Evaluation of methodology for serotyping invasive and nasopharyngeal isolates of Haemophilus influenzae in the ongoing surveillance in Brazil

To assess the magnitude of discrepant results obtained by routine Haemophilus influenzae serotyping, 258 isolates, collected by the epidemiological surveillance system in Brazil from individuals with invasive diseases or carriage, were evaluated by two slide agglutination (S1Ag) methods: SIAg method 1, by which strains were initially screened with a serotype b-specific antiserum, and SIAg method 2, by which strains were tested against all serotype-specific antisera in parallel. Investigators comparing results of the two S1Ag methods with those obtained by capsule type-specific PCR were blinded to the method used. The serotype prevalence rates found by the three methods were significantly different, involving discrepancies mainly between serotype b and noncapsulated (NC) isolates. For invasive isolates (n = 131), the overall agreement rate between S1Ag method 1 or 2 and PCR was 68.0 or 88.3%, respectively, whereas for colonizing isolates (n = 127) the corresponding rate was 46.5 or 94.2%, respectively. S1Ag method 2 improved the ascertainment of serotypes over that obtained with S1Ag method 1, demonstrating good correlation with PCR. Use of the poly-valent antiserum as a screening reagent for S1Ag for invasive and colonizing isolates showed poor discriminatory power, with a sensitivity of 65.8% and a specificity of 91.7%. We stress the importance of using a well-standardized S1Ag methodology and suggest that reference laboratories should utilize PCR routinely to confirm S1Ag results and to check all nonspecific S1Ag reactions and apparent NC isolates by S1Ag in order to provide reliable data on the prevalence of H. influenzae serotypes in the H. influenzae type b vaccine era.