Large-scale Anopheles arabiensis egg quantification methods for mass-rearing operations

Abstract

BackgroundThe success of the sterile insect technique relies, among other things, on the continuous release of over flooding numbers of sexually competitive sterile males into the target area. To produce sufficiently large quantities of sterile males, rearing protocols need to be optimized including the development and validation of a standardized egg quantification method.MethodsBatches of 1000 freshly laid eggs collected from standard rearing cages were counted, gently dried under laboratory conditions (27 ± 1 °C, 75 ± 5 % RH) and combined so that 1000–8000 eggs were weighed, to calculate the correlation between weight and number. The actual counted egg number and the egg number estimated by weighing were further compared for samples of 1000, 3000 and 4000 eggs collected from both standard and mass-rearing cages. The effect of drying, brushing and weighing on egg hatch rate was evaluated in three samples each of 1000 fresh and 1000 dried eggs, and in batches of 1000, 3000 and 4000 dried eggs. Pupal production and adult life history traits were assessed for dried eggs hatched and reared in mass-rearing trays. Expected egg numbers and actual observed mean egg numbers were compared after gentle drying, and after applying a rapid drying method exposure to wind speed of 1.8 m/s for 30 min.ResultsA significant positive relationship between the number of dried eggs and egg weight was observed and the equation ‘Weight (mg) = (0.00399 × Number of counted eggs) + 0.536 was derived. The actual counted mean egg number and the egg number estimated by weighing were similar for samples from small rearing cages but significantly lower for samples of 3000 and 4000 egg samples collected from mass-rearing cages. No negative effect of the drying, brushing and weighing process on egg hatch rate was observed. No significant difference was observed in any life history trait between adults reared from dried or from fresh eggs up to twenty-one days post emergence. The mean number of eggs counted from a given replicate’s weight was significantly higher for egg batches fast dried with a suction device compared to those dried with a gentle drying method (fast: 1075 ± 9, gentle: 1024 ± 7).ConclusionAn equation has been derived to allow accurate quantification of dried Anopheles arabiensis eggs based on weight, enabling more accurate quantification of eggs for consistent larval rearing density to be achieved. Eggs can be dried for weighing in a manner which does not impair the quality of resulting adults.

A rapid quality control test to foster the development of the sterile insect technique against Anopheles arabiensis

“…Larvae were mass-reared in plastic trays held in a mechanized stainless-steel rack developed at the IPCL [6]. Eggs were quantified using the method defined in Maiga et al [7] wherein 4000 eggs are added to each tray, containing 4 l of deionized water, within a plastic ring which floats on the water surface. Water was added to each tray 24 h before the addition of the eggs to allow the temperature of the water to acclimatize to that of the ambient air temperature.…”

“…A build-up of insecticide resistance coupled with the global spread of species, such as Aedes aegypti and Aedes albopictus, has reignited interest in developing mosquito SIT as part of an AW-IPM approach, with the technique progressing towards implementation at an operational level. Mass-rearing technology has been standardized [5][6][7] and guidelines created for key vectors including Anopheles arabiensis [8], Ae. aegypti and Ae.…”

Optimization of mosquito egg production under mass rearing setting: effects of cage volume, blood meal source and adult population density for the malaria vector, Anopheles arabiensis

“…arabiensis mass-rearing procedure developed at the IPCL [7, 16, 19, 20]. Eggs were hatched and larvae reared to pupation in the larval mass rearing rack developed by Balestrino et al [7, 8].…”

“…Each tray was filled with 4 L of de-ionized water the day before adding the eggs to allow the water to reach room temperature. Using the egg quantification method developed by Maiga et al [20], 50 aliquots of 4000 eggs were added to each tray in a plastic ring floating on the surface of the water. Larvae were fed with the IAEA liquid diet (5 g/L tuna meal, 5 g/L bovine liver powder and 4.6 g/L vitamin mix) following the published protocol [19], and 24 h after the first pupae were observed the rack was tilted to collect larvae and pupae.…”

“…The hosepipe is used to fill the trough with sufficient dechlorinated water (but without overfilling otherwise eggs get stuck under the internal pipe). The following day eggs were collected, rinsed and placed on a piece of sterile filter paper, and allowed to air dry for 4 h [23] and then quantified using the method developed by Maiga et al [20].

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