qPCR method and reagent for quickly detecting MRSA (Methicillin Resistant Staphylococcus Aureus)

Abstract

The invention provides a qPCR method and a reagent for quickly detecting MRSA (Methicillin Resistant Staphylococcus Aureus). The qPCR method comprises the following steps: (1) preliminary preparationprocedure; (2) nucleic acid extraction and purification; (3) primer / probe design, synthesis and labeling; (4) qPCR amplification and result judgment; (5) detection lower limit and amplification linearrange test; (6) gene fragment sensitivity and specificity calculation; (7) diagnostic sensitivity and specificity calculation; (8) a statistical method. The qPCR method and the reagent provided by the invention have the benefits that the detection lower limit is as low as 4cfu / reaction, and the amplification linear range reaches 2.0*10<2-8> Copies / ml; the sensitivity and the specificity of MRSA identification are 100 percent (352 / 352) and are 100 percent (77 / 77) respectively, and the sensitivity and the specificity of MRSA detection are 100 percent (94 / 94) and 95.4 percent (94 / 98) respectively; the qPCR method is simple and convenient, quick, high in sensitivity and good in specificity and not only can improve the diagnostic ability of MRSA infection, but also can realize the quick detection, thereby gaining time for early and accurate treatment.

Description

【Technical field】 [0001] The invention relates to a qPCR method for Staphylococcus aureus MRSA, in particular to a qPCR method and reagents for rapidly detecting Staphylococcus aureus MRSA. 【Background technique】 [0002] Staphylococcus aureus (Staphylococcus aureus, SA) is an important pathogenic bacteria in humans, which can cause serious community-acquired infections and serious hospital-acquired infections[2]. Since its symptoms are the same as those of other bacterial infections, it is difficult to make an early and accurate diagnosis of this bacterial infection in clinical practice. Especially since Methicillin resistant staphylococcus aureus (MRSA) was first discovered in 1961, its drug resistance has become more and more serious. prominent, seriously affecting the therapeutic effect. The culture method is currently a commonly used detection method and is also the gold standard, but it takes 2 to 3 days or even longer, and its timeliness, simplicity, and sensitivity ...

AI Technical Summary

Problems solved by technology

Since its symptoms are the same as those of other bacterial infections, it is difficult to make an early and accurate diagnosis of this bacterial infection in clinical practice. Especially since Methicillin resistant staphylococcus aureus (MRSA) was first discovered in 1961, its drug resistance has become more and more serious. Prominent, seriously affecting the therapeutic effect

The culture method is currently a commonly used detection method and is also the gold standard, but it takes 2 to 3 days or even longer, and its timeliness, simplicity, and sensitivity cannot meet the needs of clinical rapid and accurate diagnosis

Bi Yanni et al. detected bloodstream MRSA infection by multiplex drop-down PCR method, and the result has high sensitivity and specificity, which is helpful to guide the clinical use of antibacterial drugs; however, analysis of PCR products by electrophoresis is easy to cause laboratory contamination and serious false positives

Shen Lei et al. compared the MRSA-PCR rapid detection method in Shanghai Zhijiang and found that in addition to the good negative predictive value, the sensitivity, specificity and positive predictive value were all low

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