title: Thymidine-Dependent Staphylococcus aureus Small-Colony Variants Are Induced by Trimethoprim-Sulfamethoxazole (SXT) and Have Increased Fitness during SXT Challenge.
abstract: Trimethoprim-sulfamethoxazole (SXT) is a possible alternative for the treatment of community- and hospital-acquired methicillin-resistant Staphylococcus aureus (MRSA) due to the susceptibility of most MRSA strains to the drug. However, after long-term treatment with SXT, thymidine-dependent (TD) SXT-resistant small-colony variants (SCVs) emerge. In TD-SCVs, mutations of thymidylate synthase ([TS] thyA) occur. Until now, it has never been systematically investigated that SXT is triggering the induction and/or selection of TD-SCVs. In our study, we performed induction, reversion, and competition experiments in vitro and in vivo using a chronic mouse pneumonia model to determine the impact of SXT on the emergence of TD-SCVs. SCVs were characterized by light and transmission electron microscopy (TEM) and auxotrophism testing. Short-term exposure of S. aureus to SXT induced the TD-SCV phenotype in S. aureus SH1000, while selection of TD-SCVs with thyA mutations occurred after long-term exposure. In reversion experiments with clinical and laboratory TD-SCVs, all revertants carried compensating mutations at the initially identified mutation site. Competition experiments in vitro and in vivo revealed a survival and growth advantage of the ΔthyA mutant under low-thymidine availability and SXT exposure although this advantage was less profound in vivo. Our results show that SXT induces the TD-SCV phenotype after short-term exposure, while long-term exposure selects for thyA mutations, which provide an advantage for TD-SCVs under specified conditions. Thus, our results further an understanding of the dynamic processes occurring during SXT exposure with induction and selection of S. aureus TD-SCVs.
authors.affiliation: Institute of Medical Microbiology, University Hospital of Münster, Münster, Germany.
authors.forename: Andre
authors.initials: A
authors.lastname: Kriegeskorte
authors.affiliation: Infections and Cystic Fibrosis Unit, Division of Immunology, Transplantation and Infectious Diseases, San Raffaele Scientific Institute, Milan, Italy.
authors.forename: Nicola Ivan
authors.initials: NI
authors.lastname: Lorè
authors.affiliation: Infections and Cystic Fibrosis Unit, Division of Immunology, Transplantation and Infectious Diseases, San Raffaele Scientific Institute, Milan, Italy.
authors.forename: Alessandra
authors.initials: A
authors.lastname: Bragonzi
authors.affiliation: Infections and Cystic Fibrosis Unit, Division of Immunology, Transplantation and Infectious Diseases, San Raffaele Scientific Institute, Milan, Italy.
authors.forename: Camilla
authors.initials: C
authors.lastname: Riva
authors.affiliation: Institute of Medical Microbiology, University Hospital of Münster, Münster, Germany.
authors.forename: Marco
authors.initials: M
authors.lastname: Kelkenberg
authors.affiliation: Institute of Medical Microbiology, University Hospital of Münster, Münster, Germany.
authors.forename: Karsten
authors.initials: K
authors.lastname: Becker
authors.affiliation: Department of Microbiology and Immunology, University of Wisconsin-Madison, Madison, Wisconsin, USA.
authors.forename: Richard A
authors.initials: RA
authors.lastname: Proctor
authors.affiliation: Institute of Medical Microbiology, University Hospital of Münster, Münster, Germany Interdisciplinary Centre for Clinical Research, University of Münster, Münster, Germany.
authors.forename: Georg
authors.initials: G
authors.lastname: Peters
authors.affiliation: Institute of Medical Microbiology, University Hospital of Münster, Münster, Germany kahl@uni-muenster.de.
authors.forename: Barbara C
authors.initials: BC
authors.lastname: Kahl
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000009:adverse effects*; D001426:Bacterial Proteins / Q000235:genetics* / Q000378:metabolism; D002908:Chronic Disease; D004195:Disease Models, Animal; D024881:Drug Resistance, Bacterial / Q000187:drug effects* / Q000235:genetics; D015870:Gene Expression; D056084:Genetic Fitness / Q000187:drug effects; D008297:Male; D055624:Methicillin-Resistant Staphylococcus aureus / Q000187:drug effects* / Q000201:enzymology / Q000235:genetics; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D018410:Pneumonia, Bacterial / Q000188:drug therapy; D012641:Selection, Genetic / Q000187:drug effects; D013203:Staphylococcal Infections / Q000188:drug therapy / Q000382:microbiology; D013936:Thymidine / Q000378:metabolism; D013940:Thymidylate Synthase / Q000172:deficiency / Q000235:genetics*; D015662:Trimethoprim, Sulfamethoxazole Drug Combination / Q000009:adverse effects*
chemical_list: D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D015662:Trimethoprim, Sulfamethoxazole Drug Combination; D013940:Thymidylate Synthase; D013936:Thymidine