PLASMID CONTENT EVALUATION OF ACTIVATED-SLUDGE

Plasmid content of activated sludge was evaluated by two different and complementary approaches. The first one consisted in the isolation of colonies from activated sludge samples, then isolated colonies were grown in nutrient broth and the plasmids were extracted from these cultures. The plasmid extraction procedure used a commercialized kit and the corresponding protocol. By this first approach, 40% of isolated colonies harboring plasmids were found. The second approach was an adaptation of the plasmid extraction kit directly on activated sludge samples. This direct application required several washing steps of sludges samples to avoid plasmid DNA digestion by DNases adsorbed within the sludge. This direct and rapid technique was validated in terms of reproducibility, specificity for plasmid DNA and losses. It allows the isolation of around 1 mu g of plasmid DNA per 10(11) cells in the sludge. The two approaches presented in this paper indicated that the plasmid content of activated sludge is important and its particular role within the wastewater epuration procedure should be investigated,