Abstract
BackgroundStaphylococcus aureus (S. aureus), including methicillin-susceptible S. aureus (MSSA) and methicillin-resistant S. aureus (MRSA), is an eminent human pathogen that can colonize the human host and cause severe life-threatening infections. The development of a reliable, simple and rapid assay for detecting S. aureus and identifying MRSA is important for diagnosis and follow-up treatment.MethodsA novel molecular diagnosis technique, named multiplex loop-mediated isothermal amplification linked to a nanoparticle-based lateral flow biosensor (m-LAMP-LFB), was applied to detect all S. aureus species and identify MRSA. Two sets of primers were designed based on the femA gene (S. aureus-specific gene) and the mecA gene (encoding penicillin-binding protein 2a), and the multiple-LAMP products were analyzed using LFB. The m-LAMP-LFB amplification conditions, including the target DNA concentration, reaction temperature and time, were optimized. The sensitivity and specificity of the m-LAMP-LFB method were tested in the current study, and the multiple-LAMP-LFB technology was applied to detect the MSSA and MRSA strains from clinical samples.ResultsThe S. aureus- and MRSA-specific primers based on the femA and mecA genes allowed the multiple-LAMP technology to detect S. aureus and MRSA, respectively. The multiple-LAMP conditions were optimized at 63 °C for 40 min. The full process, including genomic DNA template preparation, LAMP, and product identification, could be achieved in 80 min. The limit of detection (LoD) of the multiple-LAMP assay for femA and mecA detection was 100 fg of genomic DNA template per reaction. The specificity of m-LAMP-LFB detection was 100 %, and no cross-reactions to non-S. aureus strains were observed.ConclusionThe multiple-LAMP-LFB technique developed in the current study is a reliable, simple, rapid, specific and sensitive method to identify MSSA and MRSA infections for appropriate antibiotic therapy.
Highlights
Staphylococcus aureus (S. aureus), including methicillin-susceptible S. aureus (MSSA) and methicillinresistant S. aureus (MRSA), is an eminent human pathogen that can colonize the human host and cause severe lifethreatening infections
Verification and analysis of femA- and mecA-Loop-mediated isothermal amplification (LAMP) products To confirm the amplification with the two sets of LAMP primers, the femA, mecA, and multiplex LAMP (m-LAMP) mixtures were incubated at a constant temperature of 63 °C for 1 h
For femA-LAMP detection, two crimson red bands (CL and test line 1 (TL1)) appeared, indicating positive results, and control line (CL) and test line 2 (TL2) were visible for mecA-LAMP, indicating successful amplification, while the negative and blank controls only appeared as a crimson red line (CL) in the biosensor (Fig. 2b, d)
Summary
Staphylococcus aureus (S. aureus), including methicillin-susceptible S. aureus (MSSA) and methicillinresistant S. aureus (MRSA), is an eminent human pathogen that can colonize the human host and cause severe lifethreatening infections. The development of a reliable, simple and rapid assay for detecting S. aureus and identifying MRSA is important for diagnosis and follow-up treatment. Staphylococcus aureus, a gram-positive coccoid bacterium, is a common human pathogen that has the ability to cause a wide array of severe hospital and communityacquired infections, such as pneumonia, bacteremia, sepsis and toxic shock syndrome [1,2,3]. A methicillin-resistant S. aureus (MRSA) strain was found among clinical isolates from patients hospitalized in 1960. Developing a reliable and rapid method of detection for the accurate differentiation of methicillinsusceptible S. aureus (MSSA) and MRSA isolates is necessary for the follow-up treatment and management of patients
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
