Plasmid profile and curing analysis of Pseudomonas aeruginosa as metal resistant

Abstract

The isolate Pseudomonas aeruginosa exhibited resistance to heavy metals such as cadmium, chromium, nickel and lead. Plasmid DNA was isolated from P. aeruginosa and designated as pBC15. The size of the plasmid DNA was approximately 23 kb. Escherichia coli DH5α was transformed with plasmid pBC15 subsequired resistance to nickel and ampicillin. The same size of the plasmid was isolated from E. coli transformant and separated on 0.7 % agarose gel electrophoresis. The restriction analysis of pBC15 showed that the plasmid DNA has single site for Bam HI and Eco RI and three sites for Xho I which were compared with 1 Kb DNA and λ Hind III digest molecular markers. Therefore, the size of the plasmid DNA of pBC15 was confirmed to be 23 kb. Curing was carried out by ethidium bromide, acridine orange, novobiocin, sodium dodechyl sulphate and elevated temperature (40 °C). Transformation and curing results suggest that nickel and ampicillin resistance gene was conferred by plasmid DNA. Cadmium resistant gene was present on chromosomal DNA along with the gene for chromium resistance. Lead resistance gene was shown to be present on the chromosomal DNA rather than the plasmid DNA as the cured and uncured cultures remained similar in lead resistance. Therefore, the ability of P. aeruginosa resistant to nickel and ampicillin is plasmid mediated and transferable to other strains whereas cadmium, chromium and lead could be chromosomal encoded. The heavy metal and antibiotic resistances of P. aeruginosa can be used to exploit for clean up industrial wastewater and bioremediation of heavy metal contaminated soil.