Investigating life history characteristics of Anopheles arabiensis infected with Microsporidia MB, a Plasmodium falciparum blocking symbiont

Abstract

Introduction: Microsporidia MB is a naturally occurring symbiont found in Anopheles arabiensis that blocks the transmission of the Plasmodium parasite without any fitness effect on the mosquito. Microsporidia MB proliferates in mosquito gonads leading to its high intensity, which is linked to horizontal (sexual) and vertical (transovarial) transmission from one mosquito to another. Developing this mosquito-symbiont into a control tool is highly needed to complement the core malaria control tools. How diet affects Microsporidia MB intensity and the Microsporidia MB-infected mosquitoes, leading to the identification of diet regimes for mass production of high Microsporidia MB intensity mosquitoes is unknown. Furthermore, how Microsporidia MB spread in the infected mosquito population over generations is also unknown. The first aim of this study was to investigate how diet type and quantity affect the Microsporidia MB-An. arabiensis life table parameters, namely, larval development and mortality, adult size and survival, as well as Microsporidia MB intensity in both larvae and adults. The second aim was to monitor the spread of Microsporidia MB in infected An. arabiensis populations. Methods: The research was conducted using Microsporidia MB-infected and uninfected first filial generation (F1) larvae or adults from engorged females collected from the field (G0). The F1 larvae were reared together (group lines, GLs) or as individual families (eggs from one female also called isofemale lines, IMLs). GLs were fed on 0.3mg/larva/day of Tetramin, GoCat, and Cerelac respectively as well as 0.07 mg/larva/day of Tetramin. Adult GLs were fed on 1% or 6% glucose and their survival was determined. IMLs, on the other hand, were fed on Tetramin 0.07 and 0.3 mg/larva/d for the larvae and 1% or 6% glucose for the adult experiments respectively. Diet choices were selected based on commonly used diets and previous literature. Finally, three colonies (biological replicates) of Microsporidia MB-infected mosquitoes were established from F1 larvae and maintained until the sixth filial generation (F6) to observe the vii symbiont spread throughout the Microsporidia MB colony. Prevalence and intensity of Microsporidia MB, wing length of mosquitoes, humidity and temperature of the laboratory were recorded. Results: Microsporidia MB infected An. arabiensis fed on Tetramin at a dose of 0.3 mg/larva/day had the quickest larval growth, greatest adult emergence, largest mosquito body size, highest prevalence, and highest density of Microsporidia MB. In contrast to 6% glucose, 1% glucose did not prolong the lifespan of adult Microsporidia MB-infected mosquitoes. Microsporidia MB-infected and uninfected An. arabiensis fed on Tetramin at a dose of 0.07 mg/larva/day and 1% glucose had similar development time and adult survival respectively, showing that the fitness benefit conferred to the mosquito host was diet-dependent. Furthermore, the results on the Microsporidia MB colony showed two replicates progressed to F6 successfully while the one collapsed (died out) at F2. Prevalence of Microsporidia MB increased from F1 to F3 generation and declined subsequently. Furthermore, the intensity of Microsporidia MB was not significantly different between the F1 and F6 in both replicates. There was a slight positive correlation between Microsporidia MB prevalence and temperature. The size of the male mosquitoes was not reduced across the generations while that of females fluctuated. Conclusion: Even on restricted diets, Microsporidia MB did not adversely affect An. arabiensis developmental growth or fitness. Tetramin 0.3 mg/larva/day and 6% glucose are the best diet regimes for rearing large number of Microsporidia MB-infected mosquitoes. These findings are crucial for mass rearing of high-intensity Microsporidia MB-infected An. arabiensis mosquitoes in the laboratory for the purposes of experiments and field releases. Furthermore, this first successful Microsporidia MB colony establishment attempt shows the pattern of Microsporidia MB prevalence across subsequent generations, and this understanding offers a pathway for enhancing rearing protocols to sustain high Microsporidia MB prevalence viii mosquito colonies. The maintenance of Microsporidia MB intensity in the mosquitoes between the starting and the last generation gives credence to the potential of the Microsporidia MB malaria control strategy. Lastly, the fitness of the male Microsporidia MB-infected An. arabiensis across generations observed in this study demonstrates that a male release strategy is feasible.