A FUNCTIONAL TONB GENE IS REQUIRED FOR BOTH UTILIZATION OF HEME AND VIRULENCE EXPRESSION BY HAEMOPHILUS-INFLUENZAE TYPE-B

被引:75
作者
JAROSIK, GP
SANDERS, JD
COPE, LD
MULLEREBERHARD, U
HANSEN, EJ
机构
[1] UNIV TEXAS,SW MED CTR,DEPT MICROBIOL,DALLAS,TX 75235
[2] CORNELL UNIV,SCH MED,DEPT PEDIAT,NEW YORK,NY 10021
[3] CORNELL UNIV,SCH MED,DEPT BIOCHEM,NEW YORK,NY 10021
[4] CORNELL UNIV,SCH MED,DEPT PHARMACOL,NEW YORK,NY 10021
关键词
D O I
10.1128/IAI.62.6.2470-2477.1994
中图分类号
R392 [医学免疫学]; Q939.91 [免疫学];
学科分类号
100102 ;
摘要
Haemophilus influenzae is nearly unique among facultatively anaerobic bacteria in its absolute requirement for exogenously supplied heme for aerobic growth. In this study, a mutant analysis strategy was used to facilitate identification of H. influenzae cell envelope components involved in the uptake of heme. Chemical mutagenesis was employed to produce a mutant of a nontypeable H. influenzae strain unable to utilize either protein-bound forms of heme or low levels of free heme. This mutant was transformed with a plasmid shuttle vector-based genomic library constructed from the same wild-type nontypeable H. influenzae strain, and a growth selection technique was used to obtain a recombinant clone that could utilize heme. Analysis of the DNA insert in the recombinant plasmid revealed the presence of several open reading frames, one of which encoded a 28-kDa protein with significant similarity to the TonB protein of Escherichia coli. This H. influenzae gene product was able to complement a tonB mutation in E. call, allowing the E. coli tonB mutant to form single colonies on minimal medium containing vitamin B-12. When this H. influenzae gene was inactivated by insertional mutagenesis techniques and introduced into the chromosome of wild-type strains of H. influenzae type b, the resultant transformants lost their abilities to utilize heme and produce invasive disease in an animal model. Genetic restoration of the ability to express this TonB homolog resulted in the simultaneous acquisition of both heme utilization ability and virulence. These results indicate that the H. influenzae TonB protein is required not only for heme utilization by this pathogen in vitro, but also for virulence of H. influenzae type b in an animal model.
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页码:2470 / 2477
页数:8
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