mosquito Archives - Center for Tropical and Emerging Global Diseases

Insulin-like peptides activate egg formation in the Asian malaria mosquito Anopheles stephensi

Donna Huber on October 7, 2025

Background: The mosquito family Culicidae diverged into the subfamilies Anophelinae and Culicinae approximately 179 million years ago. Most female mosquitoes are anautogenous and must blood-feed on a vertebrate to produce eggs. Regulation of egg-producing gonotrophic cycles is best understood in the culicine Aedes aegypti. Anopheline mosquitoes encode all of the hormones that regulate gonotrophic cycles in Ae. aegypti, but the processes regulating egg formation may not be fully similar. In this study, we conducted experiments that compared egg formation in Anopheles stephensi to prior findings reported for Ae. aegypti.

Methods: Assays for yolk deposition into oocytes, ovary ecdysteroidogenesis, vitellogenin expression, nutrient storage and oviposition were used to characterize gonotrophic cycles in An. stephensi females that were mated or unmated.

Results: Yolk deposition into oocytes depended on the release of hormones produced in the head. Two insulin-like peptides, An. stephensi insulin-like peptide hormone 3 (AsILP3) and AsILP4, stimulated the vitellogenic phase in An. stephensi, as measured by several different assays, whereas ovary ecdysteroidogenic hormone (OEH) showed no stimulatory activity. Nutrient stores were lower in An. stephensi than Ae. aegypti, which was associated with females also being unresponsive to AsILP3 stimulation in the absence of a blood meal. Anopheles stephensi males transferred ecdysteroids (ECDs) to females, which was associated with mated females producing and laying more eggs than unmated females. However, mated and unmated females did not show differences in ECD production by the ovaries or vitellogenin expression at the messenger RNA level by the fat body. Most females that mated before consuming a first blood meal oviposited while most unmated females did not. Mating after consuming a first blood meal did not rescue oviposition. However, females that reabsorbed eggs and consumed a second blood meal did oviposit.

Conclusions: Regulation of gonotrophic cycles in An. stephensi shares some features with Ae. aegypti but also exhibits differences.

Benjamin L Phipps, Mark R Brown, Michael R Strand. Parasit Vectors. 2025 Oct 7;18(1):399. doi: 10.1186/s13071-025-07036-y.

Recent insights in the development and functions of insect hemocytes

Donna Huber on September 15, 2025

Figure 1. Cellular defense responses, humoral effector molecules and regulators of hemocyte functions in insects. In the center of the figure is a stylized image of an insect hemocyte. The arrow to the left points to the four major cellular defenses that are mediated by hemocytes. The arrow to the right points to five of the most important humoral effector molecules hemocytes produce. The upper arrow points to key receptors with functions in regulating hemocyte-mediated cellular defenses, while the lower arrow points to cytokines that regulate hemocyte functions or communication between hemocytes and other tissues. Small case statements below some headings provide brief explanations of terms while additional details are presented in the main text. — Figure 1. Cellular defense responses, humoral effector molecules and regulators of hemocyte functions in insects.

Hemocytes mediate defense responses that are collectively referred to as cellular immunity. Here I highlight recent progress in the study of hemocyte ontogeny and function. Studies conducted primarily in D. melanogaster, mosquitoes and Lepidoptera identify a number of transcription factors and signaling pathways that regulate hemocyte differentiation and proliferation. Recent single cell RNA sequencing studies identify genes that distinguish transitional states in different hemocyte populations and changes that occur during immune responses. Several new studies expand understanding of the functions of sessile hemocytes. Recent results also enhance understanding of the signaling pathways that regulate hemocyte functions.

Michael R Strand. Curr Opin Insect Sci. 2025 Sep 15:101434. doi: 10.1016/j.cois.2025.101434.

Protocol for detecting peptide hormones in mosquito tissues

Donna Huber on May 24, 2025

Peptide hormones in insects are primarily expressed in specialized brain, ventral nerve chord, and midgut cells. When released, peptide hormones play crucial roles in regulating physiology, reproduction, and behavior. Here, we present a protocol for detecting peptide hormones in mosquito tissues such as the brain, midgut, and hemolymph. We describe steps for tissue preparation, immunocytochemistry, fluorescent quantification, and enzyme-linked immunoassay (EIA). This protocol offers a versatile and effective approach for studying peptide hormone expression and abundance in insects. For complete details on the use and execution of this protocol, please refer to Dou et al.¹.

Xiaoyi Dou, Kangkang Chen, Mark R Brown, Michael R Strand. STAR Protoc. 2025 May 24;6(2):103850. doi: 10.1016/j.xpro.2025.103850.

In the news: Michael Strand

Donna Huber on July 8, 2024

Michael Strand is a Regents Professor in the Department of Entomology and member of the Center for Tropical and Emerging Global Diseases. His mosquito research has recently been featured in a number of news stories.

What drives mosquitoes’ bloodlust? Their hormones (Nature)

The Science Behind What Makes Mosquitoes Bite You! Explained (News 9)

Mosquito bloodlust controlled by two hormones (The Naked Scientists)

Reciprocal interactions between neuropeptide F and RYamide regulate host attraction in the mosquito Aedes aegypti

Donna Huber on July 1, 2024

NPF accumulates in EECs of the posterior midgut after adult emergence but rapidly depletes after blood feeding.

Female mosquitoes produce eggs in gonadotrophic cycles that are divided between a previtellogenic and vitellogenic phase. Previtellogenic females consume water and sugar sources like nectar while also being attracted to hosts for blood feeding. Consumption of a blood meal activates the vitellogenic phase, which produces mature eggs and suppresses host attraction. In this study, we tested the hypothesis that neuropeptide Y-like hormones differentially modulate host attraction behavior in the mosquito Aedes aegypti. A series of experiments collectively indicated that enteroendocrine cells (EECs) in the posterior midgut produce and release neuropeptide F (NPF) into the hemolymph during the previtellogenic phase which stimulates attraction to humans and biting behavior. Consumption of a blood meal, which primarily consists of protein by dry weight, down-regulated NPF in EECs until mature eggs developed, which was associated with a decline in hemolymph titer. NPF depletion depended on protein digestion but was not associated with EEC loss. Other experiments showed that neurons in the terminal ganglion extend axons to the posterior midgut and produce RYamide, which showed evidence of increased secretion into circulation after a blood meal. Injection of RYamide-1 and -2 into previtellogenic females suppressed host attraction, while coinjection of RYamides with or without short NPF-2 also inhibited the host attraction activity of NPF. Overall, our results identify NPF and RYamide as gut-associated hormones in A. aegypti that link host attraction behavior to shifts in diet during sequential gonadotrophic cycles.

Xiaoyi Dou, Kangkang Chen, Mark R Brown, Michael R Strand. Proc Natl Acad Sci U S A. 2024 Jul 9;121(28):e2408072121. doi: 10.1073/pnas.2408072121.

The influence of oviposition status on measures of transmission potential in malaria-infected mosquitoes depends on sugar availability

Donna Huber on May 23, 2024

Background: Like other oviparous organisms, the gonotrophic cycle of mosquitoes is not complete until they have selected a suitable habitat to oviposit. In addition to the evolutionary constraints associated with selective oviposition behavior, the physiological demands relative to an organism’s oviposition status also influence their nutrient requirement from the environment. Yet, studies that measure transmission potential (vectorial capacity or competence) of mosquito-borne parasites rarely consider whether the rates of parasite replication and development could be influenced by these constraints resulting from whether mosquitoes have completed their gonotrophic cycle.

Methods: Anopheles stephensi mosquitoes were infected with Plasmodium berghei, the rodent analog of human malaria, and maintained on 1% or 10% dextrose and either provided oviposition sites (‘oviposited’ herein) to complete their gonotrophic cycle or forced to retain eggs (‘non-oviposited’). Transmission potential in the four groups was measured up to 27 days post-infection as the rates of (i) sporozoite appearance in the salivary glands (‘extrinsic incubation period’ or EIP), (ii) vector survival and (iii) sporozoite densities.

Results: In the two groups of oviposited mosquitoes, rates of sporozoite appearance and densities in the salivary glands were clearly dependent on sugar availability, with shorter EIP and higher sporozoite densities in mosquitoes fed 10% dextrose. In contrast, rates of appearance and densities in the salivary glands were independent of sugar concentrations in non-oviposited mosquitoes, although both measures were slightly lower than in oviposited mosquitoes fed 10% dextrose. Vector survival was higher in non-oviposited mosquitoes.

Conclusions: Costs to parasite fitness and vector survival were buffered against changes in nutritional availability from the environment in non-oviposited but not oviposited mosquitoes. Taken together, these results suggest vectorial capacity for malaria parasites may be dependent on nutrient availability and oviposition/gonotrophic status and, as such, argue for more careful consideration of this interaction when estimating transmission potential. More broadly, the complex patterns resulting from physiological (nutrition) and evolutionary (egg-retention) trade-offs described here, combined with the ubiquity of selective oviposition behavior, implies the fitness of vector-borne pathogens could be shaped by selection for these traits, with implications for disease transmission and management. For instance, while reducing availability of oviposition sites and environmental sources of nutrition are key components of integrated vector management strategies, their abundance and distribution are under strong selection pressure from the patterns associated with climate change.

Justine C Shiau, Nathan Garcia-Diaz, Dennis E Kyle, Ashutosh K Pathak. Parasit Vectors. 2024 May 23;17(1):236. doi: 10.1186/s13071-024-06317-2.

On the origin and evolution of the mosquito male-determining factor Nix

Donna Huber on December 21, 2023

The mosquito family Culicidae is divided into two subfamilies named the Culicinae and Anophelinae. Nix, the dominant male-determining factor, has only been found in the culicines Aedes aegypti and Ae. albopictus, two important arboviral vectors that belong to the subgenus Stegomyia. Here we performed sex-specific whole-genome sequencing and RNAseq of divergent mosquito species and explored additional male-inclusive datasets to investigate the distribution of Nix. Except for the Culex genus, Nix homologs were found in all species surveyed from the Culicinae subfamily, including 12 additional species from three highly divergent tribes comprising 4 genera, suggesting Nix originated at least 133-165 MYA. Heterologous expression of one of three divergent Nix ORFs in Ae. aegypti resulted in partial masculinization of genetic females as evidenced by morphology and doublesex splicing. Phylogenetic analysis suggests Nix is related to femaleless (fle), a recently described intermediate sex-determining factor found exclusively in anopheline mosquitoes. Nix from all species has a conserved structure, including three RNA-recognition motifs (RRMs), as does fle. However, Nix has evolved at a much faster rate than fle. The RRM3 of both Nix and fle are distantly related to the single RRM of a widely distributed and conserved splicing factor transformer-2 (tra2). RRM3-based phylogenetic analysis suggests this domain in Nix and fle may have evolved from tra2 or a tra2-related gene in a common ancestor of mosquitoes. Our results provide insights into the evolution of sex-determination in mosquitoes and will inform broad applications of mosquito-control strategies based on manipulating sex ratios towards the non-biting males.

James K Biedler, Azadeh Aryan, Yumin Qi, Aihua Wang, Ellen O Martinson, Daniel A Hartman, Fan Yang, Atashi Sharma, Katherine S Morton, Mark Potters, Chujia Chen, Stephen L Dobson, Gregory D Ebel, Rebekah C Kading, Sally Paulson, Rui-De Xue, Michael R Strand, Zhijian Tu. Mol Biol Evol. 2023 Dec 21:msad276. doi: 10.1093/molbev/msad276. Online ahead of print.

Increased environmental microbial diversity reduces the disease risk of a mosquitocidal pathogen

Donna Huber on December 7, 2023

Fig 6 Ch_R13E2-SpR systemically infects A. aegypti larvae. — Fig 6 Ch_R13E2-SpR systemically infects *A. aegypti* larvae.

The host-specific microbiotas of animals can both reduce and increase disease risks from pathogens. In contrast, how environmental microbial communities affect pathogens is largely unexplored. Aquatic habitats are of interest because water enables environmental microbes to readily interact with animal pathogens. Here, we focused on mosquitoes, which are important disease vectors as terrestrial adults but are strictly aquatic as larvae. We identified a pathogen of mosquito larvae from the field as a strain of Chromobacterium haemolyticum. Comparative genomic analyses and functional assays indicate this strain and other Chromobacterium are mosquitocidal but are also opportunistic pathogens of other animals. We also identify a critical role for diversity of the environmental microbiota in disease risk. Our study characterizes both the virulence mechanisms of a pathogen and the role of the environmental microbiota in disease risk to an aquatic animal of significant importance to human health.

Zhiwei Kang, Vincent G Martinson, Yin Wang, Kerri L Coon, Luca Valzania, Michael R Strand. mBio. 2023 Dec 6:e0272623. doi: 10.1128/mbio.02726-23.

Blood meals from ‘dead-end’ vertebrate hosts enhance transmission potential of malaria-infected mosquitoes

Donna Huber on November 30, 2023

Ingestion of an additional blood meal(s) by a hematophagic insect can accelerate development of several vector-borne parasites and pathogens. Most studies, however, offer blood from the same vertebrate host species as the original challenge (for e.g., human for primary and additional blood meals). Here, we show a second blood meal from bovine and canine hosts can also enhance sporozoite migration in Anopheles stephensi mosquitoes infected with the human- and rodent-restricted Plasmodium falciparum and P. berghei, respectively. The extrinsic incubation period (time to sporozoite appearance in salivary glands) showed more consistent reductions with blood from human and bovine donors than canine blood, although the latter’s effect may be confounded by the toxicity, albeit non-specific, associated with the anticoagulant used to collect whole blood from donors. The complex patterns of enhancement highlight the limitations of a laboratory system but are nonetheless reminiscent of parasite host-specificity and mosquito adaptations, and the genetic predisposition of An. stephensi for bovine blood. We suggest that in natural settings, a blood meal from any vertebrate host could accentuate the risk of human infections by P. falciparum: targeting vectors that also feed on animals, via endectocides for instance, may reduce the number of malaria-infected mosquitoes and thus directly lower residual transmission. Since endectocides also benefit animal health, our results underscore the utility of the One Health framework, which postulates that human health and well-being is interconnected with that of animals. We posit this framework will be further validated if our observations also apply to other vector-borne diseases which together are responsible for some of the highest rates of morbidity and mortality in socio-economically disadvantaged populations.

Ashutosh K Pathak, Justine C Shiau, Rafael C S Freitas, Dennis E Kyle. One Health. 2023 Jun 9:17:100582. doi: 10.1016/j.onehlt.2023.100582. eCollection 2023 Dec.

The mosquito Aedes aegypti requires a gut microbiota for normal fecundity, longevity and vector competence

Donna Huber on November 13, 2023

Mosquitoes shift from detritus-feeding larvae to blood-feeding adults that can vector pathogens to humans and other vertebrates. The sugar and blood meals adults consume are rich in carbohydrates and protein but are deficient in other nutrients including B vitamins. Facultatively hematophagous insects like mosquitoes have been hypothesized to avoid B vitamin deficiencies by carryover of resources from the larval stage. However, prior experimental studies have also used adults with a gut microbiota that could provision B vitamins. Here, we used Aedes aegypti, which is the primary vector of dengue virus (DENV), to ask if carryover effects enable normal function in adults with no microbiota. We show that adults with no gut microbiota produce fewer eggs, live longer with lower metabolic rates, and exhibit reduced DENV vector competence but are rescued by provisioning B vitamins or recolonizing the gut with B vitamin autotrophs. We conclude carryover effects do not enable normal function.

Ruby E Harrison, Xiushuai Yang, Jai Hoon Eum, Vincent G Martinson, Xiaoyi Dou, Luca Valzania, Yin Wang, Bret M Boyd, Mark R Brown, Michael R Strand. Commun Biol. 2023 Nov 13;6(1):1154. doi: 10.1038/s42003-023-05545-z.

Tag: mosquito