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Infection and Immunity, November 2002, p. 5955-5964, Vol. 70, No. 11
0019-9567/02/$04.00+0     DOI: 10.1128/IAI.70.11.5955-5964.2002
Copyright © 2002, American Society for Microbiology. All Rights Reserved.

Characterization of the Pasteurella multocida hgbA Gene Encoding a Hemoglobin-Binding Protein

Department of Genetics and Microbiology, Universitat Autònoma de Barcelona,¹ Centre de Recerca en Sanitat Animal (CReSA), Universitat Autònoma de Barcelona-Institut de Recerca i Tecnologia Agroalimentària (UAB-IRTA), Bellaterra, 08193 Barcelona, Spain²

Received 22 March 2002/ Returned for modification 3 July 2002/ Accepted 23 July 2002

Reverse transcriptase PCR analyses have demonstrated that open reading frames (ORFs) PM0298, PM0299, and PM0300 of the animal pathogen Pasteurella multocida constitute a single transcriptional unit. By cloning and overexpression studies in Escherichia coli cells, the product of ORF PM0300 was shown to bind hemoglobin in vitro; this ORF was therefore designated hgbA. In vitro and in vivo quantitative assays demonstrated that the P. multocida hgbA mutant bound hemoglobin to the same extent as the wild-type strain, although the adsorption kinetics was slightly slower for the hgbA cells. In agreement with this, the virulence of P. multocida hgbA cells was not affected, suggesting that other functional hemoglobin receptor proteins must be present in this organism. On the other hand, P. multocida mutants defective in PM0298 and PM0299 could be isolated only when a plasmid containing an intact copy of the gene was present in the cells, suggesting that these genes are essential for the viability of this bacterial pathogen. By adapting the recombinase-based expression technology in vivo to P. multocida, we also demonstrated that the transcriptional PM0298-PM0299-hgbA unit is iron regulated and that its expression is triggered in the first 2 h following infection in a mouse model. Furthermore, hybridization experiments showed that the hgbA gene is widespread in P. multocida strains regardless of their serotype or the animal from which they were isolated.

Editor: B. B. Finlay

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