Development and Evaluation of a TaqMan Real-Time PCR Assay for the Detection of Mycoplasma pneumoniae

Mycoplasma pneumoniae is a major pathogen causing community-acquired pneumonia, often co-infecting with other pathogens and leading to severe infections that are difficult to differentiate. Current diagnostic methods have significant limitations, including the inability to accurately distinguish M. pneumoniae from other pathogens and the lack of precise quantification. To address these issues, this study developed a TaqMan-based real-time quantitative PCR (TaqMan PCR) assay. Specific primers and probes were designed based on the 16S rRNA gene sequences of M. pneumoniae retrieved from GenBank. After optimization, the established assay provided a sensitive, accurate, reliable, and cost-effective platform for M. pneumoniae detection. A comparative analysis using clinically confirmed M. pneumoniae samples demonstrated a positivity rate of 97.5% with this method, outperforming the 90% positivity rate of commercially available diagnostic kits. Additionally, the quantification results of M. pneumoniae strains showed that the 1 : 10 passaged culture entered the logarithmic growth phase on days 4 and 5, followed by a sharp decline in cell numbers. These findings highlight the potential of the TaqMan PCR assay as an improved diagnostic tool for more effective clinical detection and monitoring of M. pneumoniae infections.