The importance of bacterial multiplication in the selection, by oxytetracycline-HCl, of oxytetracycline-resistant bacteria in marine sediment microcosms

Small-scale microcosms were used to investigate the impact of oxytetracycline-HCl on both metabolic processes and the emergence of elevated frequencies of resistance in marine sediment microflora, The microcosms were designed to simulate the environment of sediments found under fish farms, and included marine mud, fish feed and sea water. Measurements of the rate of changes in gas pressure in the sealed microcosms were used as a proxy indicator of metabolic activity. In these microcosms, the sediment was the most significant source of gas-generating bacteria and the feed provided the major substrate for gas production. The frequency of resistance was monitored by differential plate counts on 2216 V media with and without 25 mu g ml(-1) oxytetracycline-HCl, After a 2-3-day adaptation phase all microcosms entered a steady-state phase during which the increase in gas pressure was linear with respect to time. The addition of oxytetracycline-HCl (0.1-600 mu g g(-1)) resulted in an increase in the length of the adaptation phase but had no effect on the rate of gas pressure increase in the steady-state phase. The frequency of oxytetracycline-resistant colony-forming units was determined after 194 h incubation of the microcosms. The frequency in microcosms containing over 25 mu g g(-1) was 90%; in those containing 6.25 mu g g(-1) or less the frequency was 7%. Investigations revealed that there was a rapid increase in cell numbers only during the first 2-3 days of the incubation, and that the majority of the increase in resistance frequency occurred during this time. If the addition of oxytetracycline-HCl was delayed until after 3 days incubation little selection for increased resistance frequency occurred.These data are compared with the data from other microcosm studies of the impact of oxytetracycline on marine sediment processes. It is suggested that the extent of cell division achieved in such systems is a critical parameter determining the results that they generate.