D-080. The Role of the Heme-Binding Lipoprotein (HbpA) of Haemophilus influenzae Type b in Heme Utilization and Virulence

D. J. Morton, P. W. Whitby, T. M. VanWagoner, T. W. Seale, T. L. Stull;
Univ. of Oklahoma Hlth. Sci. Ctr., Oklahoma City, OK.

Haemophilus influenzae has an absolute aerobic growth requirement for heme, which can be satisfied in vitro by heme, hemoglobin, hemoglobin-haptoglobin, heme-hemopexin and heme-albumin. The mechanisms used by H. influenzae to obtain heme are complex and highly redundant. One component of the H. influenzae heme acquisition pathway(s) is the heme binding lipoprotein (HbpA) which is localized to the periplasmic space. In a previous study we demonstrated that in a nontypeable (NT) H. influenzae strain mutation of hbpA significantly reduced utilization of each of the heme sources listed above. To extend these studies to a type b strain and also examine the potential role of HbpA in virulence, we constructed an insertional mutation of hbpA in the type b strain HI689. In growth studies the hbpA mutant of HI689 was impaired in utilization of heme complexed to either hemopexin or to albumin and in the utilization of low levels of free heme. Utilization of heme at high levels or of hemoglobin or hemoglobin-haptoglobin complexes was unaffected. These data contrast with that for the hbpA mutant derivative of the NT strain which was impaired in utilization of all tested heme sources. These data indicate that type b strains possess additional, as yet unidentified, periplasmic heme transport mechanisms. We additionally examined the impact of the hbpA mutation in a rat model of invasive H. influenzae disease. The hbpA mutant derivative of the type b strain established and maintained bacteremia as well as the wildtype strain in 5-day old infant rats although bacteremic titers were lower in animals infected with the mutant strain In 30-day old rats the hbpA mutant established bacteremia at a lower rate, bacteremia persisted for a shorter time and bacteremic titers were lower than in animals infected with the wildtype strain. These age-dependent differences in the impact on virulence may reflect changes in available heme sources in the developing rat, for example serum levels of both hemopexin and haptoglobin are known to increase during the first month if life in the rat. This is the first report of a role for HbpA in virulence of H. influenzae in invasive disease.