探討白線斑蚊腸道H+ V-ATPase的表現與簇蟲感染的關係

Abstract

Gregarines (Apicomplexa, Eugregarinida, Lecudinidae) are common parasites of mosquitoes. The life cycle of Ascogregarina is completed in the insect with its development synchronized with the host. The mosquito Aedes albopictus acquires the A. taiwanensis from the ingestion of its oocysts in the breeding water. Within the digestive tract, the sporozoites emerge from the oocysts and enter the epithelial cells. After a short duration of intracellular development, the trophozoite attaches to the luminal side of the midgut cell by the epimerite. During the mosquito pupation, the gregarine migrates into the Malpighian tubules to begin the gametogenous phase. The gregarine sporozoites are released by ingested oocysts in the high pH environment. In mosquito larvae, V-ATPase plays an important role in regulating luminal alkalization of midgut. The V-ATPase is a family of ATP-dependent proton pumps and usually located on the endomembrane of eukaryotic cells. Acidification of intracellular compartments such as lysosomes, endosomes, and parasitophorous vacuoles medicated by V-ATPase are essential for the entry of many enveloped viruses and invasion into or escape from host cells. We had cloned and expressed the full-length V-ATPase subunit A protein from mRNAs extracted from Ae. albopictus larval tissues. This open reading frame (ORF) and 3’ uncoding sequences were 2971 base pairs in length. The rabbit anti V-ATPase subunit A antiserum was prepared and was used to detect V-ATPase in the midgut of mosquito larvae. Antibody-binding sites were observed to be located at the basal of anterior midgut and the apical of posterior midgut. Longitudinal section of infected larva showed gregarine located in the posterior midgut. In response to the invasion of A taiwanensis, a parasitophorous vacuole confining intracellular trophozoites was formed; within which acidification was demonstrated by using acridine orange staining. Mosquito treated with bafilomycin A1, a specific inhibitors of V-ATPase, showed a dramatic reduction in intensity of infection by gregarina. Therefore, V-ATPase may play a crucial role in determining the infection of gregarine.