Properties of spontaneous Enterobacter cloacae mutants with temperature-conditional derepression of Type I β-lactamase synthesis

Abstract

Spontaneous mutants, with temperature-conditional derepression of chromosomally-encoded Type I beta-lactamase synthesis, were derived from two independent clinical isolates of Enterobacter cloacae. At the permissive temperature (28 degrees C) the mutants' beta-lactamase activity was equivalent to that of their respective parents but at restrictive temperatures (above 40 degrees C) the activity increased many hundred-fold. The increased beta-lactamase expression correlated with reduced beta-lactam susceptibility. In temperature shift-up experiments, the initial rate of beta-lactamase synthesis closely paralleled that of the parent strains induced with cefoxitin. Maximal beta-lactamase activity in the mutants was attained after about 3 h growth at restrictive temperatures and was significantly higher than that of the cefoxitin-induced parents. However, the level was not as high as that observed in isogenic temperature-stable derepressed mutants, under the same conditions. All temperature-conditional mutants showed hyper-induction of beta-lactamase synthesis at permissive temperatures. Our findings are discussed in relation to a positive control model for regulation of Type I beta-lactamase synthesis in Ent. cloacae.

Altered phenotypes associated with ampD mutations in Enterobacter cloacae

“…Although it had been originally hypothesized that the lesion resided in ampR (4), this theory arose before the existence of ampD was known. In general, the results of our studies on P-lactamase expression in E. cloacae 029 and 029-5 are in agreement with those published by Curtis et al (4). The one major discrepancy concems the inducibility of wild-type E. cloacae 029 at 42°C, which was found to be due to differences between the methods used for induction.…”

“…The one major discrepancy concems the inducibility of wild-type E. cloacae 029 at 42°C, which was found to be due to differences between the methods used for induction. Curtis et al incubated cells at 28°C and switched them to 41°C immediately before adding inducer (4). Cells were at 42°C for 1.5 h prior to exposure to inducer in our study.…”

“…These phenotypes include semiconstitutive to constitutive (11,14,17,20), hyperinducible (20), and temperature-sensitive (4) expression.…”

“…High-level constitutive or semiconstitutive production of the enzyme results from mutations within ampD (17,24), while mutations within ampG produce basal, constitutive expression of the enzyme (14). Recently, Lindquist et al (20) of wild-type strains, while expression at restrictive temperatures was similar to that seen with ampD mutants (4). In 1988, Goering et al (6) described a hyperinducible phenotype in E. cloacae 1194E that was thought to be due to an unusual ampR gene.…”

“…However, subsequent studies with this strain revealed a normal ampR ampC region upon recloning and suggested that the abnormal phenotype resulted from a mutation within the ampD region (15). Therefore, studies were designed to confirm that the hyperinducible phenotype in E. cloacae 1194E was due to a mutation within ampD and determine whether the temperature-sensitive phenotype described by Curtis et al (4) was also due to an ampD mutation.…”