Molecular analysis of chloramphenicol resistance in staphylococci

Abstract

Staphylococci are among the most important bacterial pathogens. S. aureus is a major cause of both nosocomial and community-acquired infections. Coagulase-negative staphylococci (CNS) are important nosocomial pathogens causing infections of prosthetic implants and devices. Antimicrobial resistance is an important problem for the therapy of staphylococcal infections. Staphylococci, especially methicillin-resistant staphylococci (MRS) are resistant to most antimicrobial agents, including chloramphenicol. Chloramphenicol is a broad-spectrum antibiotic that causes a bacteriostatic effect by binding to 50s subunit of ribosomal RNA and it inhibits the elongation of protein synthesis. The most important mechanism of chloramphenicol resistance in staphylococci involved chloramphenicol acetyltransferase (CAT), which acetylated the 3-hydroxy group of chloramphenicol, so the acetylated chloramphenicol cannot bind to the target site. The CAT enzymes are encoded by cat genes on plasmids. The staphylococcal cat-encoded plasmids can be divided into 3 groups, based on sequence homology: pC221, pC223 and pC194 groups. In this study, A total of 149 clinical staphylococcal isolates, which were divided into 4 groups based on methicillin susceptibility, including methicillin-resistant S. aureus (MRSA), methicillin-sensitive S. aureus (MSSA), methicillin-resistant coagulase-negative staphylococci (MRCNS) and methicillin-sensitive coagulase-negative staphylococci (MSCNS) were studied. All of the 149 staphylococcal isolates exhibited the chloramphenicol minimum inhibitory concentration (MIC), between 16-128 mg/ml. These staphylococcal isolates were resistant to chloramphenicol, according to the recommendation of NCCLS. The cat genes of all 149 staphylococcal isolates were detected by multiplex PCR. The pC221-related cat gene was observed in 2.9%, 80%, 70% and 25% of MRSA, MRCNS, MSSA and MSCNS, respectively, whereas pC223-related cat gene was found in 20%, 30% and 75% of MRCNS, MSSA and MSCNS, respectively. Surprisingly, the pC223-related cat gene was not found in MRSA, whereas the pC194-related cat gene was solely observed in MRSA, with a frequency of 95.6%. However, there is one isolate of MRSA (MRSA505) in which the cat gene could not be detected by PCR. The unique distribution of cat-specific plasmid suggests a pattern of transmission of genetic material between specific groups of bacteria that may relate to the drug resistance in some bacterial isolates. The location of the cat gene of 20 random isolates of each cat-encoded plasmid groups were studied by Southern blot hybridization. The cat genes of each cat-encoded plasmid group were observed only on plasmids. This indicated the potential of the staphylococcal cat gene in the spread of itself among staphylococcal isolates via mechanisms of gene transfer.