Biodegradation of 2,4-dinitrotoluene by a genetically modified Pseudomonas fluorescens strain

MONTI MARIELA. R.; SMANIA ANDREA M.; ARGARAÑA CARLOS E.

Córdoba-Argentina

Congreso; XXXVII Reunión Anual de SAIB; 2001

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

2,4-dinitrotoluene (DNT) is a priority pollutant present in effluents from industrial production of feedstock for polyurethane foams. The elimination of this compound by a biodegradative process would represent an economic and environmental safe alternative. It has been described an oxidative pathway for mineralization of DNT by a Burkholderia cepacia strain which involves the consecutive action of a three-plasmid encoded oxygenase enzymes. Although B. cepacia can mineralize efficiently DNT, this bacteria has been characterized as a phytopathogen and a human opportunistic pathogen which impedes its use in industrial biodegradative processes. The main objective of our work was to overcome this problem by transferring the genes that encode the DNT degradation enzymes from B. cepacia to a non-pathogenic bacteria such as Pseudomonas fluorescens. To generate a stable phenotype, the genes previously cloned from B. cepacia were transferred to the P. fluorescens chromosome by conjugation and transposition events using the pUT-miniTn5 plasmids. The insertion of the genes in the chromosome and the functionality of the encoded enzymes were verified by Southern blot and an in vivo enzymatic assay respectively. The genetically modified strain is able to grow in solid medium using DNT as a sole source of carbon, nitrogen and energy. In liquid cultures, this strain degrades DNT with the concomitant release of NO2 and oxygen uptake.