Mosquito reproduction is regulated by a suite of hormones, many acting through membrane-bound receptor proteins. The Aedes aegypti G protein-coupled receptors AAEL024199 (AeCNMaR-1a) and AAEL018316 (AeCNMaR-1b) were identified as orthologs of the Drosophila melanogaster CNMa receptor (DmCNMaR). The receptor was duplicated early in the evolution of insects, and subsequently in Culicidae, into what we refer to as CNMaR-1a and CNMaR-1b. AeCNMaR-1a is only detected in male mosquito antennae while AeCNMaR-1b is expressed at high levels in mosquito ovaries. Using a heterologous cell assay, we determined that AeCNMa activates AeCNMaR-1a with a ~10-fold lower concentration than it does AeCNMaR-1b, though both receptors displayed half maximal effective concentrations of AeCNMa in the low nanomolar range. Finally, we show that injections of AeCNMa into blood-fed mated female Ae. aegypti resulted in fewer eggs laid.
Nia I Keyes-Scott, Aryan Lajevardi, Kyle R Swade, Mark R Brown, Jean-Paul Paluzzi, Kevin J Vogel. Insects. 2022 Feb 25;13(3):230. doi: 10.3390/insects13030230.
Most mosquito species are anautogenous, which means they must blood feed on a vertebrate host to produce eggs, while a few are autogenous and can produce eggs without blood feeding. Egg formation is best understood in the anautogenous mosquito Aedes aegypti where insulin-like peptides (ILPs), ovary ecdysteroidogenic hormone (OEH) and 20-hydroxyecdysone (20E) interact to regulate gonadotrophic cycles. Circulating hemocytes also approximately double in abundance in conjunction with a gonadotrophic cycle but the factors responsible for stimulating this increase remain unclear. Focusing on Ae. aegypti, we determined that hemocyte abundance similarly increased in intact blood-fed females and decapitated blood-fed females that were injected with ILP3, whereas OEH, 20E, or heat-killed bacteria had no stimulatory activity. ILP3 upregulated insulin-insulin growth factor signaling in hemocytes but few genes, including almost no transcripts for immune factors, were differentially expressed. ILP3 also stimulated circulating hemocytes to increase in two other anautogenous (Anopheles gambiae and Culex quinquefasciatus) and two facultatively autogenous mosquitoes (Aedes atropalpus and Culex pipiens molestus), but had no stimulatory activity in the obligately autogenous mosquito Toxorhynchites amboinensis. Altogether, our results identify ILPs as the primary regulators of hemocyte proliferation in association with egg formation, but also suggest this response has been lost in the evolution of obligate autogeny.
Gut microbes and diet can both strongly affect the biology of multicellular animals, but it is often difficult to disentangle microbiota-diet interactions due to the complex microbial communities many animals harbor and the nutritionally variable diets they consume. While theoretical and empirical studies indicate that greater microbiota diversity is beneficial for many animal hosts, there have been few tests performed in aquatic invertebrates. Most mosquito species are aquatic detritivores during their juvenile stages that harbor variable microbiotas and consume diets that range from nutrient rich to nutrient poor. In this study, we produced a gnotobiotic model that allowed us to examine how interactions between specific gut microbes and diets affect the fitness of Aedes aegypti, the yellow fever mosquito. Using a simplified seven-member community of bacteria (ALL7) and various laboratory and natural mosquito diets, we allowed larval mosquitoes to develop under different microbial and dietary conditions and measured the resulting time to adulthood and adult size. Larvae inoculated with the ALL7 or a more complex community developed similarly when fed nutrient-rich rat chow or fish food laboratory diets, whereas larvae inoculated with individual bacterial members of the ALL7 community exhibited few differences in development when fed a rat chow diet but exhibited large differences in performance when fed a fish food diet. In contrast, the ALL7 community largely failed to support the growth of larvae fed field-collected detritus diets unless supplemented with additional protein or yeast. Collectively, our results indicate that mosquito development and fitness are strongly contingent on both diet and microbial community composition.
We previously determined that several diets used to rear Aedes aegypti and other mosquito species support the development of larvae with a gut microbiota but do not support the development of axenic larvae. In contrast, axenic larvae have been shown to develop when fed other diets. To understand the mechanisms underlying this dichotomy, we developed a defined diet that could be manipulated in concert with microbiota composition and environmental conditions. Initial studies showed that axenic larvae could not grow under standard rearing conditions (27 °C, 16-h light: 8-h dark photoperiod) when fed a defined diet but could develop when maintained in darkness. Downstream assays identified riboflavin decay to lumichrome as the key factor that prevented axenic larvae from growing under standard conditions, while gut community members like Escherichia coli rescued development by being able to synthesize riboflavin. Earlier results showed that conventional and gnotobiotic but not axenic larvae exhibit midgut hypoxia under standard rearing conditions, which correlated with activation of several pathways with essential growth functions. In this study, axenic larvae in darkness also exhibited midgut hypoxia and activation of growth signaling but rapidly shifted to midgut normoxia and arrested growth in light, which indicated that gut hypoxia was not due to aerobic respiration by the gut microbiota but did depend on riboflavin that only resident microbes could provide under standard conditions. Overall, our results identify riboflavin provisioning as an essential function for the gut microbiota under most conditions A. aegypti larvae experience in the laboratory and field.
Yin Wang, Jai Hoon Eum, Ruby E. Harrison, Luca Valzania, Xiushuai Yang, Jena A. Johnson, Derek T. Huck, Mark R. Brown, Michael R. Strand Proceedings of the National Academy of Sciences Apr 2021, 118 (15) e2101080118; DOI: 10.1073/pnas.2101080118
Background: Most female mosquitoes are anautogenous and must blood feed on a vertebrate host to produce eggs. Prior studies show that the number of eggs females lay per clutch correlates with the volume of blood ingested and that protein is the most important macronutrient for egg formation. In contrast, how whole blood, blood fractions and specific blood proteins from different vertebrates affect egg formation is less clear. Since egg formation is best understood in Aedes aegypti, we examined how blood and blood components from different vertebrates affect this species and two others: the malaria vector Anopheles gambiae and arbovirus vector Culex quinquefasciatus.
Methods: Adult female mosquitoes were fed blood, blood fractions and purified major blood proteins from different vertebrate hosts. Markers of reproductive response including ovary ecdysteroidogenesis, yolk deposition into oocytes and number of mature eggs produced were measured.
Results: Ae. aegypti, An. gambiae and C. quinquefasciatus responded differently to meals of whole blood, plasma or blood cells from human, rat, chicken and turkey hosts. We observed more similarities between the anthropophiles Ae. aegypti and An. gambiae than the ornithophile C. quinquefasciatus. Focusing on Ae. aegypti, the major plasma-derived proteins (serum albumin, fibrinogen and globulins) differentially stimulated egg formation as a function of vertebrate host source. The major blood cell protein, hemoglobin, stimulated yolk deposition when from pigs but not humans, cows or sheep. Serum albumins from different vertebrates also variably affected egg formation. Bovine serum albumin (BSA) stimulated ovary ecdysteroidogenesis, but more weakly induced digestive enzyme activities than whole blood. In contrast, BSA-derived peptides and free amino acids had no stimulatory effects on ecdysteroidogenesis or yolk deposition into oocytes.
Conclusions: Whole blood, blood fractions and specific blood proteins supported egg formation in three species of anautogenous mosquitoes but specific responses varied with the vertebrate source of the blood components tested.
Environmental portrait of Courtney Murdock in her laboratory at UGA’s School of Veterinary Medicine (Dorothy Kozlowski)
Malaria disease transmission models are important tools for controlling and eliminating disease spread. However, a model is only as good as the assumptions about the various variables. Dr. Courtney Murdock, a member of the UGA’s Center for Tropical and Emerging Global Diseases and professor at Cornell University, has been studying how various biological and environmental factors influence mosquito survival. In a study recently published in the Proceedings of the Royal Society B led by graduate student Kerri Miazgowicz, Murdock and her colleagues examined several life traits, such as biting, feeding, and egg production, over the course of the life span of the mosquito Anopheles stephensi in hopes of providing better data for the models.
“Due to a lack of high-quality entomological data in general, researchers are often forced to input data from multiple disease systems to inform models in a given system or use approximations of key model components,” said Murdock.
An. stephensi is the primary mosquito species that transmits malaria in India. While most of the focus on malaria is most often associated with sub-Saharan Africa, it is widespread in the Indian subcontinent and throughout southeast Asia. Several Plasmodium species cause malaria, but P. falciparum is the deadliest of them. It has also shown drug resistance to current treatments. Control of the mosquito population is an important component in malaria control and elimination programs. Researchers need to be able to more accurately predict where mosquito populations will occur as climate changes and current territories become unsuitable living and breeding grounds. Program managers need to be prepared to incorporate more northern regions in their control efforts.
Kerri Miazgowicz, a graduate student in the Murdock Laboratory at the University of Georgia, led the study on the effects of age on thermal tolerances.
Current models rely on data that are only snapshots in time and often from multiple mosquito species, particularly the African mosquitoes, which are vectors for different malaria species. Miazgowicz, Murdock, and colleagues wanted to determine if the data from a single species of mosquito and parasite, over the course of its entire lifespan, significantly influenced current models in determining disease transmission in hopes of creating more accurate models.
The single most important factor driving current models is temperature. Mosquitoes are cold-blooded animals and therefore rely on their environment to regulate their body temperature. However, temperature is not the only factor influencing life traits. Currently, data are only available as snapshots in time. These incomplete data do not take into account for changes in mosquito behavior and life traits that occur over the course of the mosquito’s life. Murdock and her colleagues have recorded changes in biological function as the mosquito ages. Just as people slow down biologically as they age – metabolism slows, reproduction ability declines, etc. – the same is true for mosquitos. They also found that various traits peak at different times depending on temperature. Importantly they found that temperature and age significantly affected the number of females taking a blood meal (this is the means in which malaria parasites are transmitted to humans) on a given day, average daily egg production, and ultimately survival.
The findings in this study indicated that the addition of An. stephensi data yielded qualitatively different temperature-transmission suitability relationships compared to models that included multiple malaria vectors. With An. stephensi data, the model predicted a broader geographical range of temperature suitability.
“Accounting for these age and species effects in models of transmission potential alters how much of South Asia is predicted to be suitable for malaria relative to models that do not account for these factors,” said Murdock.
These findings can lead to improved malaria transmission models. However, more study outside the laboratory is needed to truly understand the impact mosquito age has on life traits and thermal tolerance.
“This study highlights a critical need for more research in natural settings characterizing the effects of age on mosquito biology to improve predictions of current and future risk,” concluded Murdock.
Models predicting disease transmission are vital tools for long-term planning of malaria reduction efforts, particularly for mitigating impacts of climate change. We compared temperature-dependent malaria transmission models when mosquito life-history traits were estimated from a truncated portion of the lifespan (a common practice) versus traits measured across the full lifespan. We conducted an experiment on adult female Anopheles stephensi, the Asian urban malaria mosquito, to generate daily per capita values for mortality, egg production and biting rate at six constant temperatures. Both temperature and age significantly affected trait values. Further, we found quantitative and qualitative differences between temperature–trait relationships estimated from truncated data versus observed lifetime values. Incorporating these temperature–trait relationships into an expression governing the thermal suitability of transmission, relative R0(T), resulted in minor differences in the breadth of suitable temperatures for Plasmodium falciparum transmission between the two models constructed from only An. stephensi trait data. However, we found a substantial increase in thermal niche breadth compared with a previously published model consisting of trait data from multiple Anopheles mosquito species. Overall, this work highlights the importance of considering how mosquito trait values vary with mosquito age and mosquito species when generating temperature-based suitability predictions of transmission.
K. L. Miazgowicz, M. S. Shocket, S. J. Ryan, O. C. Villena, R. J. Hall, J. Owen, T. Adanlawo, K. Balaji, L. R. Johnson, E. A. Mordecai and C. C. Murdock. Proc Biol Sci. 2020 Jul 29;287(1931):20201093. doi: 10.1098/rspb.2020.1093.
Most species of mosquitoes are detritivores that feed on decaying plant and animal materials in their aquatic environment. Studies of several detritivorous mosquito species indicate that they host relatively low diversity communities of microbes that are acquired from the environment while feeding. Our recent results also indicate that detritivorous species normally require a living gut microbiota to grow beyond the first instar. Less well known is that some mosquitoes, including those belonging to the genus Toxorhynchites, are predators that feed on other species of mosquitoes and nektonic prey. In this study, we asked whether predaceous Toxorhynchites amboinensis larvae still require living microbes in their gut in order to develop. Using the detritivorous mosquito Aedes aegypti as prey, we found that T. amboinensis larvae harbour bacterial communities that are highly similar to that of their prey. Functional assays showed that T. amboinensis first instars provided axenic (i.e. bacteria-free) prey failed to develop, while two bacterial species present in gnotobiotic (i.e. colonized by one or more known bacterial species) prey successfully colonized the T. amboinensis gut and rescued development. Axenic T. amboinensis larvae also displayed defects in growth consistent with previously identified roles for microbe-mediated gut hypoxia in nutrient acquisition and assimilation in A. aegypti. Collectively, these results support a conserved role for gut microbes in regulating the development of mosquitoes with different feeding strategies.
When warm weather approaches, so do pesky little bloodsucking pests.
The unassuming mosquito may be smaller than a dime, but it packs a serious punch, killing more people each year than any other animal. And with average temperatures climbing around the globe, different mosquito species are making their way farther north than ever before and bringing their diseases—malaria, West Nile, dengue, and more—along for the ride.
But thanks to recent discoveries at the University of Georgia, it may soon become easier to fend off the swarm.
Regents Professor of Entomology Michael Strand’s lab found that microorganisms, or microbes, in a mosquito’s gut are essential for growth and development. Mosquito larvae spend anywhere from a few days to two weeks developing in pools of water that can be as small as an upside-down bottle cap. Microbes colonize the larvae’s digestive tracts, forming a community of microorganisms that enables the larvae to mature into adult mosquitos.
The implications of the findings could lead to new approaches for mosquito control.
“If you can disrupt their growth cycle, you could control mosquito populations,” Strand says. “Certain combinations of these organisms that exist in the digestive system of the mosquito also affect how well they are able to acquire and transmit disease-causing microorganisms to people.
Understanding how these organisms alter the mosquito’s ability to transmit diseases offers the potential for increasing resistance to certain organisms they can pass on to people.”
From a more basic science perspective, insects provide a more simplified version of a microbiome, the ecological community of microorganisms that call a space home. Researchers often discuss the roles microbiomes, such as that of the human gut, play in an individual’s health, but it’s difficult to sort through the billions of different organisms that can be present. Mosquitoes, and other insects in general, are much less complex, sometimes hosting only several hundreds of microorganisms in their digestive tracts. The smaller number of microbes make it easier for researchers to study.
“In effect, this simplicity reduces the many variables involved,” Strand says. “Some of the rules determining the importance of gut microbes in mosquito development may also have generalizable applications in how similar processes are regulated in larger animals.”
Sting like a Bee
Mosquitoes aren’t the only insects Strand studies.
His interests lie in parasitology, or how parasites interact with the animals they feed from. Parasitic wasps, comprising over a million different species, are the perfect medium to study parasite-host interactions.
Around 100 million years ago, some parasitic wasps were infected by a virus that became part of their genome. Wasps coopted that virus to deliver different types of genes into hosts.
One way wasps accomplish that is by injecting the coopted virus into other insects along with their eggs. The virus then infects the insects’ cells in much the same way as modern medicine’s gene therapies that use viruses to introduce genes into human patients for disease prevention or treatment.
The virus’ genes suppress the host insect’s immune defenses, which would otherwise destroy the foreign eggs. The wasps can then hatch and develop into adults while slowly consuming the host from the inside out.
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