Transgenic Anopheles stephensi coexpressing single-chain antibodies resist Plasmodium falciparum development

Abstract

Anopheles stephensi mosquitoes expressing m1C3, m4B7, or m2A10 single-chain antibodies (scFvs) have significantly lower levels of infection compared to controls when challenged with Plasmodium falciparum, a human malaria pathogen. These scFvs are derived from antibodies specific to a parasite chitinase, the 25 kDa protein and the circumsporozoite protein, respectively. Transgenes comprising m2A10 in combination with either m1C3 or m4B7 were inserted into previously-characterized mosquito chromosomal "docking" sites using site-specific recombination. Transgene expression was evaluated at four different genomic locations and a docking site that permitted tissue-and sex-specific expression was researched further. Fitness studies of docking site and dual scFv transgene strains detected only one significant fitness cost: adult docking-site males displayed a late-onset reduction in survival. The m4B7/m2A10 mosquitoes challenged with P. falciparum had few or no sporozoites, the parasite stage infective to humans, in three of four experiments. No sporozoites were detected in m1C3/m2A10 mosquitoes in challenge experiments when both genes were induced at developmentally relevant times. These studies support the conclusion that expression of a single copy of a dual scFv transgene can completely inhibit parasite development without imposing a fitness cost on the mosquito.mosquito vector | population replacement | genetic engineering

Concerning RNA-Guided Gene Drives for the Alteration of Wild Populations

“…These afflictions could potentially be controlled or even eradicated by altering vector species to block transmission. Several laboratories have identified candidate gene disruptions or transgenes that interfere with the transmission of malaria 97,103,104 and other well-studied diseases 105 . Depending on their effectiveness, these alterations may or may not allow the disease to be eradicated before the pathogen evolves resistance.…”

Efficient ΦC31 integrase–mediated site-specific germline transformation of Anopheles gambiae

“…stephensi 14,31 . Water-saturated Halocarbon oil Mix 9 ml Halocarbon Oil 700, 1 ml Halocarbon Oil 27 and 10 ml molecular biology grade water gently to avoid forming an emulsion.…”

“…To surmount this, several researchers have used Drosophila or Anopheles stephensi as a recipient species to analyse the function of An. gambiae genes [10][11][12][13][14] as these species are more robust and amenable to genetic transformation, and in the case of Drosophila, possess a complex genetic toolset. However, these heterologous systems have their limitations, since presumed orthologous genes may behave differently in An.…”

Gene Drive Strategies for Population Replacement

“…Several approaches are being explored to engineer mosquitoes unable to transmit human diseases, and there have been a number of notable successes. For example, Isaacs et al have engineered Anopheles stephensi mosquitoes expressing single-chain antibodies that prevent Plasmodium falciparum malaria parasites from developing in the mosquito, thus preventing onward transmission of the parasite [1]. Gene drive systems are expected to be instrumental in spreading disease-refractory genes into wild mosquito populations, given the wide geographical areas that these species inhabit and the expectation that refractory genes will be associated with at least modest fitness costs [2].…”