Characterisation of Haemophilus influenza and Haemophilus haemolyticus in Chronic Obstructive Pulmonary Disease (COPD)

Abstract

Chronic obstructive pulmonary disease (COPD) accounted for 5.71% of all global mortality in 2015. Non-typeable Haemophilus influenzae (NTHi) is a genetically diverse opportunistic pathogen that colonises the respiratory tract and is linked to acute exacerbations in COPD which eventually lead to a fatal irreversible degradation of lung function.

H. haemolyticus is the closest phylogenetic relative to NTHi and is also found to colonise the respiratory tract of subjects with COPD. However, H. haemolyticus is not thought to cause either invasive disease or exacerbations in COPD. The motivation for the work in this thesis is that there are currently no molecular or culture assays that can unequivocally differentiate these two species. The genetic diversity of NTHi has been reported. The objective of this study was to determine whether the genetic variation in atypical strains of the two species was sufficiently large to result in a genetic continuum that makes their delineation impossible and therefore questioning the current taxonomic classification.

Whole genome sequencing data generated from 1460 isolates phenotypically identified as Haemophilus spp. were used to determine the genomic heterogeneity of NTHi and H. haemolyticus. This included assessing the utility of gene markers for example, fucK, smpB, iga, sodC, lgtC, hpd, omp2 and omp6, which have previously been shown to enable differentiation, as well as more extensive comparative genomic analyses. These data revealed the presence of previously poorly recognised atypical strains of NTHi and H. haemolyticus. Atypical isolates were initially characterised as those with an unexpected genotype for NTHi or H. haemolyticus as defined by previously described markers. The use of further adhesin genes and comparative genomics supported these findings. Comparison of NTHi and H. haemolyticus demonstrated distinctive clustering based on ANI with <92% similarity; a value below current taxonomic thresholds for species definition. In conclusion, despite extensive genomic variation in strains of NTHi they may be routinely separated from H. haemolyticus using whole genome analysis and multiple gene markers.

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Identifiers

ORCID for Stuart Clarke: orcid.org/0000-0002-7009-1548

ORCID for David Cleary: orcid.org/0000-0003-4533-0700

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