Clinical relevance of topical antibiotic use in co-selecting for multidrug-resistant Staphylococcus aureus: Insights from in vitro and ex vivo models

Abstract

Topical antibiotic preparations, such as fusidic acid (FA) or mupirocin, are used in the prevention and treatment of superficial skin infections caused by staphylococci. Previous genomic epidemiology work has suggested an association between the widespread use of topical antibiotics and the emergence of methicillin resistant Staphylococcus aureus in some settings. In this study, we provide experimental proof of co-selection for multidrug resistance in S. aureus following exposure to FA or mupirocin. Through targeted mutagenesis and phenotypic analyses, we confirmed that fusC carriage confers resistance to FA, and mupA carriage confers high-level resistance to mupirocin in multiple S. aureus genetic backgrounds. In vitro experiments demonstrated that carriage of fusC and mupA confer a competitive advantage in the presence of sub-inhibitory concentrations of FA and mupirocin, respectively. Further, we used a porcine skin colonisation model to show that clinically relevant concentrations of topical antibiotics can co-select for presence of unrelated antimicrobial resistance determinants, such as mecA, blaZ, and qacA, in fusC or mupA harbouring S. aureus These findings provide valuable insights on the role of acquired FA or mupirocin resistance in co-selecting for broader antibiotic resistance in S. aureus, prompting greater need for judicious use of topical antibiotics.

FIG 1

In vitro competition assays of S. aureus reveal the selective advantage of topical antibiotic resistance gene carriage. S. aureus strains NZ14132 (orange), NZ14487 (aqua), and NZAK3 (green), wild-type or complemented strains, were paired with their respective isogenic mutants under nonselective conditions and exposure to a sub-MIC level of FA (0.03125 mg/liter) or mupirocin (0.125 mg/liter) for 7 days. (A to D) Percentages of wild-type or complemented isolates in mixed cultures of the wild type and fusC mutant (A), fusC complemented and fusC mutant (B), wild-type and mupA mutant (C), and mupA complemented and mupA mutant (D) strains were determined on days 1 and 7 postexposure. The mean percentages of three biological replicates are displayed for each condition tested, with black error bars representing the standard errors of the means (SEM). Statistically significant differences are indicated by asterisks (***, P < 0.001 by paired t test).

FIG 2

Ex vivo competition assays of S. aureus reveal the selective advantage of topical antibiotic resistance gene carriage in clinically relevant environments. S. aureus strains NZ14132 (orange), NZ14487 (aqua), and NZAK3 (green) wild-type or complemented strains paired with their respective isogenic mutants were grown on porcine skin under nonselective conditions and exposure to a single dose of 20 to 25 mg 2% Fucidin or 2% Bactroban ointment for 24 h. (A to D) Percentages of wild-type or complemented isolates within mixed cultures of wild-type and fusC mutant (A), fusC complemented and fusC mutant (B), wild-type and mupA mutant (C), and mupA complemented and mupA mutant (D) strains were determined at the conclusion of the assays. Five biological replicates were used to calculate the mean percentages and the SEM (black error bars) for each condition tested. Statistically significant differences are indicated by asterisks (**, P < 0.01; ***, P < 0.001, paired t test).