Abstract
BackgroundThe RNA binding protein Hfq of Haemophilus influenzae is highly homologous to Hfq from other bacterial species. In many of these other bacteria, Hfq affects the expression of a broad range of genes and enhances the ability to respond to stressful environments. However, the role of Hfq in H. influenzae is unknown.ResultsDeletion mutants of hfq were generated in the nontypeable H. influenzae strains R2866 and 86-028NP to assess the role of Hfq in these well characterized but genotypically and phenotypically divergent clinical isolates. A deletion mutation of hfq had no effect on growth of H. influenzae in nutrient rich media and had no effect on survival in several stressful conditions in vitro. However, the mutation resulted in a reduced ability to utilize heme from hemoglobin. The mutant and wild type strains were assessed for virulence and competitive fitness in models of invasive disease and otitis media. In the chinchilla model of otitis media, the hfq mutant of 86-028NP exhibited impaired competitive fitness when compared to its wild type progenitor but exhibited no apparent defect in virulence. In the infant rat model, deletion of hfq in R2866 resulted in reduced bacterial titers in blood and a shorter duration of infection when compared to the wild type strain in the competitive fitness study.ConclusionWe conclude that Hfq is involved in the utilization of essential nutrients and facilitates infection by H. influenzae.
Highlights
The RNA binding protein Hfq of Haemophilus influenzae is highly homologous to Hfq from other bacterial species
H. influenzae was cultured on brain heart infusion (BHI) agar or in BHI broth supplemented with 10 μg mL-1 heme and 10 μg mL-1 β-NAD or BHI supplemented with 10 μg mL-1 β-NAD
Promoter and sequence analysis of hfq in H. influenzae Hfq is encoded by the gene HI0411 in the H. influenzae reference strain Rd KW20 [GenBank: NC_000907] and consists of 91 amino acids and is at least 97% identical to Hfq in 20 sequenced strains of H. influenzae
Summary
The RNA binding protein Hfq of Haemophilus influenzae is highly homologous to Hfq from other bacterial species. H. influenzae requires iron for growth but it has an absolute requirement for a porphyrin source, in the form of protoporphyrin IX (PPIX) or heme, to grow aerobically [5]. This requirement for a porphyrin source is due to the lack of enzymes required to synthesize the protoporphyrin ring. H. influenzae has evolved multiple mechanisms to counter and exploit host mechanisms for sequestering heme from invading pathogens [9]. Many of these mechanisms are transcriptionally upregulated in response to iron and heme restriction, the specific regulation of many of these systems is largely uncharacterized in H. influenzae [10,11]
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