Survival and detection of bacteria in an aquatic environment

Abstract

A genetically engineered plasmid, pPSA131, was used as a DNA probe to detect homologous DNA in Escherichia coli HB101(pPSA131) after it was mixed with aquatic microorganisms from Lake Mead, Nevada, water samples. An isolate from the pLAFR1 chromosomal library of Pseudomonas syringae Cit 7 was used to detect parent P. syringae Cit 7 that had been mixed with Lake Mead water. E. coli(pPSA131) was kept in variously treated samples of lake water or buffer, and its survival was measured by viable cell counting on modified Luria-Bertani (LB) agar. Full-strength LB agar proved better than 0.1 x LB agar at recovering E. coli(pPSA131) after survival in low-nutrient environments. Survival of E. coli(pPSA131) remained high in filtered (0.22-μm pore size) lake water and salts buffer on both selective and nonselective agars but was lower in untreated lake water or lake water filtered with a 0.8-μm-pore-size membrane. Total recoverable colonies grown on LB agar were higher when lake water was filter treated (0.8-μm pore size) than when lake water was untreated. Microorganisms recovered from lake water alone grew rapidly on nonselective media, probably because of the 'bottle effect'. After being mixed with Lake Mead water, E. coli(pPSA131) and P. syringae were detected by colony blotting with non-radioactively labeled DNA probes. E. coli((pPSA131) were recovered at three times during 48 h from variously treated samples of lake water and from a mixture with Lake Mead water organisms. Colonies were supported on either nonselective or selective agar for comparison. Target cells were best detected when a combination of selection and DNA probing was used and when there was no competition with native lake microorganisms. Only four colonies from Lake Mead were recovered on selective agar; when DNA probed, these colonies proved to be negative for the target sequence.