Reproduction and signals regulating worker policing under identical hormonal control in social wasps

Abstract

In social Hymenoptera, fertility and fertility signalling are often under identical hormonal control, and it has been suggested that such hormonal pleiotropies can help to maintain signal honesty. In the common wasp Vespula vulgaris, for example, fertile queens have much higher juvenile hormone (JH) titers than workers, and JH also controls the production of chemical fertility cues present on the females' cuticle. To regulate reproductive division of labour, queens use these fertility cues in two distinct ways: as queen pheromones that directly suppress the workers' reproduction as well as to mark queen eggs and enable the workers to recognize and police eggs laid by other workers. Here, we investigated the hormonal pleiotropy hypothesis by testing if experimental treatment with the JH analogue methoprene could enable the workers to lay eggs that evade policing. In support of this hypothesis, we find that methoprene-treated workers laid more eggs, and that the chemical profiles of their eggs were more queen-like, thereby causing fewer of their eggs to be policed compared to in the control. Overall, our results identify JH as a key regulator of both reproduction and the production of egg marking pheromones that mediate policing behaviour in eusocial wasps.

Figure 1

JH has a gonadotropic effect in V. vulgaris workers. This is shown in the barplot by the fact that the total number of eggs laid by workers treated with the JH analogue methoprene over a period of one week were significantly higher than in the acetone solvent-treated control workers (Poisson GLMM, z = 3.171, p = 0.0015**). Error bars indicate 95% confidence intervals.

Figure 2

Workers treated with the JH analogue methoprene laid eggs that were more queen-like and were policed less. (a) PCA analysis of the chemical surfaces of the eggs, based on centered log-ratio transformed peak areas, shows that methoprene-treated workers laid eggs that slightly shifted towards the profile typical of queen-laid eggs. (b) Methoprene-treated workers laid eggs that were more queen-like and consequently were also less likely to be policed within 24 h than those laid by control workers (binomial GLMM, z = -4.185, p < 0.001***). Error bars indicate 95% confidence intervals.

Figure 3

Heatmap showing differences in the chemical profiles of queen-laid eggs (QLE) and worker-laid eggs in the control groups (control) and how treatment with the JH analogue methoprene caused workers to lay eggs that became more queen-like. Colours indicate the mean fold difference in relative abundance of each hydrocarbon on the surface of queen-laid eggs and eggs laid by methoprene-treated workers compared to the eggs laid by the acetone solvent-treated control workers. Compounds were clustered based on a UPGMA hierarchical clustering using Euclidean distance as the distance metric. Asterisks indicate the FDR-corrected significances of individual contrasts for cases where the absolute fold difference was greater than 1.5 based on ANOVA. Highlighted red the compounds previously identified queen pheromones (C27, C29 and 3-MeC29) and queen egg marking pheromones (3-MeC27 and 3-MeC29).

Figure 4

Bar plots of the relative peak areas of compounds present on the egg surface that were previously identified as egg-marking pheromones (3-MeC27 and 3-MeC29) or as sterility-inducing queen pheromones (C29, C27 and 3-MeC29) in the common wasp V. vulgaris in our different groups (control, methoprene and queen-laid eggs QLE). Asterisks indicate FDR-corrected significances in the log2-transformed relative peak areas compared to the acetone solvent-treated control group. Error bars indicate 95% confidence intervals of linear modes fit on log2 relative peak areas. The two methyl-branched hydrocarbons 3-MeC29 and 3-MeC27 were significantly more abundant on queen-laid eggs (QLE) and became significantly more abundant on worker-laid eggs if workers were treated with the JH analogue methoprene. The linear alkanes C27 and C29, by contrast, were not significantly more abundant on queen-laid eggs (QLE) than control worker-laid eggs, but previous research has shown these compounds to be characteristic for the queen cuticle and act as worker-sterility inducing queen pheromones. One of these compounds, C27, also became more abundant on worker-laid eggs following methoprene treatment.

Figure 5

Experimental setup used to test for the effect of treatment with the JH analogue methoprene on worker fertility and rates of egg policing in the common wasp V. vulgaris.