Continuous cultures of Pseudomonas putida mt-2 overcome catabolic function loss under real case operating conditions

Abstract

The long-term performance and stability of Pseudomonas putida mt-2 cultures, a toluene-sensitive strain harboring the genes responsible for toluene biodegradation in the archetypal plasmid pWW0, was investigated in a chemostat bioreactor functioning under real case operating conditions. The process was operated at a dilution rate of 0.1 h(-1) under toluene loading rates of 259 +/- 23 and 801 +/- 78 g m(-3) h(-1) (inlet toluene concentrations of 3.5 and 10.9 g m(-3), respectively). Despite the deleterious effects of toluene and its degradation intermediates, the phenotype of this sensitive P. putida culture rapidly recovered from a 95% Tol(-) population at day 4 to approx. 100% Tol(+) cells from day 13 onward, sustaining elimination capacities of 232 +/- 10 g m(-3) h(-1) at 3.5 g Tol m(-3) and 377 +/- 13 g m(-3) h(-1) at 10.9 g Tol m(-3), which were comparable to those achieved by highly tolerant strains such as P. putida DOT T1E and P. putida F1 under identical experimental conditions. Only one type of Tol(-) variant, harboring a TOL-like plasmid with a 38.5 kb deletion (containing the upper and meta operons for toluene biodegradation), was identified.