[Genotyping of ampicillin-resistant Haemophilus influenzae]

Haemophilus (H.) influenzae is a gram-negative bacillus that is a common commensal organism of the human upper respiratory tract and an important cause of human diseases such as pneumonia, meningitis, septicemia, epiglottitis and cellulitis. Strains of H. influenzae are classified according to their capsular polysaccharide. There are six serotypes, designated as a through f. In addition, there are nonencapsulated strains. Although the type of infectious diseases caused by H. influenzae has changed considerably in recent years because of the widespread and routine immunization of children against type b H. influenzae (Hib), Hib remains an important pathogen. Ampicillin is the drug of choice for treating many infections caused by H. influenzae, but its usefulness has been compromised by the increasing prevalence of ampicillin-resistant strains. The continued monitoring of resistant strains by using genotyping methods may provide insights into the epidemiology of transmission. A molecular epidemiological study of ampicillin-resistant H. influenzae derived from nasopharyngeal swabs specimens of children less than 5 years of age with respiratory tract infection were investigated in this study.

A total of 899 isolates were collected from Beijing, Shanghai, and Guangzhou during 2000-2003. Susceptibility to ampicillin was determined by using E-test. Ampicillin-resistant H. influenzae strains were selected according to National Committee for Clinical Laboratory Standards (NCCLS) 2002 breakpoints. Nested PCR method with primers specific for bexA gene and b capsulate type-specific gene was established. Genotyping by pulsed-field gel electrophoresis (PFGE) and multiplex PCR assay was performed for all ampicillin-resistant H. influenzae strains.

Seventy-four ampicillin-resistant H. influenzae strains were obtained. Two strains were positive by nested PCR, characterized as b genotype. The incidence of Hib in ampicillin-resistant H. influenzae strains was 2.7%; 38 genotypes were detected by PFGE. Detection of five types strains of clonal dissemination by PFGE accounted for 55.4% in all ampicillin-resistant H. influenzae strains. Among them eighteen H. influenzae strains belonged to one type, accounted for 24.3% in all ampicillin-resistant H. influenzae strains. Thirty one genotypes were identified by multiplex PCR assay for ampicillin-resistant H. influenzae. The identity ratio of PFGE and multiplex PCR was 63.5%.

In Beijing, Shanghai and Guangzhou areas 55.4% of ampicillin-resistant H. influenzae strains had clonal dissemination during the 4 years.