关键词: 呼吸道病原体, 多重逆转录聚合酶链反应, MassARRAY技术

Abstract:

Objective To establish a method based on multiple reverse transcription polymerase chain reaction（RT-PCR）MassARRAY for the determinations of 27 respiratory pathogens,and to investigate its clinical application value. Methods Multiple polymerase chain reaction amplification and extension primers were designed for pathogens,and the sensitivity,specificity and repeatability of multiple RT-PCR MassARRAY were evaluated with the standard plasmid of pathogens established. Totally,207 clinical specimens were determined by multiple RT-PCR MassARRAY,and the results were compared with real-time RT-PCR. Results The minimum detection limit of the established multiple RT-PCR MassARRAY for 27 respiratory pathogens was 1×10¹-1×10³ copies/μL,with a high specificity. The 57 of 207 clinical specimens were positive,and the positive rate was 27.54%. Streptococcus pneumoniae,Moraxella catarrhalis,Mycoplasma pneumoniae and Haemophilus influenzae were the most common bacterial pathogens,and parainfluenza virus,human metapneumovirus and influenza A virus had high detection rates.”Compared with real-time RT-PCR,the total consistency rate was 99.03%,with high consistency and a kappa value of 0.98. Conclusions The detection system of 27 respiratory pathogens based on multiple RT-PCR MassARRAY has high sensitivity and specificity,simple operation and high throughput,which can provide a reference for the clinical diagnosis of respiratory pathogens.

Key words: Respiratory pathogen, Multiple reverse transcription polymerase chain reaction, MassARRAY technology