Temperature-sensitive mutants ofStaphylococcus aureus: Isolation and preliminary characterization

Abstract

Temperature-sensitive (ts) mutants ofStaphylococcus aureus were isolated after mutagenesis with nitrosoguanidine and two cycles of enrichment with Penicillin G and D-Cycloserine. The mutants expressed tight, coasting, and leaky phenotypes on solid media. In broth, however, most exhibited coasting for a limited number of generations. The reversion frequency of selected ts mutants was less than 10(-6). Intraperitoneal (i.p.) immunization with ts mutant G/1/2 conferred significant protection (0 dead/6 total vs. 7/7, immunized vs. control; p=0.0006) from lethal i.p. challenge with the parental wild-type (wt)S. aureus suspended in 5% porcine mucin, performed 28 days after i.p. administration of 10(8) colony-forming units. Protection induced by mutants of coasting phenotype was higher and lasted longer than that induced by mutants of the tight phenotype. The results of this study demonstrate that ts mutants ofS. aureus can be obtained and that ts mutants are able to induce protective immunity from subsequent challenge with the parental wt strain.

Intramammary immunization with live-attenuated Staphylococcus aureus: microbiological and immunological studies in a mouse mastitis model

“…The S. aureus 8325-4 (phi-111 strain (the wt strain) used in this study was generously provided by Dr. John J. Iandolo (Department of Pathology, College of Veterinary Medicine, Kansas State University, Manhattan, KS). Temperature-sensitive derivatives were obtained from the wt strain as described earlier [3]. The mutant used in this study (A523) replicates well at low temperatures (below 32°C) but undergoes a limited number of divisions when transferred to the mammalian body temperature.…”

“…R4) as described elsewhere [5]. We have obtained temperature-sensitive (tsl mutants of S. aureus and have shown that parenteral immunization with ts-S. aureus confers protection from intraperitoneal (i.p.> challenge with the parental wild-type (wt) S. aureus [3]. We hypothesize that stimulation of the mucosal immune system with a live attenuated vaccine would be a valid approach to prevention of the disease in lactating cows.…”

Intramammary immunization with live-attenuated Staphylococcus aureus protects mice from experimental mastitis

“…Live‐attenuated, temperature‐sensitive derivatives were obtained by mutagenesis of S. aureus 8325‐4 phi11 (the wt strain) following a procedure described elsewhere [10]. The attenuated mutant used in this study (A523) replicates well at low temperature (below 30°C) but undergoes a limited number of replications when transferred to the mammalian body temperature.…”

Induction of Cell-Mediated Immunity toStaphylococcus aureusin the Mouse Mammary Gland by Local Immunization with a Live Attenuated Mutant

“…Live attenuated, temperature-sensitive derivatives were obtained from the wild-type strain by chemical mutagenesis with nitrosoguanidine (Sigma Chemical Co., St. Louis, Mo.) as described previously (29). The attenuated mutant used in this study (A523) replicates well at low temperatures (below 30°C) and only undergoes limited replication when transferred to the mammalian body temperature.…”