DEVELOPMENT OF A CONJUGATE VACCINE AGAINST STAPHYLOCOCCUS AUREUS BOVINE MASTITIS USING CAPSULAR POLYSACCHARIDES.

Introduction:

Mastitis is the most significant cause of economic loss to the dairy industry, and bacteria are the most common cause of mastitis. Staphylococcus aureus is the most ubiquitous and results in the greatest economic loss (Foster, 1986). Staphylcoccus aureus mastitis occurs despite numerous host defense mechanisms. Of these defense mechanisms, neutrophil phagocytosis is the most effective. However, 10high concentration of milk neutrophils (9 ×⁵ neutrophils/ml) is required to prevent infection. Because this concentration far exceeds the number of neutrophils in the normal healthy gland, increasing the phagocytic efficiency of neutrophils via opsonins is the most effective way of enhancing mammary gland resistance to S. aureus infection. A major obstacle to producing a protective immune response to S. aureus is the development of capsular polysaccharides (CP). It has been estimated that 94% to 100% of S. aureus isolated from cows possess CP (Norcross and Opdebeeck. 1993). The CP block neutrophil recognition of antibodies bound to the highly antigenic cell wall components of S. aureus. Polysaccharides are low in immunogenicity and do not readily elicit an antibody response. In addition to being low in immunogenicity, polysaccharides are T-cell independent antigens and thus do not stimulate immunological memory. However, it has been shown that antibodies to CP enhance phagocytosis (Guidry et al. 1994). Progress has been made toward increasing the production of antibodies to CP and stimulation of immunological memory by conjugating CP to carrier proteins (Fattom et al, 1993).

Objective:

To prepare S. aureus mastitis vaccine using capsular polysaccharides conjugated to carrier proteins and evaluate its efficacy in increasing antibody production to capsular polysaccharides, and stimulation of immunological memory.

Methodology:

S. aureus serotypes 5, 8 and 336 will be isolated from clinical and subclinical cases of mastitis. S. aureus serotypes 5, 8 and 336 capsular polysaccharides (CP) will be prepared by treartment with lysostaphin and multiple alcohol purification followed by ion exchange chromatography. S. aureus serotypes 5,8 and 336 CP will be conjugated to purified carrier protein recombinant Pseudomonas aeruginosa exotoxin A as described by Fattom et al, 1993. In this study, cows will be immunized with a vaccine consisting of the conjugated S. aureus CP serotypes 5,8 and 336 emulsified in adjuvant and conjugated CP in microspheres emulsified in adjuvant. The antibody response will be tested for specificity, isotype, enhancement of neutrophil phagocytosis of three S. aureus serotypes, and prevention of S. aureus adherence to bovine mammary epithelial cells.

References:

• Fattom, A., R. Schneerson, D. C. Watson, W. W. Karakawa, D. Fitzgerald, I. Pastan, X. R. Li, J. Shiloach, D. A. Bryla, and J. B. Robbins. 1993. Laboratory and clinical evaluation of conjugate vaccines composed of Staphylococcus aureus type-5 and type-8 capsular polysaccharides conjugated to Pseudomonas aeruginosa recombinant exoprotein-A. Infect. Immun. 61:1023–1032.
• Foster, T. J. 1986. A new genetic approach to defining the virulence determinants of Staphylococcus aureus strains that cause bovine mastitis. Ir. Vet. J. 40:110–115.
• Norcross, N. L., and J. P. Opdebeeck. 1993. Encapsulation of Staphylococcus aureus isolated from bovine milk. Vet. Microbiol. 8:397–404.
• Guidry, A. J., C. N. O’Brien, S. P. Oliver, H. H. Dowlen, and L. W. Douglass. 1994. Effect of whole Staphylococcus aureus and mode of immunization on bovine opsonizing antibodies to capsule. J. Dairy Sci. 77:2965–2974