Wolbachia inhibits ovarian formation and increases blood feeding rate in female Aedes aegypti

Abstract

Wolbachia, a gram-negative endosymbiotic bacterium widespread in arthropods, is well-known for changing the reproduction of its host in ways that increase its rate of spread, but there are also costs to hosts that can reduce this. Here we investigated a novel reproductive alteration of Wolbachia wAlbB on its host Aedes aegypti, using studies on mosquito life history traits, ovarian dissection, as well as gene expression assays. We found that an extended period of the larval stage as well as the egg stage (as previously shown) can increase the proportion of Wolbachia-infected females that become infertile; an effect which was not observed in uninfected females. Infertile females had incomplete ovarian formation and also showed a higher frequency of blood feeding following a prior blood meal, indicating that they do not enter a complete gonotrophic cycle. Treatments leading to infertility also decreased the expression of genes related to reproduction, especially the vitellogenin receptor gene whose product regulates the uptake of vitellogenin (Vg) into ovaries. Our results demonstrate effects associated with the development of infertility in wAlbB-infected Ae. aegypti females with implications for Wolbachia releases. The results also have implications for the evolution of Wolbachia infections in novel hosts.

Fig 1. Appearance of internal organs in wAlbB-infected Aedes aegypti mosquitoes.

(A) Infertile females lack observable ovaries, while ovaries are present in (B) fertile females (labelled as “ovary”). The background of the photographs was removed for clarity, with the original photographs presented in S3 Fig.

Fig 2

(A) Boxplots showing the proportion of infertile females in wAlbB-infected and uninfected mosquitoes with an egg stage that had lasted for 12 weeks (stored) or one week and a larvae stage that had been deprived of food for two weeks (starved) or that had not been deprived of food. Mosquitoes that had not been treated with "stored" or “starved” are designated as “control”. No uninfected females from all four treatments were infertile. Values above boxplots represent corresponding averaged percentages based on two replicates for controls and three replicates for treatments with each replicate containing 30 individuals. (B) Boxplots of relative Wolbachia density in 3–5 days post-emergence females with unknown fertility status. Each group is based on two consistent real-time PCR replicates.

Fig 3. Boxplots of relative expression of reproduction-related genes ecdysone receptor (ecr), eggshell organizing factor (eof) and vitellogenin receptor (vgr) in female (A) pupae and (B) young adults (1 ± 0.5 days post-emergence).

These three genes were normalized to reference gene RPS17. Values with the same letter are not significantly different based on Tukey’s HSD tests (S5 Table). Results are based on two consistent real-time PCR replicates of ten individual females from each group.

Fig 4. Boxplots of relative expression of reproduction-related genes ecdysone receptor (ecr), eggshell organizing factor (eof) and vitellogenin receptor (vgr) in fertile and infertile females one week after their second blood meal.

Genes were normalized to reference gene RPS17. Values with the same letter are not significantly different according to Tukey’s HSD tests. Results are based on two consistent real-time PCR replicates of ten individual females from each group.

Fig 5

Boxplots of (A) female proportion that blood fed three days after they had been fully fed, in which three replicates were completed with 20–30 individual females for each replicate; and (B) the weights of fed and unfed females. 20 females that had fully fed and 20 that had not been provided with a blood meal from all three replicates were weighed at random.