In vitro and in silico characterization of the anticholinesterase activity of select terpenoids against anopheles vectors

Malaria is a life-threatening plasmodial disease that is transmitted by female Anopheles mosquitoes. Major African malaria vectors include Anopheles arabiensis, An. funestus, An. gambiae and An. coluzzii. Malaria vector control programs have shown effectiveness in reducing the Anopheles populations. The main insecticide classes used in these interventions include pyrethroids, organochlorines, organophosphates, carbamates, and neonicotinoids. Nevertheless, the development of Anopheles resistance to these insecticide classes has greatly reduced the effectiveness of these interventions. A common resistance mechanism is through rapid detoxification of insecticides by overexpressed P450 monooxygenases. Although acetylcholinesterase (AChE) is a valid target in Anopheles vector, current anticholinesterase insecticides suffer from resistance and low selectivity between insect and mammal AChE targets. This indicates the urgent need to discover novel AChE inhibitors with higher affinity to Anopheles AChE compared to the mammal target, and less prone to resistance caused by the overexpressed monooxygenases. Identification of novel AChE inhibitors from natural sources and their potential to kill Anopheles during all its different life stages, presents a cost-effective approach. This PhD study aimed to identify such novel AChE inhibitors from essential oil sources and assess them for consistent activity against Anopheles species with hyperactive P450 monooxygenases. In this study, molecular differences between Anopheles and human AChEs were identified showing the opportunity to develop selective Anopheles AChE inhibitors. A novel approach was used to integrate the in silico and in vitro assays in assessing the Anopheles AChE inhibitory potential of select terpenoids and coupled these to the in vivo assays against different life stages of Anopheles. The terpenoids, farnesol, (-)-α-bisabolol, cisnerolidol, trans-nerolidol, and methyleugenol were identified as potent Anopheles AChE inhibitors and larvicidal agents with moderate adulticidal effects. Farnesol and (-)-α-bisabolol also displayed pupicidal activity, while methyleugenol inhibited the hatching of Anopheles eggs. Generally, farnesol and (-)-α-bisabolol were highly active across the Anopheles species, except in the strain with P450-based metabolic resistance. In contrast, the efficacy of cisnerolidol, trans-nerolidol, and methyleugenol was not affected by this resistance mechanism. This research suggests that cis-nerolidol, trans-nerolidol, and methyleugenol are potential candidates for further development as anticholinesterase bioinsecticides.
A thesis submitted in fulfilment of the requirements for the degree of Doctor of Philosophy to the Faculty of Health Sciences, School of Anatomical Sciences, University of the Witwatersrand, Johannesburg, 2023
Malaria, Anopheles, Life cycle, Terpenoids, Monooxygenases