Identification of Haemophilus influenzae type b isolates by use of matrix-assisted laser desorption ionization-time of flight mass spectrometry

Haemophilus influenzae type b (Hib) is, in contrast to non-b H. influenzae, associated with severe invasive disease such as meningitis and epiglottitis in small children. To date accurate H. influenzae capsule typing requires PCR, a time-consuming and cumbersome method. MALDI-TOF MS provides rapid bacterial diagnostics and is increasingly used in clinical microbiology laboratories. Here MALDI-TOF MS was evaluated as a novel approach to separate Hib from other H. influenzae. PCR-verified Hib and non-Hib reference isolates were selected based on genetic and spectral characteristics. Mass spectra of reference isolates were acquired, and used to generate different classification algorithms for Hib/non-Hib separation using both ClinProTools and MALDI Biotyper software. A test series of mass spectra from 33 Hib and 77 non-Hib isolates, all characterized by PCR, was used to evaluate the algorithms. Several algorithms yielded good results but the two best were a ClinProTools model based on 22 separating peaks and a subtyping main spectra (MSP) model using MALDI Biotyper. The ClinProTools model had a sensitivity of 100%, a specificity of 99% and the results were 98% reproducible using a different MALDI-TOF MS instrument. The Biotyper subtyping MSPs had a sensitivity of 97%, a specificity of 100% and 93% reproducibility. Our results suggest that it is possible to use MALDI-TOF MS to differentiate Hib from other H. influenzae. This is a promising method to rapidly identify Hib in unvaccinated populations, and for screening or surveillance of Hib carriage in vaccinated populations.