A multicenter clinical evaluation of polymerase chain reaction coupled with quantum dot fluorescence analysis and quantitative real-time reverse transcription polymerase chain reaction in the diagnosis of pathogens

This multi-center clinical study evaluated 17 common pathogens in 1,922 pharyngeal swab samples, comparing the diagnostic performance of polymerase chain reaction coupled with quantum dot fluorescence analysis (PCR-QDFA) with that of clinically routine quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The results were validated using Sanger sequencing as the gold standard. Our results showed that among the samples with single-pathogen infections (1037 cases, 53.95%), the three most frequently detected pathogen were influenza A virus (IAV) (296 cases, 15.40%), severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) (164 cases, 8.53%), and Mycoplasma pneumoniae (94 cases, 4.89%). Similarly, among the samples with co-infections of two or more pathogens (382 cases, 19.88%), the three most frequently detected pathogens were IAV (148 cases, 7.70%), SARS-CoV-2 (107 cases, 5.57%), and M. pneumoniae (67 cases, 3.49%). In the comprehensive evaluation of 17 respiratory pathogens, PCR-QDFA demonstrated comparable diagnostic performance to qRT-PCR, with an overall sensitivity of 99.78% (99.44–99.92%) (vs. qRT-PCR: 99.82% [99.48–99.94%]) and specificity of 99.94% (99.90–99.96%) (vs. qRT-PCR: 99.95% [99.91–99.97%]). PCR-QDFA offers significant operational advantages, including high-throughput capacity (96 samples per run) and lower cost.