Deregulation of the arginine deiminase (arc) operon in penicillin-tolerant mutants of Streptococcus gordonii

Penicillin tolerance is an incompletely understood phenomenon that allows bacteria to resist drug-induced killing. Tolerance was studied with independent Streptococcus gordonii mutants generated by cyclic exposure to 500 times the MIC of penicillin. Parent cultures lost 4 to 5 log(10) CFU/mL of viable counts/24 h, In contrast, each of four independent mutant cultures Lost less than or equal to 2 log(10) CFU/ml/24 h. The mutants had unchanged penicillin-binding proteins but contained increased amounts of two proteins with respective masses of ca, 50 and 45 kDa. One mutant (Toll) was further characterized, The two proteins showing increased levels were homologous to the arginine deiminase and ornithine carbamoyl transferase of other gram-positive bacteria and were encoded by an operon that was >80% similar to the arginine-deiminase (arc) operon of these organisms. Partial nucleotide sequencing and insertion inactivation of the S. gordonii are locus indicated that tolerance was not a direct consequence of arc alteration. On the other hand, genetic transformation of tolerance by Toll DNA always conferred are deregulation In nontolerant recipients, are was repressed during exponential growth and up-regulated during postexponential growth. In tolerant transformants, are was constitutively expressed. Toll DNA transformed tolerance at the same rate as transformation of a point mutation (10(-2) to 10(-3)). The tolerance mutation mapped on a specific chromosomal fragment but was physically distant from are, Importantly, are deregulation was observed in most (6 of 10) of additional independent penicillin-tolerant mutants. Thus, although not exclusive, the association between arc deregulation and tolerance was not fortuitous. Since penicillin selection mimicked the antibiotic pressure operating in the clinical environment, are deregulation might be an important correlate of naturally occurring tolerance and help in understanding the mechanism(s) underlying this clinically problematic phenotype.