Identification of lambda gt11 clones encoding the major antigenic determinants expressed by Theileria parva sporozoites

Abstract

An antiserum, C16, was raised in cattle against freeze-thawed extracts of sporozoites of Theileria parva (Muguga). This antiserum, which neutralizes sporozoite infectivity in vitro, identified theileria-specific antigens having approximate molecular masses of 105, 90, 85, 69, 67, 52, 47, and 43 kilodaltons (kDa) on Western blots (immunoblots) of infected tick salivary gland extracts. The antiserum was used to screen an expression library of T. parva (Muguga) genomic DNA fragments. Three recombinant bacteriophage clones carrying different theileria DNA inserts were studied. The expressed gene product from each clone was used to affinity purify antibodies from C16 antiserum for use in probing Western blots of uninfected and infected tick salivary gland extracts. The population of antibodies selected by each clone specifically recognized a subset of the antigens identified by C16 antiserum. The antigens fell into three distinct groups as defined by their reactivity with each set of selected antibodies. One group included antigens of 105, 90, 85, and 35 kDa, a second group included antigens of 69, 67, 52, 47, and 43 kDa, and the third group included an apparently distinct pair of antigens of 47 and 43 kDa. Thus, antibodies that reacted with determinants encoded by the three recombinant phage clones recognized all of the major antigens seen on Western blots probed with whole C16 antiserum. These results suggest that there may be only three immunodominant antigens expressed in T. parva (Muguga) sporozoites. Additionally, monoclonal antibodies have been raised which neutralize sporozoite infectivity in vitro. These antibodies react with epitopes of the antigens with Mrs of 69,000, 67,000, 52,000, 47,000, and 43,000 which are encoded in clone pgT-42 and have been used to localize these epitopes on the sporozoite surface.