Detection and Evaluation of Methicillin-Resistant Staphylococcus aureus by Duplex PCR

Abstract

Staphylococcusaureusis avirulence pathogenic bacterium.Detection of methicillin-resistant S. aureus(MRSA) using conventional culture and biochemical methods is labor and time consuming. Many MRSA isolates are heterogeneously resistant to ß-lactams, for example only 1 daughter cell out of 10 4 to 10 6 cells appears phenotypically resistant when routine antimicrobial susceptibility tests are performed. In this work two genes were used for detection of MRSA in urine, boil and nasal swabs using conventional PCR, mecA gene (533bp) were used for detection of methicillin resistant and femA gene (318bp) were used for S.aureus identification. It was found that not all resistant S. aureus tested with disk diffusion method carry the mecA gene that caused the resistant phenomena there were only 55% of all MRSA carrying mecA gene while femA gene gave 100% positive for S. aureus and this will lead us to appoint that penicillin-bindingprotein 2a (PBP2a) produced by mecA gene is not the only cause of resistant phenomena.

Characterization and Detection of mecA gene in Different Species of Staphylococcus, Streptococcus and Kocuria Which Isolated from Thalassemia Patients in Erbil City

“…Conventional PCR (Eppendorf/ Germany) was used to detect methicillin resistant gene mecA in all isolated bacteria. The primers were provided by macrogen and the forward primer sequence is 5 ' AAA ATC GAT GGT AAA GGT TGG C and with Reverse primer sequence 5 ' AGT TCT GCA GTA CCG GAT TTG C 3 ' ' with product size 533 bp ( 17). Macrogen synthesized the primer as lyophilized powder, thus the concentration define as pmol.…”

“…After that lyophilized blue pellet dissolved by vortexing. PCR performed for samples, procees in Initial denaturation at 95°C/5min , the thermal cycler for 30 cycles, Denaturation at 94°C/60sec Annealing at 63°C/60sec , Elongation at 72°C/60sec and Final elongation 72°C/5min then 4°C→∞ as mentioned [17].…”