Simultaneous visual diagnosis of capsulated and non-capsulated Haemophilus influenzae using a duplex RPA method

Haemophilus influenzae, particularly type b (Hib), represents a significant opportunistic pathogen that poses a substantial threat to the health and safety of vulnerable populations, including the elderly and children. To facilitate early detection and timely intervention, there is an urgent need for a simple, portable, and efficient molecular diagnostic approach. In this study, a duplex recombinase polymerase amplification assay with a lateral flow strip was developed to target the outer membrane protein P6 (omp6) of H. influenzae, and the capsule encoding gene bexA of capsulated H. influenzae. The assay is performed at 37℃ for 25 min, with a limit of detection of 10¹ CFU, comparable to that of dual PCR. The detection accuracy for clinical samples was consistent with the traditional culture and dual PCR methods. The duplex RPA assay does not require a laboratory facility and may be useful for genotyping of H. influenzae.