Haemophilus influenzae – typing, epidemiology and beta-lactam resistance

Abstract: Haemophilus influenzae is a common cause of respiratory tract infections such as acute otitis media (AOM), exacerbations of chronic obstructive pulmonary disease (COPD) and pneumonia. The species is subdivided into encapsulated and non-encapsulated strains, designated type a-f and nontypeable H. influenzae (NTHi), respectively. Prior to introduction of polysaccharide-protein conjugate vaccines against H. influenzae type b (Hib) in childhood vaccination programmes this serotype frequently caused severe invasive infections in small children. Nowadays invasive disease by Hib is rare, but cases still occur. At present, NTHi is the dominating type to cause invasive disease and invasive NTHi disease appears to be increasing. Nontypeable H. influenzae disease severity traditionally has been considered largely host dependant. In parallel, non-beta-lactamase mediated beta-lactam resistance among NTHi is also increasing. In the first two studies of this thesis, we investigated capsule typing of H. influenzae by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS), a technology routinely used for bacterial species identification. Mass spectra of the different types of encapsulated H. influenzae were highly similar within each type and separable from each other. The differences in mass spectra relied on the clonal population structure of encapsulated H. influenzae, with conserved type specific genetic lineages. Mass spectra of NTHi were diverse, due to their genetic heterogeneity. Following construction of a comprehensive reference database, MALDI-TOF MS showed high accuracy for capsule typing of H. influenzae. In the third study of the thesis, a clonal group of NTHi with resistance against beta-lactam antibiotics was investigated. The clonal group accounted for one quarter of clinical respiratory H. influenzae isolates with non-beta-lactamase mediated beta-lactam resistance in the study region. Furthermore, patients infected by isolates of the clonal group had an increased risk of hospitalization compared to patients infected by other NTHi, indicating enhanced virulence traits. The clonal group was also found among invasive isolates. In the final study of the thesis we retrospectively compared benzylpenicillin, whose effect on H. influenzae is debated, to wide spectrum beta-lactams (WSBLs) as empirical treatment of H. influenzae lower respiratory tract infections in patients requiring hospitalization. Empirical treatment with benzylpenicllin was not associated with higher mortality or increased risk of hospital readmission compared to treatment with WSBLs. The early clinical response rate was, however, lower for patients receiving benzylpenicillin, which was attributed mainly to a lower response rate in patients infected with beta-lactamase producing isolates. In conclusion this thesis shows that MALDI-TOF MS can be efficiently used for rapid capsule typing of H. influenzae. The newly developed method can be valuable for typing of invasive H. influenzae isolates and for surveillance of Hib vaccination efficacy. The studied clonal group appears to harbour enhanced virulence traits. This indicates that bacterial factors may affect NTHi disease severity more than previously considered, and possibly contribute to the increased incidence of invasive NTHi disease. Finally, although the effect of benzylpenicillin on H. influenzae is debated, empirical treatment of lower respiratory tract infections of mild to moderate severity caused by H. influenzae with the agent appears safe