Characterization of a conserved outer-membrane protein in non-typeable Haemophilus influenzae with an unidentified...

Non-typeable Haemophilus influenzae (NTHi) is an opportunistic bacterial pathogen of the human airway and is a cause of significant morbidity and mortality worldwide. No vaccine currently exists in part due to the incredible diversity and extreme variability exhibited both genetically and phenotypically between NTHi strains. For a vaccine to be successful, selection of appropriate antigens is key. Antigens need to show high conservation between strains, be present in most/all strains, and be stably expressed. The uncharacterized protein NTHI1101, initially identified from sequential isolates of NTHi from the sputum of COPD patients, demonstrated attributes of interest as a potential vaccine antigen: analysis of over 4,500 individual NTHi genomes demonstrated this protein was present in over 99% of strains, with over 80% amino acid identity. Further analysis predicted that NTHI1101 contained a lipoprotein signal sequence, meaning it was likely located in the outer membrane. An experimental determination of the location of the NTHI1101 protein demonstrated that although this protein was present in enriched outer-membrane fractions of NTHi cells, analysis with whole-cell enzyme-linked immunosorbent assays indicated it was not surface located. A variety of in vitro analyses with relevance to pathobiology demonstrated that the loss of the NTHI1101 protein had no impact on the phenotype of NTHi. Therefore, although this protein is highly conserved and almost universally present in NTHi, we conclude that the use of this protein as a target for novel therapeutics is perhaps not warranted.

Non-typeable Haemophilus influenzae (NTHi) is a major human pathogen for which there is no vaccine. Subunits for a rationally designed vaccine need to be conserved and present in almost all strains of an organism, be stably expressed, and be surface-located, so they will be "recognized" by the immune system. Our work sought to determine if a highly conserved NTHi protein, NTHI1101, was surface-located and required for pathobiology. Characterization showed that this protein was located in the outer membrane, but not present on the bacterial cell surface, and not required for key aspects of disease. We therefore conclude that NTHI1101 should not be further investigated as a vaccine candidate, so that ineffective antigens are not included in an NTHi vaccine.