Objective: To develop a rapid method for detecting Haemophilus influenzae by multiplex polymerase chain reaction (M-PCR).
Methods: Primers (Hi) were designed for amplification of p6 gene coding P6 protein of Haemophilus influenzae, which was used to identify Haemophilus influenzae species. Primers (Hi-cap) were designed for amplification of bexA gene which coding capsular polysaccharide (cap) synthesis was used for detecting whether Haemophilus influenzae isolates possess bexA gene relating to cap synthesis. Twelve primers (Hia-Hif) were designed for amplification of cap synthesis gene to identify the cap-type of Haemophilus influenzae. Other relative enteric pathogenic bacteria were amplified by M-PCR to serve as controls. 200 strains isolated from patients were identified. Results from M-PCR were compared to two methods including V and X factors grow requirement test and standard slide agglutination serotyping (SAST).
Results: The results indicated that the M-PCR assay was high specificity and sensitivity and might be valuable for differential diagnosis of Haemophilus influenzae. The sensitivity of detection was 0.935 pg. 189 strains out of the 200 belonged to Haemophilus influenzae isolates, and one isolate was cap-type f. An agreement results were seen among the V and X factors grow requirement test, SAST and M-PCR methods.
Conclusion: M-PCR method showed satisfactory sensitivity, specificity and stability for detecting and identifying Haemophilus influenzae, and could be used in clinic diagnosis, surveillance and rapid diagnosis for plague of Haemophilus influenzae.