Male contributions during mating increase female survival in the disease vector mosquito Aedes aegypti.

Aedes aegypti is a vector of medically important viruses including those causing Zika, dengue, and chikungunya. During mating, males transfer a number of proteins and other molecules to the female and these components of the male ejaculate are essential in shifting female post-mating behaviors in a number of insect species. Because these molecules are highly variable by species, and female post-mating behavior by species is also varied, behavioral assays testing the function of the ejaculate are necessary before we can develop control strategies targeting the mating system to reduce mosquito populations. Because increased survival in mosquitoes strongly increases vectorial capacity and can influence population sizes and potential risk we tested the effect of mating on female survival. The ejaculate can either promote or reduce female survival, as both have been shown in multiple insect species, yet this effect has not been directly assessed in mosquitoes. We compared survival of females in four treatment groups: mated females, virgin females, and virgin females injected with either an extract from the male reproductive glands or a saline control. Survival, blood feeding frequency, fecundity and cumulative net reproductive rate (R0) were determined after multiple feedings from a human host. Our results confirm that male reproductive gland substances increase female fecundity and blood feeding frequency, resulting in dramatic increases in fitness (R0). We also demonstrate, for the first time, an effect of male reproductive gland extracts alone on female survival, regardless of whether or not the female ingested a vertebrate blood meal. Thus, the effects of MAG extract on survival are not secondary effects from altered blood feeding. Collectively, we demonstrate a direct role for Ae. aegypti male-derived molecules on increasing female fitness, reproductive success and, ultimately, transmission potential for vector borne pathogens.

The Impact of Temperature and Body Size on Fundamental Flight Tone Variation in the Mosquito Vector Aedes aegypti (Diptera: Culicidae): Implications for Acoustic Lures.

Aedes aegypti (L.) males use female flight tone as a means of mate localization. By playing the sound of a flying female, males can be attracted to a trap to monitor mosquito populations and the progress of transgenic male releases. However, the female flight tone used to attract males needs to be optimized to maximize trap effectiveness. The fundamental frequency of female flight tone could be influenced by both body size and ambient temperature. However, no analysis yet has considered both the effect of body size and temperature on female flight tone of Ae. aegypti. Here, we present results for both these factors by recording the sounds of free-flying and tethered females across multiple temperature environments and with females reared for small, medium, and large body sizes. We demonstrate that female fundamental frequency is highly dependent on the environmental temperature, increasing ∼8-13 Hz with each °C gain. Body size and whether a female was tethered or free-flying did not impact the relationship between frequency and temperature, although further analysis is warranted. Our study highlights the importance of understanding the relationship between flight tone and temperature, and will inform the design of male mosquito traps.

Mating-Induced Transcriptome Changes in the Reproductive Tract of Female Aedes aegypti.

The Aedes aegypti mosquito is a significant public health threat, as it is the main vector of dengue and chikungunya viruses. Disease control efforts could be enhanced through reproductive manipulation of these vectors. Previous work has revealed a relationship between male seminal fluid proteins transferred to females during mating and female post-mating physiology and behavior. To better understand this interplay, we used short-read RNA sequencing to identify gene expression changes in the lower reproductive tract of females in response to mating. We characterized mRNA expression in virgin and mated females at 0, 6 and 24 hours post-mating (hpm) and identified 364 differentially abundant transcripts between mating status groups. Surprisingly, 60 transcripts were more abundant at 0 hpm compared to virgin females, suggesting transfer from males. Twenty of these encode known Ae. aegypti seminal fluid proteins. Transfer and detection of male accessory gland-derived mRNA in females at 0 hpm was confirmed by measurement of eGFP mRNA in females mated to eGFP-expressing males. In addition, 150 transcripts were up-regulated at 6 hpm and 24 hpm, while 130 transcripts were down-regulated at 6 hpm and 24 hpm. Gene Ontology (GO) enrichment analysis revealed that proteases, a protein class broadly known to play important roles in reproduction, were among the most enriched protein classes. RNAs associated with immune system and antimicrobial function were also up-regulated at 24 hpm. Collectively, our results suggest that copulation initiates broad transcriptome changes across the mosquito female reproductive tract, "priming" her for important subsequent processes of blood feeding, egg development and immune defense. Our transcriptome analysis provides a vital foundation for future studies of the consequences of mating on female biology and will aid studies seeking to identify specific gene families, molecules and pathways that support key reproductive processes in the female mosquito.

Underwater survival in the dog tick Dermacentor variabilis (Acari:Ixodidae).

Ticks are blood-feeding arthropods known for their long survivability off the host. Although ticks are terrestrial, they can survive extended periods of time submerged underwater. A plastron is an alternative respiration system that can absorb oxygen from water via a thin layer of air trapped by hydrophobic hairs or other cuticular projections. The complex spiracular plate of ticks has been postulated to serve as a plastron but that function has not been verified. This study provides evidence of plastron respiration in the American dog tick, Dermacentor variabilis, and for the first time confirmed the existence of plastron respiration in Ixodidae. Longer survival rates in oxygenated water indicate that underwater respiration requires oxygen. Wetting the spiracular plate with alcohol debilitates any potential plastron function and lowers the survival rate. Survival underwater may also be enhanced by metabolic depression and possibly anaerobic respiration. This study describes the first example of plastron respiration in the Ixodidae.