Targeting the Highly Invasive Malaria Vector Anopheles stephensi using Yeast RNAi Pesticides

Apart from widespread resistance of malaria mosquitoes to insecticides, Plasmodium parasite resistance to frontline anti-malaria drugs, and challenges in malaria diagnosis, the World Health Organization (WHO) has described the highly invasive Anopheles stephensi as a major threat to malaria control. New classes of insecticides are vitally needed for integrated control of the dangerous malaria vector that continues to spread across African countries. Yeast RNAi insecticides are promising novel pesticides that could prove effective for integrated responses to A. stephensi. Here we explore the use of RNAi yeast pesticides for control of this invasive malaria vector.

Sh.463, a modified Saccharomyces cerevisiae baker's yeast RNAi pesticide corresponding to the A. stephensi Shaker gene, was evaluated in A. stephensi. A scalable attractive targeted sugar bait (ATSB)-based system for delivery of Sh interfering RNA pesticides (IRPs) to adult A. stephensi under insectary conditions is examined, and a yeast-based system for delivery of Sh IRP to larvae is developed and evaluated. Additionally, female-specific yeast RNAi-larvicides targeting putative A. stephensi female-specific genes for male mosquito sorting are also developed and evaluated in laboratory assays.

We demonstrate that the treatment of A. stephensi larvae and adults with Sh.463-56.10R yeast silences the mosquito Shaker gene, resulting in high levels of mortality in laboratory studies. Additionally, our siRNA screens of putative female-specific genes in A. stephensi using female-specific RNAi yeast larvicides resulted in significant female mortality in cup bioassays leading to significantly higher male: female ratios in the resulting offspring.

The results of these studies suggest that an RNAi pesticide targeting mosquito Shaker genes may represent a novel biorational intervention that can be used in integrated A. stephensi control programs while also targeting other species of disease vector mosquitoes. The potential of A. stephensi female-specific RNAi yeast larvicides in male mosquito sorting is also described.