Genomic Basis of Occurrence of Cryptic Resistance among Oxacillin- and Cefoxitin-Susceptible <i>mecA</i> -Positive Staphylococcus aureus

Abstract

The oxacillin- and cefoxitin-susceptible <i>mecA</i>-positive Staphylococcus aureus is a novel "stealth" methicillin-resistant S. aureus (MRSA) type. Here, we sequenced the whole genome of two oxacillin- and cefoxitin-susceptible <i>mecA</i>-positive MRSA isolates from breast abscesses in a lactating woman and a nasal swab of a healthy student in Guangzhou for investigating the mechanism underlying its occurrence. The reversion of these isolates was selected by exposure to sub-MICs of cefoxitin with or without mupirocin. The <i>mecA</i> expression of both parental strains and their revertants was determined, and the whole genome of the revertants was sequenced. Comparative whole-genome analyses performed for both strains revealed that <i>mecA</i> of the clinical strain was mutated by a single-bp insertion at the 262nd position in the tandem repeat region of the gene, and this mutation that led to the formation of a premature stop codon. The colonizing strain was mutated by a novel G-to-A base substitution in the second promoter region (-35 bp) of <i>mecA</i>. The <i>mecA</i> expression level of strain 697 revertant was 37 times higher than that of the parental strain. Although the <i>mecA</i> expression level was even higher for parental strain 199 compared with that for its revertant, its cDNA sequence contained a single-bp insertion. Collectively, both the missense and single substitution mutations of the second promoter of <i>mecA</i> could render MRSA isolates as "stealth" MRSA, thereby emphasizing the importance of combining phenotype tests with <i>mecA</i> or penicillin-binding protein 2a detection for the identification of MRSA. <b>IMPORTANCE</b> The oxacillin- and cefoxitin-susceptible <i>mecA</i>-positive Staphylococcus aureus is a novel type of "stealth" methicillin-resistant S. aureus (MRSA), which is difficult to be detected using conventional methods. To investigate the genomic basis of their occurrence, we sequenced the whole genome of two previously recovered oxacillin- and cefoxitin-susceptible <i>mecA</i>-positive MRSA isolates from breast abscesses in a lactating woman and a nasal swab of a healthy student in Guangzhou. Complete SCC<i>mec</i> structure was absent except for <i>mecA</i> in clinical isolate 199. Additionally, a novel single-base pair insertion was observed in the clinical strain, which resulted in premature termination and a frameshift mutation. The colonizing isolate 697 had a Scc-<i>mec</i>-type IVa, and the second promoter region (-35 bp) of <i>mecA</i> was mutated by a novel G-to-A base substitution. The reversion of oxacillin- and cefoxitin-susceptible <i>mecA</i>-positive S. aureus to resistant MRSA isolates was selected by exposure to subminimum inhibitory cefoxitin with or without mupirocin.