Identification and characterization of methicillin-resistant coagulase-negative staphylococci from bovine mastitis

This study focused on the correlation between geno- and phenotypic tests in the correct assessment of mecA-mediated methicillin resistance among coagulase-negative staphylococci (CoNS) and the further characterization of mecA-positive isolates. A total of 121 CoNS from cases of bovine mastitis were investigated for oxacillin susceptibility by disc diffusion and broth microdilution. Isolates classified as methicillin resistant by either method were tested by PCR for the mecA gene and the SCCmec type. The cefoxitin disc test was also applied. PFGE served to determine the genetic relationships of the resistant isolates. Sixteen isolates were classified as methicillin resistant and 96 isolates as methicillin susceptible by both methods. The mecA gene was identified in 15 of the 16 resistant isolates. Nine mecA-negative isolates showed oxacillin MICs of 0.5 or 1 mg/L, oxacillin zone sizes of 18-23 mm and were classified as methicillin susceptible in the cefoxitin disc test. SCCmec cassettes of types V (five Staphylococcus haemolyticus), III (one Staphylococcus saprophyticus), IV (five Staphylococcus epidermidis, one Staphylococcus capitis) and IV with an additional ccrA4/B4 gene (two S. epidermidis) were seen, while one S. epidermidis carried a non-typeable SCCmec element (mec complex B + no ccr gene complex detected). All isolates with SCCmec type IV or non-typeable cassettes exhibited low oxacillin MICs of 1-4 mg/L, whereas isolates with type III or V cassettes had MICs of >= 16 mg/L. CoNS with oxacillin MICs of 0.5 and 1 mg/L should be confirmed for the presence of mecA before reporting them as methicillin resistant.