THESIS: A.M. Moura da Silva submitted this dissertation for the degree of Doctor of Microbiology and Immunology publicly examined at the Department of Microbiology and Immunology of the Paulista Medicine School, São Paulo, Brazil in 1991.
Advisor: Professor Ivan Mota
ABSTRACT. Antigenic cross-reactivity among components of venoms from nine different species of Bothrops regarding isolation and characterization of private antigens was studied. Antisera against each venom showed similar levels of antibody when either homologous or heterologous venoms were used as antigens. Transblotted antigens after SDS-PAGE fractionation were also revealed by homologous and heterologous antivenoms. Antigens with molecular weight higher than 30 kDa seemed to be the most cross-reactive. Antigens with molecular weight between 14 and 18 kDa showed cross-reactivity only for Bothrops jararacussu, Bothrops moojeni, Bothrops neuwiedi and Bothrops pradoi venoms. Venoms of the nine different species studied were fractionated using Fast Phase Liquid Column (FPLC). Basic proteins were isolated from B. jararacussu, B. moojeni, B. neuwiedi and B. pradoi venoms. Bothrops jararaca venom presented very low concentration of these proteins, which were undetectable in Bothrops alternatus, Bothrops atrox, Bothrops cotiara and Bothrops erythromelas venoms. All active fractions showed a common band of 15 kDa, which led to a rise in serum creatine kinase levels and to histopathological changes in muscle cells following intra-muscular injection in mice. Levels of phospholipase A2 activity were variable. Myotoxicity induced by crude venoms confirmed that these antigens were possibly the only major myotoxins, as the levels of creatine kinase activity in mouse serum released by intramuscular injection of B. jararacussu, B. moojeni, B. neuwiedi and B. pradoi venoms were five to eight times as high as those obtained for B. alternatus, B. atrox, B. cotiara, B. erythromelas and B. jararaca venoms. Using double immunodiffusion technique, myotoxins of B. jararacussu, B. neuwiedi and B. pradoi showed total identity, while B. moojeni myotoxin behaved as a partially identical antigen. A phospholipase myotoxin (MOO-1) and a non-phospholipase myotoxin (JSU-5) were studied for their antigenic cross-reactivity and neutralization by different antisera. Antisera against JSU-5 and MOO-1 reacted equally with both myotoxins in ELISA. Specificity of these antisera was also similar, recognizing the same 14 to 18 kDa antigens in B. jararacussu, B. moojeni, B. neuwiedi and B. pradoi venoms. Creatine kinase assays showed that JSU-5 myotoxicity was completely neutralized by B. jararacussu antivenom or anti-JSU-5 antibodies and partially neutralized by anti-MOO-1 antibodies or B. moojeni antivenom. MOO-1 myotoxicity was completely neutralized by antisera against JSU-5, MOO-1 and B. jararacussu antivenom, and only partially neutralized by B. moojeni antivenom. Among the myotoxins containing venoms, B. jararacussu venom induced the highest titers of anti-myotoxin antibodies. This antiserum completely inhibited myotoxicity of homologous venom and significantly reduced myotoxicity of B. moojeni, B. neuwiedi and B. pradoi venoms. The implication of these results in serotherapy neutralization of systemic and local effects of Bothrops venoms was discussed.
CORRESPONDENCE TO:
A.M. MOURA DA SILVA - Laboratório de Imunopatologia - Instituto Butantan - Av. Vital Brazil 1500 - CEP 05503-900 - São Paulo - SP - Brasil.
Publication Dates
-
Publication in this collection
08 Jan 1999 -
Date of issue
1995
