Development of a multiplex PCR assay for rapid detection of Clostridium perfringens, Clostridium septicum and Paeniclostridium sordellii in necrotising soft tissue infections.

BACKGROUND

Necrotising soft tissue infections (NSTIs) are fulminant infections requiring rapid diagnosis and intervention. Conventional methods, such as anaerobic culture, are slow and insensitive for clostridial pathogens.

OBJECTIVES

To develop and evaluate a multiplex PCR assay targeting three clinically relevant clostridia directly in NSTI samples.

METHODS

A multiplex conventional PCR targeting the cpa (Clostridium perfringens), csa (Clostridium septicum), and 16S rRNA (Paeniclostridium sordellii) genes was optimised and applied to tissue/pus samples from 87 clinically diagnosed NSTI patients. Anaerobic culture served as the reference. Sensitivity, specificity, and diagnostic turnaround times were compared.

RESULTS

Seven patients (8.0%, 95% CI 3.3-16.2) were positive for clostridia by PCR, compared with four (4.6%, 95% CI 1.3-11.4) by culture. PCR identified three additional culture-negative cases. While McNemar's test did not show statistical significance (p = 0.25), PCR achieved a sensitivity of 100% (95% CI 39.8-100.0), specificity of 96.4% (95% CI 89.8-99.2), and a negative predictive value of 100%. Turnaround time was 3-4 h, compared with 48-96 h for culture. Both patients with monomicrobial C. septicum NSTI succumbed rapidly, highlighting its prognostic importance.

CONCLUSION

This is the first report of a multiplex PCR assay for direct detection of C. perfringens, C. septicum, and P. sordellii in NSTIs. The assay enhances detection, reduces turnaround time, and provides clinically relevant species-level identification. Multiplex PCR may serve as a valuable adjunct to culture in early NSTI diagnostics and could be expanded to include additional anaerobic pathogens.