Consecutive mating and female refractoriness
Description
Dataset associated with the manuscript "Consecutive matings reveal differential depletion of male ejaculate functions in Aedes aegypti." The dataset contains individual-level observations from consecutive mating assays, including female remating incidence, first male paternity (P1), and sperm abundance in male seminal vesicles. Data were collected from laboratory-reared Aedes aegypti males mated to up to five consecutive females and include biological replicate identifiers and variables used in all statistical analyses reported in the manuscript.
Files
Steps to reproduce
Thai and DsRed strain Ae. aegypti, 4–5-days-old were used in our assays. Consecutive mating assays: We observed all matings by placing a single Thai male and female into an 8 L cage. After each successful mating, the mated female was removed, and a new virgin female was placed into the cage to allow the male to mate again. This was repeated until males had mated with five consecutive females. Re-mating assays: Immediately after the initial mating, females in each group (the first through fifth mated) were given the opportunity to re-mate with DsRed males for 2 h. Upon completion of the 2 h remating interval, females were blood-fed on the arm of a volunteer and males were removed. Four days after blood-feeding, females were individually aspirated into a 50 ml conical tube with a paper towel strip and 6 ml of water and given 48 h to lay eggs, after which females were frozen at -80°C until dissections commenced to identify singly or multiply mated females. Insemination status was determined by dissection of the lower female reproductive tract in 1X PBS to detect the presence (multiply mated) or absence (once mated) of DsRed sperm. Determination of larval paternity from multiply mated females: Oviposited eggs were partially dried and stored at 27°C. Five-to-seven days later, paper strips from multiply mated females were placed into a 40 ml cup, filled with type II H2O, supplemented with a pinch of active yeast, and placed under a vacuum for 30 min. Resulting third instar larvae were placed into a 24-welled cell culture plate (2-3 larvae per well) and visualized to detect the presence (sired by the second, DsRed male) or absence (sired by the first, Thai male) of GFP-expression from the larval eye. The proportion of progeny sired by the first (P1) and second mating male (P2) was determined as P1 + P2 = 1. Sperm quantification: Seminal vesicles and associated accessory glands from virgin, males mated five consecutive times (5X), and 5X males allowed to recover for 2 days (5X + 2d) or 4 days (5X + 4d) were dissected in 1X PBS, placed into a 200 μl chamber containing 100 μl of 1X PBS, ruptured with minutiae pins to release sperm and mixed by pipetting up and down. An additional 100 μl of PBS was added and the solution re-mixed. Ten 5 μl aliquots of the sperm mixture were placed onto a glass slide and dried at 50°C for 5 min. Sperm were fixed in 70% ethanol and stained with Giemsa dye. Sperm heads in each drop were counted under brightfield illumination at 200X magnification. This subsample was used to calculate total sperm.
Institutions
- Universidad de AntioquiaAntioquia, Medellín