Application of in-vitro models: development of resistance.

The development of chromosomal beta-lactam resistance of Escherichia coli, Enterobacter cloacae and Citrobacter freundii was observed by following the bacterial kill-kinetics in an in-vitro model simulating human serum antibiotic concentrations. From sensitive Escherichia coli cells mutants arose resistant to aminopenicillins, and to first and second generation cephalosporins, whereas with the Ent. cloacae and Citro. freundii mutants were also resistant to cefotaxime. Resistant mutants from all three species could also be selected on antibiotic-containing agar plates. With E. coli they occurred in two steps, the first mutation being stable but the second mutation reverting spontaneously. In Ent. cloacae, and apparently also in Citro. freundii, the mutation changes the cephalosporinase production from an inducible to a constitutive one. The different mutation rates, and the rate was extremely high for some Ent. cloacae strains, were correlated with a corresponding reduction of viable cell count and with the time of re-growth observed in the model.