R plasmid transfer of antibiotic resistance in a wastewater treatment plant

Enteric bacteria were examined for ability to transfer antibiotic resistance in a wastewater treatment plant. Resistant Salmonella enteritidis, Proteus mirabilis, and Escherichia coli were isolated from clinical specimens and from primary (1 degree) sewage effluent. Resistance to ampicillin (Ap), chloramphenicol (Cm), streptomycin (Sm), sulfadiazene (Su), and tetracycline (Tc) was demonstrated by spread plate and tube dilution techniques. Plasmid mediation of resistance was shown by acridine orange, dodecyl sodium sulfate, and ethidium bromide curing, and by direct cell transfer. Each donor was mated with sensitive E. coli and Shigella sonnei. Mating pairs (and recipient conrols) were suspended in unchlorinated, (1 degree) effluent that had been filtered and autoclaved. Suspensions were added to membrane diffusion chambers which were then placed in the primary (1 degree) and secondary (2 degree) settling tanks of the wastewater treatment plant. Resistant recombinants were detected by replica plating nutrient agar master plates onto XLD agar plates that contained either 10 micrograms Ap, 30 micrograms Cm, 10 micrograms Sm, 100 micrograms Sy or 30 micrograms Tc per ml of medium. Mean transfer frequencies for laboratory matings were 2.1 x 10 -5, while in situ matings for 1 degree and 2 degree settling were 4.3 and 7.5 x 10-5, respectively. These values revealed that a significant level of resistance transfer may occur in wastewater treatment plants. Epidemiological significance was discussed