Genetic vaccination against Coccidioides immitis: comparison of vaccine efficacy of recombinant antigen 2 and antigen 2 cDNA

Abstract

Previous studies from our laboratory established that C-ASWS, an alkali-soluble, water-soluble extract from cell walls of Coccidioides immitis, protects mice against lethal challenge with this fungus. The C-ASWS extract contains a glycosylated protein, designated antigen 2 (Ag2), and a polysaccharide antigen. We recently cloned Ag2 cDNA and showed that the recombinant fusion protein elicited strong delayed-type hypersensitivity responses in immunized mice. This investigation was undertaken to determine if the recombinant Ag2 protein, expressed as an Ag2-glutathione S-transferase (GST) fusion protein, or Ag2 cDNA would protect mice against lethal challenge with C. immitis. The recombinant Ag2-GST protein protected BALB/c mice against intraperitoneal challenge with 250 arthroconidia, as assessed by a decrease in fungal CFU in tissues. The Ag2-GST-immunized mice did not show, however, an increased survival during a 30-day period postinfection. By contrast, immunization of mice with Ag2 cDNA ligated into the pVR1012 plasmid engendered protection against intraperitoneal challenge with 2,500 arthroconidia and against pulmonary challenge with 50 arthroconidia. Vaccine efficacy paralleled the development of delayed-type hypersensitivity responses to C. immitis antigen. Whereas mice vaccinated with the recombinant Ag2-GST protein did not mount footpad hypersensitivity to C-ASWS or the recombinant Ag2-GST protein, mice vaccinated with the pVR1012-Ag2 construct mounted a strong footpad hypersensitivity and their spleen cells secreted gamma interferon upon in vitro stimulation with the Ag2-containing C-ASWS extract. This is the first investigation to show that genetic immunization can protect against lethal challenge with C. immitis.

FIG. 1

Vaccine efficacy of the recombinant Ag2 protein in BALB/c mice challenged by an i.p. route with 250 arthroconidia. Results are expressed as the number of CFU (mean ± standard error) in tissues at day 12 postinfection (A) and the mortality in mice at days 1 through 30 postinfection (B). The results in panel A are representative of those obtained in three separate experiments with groups of 13 or more mice immunized with the Ag2-GST protein or GST alone.

FIG. 2

Vaccine efficacy of the pVR1012-Ag2 construct in BALB/c mice challenged by an i.p. route with 2,500 arthroconidia, as measured by CFU in tissues from groups of 20 mice immunized with pVR1012-Ag2 or pVR1012 alone (A) and mortality at days 1 through 40 postinfection in groups of 11 mice immunized with the Ag2 gene or the pVR1012 plasmid alone (B). The results shown in panel A are representative of those obtained in three separate experiments.

FIG. 3

Vaccine efficacy of the pVR1012-Ag2 construct in BALB/c mice challenged with 50 arthroconidia via the pulmonary route, as assessed by measuring the fungal CFU in tissues. Results depict mean ± the standard error obtained in one of two experiment with groups of 9 to 10 mice.

FIG. 4

Footpad hypersensitivity response of mice immunized with the pVR1012-Ag2 construct or the pVR1012 plasmid alone. Results depict mean ± the standard error obtained in groups of 19 mice footpad tested with C-ASWS (10 μg) 12 days after the third immunization. The results are representative of those obtained in two separate experiments.