Survival of and lacZ expression in recombinant Pseudomonas strains introduced into river water microcosms

The lacZY gene cassette inserted into the chromosome of Pseudomonas aeruginosa UG2Lr and Pseudomonas aureofaciens Ps3732RNL11 was used as a genetic marker to study the fate of the Pseudomonas strains in river water microcosms. Expression of the lacZ marker in UG2Lr and Ps3732RNL11 was detected in microcosms containing as few as 12 and 14 colony-forming units (cfu)/mL of river water, respectively, by fluorimetric measurement of the beta-galactosidase activity against 4-methylumbelliferyl-beta-D-galactoside as the substrate. The persistence of and lacZ expression in the pseudomonas strains were monitored in sterile and nonsterile river water in the presence and absence of added nutrients by dilution plating and fluorimetry, respectively. After incubation for 10 d at 10 degrees C, culturable populations of strain UG2Lr in sterile water samples, with and without nutrient added, decreased from an initial density of 1.5 x 10(4) to 1.7 x 10(3) and 4.6 x 10(3) cfu/mL, respectively. Despite similar numbers of UG2Lr cells in the two treatments, expression of the lacZ marker in the surviving cells of the nutrient-supplemented sample was 24 times higher than in the cells of the unamended sample. In nonsterile water samples, the density of UG2Lr declined to less than 6 cfu/mL in 30 d regardless of the nutrient conditions. A nutrient supplement increased the growth of the native bacterial population but did not enhance growth of and lacZ expression in the bacteria. Polymerase chain reaction analysis showed a decrease in amplification signal indicating a genuine decline in viable UG2Lr cell density in the water samples.(ABSTRACT TRUNCATED AT 250 WORDS)