Mosquito Fertility Research Articles

The effect of repeat feeding on dengue virus transmission potential in Wolbachia-infected Aedes aegypti following extended egg quiescence.

As Wolbachia pipientis is more widely being released into field populations of Aedes aegypti for disease control, the ability to select the appropriate strain for differing environments is increasingly important. A previous study revealed that longer-term quiescence in the egg phase reduced the fertility of mosquitoes, especially those harboring the wAlbB Wolbachia strain. This infertility was also associated with a greater biting rate. Here, we attempt to quantify the effect of this heightened biting behavior on the transmission potential of the dengue virus using a combination of assays for fitness, probing behavior, and vector competence, allowing repeat feeding, and incorporate these effects in a model of R0. We show that Wolbachia-infected infertile mosquitoes are more interested in feeding almost immediately after an initial blood meal relative to wild type and Wolbachia-infected fertile mosquitoes and that these differences continue for up to 8 days over the period we measured. As a result, the infertile Wolbachia mosquitoes have higher virus prevalence and loads than Wolbachia-fertile mosquitoes. We saw limited evidence of Wolbachia-mediated blocking in the disseminated tissue (legs) in terms of prevalence but did see reduced viral loads. Using a previously published estimate of the extrinsic incubation period, we demonstrate that the effect of repeat feeding/infertility is insufficient to overcome the effects of Wolbachia-mediated blocking on R0. These estimates are very conservative, however, and we posit that future studies should empirically measure EIP under a repeat feeding model. Our findings echo previous work where periods of extensive egg quiescence affected the reproductive success of Wolbachia-infected Ae. aegypti. Additionally, we show that increased biting behavior in association with this infertility in Wolbachia-infected mosquitoes may drive greater vector competence. These relationships require further exploration, given their ability to affect the success of field releases of Wolbachia for human disease reduction in drier climates where longer egg quiescence periods are expected.

Gene Drive System Based on CRISPR/Cas9 in Mosquito Control for Preventing Malaria

Malaria is a disease transmitted by mosquitoes to humans and has symptoms varying from mild such as fever and headache to severe such as loss of consciousness, bleeding, or even death. Although there are a few other pathways to get infected with malaria, female mosquitoes belonging to the genus Anopheles infected by pathogen parasites belonging to Plasmodium have been considered as the predominant media of transmission, and elimination of this pathway could greatly assist in controlling the spread of malaria among populations. CRISPR/Cas9, as an emerging genome editing technology, functions by designed guide RNA (gRNA) for targeting sequence and Cas9 for cleaving targeted double-stranded DNA. Due to the great power of CRISPR/Cas9 in gene editing, it has significantly facilitated the study of mosquito control. This review describes the potential and applicable properties of the CRISPR/Cas9 system in restricting the spread of malaria via genetic screening of mosquitoes and gene-drive population suppression by modifying female mosquito fertility to reduce pathogen infection rate by targeting alleles from mosquitoes and pathogen to prohibit mosquito infections or retard the parasite development process. Furthermore, this review discusses the urgency that adjusting CRISPR/Cas9 technique to be affordable by more performers with low off-target effect. In conclusion, CRISPR/Cas9 system is expected to affirm the applicability of the technique in nature to restrict the spread of malaria among populations.

Hemp (Cannabis sativa cv. Kompolti) essential oil and its nanoemulsion: Prospects for insecticide development and impact on non-target microcrustaceans

Formulation development and non-target effect studies are two crucial steps for the registration of new botanical insecticides. Previous reports put in evidence the potential of hemp (Cannabis sativa L.) essential oil (EO) as a new candidate ingredient for developing green insecticides. However, the validation of its target and non-target effects after proper encapsulation is missing so far. In the current work, we investigated the insecticidal efficacy of the EO obtained from the hemp variety ‘Kompolti’, and we optimized its formulation development into a nanoemulsion (NE). The latter was assayed against larvae of Culex quinquefasciatus Say, a well-known vector of Wuchereria bancrofti Cobbold, avian malaria, and several arboviruses, such as Western equine encephalitis virus, St. Louis encephalitis virus, West Nile virus, and Zika virus. Considering the percentage of encapsulated EO (7.5% w/w), the developed NE showed mortality on Cx. quinquefasciatus larvae, in terms of LC50(90) values [72.2 (207.2) ppm], like those of pure EO [50.8(142.3) ppm]. Non-target toxicity experiments showed limited toxicity on the aquatic microcrustacean Daphnia magna Straus (i.e., mortality <16% after 48 h of exposure to the EO-based NE at the LC90 estimated on mosquito larvae). Considering that the LC50 values detected in this study are lower than those belonging to many EOs, and that the application at sublethal doses caused mortality between 39.1% and 40.8% as well as a reduction in mosquito fertility resulting in a reduction in natality of 40.4% and 45.1% for the EO and EO-based NE, respectively, both products may be considered as potential environmentally friendly and sustainable insecticides.

The chaperone BiP promotes dengue virus replication and mosquito vitellogenesis in Aedes aegypti

Binding immunoglobulin protein (BiP, also known as GRP78), a chaperone and master regulator of the unfolded protein response (UPR) pathway, plays an essential role in several flavivirus infections, but its functional role in regulating dengue virus replication in the mosquito remains largely unknown. We here demonstrated the interaction between a dengue virus serotype 2 (DENV2) and BiP in Aedes aegypti and report the discovery of a novel functional role of BiP in mosquito vitellogenesis. Silencing Ae. aegypti BiP (AaBiP) expression resulted in the significant inhibition of DENV2 viral genome replication, viral protein production, and infectious viral particle biogenesis. Co-immunoprecipitation assays showed that the DENV2 non-structural protein 1 (NS1) interacts with the AaBiP protein, and silencing AaBiP expression led to enhanced DENV2 NS1 aggregation, indicating that AaBiP plays a role in viral protein stability. A kinetic study focusing on pulse treatment of MG132, a proteasome inhibitor, in AaBiP-silenced mosquitoes showed that DENV2 NS1 was drastically elevated, which further suggests that AaBiP-mediated viral protein degradation is mediated by proteasomal machinery. Silencing of AaBiP also resulted in a reduction in mosquito fertility and fecundity. Depletion of AaBiP inhibited mosquito vitellogenesis due to the reduction of vitellogenin mRNA and elevated aggregation of vitellogenin protein post blood meal, further suppressing ovary development and fecundity. Overall, our results suggest that AaBiP is a dual-function protein with roles in both the regulation of dengue virus replication and mosquito reproduction. Our findings will be useful in the establishment of more efficient strategies for vector-borne disease control.

Exposure to copper sulfate impairs survival, post-embryonic midgut development and reproduction in Aedes aegypti.

Aedes aegypti is a vector of several global human viruses responsible for high human morbidity and mortality. The method to prevent the transmission of vector-borne viruses is mainly based on the control of the insect vector using insecticides. Among these chemicals, copper sulfate is a compound widely used in agriculture with the potential to be used as an alternative to control these insects. This study evaluated the effects of the exposure of A. aegypti larvae to copper sulfate on survival, midgut morphology, blood-feeding and fecundity. The exposure to CuSO4 decreased the survival of A. aegypti during the immature phase. Adults obtained from exposed larvae had their lifespan decreased at all tested concentrations. The exposure to CuSO4 impaired the development in the transition from larvae to pupae and from pupae to adult. The number of eggs laid by females developed from larvae treated with CuSO4 was significantly lower than in control. In addition, the egg hatching rates were also negatively affected. The midguts of treated larvae and pupae showed epithelial disorganization. The number of cleaved caspase-3 cells increased in the midgut of exposed pupae compared to control. Moreover, there was a reduction in proliferating cells in treated larvae and pupae compared to the control. In conclusion, the results reveal that CuSO4 exposure has insecticidal activity against A. aegypti, which may be related to the impairment of the midgut metamorphosis and reduced proliferation of stem cells, with the consequent impairment of female mosquito fertility and fecundity.

Metanetwork Transmission Model for Predicting a Malaria-Control Strategy.

Background: Mosquitoes are the primary vectors responsible for malaria transmission to humans, with numerous experiments having been conducted to aid in the control of malaria transmission. One of the main approaches aims to develop malaria parasite resistance within the mosquito population by introducing a resistance (R) allele. However, when considering this approach, some critical factors, such as the life of the mosquito, female mosquito fertility capacity, and human and mosquito mobility, have not been considered. Thus, an understanding of how mosquitoes and humans affect disease dynamics is needed to better inform malaria control policymaking.Methods: In this study, a method was proposed to create a metanetwork on the basis of the geographic maps of Gambia, and a model was constructed to simulate evolution within a mixed population, with factors such as birth, death, reproduction, biting, infection, incubation, recovery, and transmission between populations considered in the network metrics. First, the same number of refractory mosquitoes (RR genotype) was introduced into each population, and the prevalence of the R allele (the ratio of resistant alleles to all alleles) and malaria were examined. In addition, a series of simulations were performed to evaluate two different deployment strategies for the reduction of the prevalence of malaria. The R allele and malaria prevalence were calculated for both the strategies, with 10,000 refractory mosquitoes deployed into randomly selected populations or selection based on nodes with top-betweenness values. The 10,000 mosquitoes were deployed among 1, 5, 10, 20, or 40 populations.Results: The simulations in this paper showed that a higher RR genotype (resistant-resistant genes) ratio leads to a higher R allele prevalence and lowers malaria prevalence. Considering the cost of deployment, the simulation was performed with 10,000 refractory mosquitoes deployed among 1 or 5 populations, but this approach did not reduce the original malaria prevalence. Thus, instead, the 10,000 refractory mosquitoes were distributed among 10, 20, or 40 populations and were shown to effectively reduce the original malaria prevalence. Thus, deployment among a relatively small fraction of central nodes can offer an effective strategy to reduce malaria.Conclusion: The standard network centrality measure is suitable for planning the deployment of refractory mosquitoes.Importance: Malaria is an infectious disease that is caused by a plasmodial parasite, and some control strategies have focused on genetically modifying the mosquitoes. This work aims to create a model that takes into account mosquito development and malaria transmission among the population and how these factors influence disease dynamics so as to better inform malaria-control policymaking.

MODELING VACCINE PREVENTABLE VECTOR-BORNE INFECTIONS: YELLOW FEVER AS A CASE STUDY

In this paper, we propose and simulate a deterministic model for a vector-borne disease in the presence of a vaccine. The model allows the assessment of the impact of an imperfect vaccine with various characteristics, which include waning protective immunity, incomplete vaccine-induced protection and adverse events. We find three threshold parameters which govern the existence and stability of the equilibrium points. Our stability analysis suggests that the reduction in the mosquito fertility theoretically is the most effective factor of reducing disease prevalence in both low and high transmission areas. To illustrate the theoretical results, the model is simulated by the example of yellow fever. We also perform sensitivity analyses to determine the importance of both vaccine-induced mortality rate and disease-induced mortality rate for determining a control strategy. We found that there is an optimum vaccination rate, above which people die by the vaccination and below which people die by the disease.

Disruption of aminergic signalling reveals novel compounds with distinct inhibitory effects on mosquito reproduction, locomotor function and survival.

Insecticide resistance amongst disease vectors is a growing problem and novel compounds are needed. Biogenic amines are important for neurotransmission and we have recently shown a potential role for these in mosquito fertility. Here, we dissected the relative contribution of different aminergic signalling pathways to biological processes essential for vectorial capacity such as fertility, locomotion and survival by injecting agonists and antagonists and showed that octopaminergic/tyraminergic signalling is essential for oviposition and hatching rate. We show that egg melanisation is regulated by adrenergic signalling, whose disruption causes premature melanisation specifically through the action of tyramine. In addition to this, co-injection of tyramine with DOPA, the precursor of melanin, had a strong cumulative negative effect on mosquito locomotion and survival. Dopaminergic and serotonergic antagonists such as amitriptyline and citalopram recapitulate this effect. Together these results reveal potential new target sites for the development of future mosquito sterilants and insecticides.

Phenylalanine Metabolism Regulates Reproduction and Parasite Melanization in the Malaria Mosquito

The blood meal of the female malaria mosquito is a pre-requisite to egg production and also represents the transmission route for the malaria parasite. The proper and rapid assimilation of proteins and nutrients in the blood meal creates a significant metabolic challenge for the mosquito. To better understand this process we generated a global profile of metabolite changes in response to blood meal of Anopheles gambiae, using Gas Chromatography-Mass Spectrometry (GC-MS). To disrupt a key pathway of amino acid metabolism we silenced the gene phenylalanine hydroxylase (PAH) involved in the conversion of the amino acid phenylalanine into tyrosine. We observed increased levels of phenylalanine and the potentially toxic metabolites phenylpyruvate and phenyllactate as well as a reduction in the amount of tyrosine available for melanin synthesis. This in turn resulted in a significant impairment of the melanotic encapsulation response against the rodent malaria parasite Plasmodium berghei. Furthermore silencing of PAH resulted in a significant impairment of mosquito fertility associated with reduction of laid eggs, retarded vitellogenesis and impaired melanisation of the chorion. Carbidopa, an inhibitor of the downstream enzyme DOPA decarboxylase that coverts DOPA into dopamine, produced similar effects on egg melanization and hatching rate suggesting that egg chorion maturation is mainly regulated via dopamine. This study sheds new light on the role of amino acid metabolism in regulating reproduction and immunity.

Transcription Regulation of Sex-Biased Genes during Ontogeny in the Malaria Vector Anopheles gambiae

In Anopheles gambiae, sex-regulated genes are responsible for controlling gender dimorphism and are therefore crucial in determining the ability of female mosquitoes to transmit human malaria. The identification and functional characterization of these genes will shed light on the sexual development and maturation of mosquitoes and provide useful targets for genetic control measures aimed at reducing mosquito fertility and/or distorting the sex ratio.We conducted a genome wide transcriptional analysis of sex-regulated genes from early developmental stages through adulthood combined with functional screening of novel gonadal genes. Our results demonstrate that the male-biased genes undergo a major transcription turnover starting from larval stages to adulthood. The male biased genes at the adult stage include a significant high number of unique sequences compared to the rest of the genome. This is in contrast to female-biased genes that are much more conserved and are mainly activated during late developmental stages.The high frequency of unique sequences would indicate that male-biased genes evolve more rapidly than the rest of the genome. This finding is particularly intriguing because A. gambiae is a strictly female monogamous species suggesting that driving forces in addition to sperm competition must account for the rapid evolution of male-biased genes. We have also identified and functionally characterized a number of previously unknown A. gambiae testis- and ovary-specific genes. Two of these genes, zero population growth and a suppressor of defective silencing 3 domain of the histone deacetylase co-repressor complex, were shown to play a key role in gonad development.

Contribution of midgut bacteria to blood digestion and egg production in aedes aegypti (diptera: culicidae) (L.).

BackgroundThe insect gut harbors a variety of microorganisms that probably exceed the number of cells in insects themselves. These microorganisms can live and multiply in the insect, contributing to digestion, nutrition, and development of their host.Recent studies have shown that midgut bacteria appear to strengthen the mosquito's immune system and indirectly enhance protection from invading pathogens. Nevertheless, the physiological significance of these bacteria for mosquitoes has not been established to date. In this study, oral administration of antibiotics was employed in order to examine the contribution of gut bacteria to blood digestion and fecundity in Aedes aegypti.ResultsThe antibiotics carbenicillin, tetracycline, spectinomycin, gentamycin and kanamycin, were individually offered to female mosquitoes. Treatment of female mosquitoes with antibiotics affected the lysis of red blood cells (RBCs), retarded the digestion of blood proteins and reduced egg production. In addition, antibiotics did not affect the survival of mosquitoes. Mosquito fertility was restored in the second gonotrophic cycle after suspension of the antibiotic treatment, showing that the negative effects of antibiotics in blood digestion and egg production in the first gonotrophic cycle were reversible.ConclusionsThe reduction of bacteria affected RBC lysis, subsequently retarded protein digestion, deprived mosquito from essential nutrients and, finally, oocyte maturation was affected, resulting in the production of fewer viable eggs. These results indicate that Ae. aegypti and its midgut bacteria work in synergism to digest a blood meal.Our findings open new possibilities to investigate Ae. aegypti-associated bacteria as targets for mosquito control strategies.

Characterization of Aedes albopictus akirin for the control of mosquito and sand fly infestations

The control of arthropod vectors of pathogens that affect human and animal health is important for the eradication of vector-borne diseases. Recent evidences showed a reduction in the survival and/or fertility of mosquitoes, sand flies and poultry red mites fed in vitro with antibodies against the recombinant Aedes albopictus akirin. These experiments were the first step toward the development of a multi-target arthropod vaccine. In this study, we showed that the oviposition of A. albopictus and Phlebotomus perniciosus fed on mice vaccinated with recombinant A. albopictus akirin was reduced by 17% and 31%, respectively when compared to controls. However, Aedes aegypti mosquitoes were not affected after feeding on vaccinated mice. These results showed that recombinant A. albopictus akirin could be used to vaccinate hosts for the control of mosquito and sand fly infestations and suggested new experiments to develop improved vaccine formulations.

Laboratory evaluation of pyriproxifen treated bednets on mosquito fertility and fecundity. A preminary study

The study evaluated the effect of pyriproxifen treated bednets on mosquito fertility and fecundity in the laboratory. Pyriproxyfen (insect growth regulator) has been considered as a possible tool for management of pyrethroid resistance in mosquitoes. There are documentations of the effectiveness of pyriproxifen in controlling larval development when applied directly to breeding site. Considerations have been given to the use of pyriproxyfen on bed nets for sterilizing effect on mosquitoes resistant to pyrethroids. Groups of mosquitoes (Anopheles stephensi Beech) were exposed to bednet treated with 2% Pyriproxyfen and untreated netting 24 hrs after blood feeding. After oviposition, egg counts were done for each mosquito in the two groups. Larvae emerging from the laid eggs by each mosquito were counted. Three sets of experiments were successful. There was no significant difference in number of eggs laid between the two treatment groups (p=0.177). There was statistically significant difference in the number of laid eggs that hatched between the treatment types (p=0.0061). There was also statistically significant difference in the mean number of eggs that hatched between the experiments (p=0.0013). The mean number of eggs retained in the pyriproxyfen group was higher (70.3) than in the control group (41.6). This difference was not statistically significant with the small sample tested (p=0.08). The results suggest that 2% pyriproxyfen on bed nets has no effect on An. stephensi fecundity. Reduced fertility of eggs laid by mosquitoes exposed to pyriproxyfen treated bed nets was observed.

EVALUATING THE COSTS OF MOSQUITO RESISTANCE TO MALARIA PARASITES

Abstract Costly resistance mechanisms have been cited as an explanation for the widespread occurrence of parasitic infections, yet few studies have examined these costs in detail. A malaria-mosquito model has been used to test this concept by making a comparison of the fitness of highly susceptible lines of mosquitoes with lines that are resistant to infection. Malaria infection is known to cause a decrease in fecundity and fertility of mosquitoes; were thus predicted to be fitter than susceptible ones. >o?Anopheles gambiae were selected for refractoriness/resistance or for increased susceptibility to infection by Plasmodium yoelii nigeriensis. Additional lines that acted as controls for inbreeding depression were raised in parallel but not exposed to selection pressure. Selections were made in triplicate so that founder effects could be detected. Resistance mechanisms that were selected included melanotic encapsulation of parasites within 24 h postinfection and the complete disappearance of parasites from the gut. Costs of immune surveillance were assessed after an uninfected feed, and costs of immune deployment were assessed after exposure to infection and to infection and additional stresses. Mosquito survivorship was unaffected by either resistance to infection or by an increased burden of infection when compared with low levels of infection. In most cases reproductive fitness was equally affected by refractoriness or by infection. Resistant mosquitoes did not gain a fitness advantage by eliminating the parasites. Costs were consistently associated with larval production and egg hatch rate but rarely attributed to changes in blood feeding and never to changes in mosquito size. No advantages appeared to be gained by the offspring of resistant mosquitoes. Furthermore, we were unable to select for refractoriness in groups of mosquitoes in which 100% or 50% of the population were exposed to infection every generation for 22 generations. Under these selection pressures, no population had become completely refractory and only one became more resistant. Variations in fitness relative to control lines in different groups were attributed to founder effects. Our conclusion from these findings is that refractoriness to malaria is as costly as tolerance of infection.

EVALUATING THE COSTS OF MOSQUITO RESISTANCE TO MALARIA PARASITES

Costly resistance mechanisms have been cited as an explanation for the widespread occurrence of parasitic infections, yet few studies have examined these costs in detail. A malaria-mosquito model has been used to test this concept by making a comparison of the fitness of highly susceptible lines of mosquitoes with lines that are resistant to infection. Malaria infection is known to cause a decrease in fecundity and fertility of mosquitoes; resistant mosquitoes were thus predicted to be fitter than susceptible ones. Anopheles gambiae were selected for refractoriness/resistance or for increased susceptibility to infection by Plasmodium yoelii nigeriensis. Additional lines that acted as controls for inbreeding depression were raised in parallel but not exposed to selection pressure. Selections were made in triplicate so that founder effects could be detected. Resistance mechanisms that were selected included melanotic encapsulation of parasites within 24 h postinfection and the complete disappearance of parasites from the gut. Costs of immune surveillance were assessed after an uninfected feed, and costs of immune deployment were assessed after exposure to infection and to infection and additional stresses. Mosquito survivorship was unaffected by either resistance to infection or by an increased burden of infection when compared with low levels of infection. In most cases reproductive fitness was equally affected by refractoriness or by infection. Resistant mosquitoes did not gain a fitness advantage by eliminating the parasites. Costs were consistently associated with larval production and egg hatch rate but rarely attributed to changes in blood feeding and never to changes in mosquito size. No advantages appeared to be gained by the offspring of resistant mosquitoes. Furthermore, we were unable to select for refractoriness in groups of mosquitoes in which 100% or 50% of the population were exposed to infection every generation for 22 generations. Under these selection pressures, no population had become completely refractory and only one became more resistant. Variations in fitness relative to control lines in different groups were attributed to founder effects. Our conclusion from these findings is that refractoriness to malaria is as costly as tolerance of infection.

Effects of plant extracts on fecundity and fertility of mosquitoes

Effects of plant extracts on fecundity and fertility of mosquitoes

Effects of plant extracts on fecundity and fertility of mosquitoes

Rearing of Culex quinquefasciatus, Anopheles stephensi and Aedes aegypti larvae in water at concentrations less than the critical concentration for inhibition of adult emergence in 50% of treated larvae (EC50) of the ethyl acetate fractions of Calophyllum inophyllum seed and leaf, Solanum suratense and Samadera indica leaf extracts and the petrol ether fraction of Rhinocanthus nasutus leaf extract, from hatching to emergence significantly decreased the fecundity of the mosquitoes and the hatchability of their eggs. At the highest concentration of 50% EC50 of the extracts, the decrease in the fecundity over the control ranged between 62.4 and 87.4%. Sterility index of the mosquitoes reared in the media with the different extracts at this concentration ranged between 82.8 and 93.3%.

Involvement of male accessory gland substance in the fertility of mosquitoes

The male accessory gland substance is shown to be involved in the fertility of eggs for the first time in Aedes aegypti and Culex pipiens fatigans. Surgical removal of the paired accessory glands of males did not impair their mating ability. However, the eggs laid by the females after mating with operated males were found to be sterile. This condition could be reversed if the accessory gland substance was later received by the females. Evidence suggests that the accessory gland substance is essential for fertilization.