Heat Shock Factor (HSF) Expression and Its Effect on Life Table Parameters in the Main African Malaria Vector Anopheles funestus

Abstract

Anopheles funestus is a major malaria vector, which is predominantly responsible for malaria transmission in Africa and this vector needs to be controlled due to the devastation of the disease. Temperatures worldwide are rising steadily, which can affect mosquito distribution and behaviour. Heat shock factor (HSF), a transcription factor, is crucial in these physiological and behavioral processes. This is because HSF has a high degree of functional variability and is temperature dependent, enabling mosquitoes to survive the heat by repairing damaged proteins in the cell. This study aimed to understand the biological role of HSF in the African malaria vector An. funestus with regard to HSF transcript abundance and its effect on life table parameters. In this work, the HSF gene was the target of RNA interference (RNAi). RNAi was implemented using short interfering RNA (siRNA) that was previously synthesised and was performed to understand the role of HSF in An. funestus. HSF is a poorly understood gene, yet it has been found to have a variety of functions within insects and, more specifically, mosquitoes. Understanding the role of HSF in An. funestus is important to advance knowledge in this understudied vector. HSF transcript abundance was measured using real-time quantitative PCR (qPCR) throughout the four life stages in An. funestus in order to assess the average expression levels of HSF at 25°C. The HSF transcript abundance was significantly lower during the egg and pupal stages in comparison to the larval and adult stages. Inhibition, via siRNA, of the HSF transcript was subsequently used to understand its biological role in An. funestus. A system was designed to deliver the siRNA to the adult using the ingestion of sugar water. This was the chosen delivery system due to its ease of use and the potential to replicate this method in the wild because of the sugar-feeding behaviour of the adult mosquitoes. A known reference gene, RPL19, was used to evaluate the efficiency of vii a “siRNA-sugar feeding delivery system” in An. funestus. Results showed this would be an effective system for evaluating transcript inhibition or knockdown. Different concentrations of HSF-siRNA were utilised, 10 ng/μl, 20 ng/μl and 30 ng/μl at three different temperatures of 20°C, 25°C and 30°C, to evaluate the impact of different concentrations and temperatures on the fold change of HSF transcript abundance after siRNA treatments. A 20 ng/μl siRNA concentration at 25°C reduced the HSF transcript abundance. Therefore, these conditions were selected to study the phenotypic effect of HSF-siRNA on longevity and reproduction in An. funestus. Longevity, fertility, fecundity, and time-to-hatch were all reduced when mosquitoes were treated with HSF- siRNA. In addtion, the impact of decreased survivability in males was more prominent than in females. In conclusion, HSF might regulate vector development and needs to be investigated in the future. Furthermore, this study confirms the pleiotropic effect of HSF in An. funestus.