Annals of Microbiology 57 2273276 2007Detection of methicillinr - PDF document

Annals of Microbiology, 57 (2)273-276 (2007)Detection of methicillin-resistance (mec-A) gene in Staphylococcus aureusstrains by PCR and determination of antibiotic susceptibilityAlireza ZAMANI1*, Siavash SADEGHIAN2, Jalal GHADERKHANI2, Mohammad Yousef ALIKHANI2, Mohammad NAJAFIMOSLEH2 * Corresponding author. Phone: +98 811 8276295-8 ext 338; DNA extraction. A single colony was taken from eachblood agar plate which had been incubated overnight andemulsified into 100 l of phosphate buffer salt. After incu-et al.,2002).PCR for mec-Agene detection. The mec-Agene wasamplified with the following two oligonucleotides: forward1276 (5´-AAA ATC GAT GGT AAA GGT TGG C-3)and backward primer 1787 (5-AGT TCT GGA GTA CCG GATTTG C -3) which gave a PCR product of 533 bp (Merlino etal.,2002). The PCR was performed with an initial denatu-ration step of 5 min 94 ºC, followed by 35 cycles of 45 s 942006). Antibiotic-resistance tests. The isolated S. aureusstrains were tested for resistance to antimicrobial agentset al.,2002).RESULTSThe results of our study revealed that the S. aureusinfec-tions were seen frequently in urine culture (62.9%) andthen in wounds (21.5%), blood culture (5.7%), pleural fluidThe PCR analysis of 70 clinical S

. aureusisolates formec-Ashowed 35 (50%) cases were positive and 35 (50%)negative and known as MRSA and MSSA strains, respec-DISCUSSIONRapid and accurate detection of methicillin resistance S.aureusstrains is imperative for appropriate patients treat-ment and implementation of institutional programs forrecognition and management of MRSA outbreaks and crosset al.,2001).Consequently, many methods were developed over recentet al., 2002). We examined how a PCR assay thatdiscriminates MRSA from MSSA can improve patients careS. aureusandthe second to determine antibiotic susceptibility (Hallin etal.,2003).According to literatures, the first methicillin-resistantstrain of S. aureus(MRSA) was isolated in Europe in 1961(Enright et al.,2002). Then, there has been a steadyincrease in the prevalence of MRSA isolated from hospitalS. aureusweremethicillin-resistant in 1997 (Jenison et al.,2000). Twostudies in Shiraz, Iran, showed MRSA had risen up fromet al.,2000) to 43% (Japoni et al.,2004).As shown in the Table 1, vancomycin has remained themainstay of treatment for serious MRSA infections. But withS. aureusisolates must havegot the penicillin resistant gene by plasmid transformationet al.,1992). In addition, the MRSA were moreresistance to cloxac

illin, tetracycline and more sensitive toTwo MRSA isolates showed a decreased susceptibility tovancomycin which is called vancomycin-intermediate S.aureus(VISA) strain. For the first time VISA strain hasbeen described for clinical isolates of MRSA in Japan inet al.,1997) then in United States (CDC,1997) and Europe (Ploy et al.,1998) and mainly frompatients on vancomycin therapy. It is recommended, alter-In conclusion, MRSA is increasing and reaching to thecritical situation and continues to be a major cause of seri- 274A. Zamani et al. REFERENCESAlborzi A., Pourabbas B.A., Salehi H., Pourabbas B.H., Obiidi B.,Panjehshahin M.R. (2000). Prevalence and pattern of antibi-otic sensitivity of methicillin-sensitive and methicillin-resist-Staphylococcus aureusin Shiraz, Iranian. J. Med. Sci., 25(1, 2): 1-8.Boyce J.M. (1998). Are the epidemiology and microbiology ofmethicillin-resistance Staphylococcus aureuschanging? J.Am. Med. Associat., 279 (8): 623-624.CDC - Centers for Disease Control and Prevention (1997).Staphylococcus aureuswith reduced susceptibility to van-comycin - United States. JAMA, 278 (11): 891-892.Enright M.C., Robinson D.A., Randle G., Feil E.J., Grundmann H.,Spratt B.G. (2002). The evolutionary history of methicillin-Staphylo

coccus aureus(MRSA). PNAS, 99 (11):7687-7692.Hallin M., Maes N., Byl B., Jacobs F., De Gheldre Y., StruelensM.J. (2003). Clinical impact of a PCR assay for identificationof Staphylococcus aureusand determination of methicillinresistance directly from blood cultures. J. Clin. Microbiol., 41Hiramatsu K., Hanaki H., Ino T., Yabuta k., Oguri T., TenoverF.C. (1997). Methicillin-resistant Staphylococcus aureusclin-ical strain with reduced vancomycin susceptibility. J.Japoni A., Alborzi A., Rasouli M., Pourabbas B. (2004).Modification DNA extraction for rapid PCR detection of methi-Jenison R., Heaberil A., Yang S., Polisky B., Ostrosf R. (2000).Thin film bio senser for rapid detection of mec-A from methi-Staphylococcus aureus. Clin Chem., 46 (9):1501-1504. Ann. Microbiol., 57 (2), 273-276 (2007)275 TABLE 1 - Antibiotic resistance profiles of MRSA and MSSA by DDA IsolatesOx/MetCipErytTetGentSTXVanCfCzCxRaPMRSA2214242615242161832435(n=35)(62.8%)(40%)(68.5%)(74.2%)(42.8%)(68.5%)(5.7%)(45.7%)(51.4%)(91.4%)(11.4%)(100%)MSSA0066140213135 (n=35)(0%)(0%)(17.1%)(17.1%)(2.9%)(11.4%)(0%)(5.7%)(2.9%)(8.9%)(2.9%)(100%) 2214303216282181935570 FIG. 1 - Agarose gel electrophoresis analysis for the .J. Burn Care Rehabil., 13 (2): 293-297. 276A. Zamani et al.