EasyNAT MP Assay: A Simple, Rapid, and Low-Cost Method to Detect Mycoplasma pneumoniae Using Cross-Priming Amplification Technology

BACKGROUND: Mycoplasma pneumoniae (MP) is the most common pathogen of atypical pneumonia and the main cause of community-acquired pneumonia (CAP) in infants and older adults. This study aimed at investigating a method based on the cross-priming amplification (CPA) technique for the rapid detection of MP in clinical specimens collected from patients with CAP. METHODS: The sensitivity and specificity of the EasyNAT MP assay were determined. Oropharyngeal swab specimens were collected from 162 in-patients of Hangzhou First People’s Hospitals from January 2018 to December 2020. The patients were aged between 1 and 15 years with symptoms, signs, and chest radiographs consistent with CAP. This study evaluated the presence of MP in the clinical specimens using the EasyNAT method and the conventional fluorescence quantitative PCR technique. RESULTS: The limit of detection using the EasyNAT MP assay was 500 copies/mL, while the test results of the other 13 common pathogens causing CAP or colonizing in the upper respiratory tract showed no cross-reactivity. Of 162 specimens, EasyNAT MP gave a positive indication in 82 specimens. Compared with conventional fluorescence quantitative PCR, the positive coincidence rate and the negative coincidence rate of EasyNAT MP was found to be 100.00% and 97.56%, respectively. Of the 82 specimens, two specimens were determined to be negative by the conventional fluorescence quantitative PCR, but were positive for EasyNAT MP. The two samples were re-extracted and confirmed to be positive by conventional fluorescence quantitative PCR. CONCLUSION: EasyNAT MP is suitable as an initial test for MP diagnosis due to its simplicity, low turnaround time, and high sensitivity and specificity.