RHIZOSPHERE EFFECTS ON BIODEGRADATION OF 2,5-DICHLOROBENZOATE BY A BIOLUNINESCENT STRAIN OF ROOT-COLONIZING PSEUDOMONAS FLUORESCENS

A recombinant strain of bioluminescent Pseudomonas fluorescens 2-79 RL D containing a catabolic pathway for degradation of 2,5-dichlorobenzoa te (2,5-DCB) was monitored in soil microcosms to, examine the influenc e of plants on its growth and activity in a contaminated soil. Recombi nant P. fluorescens 2-79 RLD was generated by mating a versatile chlor obenzoate utilizer, P. putida P111, containing plasmid pPB111, with a bioluminescent strain of P. fluorescens that had been transformed prev iously with a Tn7-liuxCDABE marker. Plasmid pPB111 contains genes enco ding for a chlorobenzoate-1,2-dioxygenase that converts ortho-chlorobe nzoates to their corresponding catechols. DNA hybridization experiment s and cell-free extract assays with parental and recombinant P. fluore scens 2-79 RLD suggested that the reaction product of the plasmid pPB1 11 encoded chlorobenzoate dioxygenase was degraded by an endogenous ca techol dioxygenase in P. fluorescens 2-79. After introduction of P. fl uorescens 2-79 RLD into soil containing 10 mg kg(-1) 2,5-DCB, normally recalcitrant 2,5-DCB was degraded rapidly over a period of 2 to 4 day s in soil with plants. In contrast, 2,5-DCB disappearance in nonplante d soil was significantly slower, requiring 7 days in one experiment, a nd more than 2 weeks in a second experiment. Population numbers of the degrader were similar in planted and nonplanted soil for the first 7 days, but declined in nonplanted soils by day 14. Physiological status , measured using an assay based on lag-phase duration, was similar in planted and nonplanted soils at day 2, but rapidly declined in nonplan ted soil by day 7. At day 14, plasmid stability in P. fluorescens 2-79 RLD was significantly greater in rhizosphere soil; only 10% of P. flu orescens 2-79 RLD cells in rhizosphere soil had lost the ability to de grade 2,5-DCB, versus 94% of the cells in nonplanted soil. The plasmid also was transferred to indigenous bacteria in both planted and nonpl anted soils, as determined by the appearance of novel degraders. The r esults demonstrate that the presence of plants promoted rapid degradat ion of DCB and provided a niche that enhanced maintenance of plasmid p PB111 in the degrader bacterium.