Egg laying decisions in Drosophila are consistent with foraging costs of larval progeny

Abstract

Decision-making is defined as selection amongst options based on their utility, in a flexible and context-dependent manner. Oviposition site selection by the female fly, Drosophila melanogaster, has been suggested to be a simple and genetically tractable model for understanding the biological mechanisms that implement decisions. Paradoxically, female Drosophila have been found to avoid oviposition on sugar which contrasts with known Drosophila feeding preferences. Here we demonstrate that female Drosophila prefer egg laying on sugar, but this preference is sensitive to the size of the egg laying substrate. With larger experimental substrates, females preferred to lay eggs directly on sugar containing media over other (plain, bitter or salty) media. This was in contrast to smaller substrates with closely spaced choices where females preferred non-sweetened media. We show that in small egg laying chambers newly hatched first instar larvae are able to migrate along a diffusion gradient to the sugar side. In contrast, in contexts where females preferred egg laying directly on sugar, larvae were unable to migrate to find the sucrose if released on the sugar free side of the chamber. Thus, where larval foraging costs are high, female Drosophila choose to lay their eggs directly upon the nutritious sugar substrate. Our results offer a powerful model for female decision-making.

Figure 1. Drosophila show a clear preference for laying eggs on sugar containing substrates.

(A) Schematic representation of the egg laying chamber showing the relative position of both substrates. The two 1% agarose substrates (P indicates plain, S indicates Sugar) were separated by 3% agarose middle zone. (B) Flies showed a preference for sucrose over plain agarose for egg laying at most concentration tested. (5 mM (n = 5, p<0.05, average total number of eggs = 33), 100 mM (n = 13, p<0.001, average total number of eggs = 65), 250 mM (n = 5, p<0.001, average total number of eggs = 73), 500 mM (n = 10, p<0.001, average total number of eggs = 56), and 1000 mM (n = 5, p<0.05, average total number of eggs = 50)) of sucrose. (C) Glucose was also preferred to a plain agarose substrate (5 mM (n = 4, p>0.05, average total number of eggs = 37), 100 mM (n = 10, p<0.001, average total number of eggs = 56), 250 mM (n = 5, p<0.01, average total number of eggs = 105), 500 mM (n = 5, p<0.01, average total number of eggs = 66), and 1000 mM (n = 5, p<0.001, average total number of eggs = 89)) (D) Flies showed a preference for fructose over plain at each concentration tested. (5 mM (n = 5, p<0.05, average total number of eggs = 28), 100 mM (n = 5, p<0.05, average total number of eggs = 45), 250 mM (n = 3, p<0.01, average total number of eggs = 43), 500 mM (n = 6, p<0.05, average total number of eggs = 42), and 1000 mM (n = 5 m, p<0.001, average total number of eggs = 48)) (E) Flies showed a nearly absolute preference for sucrose (100 mM) over caffeine (10 mM) as an egg laying substrate (n = 4, p<0.001, average total number of eggs = 83). 1f) Flies showed strong preference for sucrose (100 mM) over NaCl (100 mM) as an egg laying substrate(n = 5, p<0.05, average total number of eggs = 39). Student t-tests were performed for statistical analysis with one-tail p value.

Figure 2. Substrate size and diffusion gradients affect Drosophila egg laying choices.

In each panel the egg laying chamber is schematically shown on the left and the oviposition preference observed for the indicated chamber is shown on the right. (A) Flies preferred plain over sucrose substrate for egg-laying in the chambers that were used in Yang et al. (n = 3, p<0.05, average total number of eggs = 67) (B) Flies also preferred plain over sucrose substrate for egg-laying in small cylindrical chambers (n = 22, p<0.01, average total number of eggs = 58). (C) Flies preferred sucrose over plain substrate for egg-laying in small cylindrical chambers in an experimental setup without diffusion (n = 11, p<0.05, average total number of eggs = 76) (D) The flies preferred sucrose over plain substrates for egg-laying in chambers in which sucrose and plain islands were spaced 5 mm apart when there was a barrier to diffusion(n = 13, p<0.001, average total number of eggs = 25). Student t-tests were performed for statistical analysis with one-tail p value.

Figure 3. In large chambers, Drosophila prefer egg laying on sugar even when given the option to lay eggs close to the sugar or with diffusion gradients.

In these single island experiments, flies had the options of laying eggs at any location on the substrate. For the purpose of quantification, the petri dish was divided into 7 zones. Relative densities of eggs were calculated for each zone. (A) With 100 mM sucrose island experiments and no diffusion, females laid a significantly greater fraction of eggs in the sucrose island than in any other zone (as determined by a one-way ANOVA and Tukey's Multiple Comparison Test; * indicates p<0.05 as compared to each other zone) (n = 13, average total number of eggs = 24). (B) The island itself was only mildly attractive when filled with plain agarose (no significant differences by one-way ANOVA and Tukey's Multiple Comparison Test) (n = 5, average total number of eggs = 37). (C) In a 100 mM sucrose island with diffusion, females strongly preferred the island as determined by a one-way ANOVA and Tukey's Multiple Comparison Test; * indicates p<0.05 as compared to each other zone; # indicates p<0.05 as compared to zone 3–7) (n = 17, average total number of eggs = 41). (D) A plain island was not attractive in comparison to the sucrose island used in C (^ indicates p<0.05 as compared to zones 1–4, and 7) (n = 8, average total number of eggs = 15).

Figure 4. Visualization of diffusion in different experimental settings.

The three different sized chambers (19 mm, 35 mm and 50 mm) were set up as described in the methods except that the 100 mM sucrose was substituted with 1% agarose with 1% ethanol, 0.025% w/v Bromophenol Blue and 0.025% Xylene Cyanol. The molecular weights of these dyes are approximately twice that of sucrose so these dyes will underestimate the diffusion of sucrose. The photos were taken at time points 0, 1, 2, 4 and 24 hours.

Figure 5. Drosophila larvae crawl towards a sucrose source in the presence of a diffusion gradient.

In these experiments larvae were placed in 19 mm chambers containing plain and 100 mM sucrose egg-laying substrate. In chambers where diffusion was not allowed, no larvae placed on the plain substrate crossed over to the sucrose substrate within 30 minutes of observation (n = 15). In chambers where diffusion was allowed, 73% (SEM = 11.42%) of larvae crossed from the plain substrate to the sucrose substrate during the 30-minute observation period (n = 15). In chambers where both halves contained plain substrate (a control for the diffusion chamber) no larvae crossed the midline within 30 minutes (n = 15).