title: Genetic and metabolic analysis of the carbofuran catabolic pathway in Novosphingobium sp. KN65.2.
abstract: The widespread agricultural application of carbofuran and concomitant contamination of surface and ground waters has raised health concerns due to the reported toxic effects of this insecticide and its degradation products. Most bacteria that degrade carbofuran only perform partial degradation involving carbamate hydrolysis without breakdown of the resulting phenolic metabolite. The capacity to mineralize carbofuran beyond the benzofuran ring has been reported for some bacterial strains, especially sphingomonads, and some common metabolites, including carbofuran phenol, were identified. In the current study, the catabolism of carbofuran by Novosphingobium sp. KN65.2 (LMG 28221), a strain isolated from a carbofuran-exposed Vietnamese soil and utilizing the compound as a sole carbon and nitrogen source, was studied. Several KN65.2 plasposon mutants with diminished or abolished capacity to degrade and mineralize carbofuran were generated and characterized. Metabolic profiling of representative mutants revealed new metabolic intermediates, in addition to the initial hydrolysis product carbofuran phenol. The promiscuous carbofuran-hydrolyzing enzyme Mcd, which is present in several bacteria lacking carbofuran ring mineralization capacity, is not encoded by the Novosphingobium sp. KN65.2 genome. An alternative hydrolase gene required for this step was not identified, but the constitutively expressed genes of the unique cfd operon, including the oxygenase genes cfdC and cfdE, could be linked to further degradation of the phenolic metabolite. A third involved oxygenase gene, cfdI, and the transporter gene cftA, encoding a TonB-dependent outer membrane receptor with potential regulatory function, are located outside the cfd cluster. This study has revealed the first dedicated carbofuran catabolic genes and provides insight in the early steps of benzofuran ring degradation.
authors.affiliation: Division of Soil and Water Management, Department of Earth and Environmental Sciences, KU Leuven, Kasteelpark Arenberg 20, Box 2459, 3001, Heverlee, Leuven, Belgium.
authors.forename: Thi Phi Oanh
authors.initials: TP
authors.lastname: Nguyen
authors.forename: Damian E
authors.initials: DE
authors.lastname: Helbling
authors.forename: Karolien
authors.initials: K
authors.lastname: Bers
authors.forename: Tekle Tafese
authors.initials: TT
authors.lastname: Fida
authors.forename: Ruddy
authors.initials: R
authors.lastname: Wattiez
authors.forename: Hans-Peter E
authors.initials: HP
authors.lastname: Kohler
authors.forename: Dirk
authors.initials: D
authors.lastname: Springael
authors.forename: René
authors.initials: R
authors.lastname: De Mot
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002235:Carbofuran / Q000737:chemistry / Q000378:metabolism*; D007306:Insecticides / Q000737:chemistry / Q000378:metabolism*; D053858:Metabolic Networks and Pathways; D012988:Soil Microbiology; D042301:Sphingomonadaceae / Q000201:enzymology / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D007306:Insecticides; D002235:Carbofuran