Zimmermann CR, Johnson SM, Kerekes KM, Ampel NM, Christian L, Pappagianis D; American Society for Microbiology. General Meeting.

Abstr Gen Meet Am Soc Microbiol. 1999 May 30-Jun 3; 99: 314 (abstract no. F-93).

University of California, Davis.

Previously, we have shown that a soluble vaccine (F- 27K)(extracted from formalin-killed, endosporulating whole spherules) when combined with an alum adjuvant, protected mice from lethal infection as well as the whole spherule starting material. We fractionated the F-27K vaccine in an attempt to isolate and identify the protective protein component(s) of this preparation. However, the fractionation methods we used that were expected to work were unable to resolve any individual protective components. We hypothesized that the formaldehyde used to kill the endosporulating spherules was causing an irreversible crosslinking of the contents of the spherules, and, therefore, inhibiting our ability to separate individual components from the F-27K vaccine mixture. We therefore changed our killing and preserving procedure to a thimerosal-killing/ thimerosal preserving procedure. A vaccine (T-27K) was prepared from the thimerosal-killed spherules using the same procedure used to produce the F-27K vaccine and used to vaccinate groups of mice. In addition, other groups of mice were vaccinated with thimerosal-killed spherules, formalin-killed spherules, and saline alone. Thirty days following intranasal challenge with 1,500 and 15,000 arthroconidia, we found that the thimerosal-killed spherules (7/7 and 5/7, respectively) and the T-27K vaccine (7/7 and 4/7, respectively) protected mice from lethal infection as well as the formalin-killed spherules (7/7 and 4/7, respectively). All the saline vaccinated mice died at both challenge levels. When lymphocyte proliferation assays were performed using PBMCs from either skin-test positive or negative human donors, we obtained similar proliferation responses from the positive donor PBMCs when using either the F-27K or T-27K vaccine. PBMCs from negative donors did not produce a significant proliferation response due to either vaccine. When the T-27K vaccine was fractionated on SDS-PAGE gels, numerous individual, well-defined protein bands were present compared to the continuous, unresolved smear produced by the F-27K vaccine. We have also used FPLC chromatofocusing and FPLC gel filtration to separate the T- 27K vaccine into fractions that show resolved, individual protein bands upon separation by SDS-PAGE. In addition, we have been able to detect chitobiase, alkaline phosphatase, chitinase, and protease activity in separate fractions derived from our chromatographic separations. We conclude that we have developed a new approach to the isolation of individual immunogens from the endosporulating spherules of C. immitis.

Publication Types:

• Meeting Abstracts

Keywords:

• Animals
• Coccidioides
• Communicable Diseases
• Fungal Vaccines
• Humans
• Infection
• Mice
• Mice, Inbred BALB C
• Mice, Inbred C57BL
• Mice, Inbred DBA
• Muridae
• Pregnancy Complications, Infectious
• Vaccines
• Vaccines, Inactivated
• Viral Vaccines

Other ID:

• 20712173

UI: 102195703

From Meeting Abstracts