Int J Med Microbiol. 2009 Nov;299(7):479-88. doi: 10.1016/j.ijmm.2009.03.004. Epub 2009 May 17.

The heme-binding protein (HbpA) of Haemophilus influenzae as a virulence determinant.

Morton DJ, Seale TW, Bakaletz LO, Jurcisek JA, Smith A, VanWagoner TM, Whitby PW, Stull TL.

Source

Department of Pediatrics, University of Oklahoma Health Sciences Center, Oklahoma City, OK 73104, USA. Daniel-morton@ouhsc.edu

Abstract

Haemophilus influenzae has an absolute growth requirement for heme and the heme-binding lipoprotein (HbpA) and has been implicated in the utilization of this essential nutrient. We constructed an insertional mutation of hbpA in a type b and a nontypeable H. influenzae strain. In the type b strain, the hbpA mutant was impaired in utilization of heme complexed to either hemopexin or to albumin and in the utilization of low levels of heme but not in the utilization of heme at high levels or of hemoglobin or hemoglobin-haptoglobin complexes. In contrast, the hbpA mutant derivative of the nontypeable strain was impaired in utilization of all tested heme sources. We further examined the impact of the hbpA mutation in animal models of H. influenzae disease. The hbpA mutant of the nontypeable strain was indistinguishable from the wild-type strain in the chinchilla model of otitis media. The hbpA mutant derivative of the type b strain caused bacteremia as well as the wild-type strain in 5-day old infant rats. However, in 30-day old rats the hbpA caused significantly lower rates of bacteremia than the wild-type strain indicating a role for hbpA and heme acquisition in virulence in this model of H. influenzae disease. In conclusion, HbpA is important for heme utilization by multiple H. influenzae strains and is a virulence determinant in a model of H. influenzae invasive disease.

PMID:: 19451029; [PubMed - indexed for MEDLINE]
PMCID:: PMC2749905

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Fig. 2

A) Growth of the H. influenzae type b strain HI689 and the hbpA insertion mutant strain HI1886 in hdBHI supplemented with heme–human serum albumin complexes as the sole heme source. Wild-type strain HI689 in heme–albumin 100 ng ml⁻¹ heme equivalent (solid circles), in heme–albumin 50 ng ml⁻¹ heme equivalent (solid triangles), and in heme–albumin 25 ng ml⁻¹ heme equivalent (solid squares). The hbpA insertion mutant strain HI1886 in heme–albumin 100 ng ml⁻¹ heme equivalent (open circles), in heme–albumin 50 ng ml⁻¹ heme equivalent (open triangles), and in heme–albumin 25 ng ml⁻¹ heme equivalent (open squares). B) Growth of the H. influenzae type b strain HI689 and the hbpA insertion mutant strain HI1886 in hdBHI supplemented with heme–hemopexin complexes or hemoglobin–haptoglobin complexes as the sole heme source. Wild-type strain HI689 in heme–hemopexin 10 μg ml⁻¹ heme equivalent (solid circles) and in heme–hemopexin 5 μg ml⁻¹ heme equivalent (solid triangles). The hbpA insertion mutant strain HI1886 in heme–hemopexin 10 μg ml⁻¹ heme equivalent (open circles) and in heme–hemopexin 5 μg ml⁻¹ heme equivalent (open triangles). Wild-type strain HI689 in hemoglobin–haptoglobin 2 μg ml⁻¹ hemoglobin equivalent (solid squares). The hbpA insertion mutant strain HI1886 in hemoglobin–haptoglobin 2 μg ml⁻¹ hemoglobin equivalent (open squares). Results are mean ± SD for quintuplicate results from representative experiments. Using the Kruskal-Wallis test to compare growth of HI689 and HI1886 in heme-albumin at 100 ng ml⁻¹ heme equivalent P=0.4156 over the entire growth curve and P=0.0036 for growth between 6 hours and 18 hour; at all other concentrations of heme-albumin P<0.0001. The Kruskal-Wallis test was used to compare growth of HI689 and HI1886 in heme–hemopexin at all other concentrations p<0.0001. The Kruskal-Wallis test was used to compare growth of HI689 and HI1886 in hemoglobin–haptoglobin at 2 μg ml⁻¹ hemoglobin equivalent p=0.3213.

The heme-binding protein (HbpA) of Haemophilus influenzae as a virulence determinant

Int J Med Microbiol. 2009 November;299(7):479-488.