Assessing survival and detection of catechol-biodegrading strains in waste water microcosms

Catechol-degrading bacteria, Escherichia coli JM101 carrying pIB1343 (strain pIB1343) and Flavimonas orizihabitans KH1 (strain KH1), were chosen as model bacteria to estimate the survival and catechol-degradability in waste water microcosms. Before their application to ecosystem, survival and catechol degradation of these bacteria were investigated in waste water microcosms. It was found that strain pIB1343 adapted much faster to waste water environments and degraded catechol in the case of Namdong samples, whereas the strain KH1 degraded catechol much faster in Sihwa samples. When catechol was added to the microcosms, indigenous microorganisms in Sihwa samples used catechol as a carbon and energy source much better than those in Namdong samples. A modified filter extraction technique was used to obtain the high-yield purified DNA from 50 ml of waste water samples and the extracted DNA (polymorphic DNA [20-23 kb]) was of sufficient quantity and quality for the amplification. PCR was performed with catA-specific primers, C12OU and C12OL, which specifically detected the catA gene encoding catechol 1,2-dioxygenase in waste water microcosms, and then the 320 bp PCR products were amplified. PCR products were quantified by densitometer. Using the standard curve of detection limit by catA-specific PCR products, the number of catA genes for their corresponding intensities of PCR products was obtained. The number of total catA genes PCR in waste water microcosms was correalated with catechol degradation.