3-Chloro-, 2,3- and 3,5-dichlorobenzoate co-metabolism in a 2-chlorobenzoate-degrading consortium: role of 3,5-dichlorobenzoate as antagonist of 2-chlorobenzoate degradation

Abstract

A study was made of the metabolic and co-metabolic intermediates of 2- and 3-chlorobenzoate, 2,3- and 3,5-dichlorobenzoate to elucidate the mechanism(s) involved in the negative effects observed on the growth of a chlorobenzoate-degrading microbial consortium in the presence of mixed chlorobenzoates. 2-Chloro-muconate accumulated as the end-product in the cultural broths of the microbial consortium during growth on 2-chlorobenzoate; the same 2-chloromuconate was identified in the reaction mixtures of resting cells pre-grown on 2-chlorobenzoate and exposed to 3-chloro- and 2,3-dichlorobenzoate, while in similar experiments 1,2-dihydroxy-3,5-dichloro-cyclohexa-3,5-dienoate was detected as dead-end product of 3,5-dichlorobenzoate co-metabolism. These results suggest an initial degradative attack by 2-chlorobenzoate induced dioxygenase(s). The role of 3,5-dichlorobenzoate as an antagonist of 2-chlorobenzoate degradation was also studied: in the presence of mixed 2-chloro- and 3,5-dichlorobenzoate, the 3,5-dichlorobenzoate preferential uptake by the resting cells of the chlorobenzoate-degrading consortium was observed. 2-Chlorobenzoate entered the cells only after the complete removal of the co-substrate. In growing cells experiments, the addition of 1,2-dihydroxy-3,5-dichloro-cyclohexa-3,5-dienoate, the 3,5-dichlorobenzoate co-metabolite, to 2-chlorobenzoate exerted the same antagonistic effect of the parent compound, inhibiting both the microbial growth and the degradative process. These data are discussed, allowing us to attribute the inhibitory effects observed to a substrate/co-substrate competition, though other additional causes may not be totally excluded.