Transgenic mosquitoes in controlling malaria transmission.

Abstract

The advent of molecular tools to manipulate insects, such as Drosophila melanogaster, genetically presents researchers with the possibility to apply these techniques to vectors of insect-borne diseases, such as malaria. Also, with these techniques, in theory, researchers could generate a mosquito that blocks development and transmission of the malaria parasite. Furthermore, these transgenic mosquitoes could cross with wild-type mosquitoes in natural environments, dispersing the antiparasitic genes in the population. To begin such a tour de force effort, emphasis was first placed on developing successful genetic transformation of the insect vector, using strong promoters that could drive expression of exogenous genes in abundant quantities and in a tissue-specific manner. The development of sterile insect techniques and dominant lethals also provided possible avenues for blocking disease transmission [1xDominant lethality and insect population control. Alphey, L. and Andreasen, M. Mol. Biochem. Parasitol. 2002; 121: 173–178Crossref | PubMed | Scopus (85)See all References][1].When an Anopheles mosquito ingests Plasmodium gametocytes during a bloodmeal, the cells develop into gametes, mate, and differentiate into zygotes. The zygote stage develops further into ookinetes, which pass through the midgut epithelium and develop into oocysts. This process takes ∼10–15 days and leads to the production of sporozoites. The development of the parasite in the mosquito is then completed when sporozoites cross the salivary gland epithelium. The mechanism of penetration of the salivary gland epithelium is not clear, but there are findings that suggest that the process might be specific, involving receptor–ligand interactions. To this end, a 12-amino acid peptide (called SM1) has been identified from a bacteriophage library, and binds in a specific manner to two epithelia types in mosquitoes (salivary glands and midgut). This peptide also inhibited the crossing of malaria parasites into these two cell types. This suggests that if SM1 could be synthesized and secreted into the mosquito midgut lumen, then perhaps Plasmodium development could be halted when a bloodmeal is ingested.This hypothesis was tested by J. Ito et al. using Anopheles stephansi transformed with a DNA cassette containing a chimeric gene with four SM1 units joined by four amino acid linkers attached to the carboxypeptidase signal sequence and a hemagluttinin (HA) epitope tag [2xTransgenic anopheline mosquitoes impaired in transmission of a malaria parasite. Ito, J. et al. Nature. 2002; 417: 452–455Crossref | PubMed | Scopus (361)See all References][2]. The cassette was flanked by a gut-specific promoter from the carboxypeptidase gene (to drive robust expression of the SM1 peptide) and necessary selectable marker genes. Carboxypeptidase is an abundant protease present in the lumen of the mosquito midgut. The luminal-targeting signal sequence present in the carboxypeptidase protein also served to sort the peptide to the midgut lumen. Ito et al. found that these transgenic mosquitoes inhibited oocyst formation by ∼80% compared with mosquitoes containing control plasmids and wild-type mosquitoes. Integration of the gene cassette was confirmed by Southern blot analysis, and expression of the chimeric gene was confirmed by mRNA analysis. The presence of the SM1 peptide was detected using differential interference contrast microscopy with an anti-HA antibody. Sporozoite prevalence, sporozoite intensity and vector competence (ability to transmit parasites to a naive mouse) were all dramatically reduced in transgenic mosquitoes compared with control mosquitoes.This marked reduction in parasite development and transmission by expressing an antiparasitic gene represents a significant stride forward in our potential to control malaria. However, it must be noted that this study used a rodent malaria parasite under a controlled environment. Extrapolating this finding to human malaria and controlling the disease in natural ecosystems will require further studies of the development of malaria parasites of humans in the mosquito, as well as the ecology of mosquito population control in the field.