A Chimeric Influenza Virus Expressing an Epitope of Outer Membrane Protein F ofPseudomonas aeruginosaAffords Protection against Challenge withP. aeruginosain a Murine Model of Chronic Pulmonary Infection

Abstract

The ability of a chimeric influenza virus containing, within the antigenic B site of its hemagglutinin, an 11-amino-acid (AEGRAINRRVE) insert from the peptide 10 epitope of outer membrane (OM) protein F of Pseudomonas aeruginosa to serve as a protective vaccine against P. aeruginosa was tested by using the murine chronic pulmonary infection model. Mice immunized with the chimeric virus developed antibodies that reacted in an enzyme-linked immunosorbent assay with peptide 10, with purified protein F, and with whole cells of various immunotype strains ofP. aeruginosa but failed to react with a protein F-deficient strain of P. aeruginosa. The chimeric-virus antisera reacted specifically with protein F alone when immunoblotted against proteins extracted from cell envelopes of each of the seven Fisher-Devlin immunotype strains and had significantly greater in vitro opsonic activity for P. aeruginosa than did antisera from wild-type influenza virus-immunized mice. Subsequent to intratracheal challenge with agar-encased cells of P. aeruginosa, chimeric-virus-immunized mice developed significantly fewer severe lung lesions than did control mice immunized with the wild-type influenza virus. Furthermore, the chimeric influenza virus-immunized group had a significantly smaller percentage of mice with >5 × 103 CFU of P. aeruginosa in their lungs upon bacterial quantitation than did the control group. These data indicate that chimeric influenza viruses expressing epitopes of OM protein F warrant continued development as vaccines to prevent pulmonary infections caused by P. aeruginosa.

Enhancement of the protective efficacy of an oprF DNA vaccine against Pseudomonas aeruginosa

“…The chimeric HG10-11 in£uenza virus and its parent WSN in£uenza virus have been described previously [24] and were each propagated in Madin^Darby canine kidney cells.…”

“…Sera recovered from the various groups of vaccine-immunized and control mice were analyzed by ELISA for reactivity against bacterial cells, peptide 10^14-mer [26], puri¢ed protein F [27], and the WSN and HG10-11 viruses [24] according to published procedures [26,27]. The wells of microtiter plates (polyvinyl chloride or Dynatech, Immulon 4) were coated with either approximately 5U10 6 bacterial whole cells of KG1077 or each FD immunotype strain, 2.5 Wg of synthetic peptide 10^14-mer, 10 Wg of protein F, or approximately 10 4 pfu of the appropriate virus.…”

“…Eight days after the challenge, protection a¡orded to the immunized mice by the DNA vaccine was assessed by two methods. First, the lungs were examined macroscopically for the presence of lesions [24,31,32]. Lesions were scored as 0 to 4+ as follows : 0, absence of any macroscopic lesion ; 1+, presence of one or two small lesions not exceeding 1 mm in diameter ; 2+, presence of three or more small lesions not exceeding 1 mm in diameter; 3+, presence of a medium lesion 2^5 mm in diameter ; 4+ presence of a large lesion exceeding 5 mm in diameter.…”

“…Lesions scored as 2+ to 4+ were considered to be severe lesions. Second, bacterial quantitation of the number of P. aeruginosa present in the lungs was performed as described previously [24,31,32]. Brie£y, both lungs were aseptically removed and homogenized in a 1:10 (w/v) dilution of sterile saline with a tissue grinder.…”

“…Brie£y, both lungs were aseptically removed and homogenized in a 1:10 (w/v) dilution of sterile saline with a tissue grinder. Dilutions of these lung tissues were plated onto nutrient agar for quantitative determination of the numbers of Pseudomonas present [24]. Duplicate challenge experiments were performed and the results compiled for analysis.…”