ISSN 2736-1756
Advanced Journal of Microbiology Research ISSN 2241-9837 Vol. 13 (3), pp. 001-005, March, 2019. © International Scholars Journals
Full Length Research Paper
Development and application of a novel multiplex polymerase chain reaction (PCR) assay for rapid detection of various types of staphylococci strains
Zhenbo Xu1,2, Lin Li1, Xihong Zhao3*, Jin Chu4, Bing Li1, Lei Shi1, Jianyu Su1* and Mark E. Shirtliff2,5
1College of Light Industry and Food Sciences, South China University of Technology, Guangzhou 510640, China.
2Department of Microbial Pathogenesis, Dental School, University of Maryland, Baltimore, MD 21201, USA.
3Key Laboratory for Green Chemical Process of Ministry of Education, School of Chemical Engineering and Pharmacy,
Wuhan Institute of Technology, Wuhan 430073, Hubei, China.
4School of Food Science and Nutrition, Faculty of Mathematics and Physical Sciences, University of Leeds, Leeds LS2 9JT, United Kingdom.
5Department of Microbiology and Immunology, School of Medicine, University of Maryland, Baltimore, MD 21201, USA.
Accepted 15 January, 2019
Abstract
In this study, a novel multiplex- polymerase chain reaction (PCR) for rapid detection of various staphylococci strains, including methicillin-resistant Staphylococcus aureus (MRSA), methicillin-sensitive Staphylococcus aureus, (MSSA) methicillin-resistant coagulase-negative staphylococci (MRCNS), methicillin-sensitive coagulase-negative staphylococci (MSCNS) and non-staphylococci strains, had been developed and applied. Six primers were specially designed on three target genes, which were mecA, 16S Ribosomal ribonucleic acid (rRNA) and femA. The specific amplification generated 3 bands on agarose gel, with sizes 374 bp for mecA, 542 bp for 16S rRNA and 823 bp for femA, respectively. The PCR product showed highest levels of resolution of DNA when 250 M of dNTP, primer concentration of mecA, 16S rRNA and femA reaching 1, 1 and 3 M respectively. No false positive amplification was observed, indicating the high specificity of the established multiplex PCR assay. Application of this multiplex-PCR had been further performed on detection for 262 MRSA and MRCNS strains with primers pairs M1 with M2 and F1 and F2. According to the results, multiplex-PCR results showed expected products for either MRSA or MRCNS strains, demonstrating the multiplex-PCR assays established in this study to be useful and powerful methods for differentiation of MRSA, MSSA, MRCNS, MSCNS and non-staphylococci strains.
Key words: Staphylococcus, multiplex-PCR, rapid detection.