application abstract): Within the Anopheles vector, two forms of Plasmodium parasites are both termed """"""""sporozoites"""""""". Oocyte- associated sporozoites are extremely infectious for the salivary gland of the mosquito. Salivary gland-associated sporozoites are, in turn, extremely infectious for the target organs in the vertebrate host. Upon commencing her postdoctoral work, the candidate wrote an extremely successful application proposing the construction of cDNA libraries from the oocyte-associated and salivary gland-associated sporozoites of Plasmodium gallinaceum, whose life cycle oscillates between Anopheles mosquitoes and chickens. Differential screening of these two libraries was proposed, with the goal of identifying genes that would discriminate between these two sporozoite stages. The applicant began postdoctoral work in 1985. Oocyte-associated and salivary gland-associated cDNA libraries were constructed, and shown to contain previously characterized sporozoite-specific transcripts. Differential screening identified two cDNAs, termed Pg19 and Pg93, that were potentially differentially expressed in the oocyte-associated sporozoites. Hybridization to a known P. falciparum surface protein identified another gene, Pg4. Subsequent characterization convinced the applicant that Pg19 and Pg93 were not differentially expressed in the two sporozoite forms, and that Pg4 was not a homolog of the P. falciparum surface protein. However, sequence analysis of the three cDNAs revealed that Pg4 is likely to have a signal sequence and therefore is possibly expressed on the cell surface. In the current proposal for a K22 award, the applicant plans to complete the sequencing characterization of Pg4, Pg19 and Pg93 to obtain complete sequence information of both cDNAs and genomic DNAs. RT-PCR, Northern analysis and in situ hybridization will be performed to determine the size and abundance of the mRNAs in various infected tissues of the mosquito. The second specific aim is to purify recombinant proteins and to make antibodies. The antibodies will be used for immunolocalization studies, to determine the intracellular location of the Pg4, Pg19 and Pg93 proteins and to determine their localization within the infected mosquito and chicken. The third specific aim is to test whether the purified proteins or antibodies can block sporozoite infectivity either into the salivary gland of the mosquito or the target tissues of the chicken.
| Beerntsen, Brenda T; Li, Jianyong (2006) Plasmodium development in white-eye (kh(w)) and transformed strains (kh43) of Aedes aegypti (Diptera: Culicidae). J Med Entomol 43:318-22 |
| LaCrue, Alexis N; Sivaguru, Mayandi; Walter, Marika F et al. (2006) A ubiquitous Plasmodium protein displays a unique surface labeling pattern in sporozoites. Mol Biochem Parasitol 148:199-209 |
| Lacrue, Alexis N; James, Anthony A; Beerntsen, Brenda T (2005) The novel Plasmodium gallinaceum sporozoite protein, Pg93, is preferentially expressed in the nucleus of oocyst sporozoites. Am J Trop Med Hyg 73:634-43 |
