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Staphylococcus aureus  ( S. aureus ), including methicillin-resistant  S. aureus  (MRSA), is one of the most important human pathogens, which is responsible for bacteremia, soft-tissue infections, and food poisoning. Hence, multiple cross displacement amplification (MCDA) is employed to detect all  S. aureus  strains, and differentiates MRSA from methicillin-sensitive  S. aureus . Multiplex MCDA (m-MCDA), which targets the  nuc  gene ( S. aureus -specific gene) and  mecA  gene (encoding penicillin-binding protein-2′), could detect  S. aureus  strains and identify MRSA within 85 min. Detection of the m-MCDA products is achieved using disposable lateral flow biosensors. A total of 58 strains, including various species of Gram-positive and Gram-negative strains, are used for evaluating and optimizing m-MCDA assays. The optimal amplification condition is found to be 63°C for 40 min, with detection limits at 100 fg DNA/reaction for  nuc  and  mecA  genes in the pure cultures, and 10 CFU/tube for  nuc  and  mecA  genes in the blood samples. The analytical specificity of m-MCDA assay is of 100%, and no cross-reactions to non- S. aureus  strains are produced according to the specificity testing. Particularly, two additional components, including AUDG enzyme and dUTP, are added into the m-MCDA amplification mixtures, which are used for eliminating the unwanted results arising from carryover contamination. Thus, the m-MCDA technique appears to be a simple, rapid, sensitive, and reliable assay to detect all  S. aureus  strains, and identify MRSA infection for appropriate antibiotic therapy.

frontiers in microbiology

Multiple Cross Displacement Amplification Coupled With Nanoparticles-Based Lateral Flow Biosensor for Detection of Staphylococcus aureus and Identification of Methicillin-Resistant S. aureus