells/g into nonsterile soil amended with 4-CB grew to 6-10(5) cells/g, a density consistent with the 4-CB consumed. 4-CB was removed only in the inoculated soil, and the recombinant strain did not grow in the same soil when it was not amended with 4-CB. The fcb operon remained stable in the recombinant strain reisolated from soil after 60 days. This work provides proof of concept that a Rhodococcus strain constructed to grow on a PCB would grow in nonsterile soil if the appropriate chlorobiphenyl is available.