Optimization of the Feeding rate of Anopheles Farauti Colony Mosquitoes in Direct Membrane Feeding Assays

Abstract

Background: Direct membrane feeding assays (DMFA) are an important tool to study parasite transmission to mosquitoes. Mosquito feeding rates in these artificial systems require optimization as there are a number of factors that potentially influence the feeding rates and there are no standardized methods that apply to all Anopheline species.Methods: A range of parameters prior to and during direct membrane feeding (DMF) were evaluated for their impact on Anopheles farauti feeding rates; including the starving conditions and duration of starving prior to feeding, membrane type, DMF exposure time, mosquito age, illumination level, blood volume, mosquito density and temperature of water bath. Results: The average successful DMFA feeding rate for An. farauti colony mosquitoes increased from 50 % to 85 % when assay parameters were varied. Overnight starvation and Baudruche membrane yielded the highest feeding rates but rates were also affected by blood volume in the feeder and the mosquito density in the feeding cups. Availability of water during the pre-feed starvation period did not significantly impact feeding rates, nor did the exposure duration to blood in membrane feeders nor, the age of mosquitoes (3, 5 and 7 days post-emergence), illumination during feeding or the temperature (34 °C, 38 °C, 42 °C and 46 °C) of the water bath.Conclusion: Optimal feeding conditions in An. farauti DMFA were to offer 50 female mosquitoes in a cup (with a total surface area of ~340 cm2 with 1 mosquito / 6.8 cm2) that were starved overnight 350-500 µL of blood (collected in heparin coated vacutainers) per feeder in feeders with a surface area ~5 cm2 (with a maximum capacity of 1.5 mL of blood) via a Baudruche membrane, for at least 10-20 min.

Modified who tunnel test for high through put evaluation of insecticide-treated nets considering the effects of hosts, exposure time and mosquito density

“…The experimental time for arm feeding was 1 h to allow for standardized evaluation and to minimize volunteer discomfort. Previous work has shown that 30 minutes of exposure resulted in high blood feeding (Timinao et al, 2021).…”

“…Membrane feeding has been widely deployed for evaluating topical mosquito repellents (Debboun, 2004), transmission-blocking drugs and vaccines (Vallejo, et al, 2016), endoctocides (Smit, 2018) as well as for mosquito rearing (Awono- Damiens, 2013;Kaufman, 2014;Phasomkusolsil et al, 2013;Robert, 1998;Timinao et al, 2021). Moreover, the use of an artificial membrane has several advantages including reduced animal welfare or ethical concerns, reduced chance of accidental disease transmission, simple logistics, and reproducibility Kaufman, 2014;Phasomkusolsil et al, 2013;Timinao et al, 2021). Given the significance of host kairomones to encourage mosquito feeding, worn socks added to augment the attractiveness of the membrane to mosquitoes (Pates et al, 2001) during host seeking process.…”

“…As would be expected, with 50 mosquitoes there is a slight increase in blood feeding success and a consequent slight decrease in mortality compared to assays using 100 mosquitoes. Higher feeding success at lower density is likely due to less competition between mosquitoes on the membrane during host-seeking (Timinao et al, 2021), which may also reduce the host defensiveness of the rabbit (Anderson & Brust, 1996;Edward, 1986). Increasing the number of mosquitoes in the tunnel may lead to density-dependent mortality effects of crowding as mosquitoes can collide each other when at high density (Styer et al, 2007).…”