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

Penicillin tolerance is an incompletely understood phenomenon that allows b acteria to resist drug-induced killing. Tolerance was studied with independ ent Streptococcus gordonii mutants generated by cyclic exposure to 500 time s the MIC of penicillin. Parent cultures lost 4 to 5 log(10) CFU/mL of viab le 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 peni cillin-binding proteins but contained increased amounts of two proteins wit h respective masses of ca, 50 and 45 kDa. One mutant (Toll) was further cha racterized, The two proteins showing increased levels were homologous to th e arginine deiminase and ornithine carbamoyl transferase of other gram-posi tive bacteria and were encoded by an operon that was >80% similar to the ar ginine-deiminase (arc) operon of these organisms. Partial nucleotide sequen cing and insertion inactivation of the S. gordonii are locus indicated that tolerance was not a direct consequence of arc alteration. On the other han d, genetic transformation of tolerance by Toll DNA always conferred are der egulation In nontolerant recipients, are was repressed during exponential g rowth and up-regulated during postexponential growth. In tolerant transform ants, are was constitutively expressed. Toll DNA transformed tolerance at t he same rate as transformation of a point mutation (10(-2) to 10(-3)). The tolerance mutation mapped on a specific chromosomal fragment but was physic ally distant from are, Importantly, are deregulation was observed in most ( 6 of 10) of additional independent penicillin-tolerant mutants. Thus, altho ugh not exclusive, the association between arc deregulation and tolerance w as not fortuitous. Since penicillin selection mimicked the antibiotic press ure operating in the clinical environment, are deregulation might be an imp ortant correlate of naturally occurring tolerance and help in understanding the mechanism(s) underlying this clinically problematic phenotype.