Efficacy of fluralaner, a long acting acaricide against three species of vector mosquitoes (Diptera: Culicidae) under laboratory conditions – a potential candidate for drug based vector control

Efficacy of fluralaner, a long acting acaricide against three species of vector mosquitoes (Diptera: Culicidae) under laboratory conditions – a potential candidate for drug based vector control | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Efficacy of fluralaner, a long acting acaricide against three species of vector mosquitoes (Diptera: Culicidae) under laboratory conditions – a potential candidate for drug based vector control Harish Kumar Shah, Vaishnavi Srinivasan, Shakila Venkatesan, Vijayakumar Balakrishnan, and 4 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-2803950/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Vector control is a key intervention against mosquito borne diseases such as dengue, chikungunya and malaria. However, conventional methods have several limitations and alternate strategies are in urgent need. Vector control with endectocides or systemic insecticides such as ivermectin is emerging as a novel strategy. The short half-life of ivermectin is a limiting factor for its application as a mass therapy tool for vector control. Isoxazoline compounds like fluralaner, a class of veterinary acaricides with long half-life hold promise as alternatives to ivermectin. These drugs persist in the animal body at detectable blood levels up to three months or more, after a one-time oral administration. However, information about their mosquitocidal effect is very much limited. In the current study, we explored the efficacy of fluralaner against laboratory reared vector mosquitoes of Aedes aegypti (Linnaeus, 1762), Anopheles stephensi (Liston, 1901) and Culex quinquefasciatus (Say, 1823) (Diptera: Culicidae) by blood feeding. Fluralaner showed significant mosquitocidal effect with the estimated 24 h LC 50 values in the range of 24.04–49.82 ng/mL for the three different mosquito species tested. Effects on life history characteristics (fecundity, egg hatch success, immature development and adult emergence success) were also observed at drug concentrations nearing 24 h LC 50 . At higher drug concentration of 250 ng/mL, significant knock down effect was observed within 1–2 h post blood feeding. Potent mosquitocidal effect coupled with its long half-life makes fluralaner an excellent candidate for drug based vector control strategies especially in the context of “One Health” approach. In this regard, further studies on the safety aspects of fluralaner in humans are desirable. fluralaner endectocide drug based vector control mosquitocidal effect life history characteristics Figures Figure 1 Figure 2 Introduction Vector borne diseases such as malaria, lymphatic filariasis, dengue, zika and others pose significant public health threats globally (WHO 2020). Effective vaccines or specific treatment are not available for many of these diseases and vector control is the only option for prevention and control (WHO 2017) . However, conventional vector control methods have several limitations and there is an urgent need for alternative control strategies. With the discovery of the potent mosquitocidal effect of the anti-parasitic drug ivermectin, vector control with endectocides is emerging as an alternative strategy (Foy et al., 2011, Imbahale et al., 2019). This approach involves mass administration of an oral or systemic insecticidal drug to animal or human population for killing the blood feeding arthropod vectors. Ivermectin is now being explored as a mass drug administration (MDA) tool in humans for malaria control in many settings (Foley et al., 2000, Chaccour et al., 2010, Foy et al., 2019, Slater et al., 2020). As we progress towards malarial elimination, due to the extensive use of LLIN and IRS, mosquitoes’ feeding behavior may change and they may prefer to feed on animals rather than humans. Hence mass therapy of domestic animals with ivermectin is also suggested (Chaccour et al., 2013, Pooda et al., 2015, Ōmura and Crump 2017, Chaccour and Rabinovich 2019). However, one major drawback for application of ivermectin for vector control is its short half-life (2.07±0.71 days) (Walther et al., 2015). This would necessitate its very frequent administration to human or animal population which may limit its utility in mass therapy. But drugs with shorter half-life tend to be safer and slow release formulation could bypass the need of frequent administration. Alternative endectocides with longer half-life which are proved to be safe or having only minimal side effects in animals and humans are highly desirable. Isoxazoline compounds like fluralaner (Bravecto®) and afoxolaner (NexGard®) used as acaricides in veterinary practice have an unusually long half-life (14.27±2.53 days) (Kilp et al., 2014, Comas and Armstrong 2018). After a single oral administration (50 mg/kg) of fluralaner in dogs, maximum plasma concentrations (C max ) of 5419 ± 2086 ng/mL have been achieved within 24 h and detectable plasma levels were found up to 112 days post administration (Kilp et al.,2014, Kilp et al.,2016). The mechanism of action of fluralaner is similar to that of ivermectin. It is a potent inhibitor of γ-amino butyric acid (GABA) and L-glutamate-gated chloride channels, but binds at a site different from that of ivermectin (Gassel et al.,2014). Isoxazoline compounds may have minimal or no effect on the vertebrate nervous system (Kilp et al., 2014) . With its excellent safety profile in animals, long half-life and the potent acaricidal effect, fluralaner can be an alternative choice for drug based vector control strategies. However, information on its mosquitocidal effect is very limited (Jiang S et al., 2016, Miglianico et al., 2018). A recent study from USA (United States of America) reported its significant lethal effect in different mosquito vector species such as Anopheles stephensi (Liston, 1901), Anopheles gambiae Kisumu, Anopheles gambiae Tiassalé (Gile, 1902), Aedes aegypti New Orleans, Aedes aegypti Cayman (Linnaeus, 1762) and Culex pipiens (Linnaeus, 1758) (Diptera: Culicidae) species (Miglianico et al., 2018). In the present study, the adulticidal effect of fluralaner was studied against three vector mosquito species from India: 1) Aedes aegypti , the primary vector of dengue, 2) Anopheles stephensi , the vector of urban malaria, and 3) Culex quinquefasciatus (Say, 1823) (Diptera: Culicidae), the vector of lymphatic filariasis under laboratory conditions. In addition, the effect of fluralaner on fecundity, egg hatch success, immature development and adult emergence success were investigated on mosquitoes fed with different concentration of the drug. Materials and Methods Extraction of fluralaner from the commercial tablet Fluralaner was extracted from the commercially available tablets for veterinary use (Bravecto®- purchased from local veterinary drug shop; manufactured by Merck Animal Health, Madison, Vienna, Austria) using the method described by Miglianico et al., 2018. The tablet was crushed into fine powder using a pestle and mortar and then dissolved in a solvent prepared by mixing dichloromethane and methanol in a 1:1 ratio. The whole mixture was then agitated at room temperature for 1 h with a magnetic stirrer and filtered using Whatman® grade 1 filter paper (purchased from Sigma-Aldrich, St. Louis, MO). The filtrate was left overnight for evaporation and the concentrated final product was obtained. Stock solution (5 mg/mL) was prepared in dimethyl sulfoxide (DMSO) (purchased from HiMedia, Mumbai, India) and aliquots were frozen at −20°C. Prior to each experiment, aliquots were dissolved in phosphate buffer saline (PBS) (purchased from Sigma-Aldrich, St. Louis, MO) and added to heparinized chicken blood to achieve the desired final drug concentrations required for the mosquito blood feeding experiments. Vector mosquitoes and colony maintenance Laboratory reared mosquito species of Ae. aegypti , An. Stephensi and Cx. quinquefasciatus were obtained from the rearing and colonization facility at the Indian Council of Medical Research - Vector Control Research Centre (ICMR-VCRC), Puducherry, India. These strains were originally collected from the Union territory of Puducherry, India and the colonies have been maintained in the insectary since 1975. Cx. quinquefasciatus were collected in 1975 and Ae. aegypti and An. stephensi were collected in 1976. They were maintained at a constant temperature of 27±2°C and relative humidity of 80±10% in one-foot Barraud cages (30cm L X 30cm W X 30cm H) and provided with 10% sucrose solution. Laboratory bioassays with fluralaner To determine the activity range of fluralaner, initially, bioassays were performed to assess the mortality effect at 24 h using wide range concentrations (20 ng/mL-120 ng/mL). Based on the results of these experiments, a narrow range of four drug concentrations were selected for bioassays to determine LC 50 (concentration resulting in 50% mortality after taking a blood meal) value at 24 h of post blood feeding. Drug concentrations of 5 ng/mL, 10 ng/mL, 20 ng/mL and 35 ng/mL were selected for the feeding experiments with Ae aegypti; 5 ng/mL, 15 ng/mL, 25 ng/mL and 40 ng/mL for An. stephensi ; and 15 ng/mL, 30 ng/mL, 45 ng/mL and 60 ng/mL for Cx. quinquefasciatus respectively (two drug concentration above and below LC 50 value of initial bioassay) . Five days old, non-blood fed female mosquitoes from same batch were used for blood feeding experiments. Prior to blood feeding, cotton pads soaked with 10% sucrose solution were removed and the adult mosquitoes were allowed to starve at least for 12 h to improve the feeding rate. Five mL of heparinized chicken blood containing desired concentration of fluralaner was used for feeding the mosquitoes in the treatment group via artificial blood feeding apparatus made of glass fitted with blood mixing rotor and parafilm for about 60-90 minutes. Treatment group consists of 4 drug concentration and each concentration constituted of three technical replicate cages (each with 50 females). 50 adult female from the same batch of mosquitoes fed on chicken blood with only solvent were used as controls. After blood feeding, mosquitoes (fully engorged) were transferred to clean cages, provided with 10% sucrose solution and the mortality effect was observed at 4, 8, 18, 24, 48, 72 and 96 h of post feeding. 48 h ( Ae. aegypti , An. stephensi ) and 72 h ( Cx. quinquefasciatus ) of post blood feeding, survived mosquitoes were observed for fecundity. The experiment was repeated three times on different days using fresh batches of mosquitoes and freshly prepared fluralaner concentration as biological replicates. In continuation to the bioassay experiments for mortality effect of fluralaner, life history characteristics of survived mosquitoes in each drug treatment groups were assessed in comparison to the control groups. Number of survived mosquitoes in each replicate cage varied with the species and also with the drug concentration. Effect of fluralaner on fecundity, egg hatch success, immature development and adult emergence success For fecundity assessment of survived Ae. aegypti and An. stephensi , oviposition cups filled with 75-90 mL water lined with filter paper were introduced into the cages at 48 h of post blood feeding. For Cx. quinquefasciatus , oviposition cups without filter paper were introduced into the cages at 72 h as it takes 3 days to become fully gravid. To assess the effect of fluralaner in egg hatch success, eggs laid by survived mosquitoes in treatment group were compared with that of eggs laid by mosquitoes in the control group. For hatching experiment of Ae. aegypti and An. stephensi , five hundred eggs counted under microscope were floated in trays filled with water using a fine tip brush. In case of Cx. quinquefasciatus , as eggs were laid in rafts, numbers were slightly more than five hundred. The rafts were carefully transferred to tray using brush without damaging it. Hundred first instar larvae hatched from eggs laid by survived mosquitoes were observed for development up to pupal stage by providing mosquito larval food containing yeast (40%) and dog biscuit (60%) and were compared with the control group. Daily mortality was recorded (if any) in both treatment and control group larvae. Similarly fifty pupae from survived mosquitoes were observed for adult emergence by transferring them into containers filled with 3/4 th water and covered with mosquito net having hole in middle plugged with cotton. The results were compared with control group. Estimation of speed of killing at high concentration We performed a set of experiment to assess the speed of killing of the mosquitoes when they were fed with a high concentration of fluralaner. In initial weeks of drug administration the concentration is expected to be higher which may result in fast mortality within hours of blood feeding. For this, 50 females in triplicates were fed at 250 ng/mL and were observed at hourly interval until complete (100%) mortality was noted in the treatment groups. The experiment was replicated three times on different days with fresh batch of mosquitoes. Statistical analysis Mean and standard deviation value from three replicates of different concentration were taken to obtain data, which was further used for various statistical analysis. Mortality data over time were subjected to Probit regression analysis to estimate LC 50 value. Mean (SD) and Frequency (%) were used to describe summary values. Mann Whitney U test was used to test the difference in fecundity, egg hatch success, immature survival and adult emergence success between the drug treated and control groups. P-value <0.05 was considered as statistically significant. SPSS 16.0 was used for carrying out probit analysis whereas, STATA 14.2 software for other statistical analysis. Results Estimated LC 50 value of fluralaner at different time points Based on the preliminary results of initial bioassay, experiments were performed with narrow range drug concentrations (with three technical replicates), as described in methodology section for each of the species and the 24 h LC 50 (95% CI) value was estimated. The results are summarized in (Table 1). Since the upper limit for 95% CI of 24 h LC 50 was exceeding the maximum dosage used for Ae. aegypti and An. stephensi , the mortality data was subjected to cox-regression analysis to compare the survivorship or risk of death between different treatment and control group. Cox-regression analysis Ae. aegypti : The risk of death was significantly higher in dosage 35 (HR=150.45 (95% CI:94.76 – 238.87), P-value<0.001), dosage 20 (HR=40.61 (95% CI:25.77 – 64.01), P-value<0.001), dosage 10 (HR=17.19 (95% CI: 10.86 – 27.22), P-value<0.001) and dosage 5 (HR=3.45 (95% CI: 2.07 – 5.74), P-value<0.001) when compared to control i.e., dosage 0 (Table 2) (Figure 1-A). An. stephensi : The risk of death was significantly higher in dosage 40 (HR=231.94 (95% CI:142.68 – 377.05), P-value<0.001), dosage 25 (HR=39.72 (95% CI:24.73 – 63.80), P-value<0.001), dosage 15 (HR=17.21 (95% CI: 10.63 – 27.88), P-value<0.001) and dosage 5 (HR=3.48 (95% CI: 2.02 – 5.97), P-value<0.001) when compared to control i.e., dosage 0 (Table 3) (Figure 1-B). Cx. quinquefasciatus : The risk of death was significantly higher in dosage 60 (HR=516.41 (95% CI:292.07 – 913.07), P-value<0.001), dosage 45 (HR=91.53 (95% CI:52.45 – 159.75), P-value<0.001), dosage 30 (HR=20.82 (95% CI: 11.88 – 36.49), P-value<0.001) and dosage 15 (HR=3.72 (95% CI: 1.99 – 7.00), P-value<0.001) when compared to control i.e., dosage 0 (Table 4) (Figure 1-C). Effects on life history characteristics of mosquitoes fed with fluralaner Fecundity The overall fecundity (average number of eggs laid per female mosquito) of Ae. aegypti, An. Stephensi and Cx. quinquefasciatus females treated with fluralaner at different drug concentrations are given in (Table 5). The mean fecundity differed significantly between treatment and control groups at drug concentrations nearing 24 h LC 50 value for all the three species tested (P<0.05) (Table 5). At higher drug concentration, dead adults with half abdomen filled with eggs were found in the oviposition cups in case of An. stephensi and Cx. quinquefasciatus. In the case of Cx. quinquefasciatus distorted egg rafts were found in the oviposition cups i.e., many eggs were scattered. Egg hatch success The egg hatch success of Ae. aegypti, An. Stephensi and Cx. quinquefasciatus mosquitoes treated with fluralaner at different drug concentrations are given in Table 6. The mean egg hatch success differed significantly between the treatment and control groups (P<0.05) at higher drug concentrations, compared to the effect observed at lower concentrations (Table 6). Immature development The overall immature development of Ae. aegypti, An. Stephensi and Cx. quinquefasciatus mosquitoes treated with fluralaner at different drug concentrations are given in Table 7. The mean immature development varied significantly between control and treatment groups at higher drug concentrations tested – for Ae. aegypti – 20 ng/mL; for An. stephensi – 25 ng/mL; for Cx. quinquefasciatus – 45 ng/mL (P<0.05 for all species). Adult emergence success The overall adult emergence success of Ae. aegypti, An. Stephensi and Cx. quinquefasciatus mosquitoes exposed to fluralaner at different drug concentrations are given in Table 8. Certain morphological abnormalities were observed in a few drug exposed mosquitoes such as partially emerged adults and emerged adult mosquitoes with their legs attached to the pupal exuviae (Figure 2A-D). Estimation of mortality over time Estimation of mortality at different time points on feeding with different concentrations of the drug in all the three species is depicted in Graphs 1-3. Speed of killing was also studied at a concentration 4-5 folds higher (250 ng/mL) (Graph 4). When treated with this concentration, An. stephensi showed 88.00% mortality at 2 h post blood feeding; mortality rate increased to 100.00% at 3 h of post blood feeding. In the case of Ae. aegypti , 10.00% mortality was recorded at 2 h post blood feeding which increased to 99.00% at 6 h of post blood feeding. Cx. quinquefasciatus showed only 1.00% mortality at 2 h post blood feeding and 100.00% mortality at 7 h post blood feeding. Discussion An ideal endectocide for administration as a mass therapy tool for vector control should have potent mosquitocidal effect along with long half-life property so that the drug effect lasts for weeks to months after one-time drug administration. In the present study, oral treatment with fluralaner resulted in significant mosquitocidal effect against the three vector species studied, with the estimated 24 h LC 50 values ranging from 24.04 ng/mL to 49.82 ng/mL. The current study is the first attempt that examined the effect of fluralaner against Cx. quinquefasciatus . Mosquitocidal effect of fluralaner observed in the current study was similar to that reported in an earlier study, which reported 24 h LC 50 values of 18.48-51.52 ng/mL (Miglianico et al., 2018). The 24 h LC 50 value of fluralaner observed in the earlier study as well as in the current study was lower than that reported for ivermectin (LC 50 : 44.68-49.40 ng/mL) (Sampaio et al., 2016) . After a single oral administration of fluralaner at 50 mg/kg body weight in dogs, 100 ng/mL concentration of fluralaner was maintained in the blood for a period of 3 months (Kilp et al., 2014). Mosquito fed on dog treated with fluralaner reported to have significant reduction in their survival and fecundity for up to 13 weeks of post-treatment (Evans et al., 2022). It is noteworthy that the blood level concentration of fluralaner observed in dogs for three months is well above the 24 h LC 50 values (24.04-49.82 ng/mL) of fluralaner observed against three species in the current study. Fluralaner affected the fecundity, egg hatch success, immature development and adult emergence success in all the three species when they were exposed to drug concentrations nearing 24 h LC 50 values of 24.04 ng/mL for Ae. aegypti , 33.22 ng/mL for An. stephensi and 49.82 ng/mL for Cx. quinquefasciatus with significant P value. Although these differences were statistically significant but may not hold any importance under field conditions. However, there was no effect or only minimal effect at lower drug concentrations. This was in contrast to that reported for ivermectin which showed significant effect on fecundity and egg hatch success at sub lethal concentrations – LC 5 value 18.28 ng/mL (Sampaio et al., 2016). It is curious that fluralaner’s anti-fecundity activity is relatively lesser to that of ivermectin, even though their mortality effects are similar post blood feeding. One possible reason for this effect could be that ivermectin and isoxazoline compounds may have different mechanism of action regarding the anti-fecundity effect. Further research may be desirable in understanding this differential effect. Though fluralaner did not cause any visible impact on the development of the pupae into adults, certain morphological abnormalities like partially emerged adults and emerged adults with their legs attached to the pupal exuviae were observed. In an earlier study, fluralaner has been shown to affect the adult emergence success in Spodoptera litura Fabricius (Lepidoptera: Noctuidae), even at sub-lethal dose of LC 15 morphological abnormalities like notched wings were observed in the adults (Liu et al., 2018). Studies on susceptible and resistant strains of horn and house flies have shown high level of mortality at lower concentration with fluralaner when compared with imidacloprid (Burgess et al., 2020). In a similar study on fipronil and pyrethroid resistant bed bugs, fluralaner outperformed ivermectin with a high mortality rate up to 28 days of post-treatment (González-Morales et al., 2022). In the current study complete mortality was observed within 7 h of post blood feeding at 250 ng/mL drug concentration in all the three species tested. Blood concentration much above this level is expected during the initial weeks in animals treated with fluralaner. This is a notable advantage over ivermectin which takes 72 h to produce 70.00-100.00% morality in mosquitoes at a dose of 93 ng/mL in animal experiments (Bastiaenset al., 2012, Walther al. 2015, Mekuriawet al., 2019) and also this plasma level lasts for less than one day only. Currently, ivermectin mass therapy is being investigated in peri-domestic animals and humans in field studies for its endectocidal effect and as a complementary tool for malaria vector control (Pooda et al., 2015, Chaccouret al., 2013) . Fluralaner, with its comparable mosquitocidal effect to ivermectin coupled with the long half–life may be a good alternative to ivermectin. With several emerging and remerging vector borne and zoonotic diseases, there is great interest in ‘One-Health’ efforts being enforced round the globe. This would be particularly relevant in Africa and Asia where humans and animals live in close proximity and the drug based vector control strategy holds promise in such settings (Ruiz-Castillo et al., 2022). Moreover, it was found to be useful in insecticide resistance management plans for other active ingredients such as DDT, pyrethroid insecticides and other insecticides with possible reason of its distinct binding site from the targets of known modulators of ionotropic GABA receptors (Miglianico et al., 2018, Wang et al., 2022). Though fluralaner appears to be a promising candidate for vector control there are some limitations that are to be resolved before proceeding further. High cost of fluralaner is a barrier. Another issue is the safety aspects of fluralaner in human beings. It is not licensed for use in human beings. However, no adverse events were reported when fluralaner and afoxolaner were used for treating scabies in human clinical trials (Goldust 2018, Goldust and Alipour 2019). There are some limitations of the current study. We used chicken blood, not human blood for carrying out the efficacy trials for convenience. Another limitation is that we have carried out the study only in laboratory maintained colony and not on field caught mosquitoes. The survived mosquitoes assessed for life history characteristics were not comparable between treatment and control groups. The lab obtained results may not be exactly replicable in field conditions. Another limitation is that we did not perform direct blood feeding experiments with mosquitoes fed upon animals treated with fluralaner. This experiment is necessary to understand the effect of the drug on mosquitoes in the context of its metabolism in the treated animals. In conclusion, our study showed significant oral toxic effect of fluralaner in adult vector mosquitoes with additional effect on life history characteristics (reduced fecundity, egg hatch success, larval development and adult emergence). Fluralaner may be a suitable candidate for future drug based mosquito control strategies. It is required to carry out future safety studies of fluralaner in human beings. Declarations Conflicting Interest Authors hereby declared that no conflict of interest is involved in this study. Ethics approval Not applicable Consent to participate Not applicable Consent for publication All authors approved and gave their consent to publish the manuscript. Availability of data and material The datasets generated and/or analyzed during the current study are available from the corresponding author on request. Acknowledgments We gratefully acknowledge the technical assistance of Mr. Shrihari Murmu, Senior Technician–2 and Mr. K. Manimaran, Technical Officer-A, ICMR-Vector Control Research Centre for maintenance of mosquito colonies and blood feeding. We thank Mr. M. Sundharesan, Technical Assistant, ICMR-Vector Control Research Centre for his assistance in extraction of active compound from the commercially available tablet. We acknowledge the financial support by ICMR-Vector Control Research Centre, Puducherry. Funding: We acknowledge the institutional funding provided to carry out the study. Contributors’ list: VSK, SC and HKS contributed in study design. HKS, VS and SV executed the work, wrote the initial draft of the manuscript. NM helped for extraction of active compound. VB worked out statistical analysis for the data. VSK and AK critically reviewed the final version of manuscript. All authors read and approved the final version of the manuscript. References Bastiaens, G. J. H., G.-J. van Gemert, J. Hooghof, S. W. Lindsay, C. Drakeley, T. S. Churche, J. P. Verhave, C. H.M. Kocken, R. W. Sauerwein, and T. Bousema (2012) Duration of the mosquitocidal effect of ivermectin. Malariaworld J 3(10). Burgess, E. R., C. J. Geden, K. H. Lohmeyer, B. H. King, E. T. Machtinger, and J. G. 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Bloomquist (2017) Mosquitocidal Activity and Mode of Action of the Isoxazoline Fluralaner. Int J Environ Res Public Health. 14(2):154. https://doi.org/10.3390/ijerph14020154 Kilp, S., D. Ramirez, M. J. Allan, and R. K. Roepke (2016) Comparative pharmacokinetics of fluralaner in dogs and cats following single topical or intravenous administration. Parasit Vectors. 9(1):296. https://doi.org/10.1186/s13071-016-1564-8 Kilp, S., D. Ramirez, M. J. Allan, R. K. A. Roepke, and M. C. Nuernberger (2014) Pharmacokinetics of fluralaner in dogs following a single oral or intravenous administration. Parasit Vectors. 7(85). https://doi.org/10.1186/1756-3305-7-85 Liu, D., Z.-Q. Jia, Y.-C. Peng, C.-W. Sheng, T. Tang, L. Xu, Z.-J. Han, and C.-Q. Zhao (2018) Toxicity and sublethal effects of fluralaner on Spodoptera litura Fabricius (Lepidoptera: Noctuidae). Pestic Biochem Physiol. 152:8–16. https://doi.org/10.1016/j.pestbp.2018.08.004 Mekuriaw, W., M. Balkew, L. A. Messenger, D. Yewhalaw, A. Woyessa, and F. Massebo (2019) The effect of ivermectin® on fertility, fecundity and mortality of Anopheles arabiensis fed on treated men in Ethiopia. Malar J. 18(1):357. https://doi.org/10.1186/s12936-019-2988-3 Miglianico, M., M. Eldering, H. Slater, N. Ferguson, P. Ambrose, R. S. Lees, K. M. J. Koolen, K. Pruzinova, M. Jancarova, P. Volf, C. J. M. Koenraadt, H.-P. Duerr, G. Trevitt, B. Yang, A. K. Chatterjee, J. Wisler, A. Sturm, T. Bousema, R. W. Sauerwein, P. G. Schultz, M. S. Tremblay, and K. J. Dechering (2018) Repurposing isoxazoline veterinary drugs for control of vector-borne human diseases. Proc Natl Acad Sci U S A. 115(29):E6920-E6926. https://doi.org/10.1073/pnas.1801338115 Ōmura, S., and A. Crump (2017) Ivermectin and malaria control. Malar J. 16(1):172. https://doi.org/10.1186/s12936-017-1825-9 Pooda, H. S., J.-B. Rayaisse, D. F. de S. Hien, T. Lefèvre, S. R. Yerbanga, Z. Bengaly, R. K. Dabiré, A. M. G. Belem, I. Sidibé, P. Solano, and K. Mouline (2015) Administration of ivermectin to peridomestic cattle: a promising approach to target the residual transmission of human malaria. Malar J. 13 Suppl 1:496. https://doi.org/10.1186/s12936-015-1001-z Ruiz-Castillo, P., C. Rist, R. Rabinovich, and C. Chaccour (2022) Insecticide-treated livestock: a potential One Health approach to malaria control in Africa. Trends in Parasitol. 38(2):112-123. https://doi.org/10.1016/j.pt.2021.09.006 Sampaio, V. S., T. P. Beltrán, K. C. Kobylinski, G. C. Melo, J. B. P. Lima, S. G. M. Silva, Í. C. Rodriguez, H. Silveira, M. G. V. B. Guerra, Q. Bassat, P. F. P. Pimenta, M. V. G. Lacerda, and W. M. Monteiro (2016) Filling gaps on ivermectin knowledge: effects on the survival and reproduction of Anopheles aquasalis, a Latin American malaria vector. Malar J. 15(1):491. https://doi.org/10.1186/s12936-016-1540-y Slater, H. C., B. D. Foy, K. Kobylinski, C. Chaccour, O. J. Watson, J. Hellewell, G. Aljayyoussi, T. Bousema, J. Burrows, U. D’Alessandro, H. Alout, F. O. Ter Kuile, P. G. T. Walker, A. C. Ghani, and M. R. Smit (2020) Ivermectin as a novel complementary malaria control tool to reduce incidence and prevalence: a modelling study. Lancet Infect Dis. 20(4):498-508. https://doi.org/10.1016/s1473-3099(19)30633-4 Walther, F. M., M. J. Allan, and R. K. A. Roepke (2015) Plasma pharmacokinetic profile of fluralaner (Bravecto TM ) and ivermectin following concurrent administration to dogs. Parasit Vectors. 8:508. https://doi.org/10.1186/s13071-015-1123-8 Wang, Q., H. Wang, Y. Zhang, J. Chen, A. Upadhyay, B. Bhowmick, J. Hang, S. Wu, C. Liao, and Q. Han (2022) Functional analysis reveals ionotropic GABA receptor subunit RDL is a target site of ivermectin and fluralaner in the yellow fever mosquito, Aedes aegypti . Pest Manag Sci. 78(10):4173-82. https://doi.org/10.1002/ps.7035 World Health Organization (2017) Global vector control response 2017–2030. https://www.who.int/publications-detail-redirect/9789241512978. Accessed 22 December 2022 World Health Organization (2020) Vector-borne diseases. https://www.who.int/news-room/fact-sheets/detail/vector-borne-diseases. Accessed 22 December 2022 Tables Table 1. Lethal concentration values Species Post blood feeding time (h) Slope (±SE) Chi 2 goodness of fit (P- value) LC 50 (95% CI) Heterogeneity factor Aedes aegypti 24 h 0.11 (0.004) 3.13 (0.08) 24.04 (14.74 – 45.27) 3.13 Anopheles stephensi 24 h 0.12 (0.005) 2.40 (0.12) 33.22 (24.30 – 51.56) 2.40 Culex quinquefasciatus 24 h 0.17 (0.01) 0.11 (0.90) 49.82 (49.35 – 50.29) 0.11 Probit regression analysis: SE – Standard error; CI – Confidence interval; LC – Lethal Concentration Table 2. Effect of fluralaner on survivorship of Aedes aegypti Dose Median Survival duration Cox-regression analysis HR (95% CI) P-value 0 (Control) - 1.00 - 5 ng/mL - 3.45(2.07 – 5.74) <0.001 10 ng/mL 96 h 17.19(10.86 – 27.22) <0.001 20 ng/mL 48 h 40.61(25.77 – 64.01) <0.001 35 ng/mL 24 h 150.45(94.76 – 238.87) <0.001 Cox-regression analysis: HR – hazard ratios; CI – Confidence interval Table 3. Effect of fluralaner on survivorship of Anopheles stephensi Dose Median Survival duration Cox-regression analysis HR (95% CI) P-value 0 (Control) - 1.00 - 5 ng/mL - 3.48(2.02 – 5.97) <0.001 15 ng/mL - 17.21(10.63 – 27.88) <0.001 25 ng/mL 72 h 39.72(24.73 – 63.80) <0.001 40 ng/mL 24 h 231.94(142.68 – 377.05) <0.001 Cox-regression analysis: HR – hazard ratios; CI – Confidence interval Table 4. Effect of fluralaner on survivorship of Culex quinquefasciatus Dose Median Survival duration Cox-regression analysis HR (95% CI) P-value 0 (Control) - 1.00 - 15 ng/mL - 3.72(1.99 – 7.00) <0.001 30 ng/mL - 20.82(11.88 – 36.49) <0.001 45 ng/mL 48 h 91.53(52.45 – 159.75) <0.001 60 ng/mL 18 h 516.41(292.07 – 913.07) <0.001 Cox-regression analysis: HR – hazard ratios; CI – Confidence interval Table 5. Fecundity success Mosquito species Fluralaner (ng/mL) Number of eggs laid per mosquito Mean(SD) Mean difference P-value n Test n Control Aedes aegypti 5 48 95.10(1.70) 49 97.70(0.60) 2.60 0.018 10 39 91.90(1.30) 49 96.70(0.60) 4.80 0.012 20 16 78.70(1.00) 48 96.70(1.10) 18.00 0.011 Anopheles stephensi 5 48 88.30(0.50) 50 90.30(0.60) 2.00 0.007 15 40 84.20(1.50) 50 90.00(0.00) 5.80 0.011 25 31 72.00(2.10) 50 89.30(0.60) 17.30 0.012 Culex quinquefasciatus 15 47 124.20(2.60) 49 136.00(3.60) 11.80 0.012 30 40 120.30(2.10) 49 134.30(2.50) 14.00 0.011 45 7 117.30(3.00) 49 135.30(1.50) 18.00 0.012 n= number kept for fecundity success Table 6. Egg hatch success Mosquito species Fluralaner (ng/mL) Number of eggs hatched Mean(SD) Mean difference P-value n Test n Control Aedes aegypti 5 500 403.67(4.74) 500 414.00(6.24) 10.33 0.026 10 500 395.67(5.29) 500 419.67(4.04) 24.00 0.013 20 500 386.00(5.17) 500 416.00(3.60) 30.00 0.012 Anopheles stephensi 5 500 435.22(6.96) 500 456.33(4.51) 21.11 0.013 15 500 420.22(7.05) 500 452.67(3.05) 32.44 0.013 25 500 408.89(4.94) 500 453.33(12.86) 44.44 0.013 Culex quinquefasciatus 15 517 426.22(9.97) 533 462.66(8.14) 36.44 0.013 30 524 425.11(11.88) 515 446.00(18.68) 20.89 0.060 45 518 417.22(4.82) 531 457.33(24.68) 40.11 0.012 n= number kept for egg hatch success Table 7. Immature survival Mosquito species Fluralaner (ng/mL) Number of pupae developed Mean(SD) Mean difference P-value n Test n Control Aedes aegypti 5 100 98.80(0.70) 100 99.70(0.60) 0.90 0.07 10 100 97.40(1.10) 100 99.00(1.00) 1.60 0.07 20 100 91.90(2.60) 100 99.00(0.00) 7.10 0.01 Anopheles stephensi 5 100 99.10(0.80) 100 100.00(0.00) 0.90 0.07 15 100 97.80(1.40) 100 99.00(0.00) 1.20 0.11 25 100 96.40(1.30) 100 98.70(0.60) 2.20 0.01 Culex quinquefasciatus 15 100 98.20(0.70) 100 99.30(1.10) 1.10 0.11 30 100 93.80(1.90) 100 99.30(0.60) 5.60 0.01 45 100 89.20(3.00) 100 99.30(0.60) 10.10 0.01 n= number kept for immature survival Table 8. Adult emergence success Mosquito species Fluralaner (ng/mL) Number of adults emerged Mean(SD) Mean difference P-value n Test n Control Aedes aegypti 5 50 49.56(0.53) 50 50.00(0.00) 0.44 0.17 10 50 49.22(0.67) 50 49.67(0.58) 0.44 0.30 20 50 48.44(0.88) 50 50.00(0.00) 1.56 0.02 Anopheles stephensi 5 50 49.44(0.53) 50 50.00(0.00) 0.56 0.10 15 50 48.00(0.71) 50 49.67(0.58) 1.67 0.01 25 50 46.78(1.30) 50 48.67(0.58) 1.89 0.04 Culex quinquefasciatus 15 50 49.56(0.53) 50 50.00(0.00) 0.44 0.17 30 50 48.78(0.44) 50 49.67(0.58) 0.89 0.02 45 50 47.67(1.32) 50 49.67(0.58) 2.00 0.02 n= number kept for adult emergence success Graphs Graphs 1 to 4 are available in the Supplementary Files section. Supplementary Files Supplementarydata.docx Supplementary graphs S1: Dose response curve for Ae. aegypti at 24 h post blood feeding S2: Dose response curve for An. stephensi at 24 h post blood feeding S3: Dose response curve for Cx. quinquefasciatus at 24 h post blood feeding Graph1.png Graph 1: Mean percent mortality observed with Ae. aegypti adults treated with different concentration of fluralaner at different time points Graph2.png Graph 2: Mean percent mortality observed with An. stephensi adults treated with different concentration of fluralaner at different time point Graph3.png Graph 3: Mean percent mortality observed with Cx. quinquefasciatus adults treated with different concentration of fluralaner at different time point Graph4.png Graph 4: Mean percent mortality at different time point observed with three species of vector mosquitoes treated with fluralaner at a concentration of 250 ng/mL Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. 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1\u003c/strong\u003e: Mean percent mortality observed with \u003cem\u003eAe. aegypti\u003c/em\u003e adults treated with different concentration of fluralaner at different time points\u003c/p\u003e","description":"","filename":"Graph1.png","url":"https://assets-eu.researchsquare.com/files/rs-2803950/v1/efb944cf88123f2320a86ddc.png"},{"id":37444431,"identity":"655fe538-0063-400f-a22d-d643702777f2","added_by":"auto","created_at":"2023-05-24 15:31:15","extension":"png","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":428153,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGraph 2\u003c/strong\u003e: Mean percent mortality observed with \u003cem\u003eAn. stephensi\u003c/em\u003e adults treated with different concentration of fluralaner at different time point\u003c/p\u003e","description":"","filename":"Graph2.png","url":"https://assets-eu.researchsquare.com/files/rs-2803950/v1/7985ee0d50167c46ab1695b6.png"},{"id":37444436,"identity":"2fbacd92-65f9-4d7b-8f68-d92f70c6cb38","added_by":"auto","created_at":"2023-05-24 15:31:15","extension":"png","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":412954,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGraph 3\u003c/strong\u003e: Mean percent mortality observed with \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e adults treated with different concentration of fluralaner at different time point\u003c/p\u003e","description":"","filename":"Graph3.png","url":"https://assets-eu.researchsquare.com/files/rs-2803950/v1/323e2a28a7c9a1d5d61552a2.png"},{"id":37444430,"identity":"0aa62ec6-7025-4441-9029-2be0ad479fae","added_by":"auto","created_at":"2023-05-24 15:31:15","extension":"png","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":413259,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eGraph 4\u003c/strong\u003e: Mean percent mortality at different time point observed with three species of vector mosquitoes treated with fluralaner at a concentration of 250 ng/mL\u003c/p\u003e","description":"","filename":"Graph4.png","url":"https://assets-eu.researchsquare.com/files/rs-2803950/v1/6bc3ea85b9e6ac979fc96896.png"}],"financialInterests":"","formattedTitle":"Efficacy of fluralaner, a long acting acaricide against three species of vector mosquitoes (Diptera: Culicidae) under laboratory conditions – a potential candidate for drug based vector control","fulltext":[{"header":"Introduction","content":"\u003cp\u003eVector borne diseases such as malaria, lymphatic filariasis, dengue, zika and others pose significant public health threats globally\u0026nbsp;(WHO 2020). Effective vaccines or specific treatment are not available for many of these diseases and vector control is the only option for \u0026nbsp; prevention and control (WHO 2017)\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eHowever, conventional vector control methods have \u0026nbsp; several limitations and there is an urgent need for alternative control strategies. With the discovery of the potent mosquitocidal effect of the anti-parasitic drug ivermectin, vector control with endectocides is emerging as an alternative strategy (Foy et al., 2011,\u0026nbsp;Imbahale et al., 2019). This approach involves mass administration of an oral or systemic insecticidal drug to animal or human population for killing the blood feeding arthropod vectors. Ivermectin is now being explored as a mass drug administration (MDA) tool in humans for malaria control in many settings (Foley\u0026nbsp;et al., 2000,\u0026nbsp;Chaccour\u0026nbsp;et al., 2010, Foy et al., 2019,\u0026nbsp;Slater et al., 2020).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003eAs we progress towards malarial elimination, due to the extensive use of LLIN and IRS, mosquitoes\u0026rsquo; feeding behavior may change and they may prefer to feed on animals rather than humans. Hence mass therapy of domestic animals with ivermectin is also suggested (Chaccour\u0026nbsp;et al., 2013,\u0026nbsp;Pooda et al., 2015, Ōmura and Crump 2017, Chaccour and Rabinovich 2019). However, one major drawback for application of ivermectin for vector control is its short half-life (2.07\u0026plusmn;0.71 days) (Walther\u0026nbsp;et al., 2015). This would necessitate its very frequent administration to human or animal population which may limit its utility in mass therapy. But drugs with shorter half-life tend to be safer and slow release formulation could bypass the need of frequent administration.\u003c/p\u003e\n\u003cp\u003eAlternative endectocides with longer half-life which are proved to be safe or having only minimal side effects in animals and humans are highly desirable. Isoxazoline compounds like fluralaner (Bravecto\u0026reg;) and afoxolaner (NexGard\u0026reg;) used as acaricides in veterinary practice have an unusually long half-life (14.27\u0026plusmn;2.53 days) (Kilp et al., 2014, Comas and Armstrong 2018).\u0026nbsp;After a single oral administration (50 mg/kg) of fluralaner in dogs, maximum plasma concentrations (C\u003csub\u003emax\u003c/sub\u003e) of 5419 \u0026plusmn; 2086 ng/mL have been achieved within 24 h and detectable plasma levels were found up to 112 days post administration (Kilp et al.,2014, Kilp et al.,2016). The mechanism of action of fluralaner is similar to that of ivermectin. It is a potent inhibitor of \u0026gamma;-amino butyric acid (GABA) and L-glutamate-gated chloride channels, but binds at a site different from that of ivermectin (Gassel et al.,2014). Isoxazoline compounds may have minimal or no effect on the vertebrate nervous system (Kilp et al., 2014)\u003cstrong\u003e.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWith its excellent safety profile in animals, long half-life and the potent acaricidal effect, fluralaner can be an alternative choice for drug based vector control strategies. However, information on its mosquitocidal effect is very limited (Jiang S et al., 2016, Miglianico et al., 2018). A recent study from USA (United States of America) reported its significant lethal effect in different mosquito vector species such as \u003cem\u003eAnopheles stephensi\u0026nbsp;\u003c/em\u003e(Liston, 1901), \u003cem\u003eAnopheles gambiae\u003c/em\u003e Kisumu, \u003cem\u003eAnopheles gambiae\u003c/em\u003e Tiassal\u0026eacute; (Gile, 1902), \u003cem\u003eAedes aegypti\u003c/em\u003e New Orleans, \u003cem\u003eAedes aegypti\u003c/em\u003e Cayman (Linnaeus, 1762) and \u003cem\u003eCulex pipiens\u0026nbsp;\u003c/em\u003e(Linnaeus, 1758) (Diptera: Culicidae) species (Miglianico et al., 2018).\u003c/p\u003e\n\u003cp\u003eIn the present study, the adulticidal effect of fluralaner was studied against three vector mosquito species from India: 1) \u003cem\u003eAedes aegypti\u003c/em\u003e, the primary vector of dengue, 2) \u003cem\u003eAnopheles stephensi\u003c/em\u003e, the vector of urban malaria, and 3) \u003cem\u003eCulex quinquefasciatus\u0026nbsp;\u003c/em\u003e(Say, 1823)\u003cem\u003e\u0026nbsp;\u003c/em\u003e(Diptera: Culicidae), the vector of lymphatic filariasis under laboratory conditions. In addition, the effect of fluralaner on fecundity, egg hatch success, immature development and adult emergence success were investigated on mosquitoes fed with different concentration of the drug.\u0026nbsp;\u003c/p\u003e"},{"header":"Materials and Methods","content":"\u003cp\u003e\u003cstrong\u003e\u003cem\u003eExtraction of fluralaner from the commercial tablet\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFluralaner was extracted from the commercially available tablets for veterinary use (Bravecto\u0026reg;- purchased from local veterinary drug shop; manufactured by Merck Animal Health, Madison, Vienna, Austria) using the method described by\u0026nbsp;Miglianico et al., 2018. The tablet was crushed into fine powder using a pestle and mortar and then dissolved in a solvent prepared by mixing dichloromethane and methanol in a 1:1 ratio. The whole mixture was then agitated at room temperature for 1 h with a magnetic stirrer and filtered using Whatman\u0026reg; grade 1 filter paper (purchased from Sigma-Aldrich, St. Louis, MO). The filtrate was left overnight for evaporation and the concentrated final product was obtained. Stock solution (5 mg/mL) was prepared in dimethyl sulfoxide (DMSO) (purchased from HiMedia, Mumbai, India) and aliquots were frozen at \u0026minus;20\u0026deg;C. Prior to each experiment, aliquots were dissolved in phosphate buffer saline (PBS) (purchased from Sigma-Aldrich, St. Louis, MO) and added to heparinized chicken blood to achieve the desired final drug concentrations required for the mosquito blood feeding experiments.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eVector mosquitoes and colony maintenance\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eLaboratory reared mosquito species of \u003cem\u003eAe. aegypti\u003c/em\u003e, \u003cem\u003eAn. Stephensi\u003c/em\u003e and\u003cem\u003e\u0026nbsp;Cx. quinquefasciatus\u0026nbsp;\u003c/em\u003e were obtained from the rearing and colonization facility at the Indian Council of Medical Research - Vector Control Research Centre (ICMR-VCRC), Puducherry, India. These strains were originally collected from the Union territory of Puducherry, India and the colonies have been maintained in the insectary since 1975. \u0026nbsp;\u003cem\u003eCx. quinquefasciatus\u003c/em\u003e were collected in 1975 and \u003cem\u003eAe. aegypti\u003c/em\u003e and \u003cem\u003eAn. stephensi\u003c/em\u003e were collected in 1976. They were maintained at a constant temperature of 27\u0026plusmn;2\u0026deg;C and relative humidity of 80\u0026plusmn;10% in one-foot Barraud cages (30cm L X 30cm W X 30cm H) and provided with 10% sucrose solution.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eLaboratory bioassays with fluralaner\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eTo determine the activity range of fluralaner, initially, bioassays were performed to assess the mortality effect at 24 h using wide range concentrations (20 ng/mL-120 ng/mL). Based on the results of these experiments, a narrow range of four drug concentrations were selected for bioassays to determine LC\u003csub\u003e50\u003c/sub\u003e (concentration resulting in 50% mortality after taking a blood meal) value at 24 h of post blood feeding. Drug concentrations of 5 ng/mL, 10 ng/mL, 20 ng/mL and 35 ng/mL were selected for the feeding experiments with \u003cem\u003eAe aegypti;\u0026nbsp;\u003c/em\u003e5 ng/mL, 15 ng/mL, 25 ng/mL and 40 ng/mL for \u003cem\u003eAn. stephensi\u003c/em\u003e; and\u003cem\u003e\u0026nbsp;\u003c/em\u003e15 ng/mL, 30 ng/mL, 45 ng/mL and 60 ng/mL for \u003cem\u003eCx. quinquefasciatus\u0026nbsp;\u003c/em\u003erespectively (two drug concentration above and below LC\u003csub\u003e50\u0026nbsp;\u003c/sub\u003evalue of initial bioassay)\u003cem\u003e.\u003c/em\u003e\u003c/p\u003e\n\u003cp\u003eFive days old, non-blood fed female mosquitoes from same batch were used for blood feeding experiments. Prior to blood feeding, cotton pads soaked with 10% sucrose solution were removed and the adult mosquitoes were allowed to starve at least for 12 h to improve the feeding rate. Five mL of heparinized chicken blood containing desired concentration of fluralaner was used for feeding the mosquitoes in the treatment group via artificial blood feeding apparatus made of glass fitted with blood mixing rotor and parafilm for about 60-90 minutes. Treatment group consists of 4 drug concentration and each concentration constituted of three technical replicate cages (each with 50 females). 50 adult female from the same batch of mosquitoes fed on chicken blood with only solvent were\u0026nbsp;used as controls. After blood feeding, mosquitoes (fully engorged) were transferred to clean cages, provided with 10% sucrose solution\u0026nbsp;and the mortality effect was observed\u0026nbsp;at 4, 8, 18, 24, 48, 72 and 96 h of post feeding. 48 h (\u003cem\u003eAe. aegypti\u003c/em\u003e,\u003cem\u003e\u0026nbsp;An. stephensi\u003c/em\u003e) and 72 h (\u003cem\u003eCx. quinquefasciatus\u003c/em\u003e) of post blood feeding, survived mosquitoes were observed for fecundity. The experiment was repeated three times on different days using fresh batches of mosquitoes and freshly prepared fluralaner concentration as biological replicates.\u003c/p\u003e\n\u003cp\u003eIn continuation to the bioassay experiments for mortality effect of fluralaner, life history characteristics of survived mosquitoes in each drug treatment groups were assessed in comparison to the control groups. Number of survived mosquitoes in each replicate cage varied with the species and also with the drug concentration. \u0026nbsp; \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eEffect of fluralaner on fecundity, egg hatch success, immature development and adult emergence success\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eFor fecundity assessment of survived \u003cem\u003eAe. aegypti\u0026nbsp;\u003c/em\u003eand\u003cem\u003e\u0026nbsp;An. stephensi\u003c/em\u003e, oviposition cups filled with 75-90 mL water lined with filter paper were introduced into the cages at 48 h of post blood feeding. For \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e, oviposition cups without filter paper were introduced into the cages at 72 h as it takes 3 days to become fully gravid. To assess the effect of fluralaner in egg hatch success, eggs laid by survived mosquitoes in treatment group were compared with that of eggs laid by mosquitoes in the control group. For hatching experiment of \u003cem\u003eAe. aegypti\u0026nbsp;\u003c/em\u003eand\u003cem\u003e\u0026nbsp;An. stephensi\u003c/em\u003e, five hundred eggs counted under microscope were floated in trays filled with water using a fine tip brush. In case of \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e, as eggs were laid in rafts, numbers were slightly more than five hundred. The rafts were carefully transferred to tray using brush without damaging it. Hundred first instar larvae hatched from eggs laid by survived mosquitoes were observed for development up to pupal stage by providing mosquito larval food containing yeast (40%) and dog biscuit (60%) and were compared with the control group. Daily mortality was recorded (if any) in both treatment and control group larvae. Similarly fifty pupae from survived mosquitoes were observed for adult emergence by transferring them into containers filled with 3/4\u003csup\u003eth\u003c/sup\u003e water and covered with mosquito net having hole in middle plugged with cotton. The results were compared with control group. \u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eEstimation of speed of killing at high concentration\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;We performed a set of experiment to assess the speed of killing of the mosquitoes when they were fed with a high concentration of fluralaner. In initial weeks of drug administration the concentration is expected to be higher which may result in fast mortality within hours of blood feeding. For this, 50 females in triplicates were fed at 250 ng/mL and were observed at hourly interval until complete (100%) mortality was noted in the treatment groups. The experiment was replicated three times on different days with fresh batch of mosquitoes.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eStatistical analysis\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eMean and standard deviation value from three replicates of different concentration were taken to obtain data, which was further used for various statistical analysis.\u0026nbsp;Mortality data over time were subjected to Probit regression analysis to estimate LC\u003csub\u003e50\u003c/sub\u003e value. Mean (SD) and Frequency (%) were used to describe summary values. \u0026nbsp;Mann Whitney U test was used to test the difference in fecundity, egg hatch success, immature survival and adult emergence success between the drug treated and control groups. P-value \u0026lt;0.05 was considered as statistically significant. SPSS 16.0 was used for carrying out probit analysis whereas, STATA 14.2 software for other statistical analysis.\u003c/p\u003e"},{"header":"Results","content":"\u003cp\u003e\u003cstrong\u003e\u003cem\u003eEstimated LC\u003csub\u003e50\u0026nbsp;\u003c/sub\u003evalue of fluralaner at different time points\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eBased on the preliminary results of initial bioassay, experiments were performed with narrow range drug concentrations (with three technical replicates), as described in methodology section for each of the species and the 24 h LC\u003csub\u003e50\u003c/sub\u003e (95% CI)\u003csub\u003e\u0026nbsp;\u003c/sub\u003evalue was estimated. The results are summarized in (Table 1). Since the upper limit for 95% CI of 24 h LC\u003csub\u003e50\u003c/sub\u003e was exceeding the maximum dosage used for\u0026nbsp;\u003cem\u003eAe. aegypti\u0026nbsp;\u003c/em\u003eand\u003cem\u003e\u0026nbsp;An. stephensi\u003c/em\u003e, the mortality data was subjected to cox-regression analysis to compare the survivorship or risk of death between different treatment and control group.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eCox-regression analysis\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAe. aegypti\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e:\u003c/strong\u003e The risk of death was significantly higher in dosage 35 (HR=150.45 (95% CI:94.76 \u0026ndash; 238.87), P-value\u0026lt;0.001), dosage 20 (HR=40.61 (95% CI:25.77 \u0026ndash; 64.01), P-value\u0026lt;0.001), dosage 10 (HR=17.19 (95% CI: 10.86 \u0026ndash; 27.22), P-value\u0026lt;0.001) and dosage 5 (HR=3.45 (95% CI: 2.07 \u0026ndash; 5.74), P-value\u0026lt;0.001) when compared to control i.e., dosage 0 (Table 2) (Figure 1-A).\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003cstrong\u003e\u003cem\u003eAn. stephensi\u003c/em\u003e:\u0026nbsp;\u003c/strong\u003eThe risk of death was significantly higher in dosage 40 (HR=231.94 (95% CI:142.68 \u0026ndash; 377.05), P-value\u0026lt;0.001), dosage 25 (HR=39.72 (95% CI:24.73 \u0026ndash; 63.80), P-value\u0026lt;0.001), dosage 15 (HR=17.21 (95% CI: 10.63 \u0026ndash; 27.88), P-value\u0026lt;0.001) and dosage 5 (HR=3.48 (95% CI: 2.02 \u0026ndash; 5.97), P-value\u0026lt;0.001) when compared to control i.e., dosage 0 (Table 3) (Figure 1-B).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eCx. quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003cstrong\u003e:\u0026nbsp;\u003c/strong\u003eThe risk of death was significantly higher in dosage 60 (HR=516.41 (95% CI:292.07 \u0026ndash; 913.07), P-value\u0026lt;0.001), dosage 45 (HR=91.53 (95% CI:52.45 \u0026ndash; 159.75), P-value\u0026lt;0.001), dosage 30 (HR=20.82 (95% CI: 11.88 \u0026ndash; 36.49), P-value\u0026lt;0.001) and dosage 15 (HR=3.72 (95% CI: 1.99 \u0026ndash; 7.00), P-value\u0026lt;0.001) when compared to control i.e., dosage 0 (Table 4) (Figure 1-C).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eEffects on life history characteristics of mosquitoes fed with fluralaner \u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eFecundity\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe overall fecundity (average number of eggs laid per female mosquito) of \u003cem\u003eAe. aegypti, An. Stephensi\u0026nbsp;\u003c/em\u003eand \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e females treated with fluralaner at different drug concentrations are given in (Table 5). The mean fecundity differed significantly between treatment and control groups at drug concentrations nearing 24 h LC\u003csub\u003e50\u0026nbsp;\u003c/sub\u003evalue for all the three species tested (P\u0026lt;0.05) (Table 5). \u0026nbsp;At higher drug concentration, dead adults with half abdomen filled with eggs were found in the oviposition cups in case of \u003cem\u003eAn. stephensi and Cx. quinquefasciatus.\u003c/em\u003e In the case of \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e distorted egg rafts were found in the oviposition cups i.e., many eggs were scattered.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eEgg hatch success\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe egg hatch success of \u003cem\u003eAe. aegypti, An. Stephensi\u0026nbsp;\u003c/em\u003eand \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e mosquitoes treated with fluralaner at different drug concentrations \u0026nbsp;are given in Table 6. The mean egg hatch success differed significantly between the treatment and control groups (P\u0026lt;0.05) at higher drug concentrations, compared to the effect observed at lower concentrations (Table 6).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eImmature development\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe overall immature development of \u003cem\u003eAe. aegypti, An. Stephensi\u0026nbsp;\u003c/em\u003eand \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e mosquitoes treated with fluralaner at different drug concentrations are given in Table 7. The mean immature development varied significantly between control and treatment groups at higher drug concentrations tested \u0026ndash; for \u003cem\u003eAe. aegypti \u0026ndash;\u0026nbsp;\u003c/em\u003e20 ng/mL; for \u003cem\u003eAn. stephensi \u0026ndash;\u0026nbsp;\u003c/em\u003e25 ng/mL; for \u003cem\u003eCx. quinquefasciatus \u0026ndash;\u003c/em\u003e 45 ng/mL (P\u0026lt;0.05 for all species).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eAdult emergence success\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe overall adult emergence success of \u003cem\u003eAe. aegypti, An. Stephensi\u0026nbsp;\u003c/em\u003eand \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e mosquitoes exposed to fluralaner at different drug concentrations are given in Table 8. Certain morphological abnormalities were observed in a few drug exposed mosquitoes such as partially emerged adults and emerged adult mosquitoes with their legs attached to the pupal exuviae (Figure 2A-D).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u003cem\u003eEstimation of mortality over time\u0026nbsp;\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eEstimation of mortality at different time points on feeding with different concentrations of the drug in all the three species is depicted in Graphs 1-3. Speed of killing was also studied at a concentration 4-5 folds higher (250 ng/mL) (Graph 4). When treated with this concentration, \u003cem\u003eAn. stephensi\u0026nbsp;\u003c/em\u003eshowed 88.00% mortality at 2 h post blood feeding; mortality rate increased to 100.00% at 3 h of post blood feeding. In the case of \u003cem\u003eAe. aegypti\u003c/em\u003e, 10.00% mortality was recorded at 2 h post blood feeding which increased to 99.00% at 6 h of post blood feeding. \u003cem\u003eCx. quinquefasciatus\u0026nbsp;\u003c/em\u003eshowed only 1.00% mortality at 2 h post blood feeding and 100.00% mortality at 7 h post blood feeding.\u003c/p\u003e"},{"header":"Discussion","content":"\u003cp\u003eAn ideal endectocide for administration as a mass therapy tool for vector control should have potent mosquitocidal effect along with long half-life property so that the drug effect lasts for weeks to months after one-time drug administration. In the present study, oral treatment with fluralaner resulted in significant mosquitocidal effect against the three vector species studied, with the estimated 24 h LC\u003csub\u003e50\u0026nbsp;\u003c/sub\u003evalues ranging from 24.04\u0026nbsp;ng/mL\u0026nbsp;to 49.82 ng/mL. The current study is the first attempt that examined the effect of fluralaner against \u003cem\u003eCx. quinquefasciatus\u003c/em\u003e.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eMosquitocidal effect of fluralaner observed in the current study was similar to that reported in an earlier study, which reported\u0026nbsp;24 h LC\u003csub\u003e50\u003c/sub\u003e values of\u0026nbsp;18.48-51.52 ng/mL (Miglianico et al., 2018). The 24 h LC\u003csub\u003e50\u0026nbsp;\u003c/sub\u003evalue of fluralaner observed in the earlier study as well as in the current study was\u0026nbsp;lower\u0026nbsp;than that reported for ivermectin (LC\u003csub\u003e50\u003c/sub\u003e:\u0026nbsp;44.68-49.40 ng/mL) (Sampaio et al., 2016)\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003e After a single oral administration of fluralaner at 50 mg/kg body weight in dogs, 100 ng/mL concentration of fluralaner was maintained in the blood for a period of 3 months\u0026nbsp;(Kilp et al., 2014).\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e Mosquito fed on dog treated with fluralaner reported to have significant reduction in their survival and fecundity for up to 13 weeks of post-treatment (Evans et al., 2022). It is noteworthy that the blood level concentration of fluralaner observed in dogs for three months is well above the 24 h LC\u003csub\u003e50\u0026nbsp;\u003c/sub\u003evalues (24.04-49.82 ng/mL) of fluralaner observed against three species in the current study.\u003c/p\u003e\n\u003cp\u003eFluralaner affected the fecundity, egg hatch success, immature development and adult emergence success in all the three species when they were exposed to drug concentrations nearing 24 h LC\u003csub\u003e50\u003c/sub\u003e values of 24.04 ng/mL for \u003cem\u003eAe. aegypti\u003c/em\u003e, 33.22 ng/mL for \u003cem\u003eAn. stephensi\u003c/em\u003e and 49.82 ng/mL for \u003cem\u003eCx. quinquefasciatus\u0026nbsp;\u003c/em\u003ewith significant P value. Although these differences were statistically significant but may not hold any importance under field conditions. However, there was no effect or only minimal effect at lower drug concentrations. This was in contrast to that reported for ivermectin which showed significant effect on fecundity and egg hatch success at sub lethal concentrations \u0026ndash; LC\u003csub\u003e5\u003c/sub\u003e value 18.28 ng/mL (Sampaio et al., 2016).\u0026nbsp;It is curious that fluralaner\u0026rsquo;s anti-fecundity activity is relatively lesser to that of ivermectin, even though their mortality effects are similar post blood feeding. One possible reason for this effect could be that ivermectin and isoxazoline compounds may have different mechanism of action regarding the anti-fecundity effect. Further research may be desirable in understanding this differential effect. Though fluralaner did not cause any visible impact on the development of the pupae into adults, certain morphological abnormalities like partially emerged adults and emerged adults with their legs attached to the pupal exuviae were observed. In an earlier study, fluralaner has been shown to affect the adult emergence success in \u003cem\u003eSpodoptera litura\u003c/em\u003e Fabricius (Lepidoptera: Noctuidae), even at sub-lethal dose of LC\u003csub\u003e15\u0026nbsp;\u003c/sub\u003emorphological abnormalities like notched wings were observed in the adults (Liu et al., 2018).\u003c/p\u003e\n\u003cp\u003eStudies on susceptible and resistant strains of horn and house flies have shown high level of mortality at lower concentration with fluralaner when compared with imidacloprid (Burgess et al., 2020). In a similar study on fipronil and pyrethroid resistant bed bugs, fluralaner outperformed ivermectin with a high mortality rate up to 28 days of post-treatment (Gonz\u0026aacute;lez-Morales\u0026nbsp;et al., 2022).\u003c/p\u003e\n\u003cp\u003eIn the current study complete mortality was observed within 7 h of post blood feeding at 250 ng/mL drug concentration in all the three species tested. Blood concentration much above this level is expected during the initial weeks in animals treated with fluralaner. This is a notable advantage over ivermectin which takes 72 h to produce 70.00-100.00% morality in mosquitoes at a dose of 93 ng/mL in animal experiments (Bastiaenset al., 2012,\u0026nbsp;Walther\u0026nbsp;al. 2015,\u0026nbsp;Mekuriawet al., 2019) and also this plasma level lasts for less than one day only.\u003c/p\u003e\n\u003cp\u003eCurrently, ivermectin mass therapy is being investigated in peri-domestic animals and humans in field studies for its endectocidal effect and as a complementary tool for malaria vector control (Pooda et al., 2015, Chaccouret al., 2013)\u003cstrong\u003e.\u0026nbsp;\u003c/strong\u003eFluralaner, with its comparable mosquitocidal effect to ivermectin coupled with the long half\u0026ndash;life may be a good alternative to ivermectin.\u003c/p\u003e\n\u003cp\u003eWith several emerging and remerging vector borne and zoonotic diseases, there is great interest in \u0026lsquo;One-Health\u0026rsquo; efforts being enforced round the globe. This would be particularly relevant in Africa and Asia where humans and animals live in close proximity and the drug based vector control strategy holds promise in such settings (Ruiz-Castillo et al., 2022). Moreover, it was found to be useful in insecticide resistance management plans for other active ingredients such as DDT, pyrethroid insecticides and other insecticides with possible reason of its distinct binding site from the targets of known modulators of ionotropic GABA receptors (Miglianico et al., 2018,\u0026nbsp;Wang et al., 2022). Though fluralaner appears to be a promising candidate for vector control there are some limitations that are to be resolved before proceeding further.\u0026nbsp;High cost of fluralaner is a barrier. Another issue is the safety aspects of fluralaner in human beings. It is not licensed for use in human beings. However, no adverse events were reported when fluralaner and afoxolaner were used for treating scabies in human clinical trials (Goldust\u0026nbsp;2018,\u0026nbsp;Goldust\u0026nbsp;and\u0026nbsp;Alipour 2019).\u0026nbsp;\u003c/p\u003e\n\u003cp\u003eThere are some limitations of the current study. We used chicken blood, not human blood for carrying out the efficacy trials for convenience. Another limitation is that we have carried out the study only in laboratory maintained colony and not on field caught mosquitoes. The survived mosquitoes assessed for life history characteristics were not comparable between treatment and control groups. The lab obtained results may not be exactly replicable in field conditions. \u0026nbsp;Another limitation is that we did not perform direct blood feeding experiments with mosquitoes fed upon animals treated with fluralaner. This experiment is necessary to understand the effect of the drug on mosquitoes in the context of its metabolism in the treated animals.\u003c/p\u003e\n\u003cp\u003eIn conclusion, our study showed significant oral toxic effect of fluralaner in adult vector mosquitoes with additional effect on life history characteristics (reduced fecundity, egg hatch success, larval development and adult emergence). Fluralaner may be a suitable candidate for future drug based mosquito control strategies. It is required to carry out future safety studies of fluralaner in human beings.\u0026nbsp;\u003c/p\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eConflicting Interest\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAuthors hereby declared that no conflict of interest is involved in this study.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eEthics approval\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eAll authors approved and gave their consent to publish the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAvailability of data and material\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets generated and/or analyzed during the current study are available from the corresponding author on request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgments\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe gratefully acknowledge the technical assistance of Mr. Shrihari Murmu, Senior Technician\u0026ndash;2 and Mr. K. Manimaran, Technical Officer-A, ICMR-Vector Control Research Centre for maintenance of mosquito colonies and blood feeding. We thank Mr. M. Sundharesan, Technical Assistant, ICMR-Vector Control Research Centre for his assistance in extraction of active compound from the commercially available tablet. We acknowledge the financial support by ICMR-Vector Control Research Centre, Puducherry.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding:\u0026nbsp;\u003c/strong\u003eWe\u0026nbsp;acknowledge the institutional funding provided to carry out the study.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eContributors\u0026rsquo; list:\u003c/strong\u003e VSK, SC and HKS contributed in study design. HKS, VS and SV executed the work, wrote the initial draft of the manuscript. NM helped for extraction of active compound. VB worked out statistical analysis for the data. VSK and AK critically reviewed the final version of manuscript. All authors read and approved the final version of the manuscript.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eBastiaens, G. J. H., G.-J. van Gemert, J. Hooghof, S. W. Lindsay, C. Drakeley, T. S. Churche, J. P. Verhave, C. H.M. Kocken, R. W. Sauerwein, and T. Bousema (2012) Duration of the mosquitocidal effect of ivermectin. Malariaworld J 3(10).\u003c/li\u003e\n\u003cli\u003eBurgess, E. R., C. J. Geden, K. H. Lohmeyer, B. H. King, E. T. Machtinger, and J. G. Scott (2020) Toxicity of fluralaner, a companion animal insecticide, relative to industry-leading agricultural insecticides against resistant and susceptible strains of filth flies. Sci Rep. 10:11166. https://doi.org/10.1038/s41598-020-68121-z\u003c/li\u003e\n\u003cli\u003eChaccour, C. J., K. C. Kobylinski, Q. Bassat, T. Bousema, C. Drakeley, P. Alonso, and B. D. Foy (2013) Ivermectin to reduce malaria transmission: a research agenda for a promising new tool for elimination. 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Bloomquist (2017) Mosquitocidal Activity and Mode of Action of the Isoxazoline Fluralaner. Int J Environ Res Public Health. 14(2):154. https://doi.org/10.3390/ijerph14020154\u003c/li\u003e\n\u003cli\u003eKilp, S., D. Ramirez, M. J. Allan, and R. K. Roepke (2016) Comparative pharmacokinetics of fluralaner in dogs and cats following single topical or intravenous administration. Parasit Vectors. 9(1):296. https://doi.org/10.1186/s13071-016-1564-8\u003c/li\u003e\n\u003cli\u003eKilp, S., D. Ramirez, M. J. Allan, R. K. A. Roepke, and M. C. Nuernberger (2014) Pharmacokinetics of fluralaner in dogs following a single oral or intravenous administration. Parasit Vectors. 7(85). https://doi.org/10.1186/1756-3305-7-85\u003c/li\u003e\n\u003cli\u003eLiu, D., Z.-Q. Jia, Y.-C. Peng, C.-W. Sheng, T. Tang, L. Xu, Z.-J. Han, and C.-Q. Zhao (2018) Toxicity and sublethal effects of fluralaner on Spodoptera litura Fabricius (Lepidoptera: Noctuidae). Pestic Biochem Physiol. 152:8\u0026ndash;16. https://doi.org/10.1016/j.pestbp.2018.08.004\u003c/li\u003e\n\u003cli\u003eMekuriaw, W., M. Balkew, L. A. Messenger, D. Yewhalaw, A. Woyessa, and F. Massebo (2019) The effect of ivermectin\u0026reg; on fertility, fecundity and mortality of Anopheles arabiensis fed on treated men in Ethiopia. Malar J. 18(1):357. https://doi.org/10.1186/s12936-019-2988-3\u003c/li\u003e\n\u003cli\u003eMiglianico, M., M. Eldering, H. Slater, N. Ferguson, P. Ambrose, R. S. Lees, K. M. J. Koolen, K. Pruzinova, M. Jancarova, P. Volf, C. J. M. Koenraadt, H.-P. Duerr, G. Trevitt, B. Yang, A. K. Chatterjee, J. Wisler, A. Sturm, T. Bousema, R. W. Sauerwein, P. G. Schultz, M. S. Tremblay, and K. J. Dechering (2018) Repurposing isoxazoline veterinary drugs for control of vector-borne human diseases. Proc Natl Acad Sci U S A. 115(29):E6920-E6926. https://doi.org/10.1073/pnas.1801338115\u003c/li\u003e\n\u003cli\u003eŌmura, S., and A. Crump (2017) Ivermectin and malaria control. Malar J. 16(1):172. https://doi.org/10.1186/s12936-017-1825-9\u003c/li\u003e\n\u003cli\u003ePooda, H. S., J.-B. Rayaisse, D. F. de S. Hien, T. Lef\u0026egrave;vre, S. R. Yerbanga, Z. Bengaly, R. K. Dabir\u0026eacute;, A. M. G. Belem, I. Sidib\u0026eacute;, P. Solano, and K. Mouline (2015) Administration of ivermectin to peridomestic cattle: a promising approach to target the residual transmission of human malaria. Malar J. 13 Suppl 1:496. https://doi.org/10.1186/s12936-015-1001-z\u003c/li\u003e\n\u003cli\u003eRuiz-Castillo, P., C. Rist, R. Rabinovich, and C. Chaccour (2022) Insecticide-treated livestock: a potential One Health approach to malaria control in Africa. Trends in Parasitol. 38(2):112-123. https://doi.org/10.1016/j.pt.2021.09.006\u003c/li\u003e\n\u003cli\u003eSampaio, V. S., T. P. Beltr\u0026aacute;n, K. C. Kobylinski, G. C. Melo, J. B. P. Lima, S. G. M. Silva, \u0026Iacute;. C. Rodriguez, H. Silveira, M. G. V. B. Guerra, Q. Bassat, P. F. P. Pimenta, M. V. G. Lacerda, and W. M. Monteiro (2016) Filling gaps on ivermectin knowledge: effects on the survival and reproduction of Anopheles aquasalis, a Latin American malaria vector. Malar J. 15(1):491. https://doi.org/10.1186/s12936-016-1540-y\u003c/li\u003e\n\u003cli\u003eSlater, H. C., B. D. Foy, K. Kobylinski, C. Chaccour, O. J. Watson, J. Hellewell, G. Aljayyoussi, T. Bousema, J. Burrows, U. D\u0026rsquo;Alessandro, H. Alout, F. O. Ter Kuile, P. G. T. Walker, A. C. Ghani, and M. R. Smit (2020) Ivermectin as a novel complementary malaria control tool to reduce incidence and prevalence: a modelling study. Lancet Infect Dis. 20(4):498-508. https://doi.org/10.1016/s1473-3099(19)30633-4\u003c/li\u003e\n\u003cli\u003eWalther, F. M., M. J. Allan, and R. K. A. Roepke (2015) Plasma pharmacokinetic profile of fluralaner (Bravecto\u003csup\u003eTM\u003c/sup\u003e) and ivermectin following concurrent administration to dogs. Parasit Vectors. 8:508. https://doi.org/10.1186/s13071-015-1123-8\u003c/li\u003e\n\u003cli\u003eWang, Q., H. Wang, Y. Zhang, J. Chen, A. Upadhyay, B. Bhowmick, J. Hang, S. Wu, C. Liao, and Q. Han (2022) Functional analysis reveals ionotropic GABA receptor subunit RDL is a target site of ivermectin and fluralaner in the yellow fever mosquito, \u003cem\u003eAedes aegypti\u003c/em\u003e. Pest Manag Sci. 78(10):4173-82. https://doi.org/10.1002/ps.7035\u003c/li\u003e\n\u003cli\u003eWorld Health Organization (2017) Global vector control response 2017\u0026ndash;2030. https://www.who.int/publications-detail-redirect/9789241512978. Accessed 22 December 2022\u003c/li\u003e\n\u003cli\u003eWorld Health Organization (2020) Vector-borne diseases. https://www.who.int/news-room/fact-sheets/detail/vector-borne-diseases. Accessed 22 December 2022\u003c/li\u003e\n\u003c/ol\u003e"},{"header":"Tables","content":"\u003cp\u003eTable\u0026nbsp;1. Lethal concentration values\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"100%\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.833333333333332%\"\u003e\n \u003cp\u003e\u003cstrong\u003eSpecies\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e\u003cstrong\u003ePost blood feeding time (h)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.416666666666666%\"\u003e\n \u003cp\u003e\u003cstrong\u003eSlope (\u0026plusmn;SE)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e\u003cstrong\u003eChi\u003csup\u003e2\u003c/sup\u003e goodness of fit (P- value)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e\u003cstrong\u003eLC\u003csub\u003e50\u003c/sub\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e(95% CI)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e\u003cstrong\u003eHeterogeneity factor\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.833333333333332%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAedes aegypti\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e24 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.416666666666666%\"\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003cp\u003e(0.004)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e3.13\u003c/p\u003e\n \u003cp\u003e(0.08)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e24.04\u003c/p\u003e\n \u003cp\u003e(14.74 \u0026ndash; 45.27)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e3.13\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.833333333333332%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e24 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.416666666666666%\"\u003e\n \u003cp\u003e0.12\u003c/p\u003e\n \u003cp\u003e(0.005)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e2.40\u003c/p\u003e\n \u003cp\u003e(0.12)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e33.22\u003c/p\u003e\n \u003cp\u003e(24.30 \u0026ndash; 51.56)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e2.40\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.833333333333332%\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e24 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.416666666666666%\"\u003e\n \u003cp\u003e0.17\u003c/p\u003e\n \u003cp\u003e(0.01)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.625%\"\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003cp\u003e(0.90)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e49.82\u003c/p\u003e\n \u003cp\u003e(49.35 \u0026ndash; 50.29)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.75%\"\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eProbit regression analysis: \u0026nbsp;SE \u0026ndash; Standard error; CI \u0026ndash; Confidence interval; LC \u0026ndash; Lethal Concentration\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 2. Effect of fluralaner on survivorship of\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eAedes aegypti\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"100%\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.161616161616163%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eDose\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.252525252525253%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMedian Survival duration\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"58.58585858585859%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eCox-regression analysis\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"68.42105263157895%\" valign=\"top\" style=\"width: 38.1035%;\"\u003e\n \u003cp\u003eHR (95% CI)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"31.57894736842105%\" valign=\"top\" style=\"width: 22.0215%;\"\u003e\n \u003cp\u003eP-value\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e0 (Control)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 38.1035%;\"\u003e\n \u003cp\u003e1.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 22.0215%;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e5 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 38.1035%;\"\u003e\n \u003cp\u003e3.45(2.07 \u0026ndash; 5.74)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 22.0215%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e10 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e96 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 38.1035%;\"\u003e\n \u003cp\u003e17.19(10.86 \u0026ndash; 27.22)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 22.0215%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e20 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e48 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 38.1035%;\"\u003e\n \u003cp\u003e40.61(25.77 \u0026ndash; 64.01)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 22.0215%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e35 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e24 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 38.1035%;\"\u003e\n \u003cp\u003e150.45(94.76 \u0026ndash; 238.87)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 22.0215%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eCox-regression analysis: HR \u0026ndash; hazard ratios;\u0026nbsp;CI \u0026ndash; Confidence interval\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 3. Effect of fluralaner on survivorship of\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"100%\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.161616161616163%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eDose\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.252525252525253%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMedian Survival duration\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"58.58585858585859%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eCox-regression analysis\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"68.42105263157895%\" valign=\"top\" style=\"width: 39.9434%;\"\u003e\n \u003cp\u003eHR (95% CI)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"31.57894736842105%\" valign=\"top\" style=\"width: 20.3066%;\"\u003e\n \u003cp\u003eP-value\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e0 (Control)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 39.9434%;\"\u003e\n \u003cp\u003e1.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 20.3066%;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e5 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 39.9434%;\"\u003e\n \u003cp\u003e3.48(2.02 \u0026ndash; 5.97)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 20.3066%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e15 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 39.9434%;\"\u003e\n \u003cp\u003e17.21(10.63 \u0026ndash; 27.88)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 20.3066%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e25 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e72 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 39.9434%;\"\u003e\n \u003cp\u003e39.72(24.73 \u0026ndash; 63.80)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 20.3066%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e40 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e24 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 39.9434%;\"\u003e\n \u003cp\u003e231.94(142.68 \u0026ndash; 377.05)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 20.3066%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eCox-regression analysis: HR \u0026ndash; hazard ratios;\u0026nbsp;CI \u0026ndash; Confidence interval\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003e\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 4. Effect of fluralaner on survivorship of\u0026nbsp;\u003c/strong\u003e\u003cstrong\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"100%\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.161616161616163%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eDose\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.252525252525253%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMedian Survival duration\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"58.58585858585859%\" colspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eCox-regression analysis\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"68.42105263157895%\" valign=\"top\" style=\"width: 41.5684%;\"\u003e\n \u003cp\u003eHR (95% CI)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"31.57894736842105%\" valign=\"top\" style=\"width: 18.6816%;\"\u003e\n \u003cp\u003eP-value\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e0 (Control)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 41.5684%;\"\u003e\n \u003cp\u003e1.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 18.6816%;\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e15 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 41.5684%;\"\u003e\n \u003cp\u003e3.72(1.99 \u0026ndash; 7.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 18.6816%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e30 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e-\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 41.5684%;\"\u003e\n \u003cp\u003e20.82(11.88 \u0026ndash; 36.49)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 18.6816%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e45 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e48 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 41.5684%;\"\u003e\n \u003cp\u003e91.53(52.45 \u0026ndash; 159.75)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 18.6816%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.3265306122449%\" valign=\"top\"\u003e\n \u003cp\u003e60 ng/mL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"25.510204081632654%\" valign=\"top\"\u003e\n \u003cp\u003e18 h\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.795918367346935%\" valign=\"top\" style=\"width: 41.5684%;\"\u003e\n \u003cp\u003e516.41(292.07 \u0026ndash; 913.07)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.367346938775512%\" valign=\"top\" style=\"width: 18.6816%;\"\u003e\n \u003cp\u003e\u0026lt;0.001\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003eCox-regression analysis: HR \u0026ndash; hazard ratios;\u0026nbsp;CI \u0026ndash; Confidence interval\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 5. Fecundity success\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMosquito species\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eFluralaner (ng/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"43.739837398373986%\" colspan=\"4\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eNumber of eggs laid per mosquito\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003e\u0026nbsp; Mean(SD)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMean difference\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.78048780487805%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"11.524163568773234%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"39.77695167286245%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTest\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.639405204460967%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"36.059479553903344%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAedes aegypti\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.040650406504065%\" valign=\"top\"\u003e\n \u003cp\u003e48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.398373983739837%\" valign=\"top\"\u003e\n \u003cp\u003e95.10(1.70)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.528455284552845%\" valign=\"top\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.772357723577235%\" valign=\"top\"\u003e\n \u003cp\u003e97.70(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\"\u003e\n \u003cp\u003e2.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.78048780487805%\"\u003e\n \u003cp\u003e0.018\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.352459016393443%\" valign=\"top\"\u003e\n \u003cp\u003e39\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"21.92622950819672%\" valign=\"top\"\u003e\n \u003cp\u003e91.90(1.30)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.967213114754099%\" valign=\"top\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.87704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e96.70(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e4.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.065573770491802%\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.352459016393443%\" valign=\"top\"\u003e\n \u003cp\u003e16\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"21.92622950819672%\" valign=\"top\"\u003e\n \u003cp\u003e78.70(1.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.967213114754099%\" valign=\"top\"\u003e\n \u003cp\u003e48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.87704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e96.70(1.10)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e18.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.065573770491802%\"\u003e\n \u003cp\u003e0.011\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.040650406504065%\" valign=\"top\"\u003e\n \u003cp\u003e48\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.398373983739837%\" valign=\"top\"\u003e\n \u003cp\u003e88.30(0.50)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.528455284552845%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.772357723577235%\" valign=\"top\"\u003e\n \u003cp\u003e90.30(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\"\u003e\n \u003cp\u003e2.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.78048780487805%\"\u003e\n \u003cp\u003e0.007\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.352459016393443%\" valign=\"top\"\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"21.92622950819672%\" valign=\"top\"\u003e\n \u003cp\u003e84.20(1.50)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.967213114754099%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.87704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e90.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e5.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.065573770491802%\"\u003e\n \u003cp\u003e0.011\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.352459016393443%\" valign=\"top\"\u003e\n \u003cp\u003e31\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"21.92622950819672%\" valign=\"top\"\u003e\n \u003cp\u003e72.00(2.10)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.967213114754099%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.87704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e89.30(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e17.30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.065573770491802%\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.040650406504065%\" valign=\"top\"\u003e\n \u003cp\u003e47\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.398373983739837%\" valign=\"top\"\u003e\n \u003cp\u003e124.20(2.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.528455284552845%\" valign=\"top\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.772357723577235%\" valign=\"top\"\u003e\n \u003cp\u003e136.00(3.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\"\u003e\n \u003cp\u003e11.80\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.78048780487805%\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.352459016393443%\" valign=\"top\"\u003e\n \u003cp\u003e40\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"21.92622950819672%\" valign=\"top\"\u003e\n \u003cp\u003e120.30(2.10)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.967213114754099%\" valign=\"top\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.87704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e134.30(2.50)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e14.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.065573770491802%\"\u003e\n \u003cp\u003e0.011\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.352459016393443%\" valign=\"top\"\u003e\n \u003cp\u003e7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"21.92622950819672%\" valign=\"top\"\u003e\n \u003cp\u003e117.30(3.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.967213114754099%\" valign=\"top\"\u003e\n \u003cp\u003e49\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.87704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e135.30(1.50)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e18.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.065573770491802%\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003en= number kept for fecundity success\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 6. Egg hatch success\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\" width=\"709\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"17.937853107344633%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMosquito species\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.581920903954803%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eFluralaner (ng/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"45.19774011299435%\" colspan=\"4\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eNumber of eggs hatched\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eMean(SD) \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.27683615819209%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMean difference\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.005649717514125%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"12.539184952978056%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"32.9153605015674%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTest\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.16614420062696%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"41.37931034482759%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"17.963224893917964%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAedes aegypti\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.598302687411598%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.657708628005658%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.851485148514852%\" valign=\"top\"\u003e\n \u003cp\u003e403.67(4.74)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.9405940594059405%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.67043847241867%\" valign=\"top\"\u003e\n \u003cp\u003e414.00(6.24)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.295615275813295%\"\u003e\n \u003cp\u003e10.33\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.022630834512023%\"\u003e\n \u003cp\u003e0.026\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"14.137931034482758%\" valign=\"top\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.896551724137931%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.103448275862068%\" valign=\"top\"\u003e\n \u003cp\u003e395.67(5.29)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.241379310344827%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.75862068965517%\" valign=\"top\"\u003e\n \u003cp\u003e419.67(4.04)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.20689655172414%\"\u003e\n \u003cp\u003e24.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.655172413793103%\"\u003e\n \u003cp\u003e0.013\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"14.137931034482758%\" valign=\"top\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.896551724137931%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.103448275862068%\" valign=\"top\"\u003e\n \u003cp\u003e386.00(5.17)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.241379310344827%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.75862068965517%\" valign=\"top\"\u003e\n \u003cp\u003e416.00(3.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.20689655172414%\"\u003e\n \u003cp\u003e30.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.655172413793103%\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"17.963224893917964%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.598302687411598%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.657708628005658%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.851485148514852%\" valign=\"top\"\u003e\n \u003cp\u003e435.22(6.96)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.9405940594059405%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.67043847241867%\" valign=\"top\"\u003e\n \u003cp\u003e456.33(4.51)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.295615275813295%\"\u003e\n \u003cp\u003e21.11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.022630834512023%\"\u003e\n \u003cp\u003e0.013\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"14.137931034482758%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.896551724137931%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.103448275862068%\" valign=\"top\"\u003e\n \u003cp\u003e420.22(7.05)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.241379310344827%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.75862068965517%\" valign=\"top\"\u003e\n \u003cp\u003e452.67(3.05)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.20689655172414%\"\u003e\n \u003cp\u003e32.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.655172413793103%\"\u003e\n \u003cp\u003e0.013\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"14.137931034482758%\" valign=\"top\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.896551724137931%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.103448275862068%\" valign=\"top\"\u003e\n \u003cp\u003e408.89(4.94)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.241379310344827%\" valign=\"top\"\u003e\n \u003cp\u003e500\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.75862068965517%\" valign=\"top\"\u003e\n \u003cp\u003e453.33(12.86)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.20689655172414%\"\u003e\n \u003cp\u003e44.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.655172413793103%\"\u003e\n \u003cp\u003e0.013\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"17.963224893917964%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"11.598302687411598%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.657708628005658%\" valign=\"top\"\u003e\n \u003cp\u003e517\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.851485148514852%\" valign=\"top\"\u003e\n \u003cp\u003e426.22(9.97)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.9405940594059405%\" valign=\"top\"\u003e\n \u003cp\u003e533\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.67043847241867%\" valign=\"top\"\u003e\n \u003cp\u003e462.66(8.14)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.295615275813295%\"\u003e\n \u003cp\u003e36.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"12.022630834512023%\"\u003e\n \u003cp\u003e0.013\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"14.137931034482758%\" valign=\"top\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.896551724137931%\" valign=\"top\"\u003e\n \u003cp\u003e524\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.103448275862068%\" valign=\"top\"\u003e\n \u003cp\u003e425.11(11.88)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.241379310344827%\" valign=\"top\"\u003e\n \u003cp\u003e515\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.75862068965517%\" valign=\"top\"\u003e\n \u003cp\u003e446.00(18.68)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.20689655172414%\"\u003e\n \u003cp\u003e20.89\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.655172413793103%\"\u003e\n \u003cp\u003e0.060\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"14.137931034482758%\" valign=\"top\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.896551724137931%\" valign=\"top\"\u003e\n \u003cp\u003e518\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.103448275862068%\" valign=\"top\"\u003e\n \u003cp\u003e417.22(4.82)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.241379310344827%\" valign=\"top\"\u003e\n \u003cp\u003e531\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"22.75862068965517%\" valign=\"top\"\u003e\n \u003cp\u003e457.33(24.68)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.20689655172414%\"\u003e\n \u003cp\u003e40.11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.655172413793103%\"\u003e\n \u003cp\u003e0.012\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003en= number kept for egg hatch success\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 7. Immature survival\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.616883116883116%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMosquito species\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.311688311688311%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eFluralaner (ng/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"44.15584415584416%\" colspan=\"4\"\u003e\n \u003cp\u003e\u003cstrong\u003eNumber of pupae developed\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eMean(SD)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.474025974025974%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMean difference\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.441558441558442%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"17.647058823529413%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.455882352941174%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTest\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.235294117647058%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"35.661764705882355%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.616883116883116%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAedes aegypti\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.311688311688311%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.792207792207792%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.772727272727273%\" valign=\"top\"\u003e\n \u003cp\u003e98.80(0.70)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.8441558441558445%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.746753246753247%\" valign=\"top\"\u003e\n \u003cp\u003e99.70(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.474025974025974%\"\u003e\n \u003cp\u003e0.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.441558441558442%\"\u003e\n \u003cp\u003e0.07\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.768916155419223%\" valign=\"top\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.815950920245399%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.609406952965234%\" valign=\"top\"\u003e\n \u003cp\u003e97.40(1.10)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.361963190184049%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.83640081799591%\" valign=\"top\"\u003e\n \u003cp\u003e99.00(1.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.973415132924334%\"\u003e\n \u003cp\u003e1.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.633946830265849%\"\u003e\n \u003cp\u003e0.07\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.768916155419223%\" valign=\"top\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.815950920245399%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.609406952965234%\" valign=\"top\"\u003e\n \u003cp\u003e91.90(2.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.361963190184049%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.83640081799591%\" valign=\"top\"\u003e\n \u003cp\u003e99.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.973415132924334%\"\u003e\n \u003cp\u003e7.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.633946830265849%\"\u003e\n \u003cp\u003e0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.616883116883116%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.311688311688311%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.792207792207792%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.772727272727273%\" valign=\"top\"\u003e\n \u003cp\u003e99.10(0.80)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.8441558441558445%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.746753246753247%\" valign=\"top\"\u003e\n \u003cp\u003e100.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.474025974025974%\" valign=\"top\"\u003e\n \u003cp\u003e0.90\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.441558441558442%\"\u003e\n \u003cp\u003e0.07\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.768916155419223%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.815950920245399%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.609406952965234%\" valign=\"top\"\u003e\n \u003cp\u003e97.80(1.40)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.361963190184049%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.83640081799591%\" valign=\"top\"\u003e\n \u003cp\u003e99.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.973415132924334%\" valign=\"top\"\u003e\n \u003cp\u003e1.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.633946830265849%\"\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.768916155419223%\" valign=\"top\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.815950920245399%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.609406952965234%\" valign=\"top\"\u003e\n \u003cp\u003e96.40(1.30)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.361963190184049%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.83640081799591%\" valign=\"top\"\u003e\n \u003cp\u003e98.70(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.973415132924334%\"\u003e\n \u003cp\u003e2.20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.633946830265849%\"\u003e\n \u003cp\u003e0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.616883116883116%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.311688311688311%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.792207792207792%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.772727272727273%\" valign=\"top\"\u003e\n \u003cp\u003e98.20(0.70)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"5.8441558441558445%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.746753246753247%\" valign=\"top\"\u003e\n \u003cp\u003e99.30(1.10)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.474025974025974%\"\u003e\n \u003cp\u003e1.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.441558441558442%\"\u003e\n \u003cp\u003e0.11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.768916155419223%\" valign=\"top\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.815950920245399%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.609406952965234%\" valign=\"top\"\u003e\n \u003cp\u003e93.80(1.90)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.361963190184049%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.83640081799591%\" valign=\"top\"\u003e\n \u003cp\u003e99.30(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.973415132924334%\"\u003e\n \u003cp\u003e5.60\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.633946830265849%\"\u003e\n \u003cp\u003e0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.768916155419223%\" valign=\"top\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.815950920245399%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.609406952965234%\" valign=\"top\"\u003e\n \u003cp\u003e89.20(3.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.361963190184049%\" valign=\"top\"\u003e\n \u003cp\u003e100\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"19.83640081799591%\" valign=\"top\"\u003e\n \u003cp\u003e99.30(0.60)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"16.973415132924334%\"\u003e\n \u003cp\u003e10.10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.633946830265849%\"\u003e\n \u003cp\u003e0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003en= number kept for immature survival\u003c/p\u003e\n\u003cp\u003e\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eTable 8. Adult emergence success\u003c/strong\u003e\u003c/p\u003e\n\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMosquito species\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eFluralaner (ng/mL)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"43.90243902439025%\" colspan=\"4\"\u003e\n \u003cp\u003e\u003cstrong\u003eNumber of adults emerged\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eMean(SD) \u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eMean difference\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.617886178861788%\" rowspan=\"2\"\u003e\n \u003cp\u003e\u003cstrong\u003eP-value\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"17.77777777777778%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"33.7037037037037%\"\u003e\n \u003cp\u003e\u003cstrong\u003eTest\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"15.555555555555555%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003en\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"32.96296296296296%\" valign=\"top\"\u003e\n \u003cp\u003e\u003cstrong\u003eControl\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAedes aegypti\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.804878048780488%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.796747967479675%\" valign=\"top\"\u003e\n \u003cp\u003e49.56(0.53)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.829268292682927%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.471544715447154%\" valign=\"top\"\u003e\n \u003cp\u003e50.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\"\u003e\n \u003cp\u003e0.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.617886178861788%\"\u003e\n \u003cp\u003e0.17\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.836065573770492%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.647540983606557%\" valign=\"top\"\u003e\n \u003cp\u003e49.22(0.67)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.60655737704918%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.237704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e49.67(0.58)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e0.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.860655737704919%\"\u003e\n \u003cp\u003e0.30\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e20\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.836065573770492%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.647540983606557%\" valign=\"top\"\u003e\n \u003cp\u003e48.44(0.88)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.60655737704918%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.237704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e50.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e1.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.860655737704919%\"\u003e\n \u003cp\u003e0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eAnopheles stephensi\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" valign=\"top\"\u003e\n \u003cp\u003e5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.804878048780488%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.796747967479675%\" valign=\"top\"\u003e\n \u003cp\u003e49.44(0.53)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.829268292682927%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.471544715447154%\" valign=\"top\"\u003e\n \u003cp\u003e50.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\" valign=\"top\"\u003e\n \u003cp\u003e0.56\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.617886178861788%\"\u003e\n \u003cp\u003e0.10\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.836065573770492%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.647540983606557%\" valign=\"top\"\u003e\n \u003cp\u003e48.00(0.71)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.60655737704918%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.237704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e49.67(0.58)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\" valign=\"top\"\u003e\n \u003cp\u003e1.67\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.860655737704919%\"\u003e\n \u003cp\u003e0.01\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e25\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.836065573770492%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.647540983606557%\" valign=\"top\"\u003e\n \u003cp\u003e46.78(1.30)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.60655737704918%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.237704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e48.67(0.58)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e1.89\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.860655737704919%\"\u003e\n \u003cp\u003e0.04\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"20.650406504065042%\" rowspan=\"3\"\u003e\n \u003cp\u003e\u003cstrong\u003e\u003cem\u003eCulex quinquefasciatus\u003c/em\u003e\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.333333333333334%\" valign=\"top\"\u003e\n \u003cp\u003e15\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"7.804878048780488%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.796747967479675%\" valign=\"top\"\u003e\n \u003cp\u003e49.56(0.53)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"6.829268292682927%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"14.471544715447154%\" valign=\"top\"\u003e\n \u003cp\u003e50.00(0.00)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"13.495934959349594%\"\u003e\n \u003cp\u003e0.44\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.617886178861788%\"\u003e\n \u003cp\u003e0.17\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e30\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.836065573770492%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.647540983606557%\" valign=\"top\"\u003e\n \u003cp\u003e48.78(0.44)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.60655737704918%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.237704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e49.67(0.58)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e0.89\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.860655737704919%\"\u003e\n \u003cp\u003e0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd width=\"16.80327868852459%\" valign=\"top\"\u003e\n \u003cp\u003e45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"9.836065573770492%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.647540983606557%\" valign=\"top\"\u003e\n \u003cp\u003e47.67(1.32)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"8.60655737704918%\" valign=\"top\"\u003e\n \u003cp\u003e50\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"18.237704918032787%\" valign=\"top\"\u003e\n \u003cp\u003e49.67(0.58)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"17.008196721311474%\"\u003e\n \u003cp\u003e2.00\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd width=\"10.860655737704919%\"\u003e\n \u003cp\u003e0.02\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e\n\u003cp\u003en= number kept for adult emergence success\u003c/p\u003e"},{"header":"Graphs","content":"\u003cp\u003eGraphs 1 to 4 are available in the Supplementary Files section.\u003c/p\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":true,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"fluralaner, endectocide, drug based vector control, mosquitocidal effect, life history characteristics","lastPublishedDoi":"10.21203/rs.3.rs-2803950/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-2803950/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eVector control is a key intervention against mosquito borne diseases such as dengue, chikungunya and malaria. However, conventional methods have several limitations and alternate strategies are in urgent need. Vector control with endectocides or systemic insecticides such as ivermectin is emerging as a novel strategy. The short half-life of ivermectin is a limiting factor for its application as a mass therapy tool for vector control. Isoxazoline compounds like fluralaner, a class of veterinary acaricides with long half-life hold promise as alternatives to ivermectin. These drugs persist in the animal body at detectable blood levels up to three months or more, after a one-time oral administration. However, information about their mosquitocidal effect is very much limited. In the current study, we explored the efficacy of fluralaner against laboratory reared vector mosquitoes of \u003cem\u003eAedes aegypti\u003c/em\u003e (Linnaeus, 1762), \u003cem\u003eAnopheles stephensi\u003c/em\u003e (Liston, 1901) \u003cem\u003eand Culex quinquefasciatus\u003c/em\u003e (Say, 1823) (Diptera: Culicidae) by blood feeding. Fluralaner showed significant mosquitocidal effect with the estimated 24 h LC\u003csub\u003e50\u003c/sub\u003e values in the range of 24.04\u0026ndash;49.82 ng/mL for the three different mosquito species tested. Effects on life history characteristics (fecundity, egg hatch success, immature development and adult emergence success) were also observed at drug concentrations nearing 24 h LC\u003csub\u003e50\u003c/sub\u003e. At higher drug concentration of 250 ng/mL, significant knock down effect was observed within 1\u0026ndash;2 h post blood feeding. Potent mosquitocidal effect coupled with its long half-life makes fluralaner an excellent candidate for drug based vector control strategies especially in the context of \u0026ldquo;One Health\u0026rdquo; approach. In this regard, further studies on the safety aspects of fluralaner in humans are desirable.\u003c/p\u003e","manuscriptTitle":"Efficacy of fluralaner, a long acting acaricide against three species of vector mosquitoes (Diptera: Culicidae) under laboratory conditions – a potential candidate for drug based vector control","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2023-05-24 15:31:10","doi":"10.21203/rs.3.rs-2803950/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"3a18156d-f457-434c-a9cf-919a67a0c80e","owner":[],"postedDate":"May 24th, 2023","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[],"tags":[],"updatedAt":"2023-06-21T20:52:45+00:00","versionOfRecord":[],"versionCreatedAt":"2023-05-24 15:31:10","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-2803950","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-2803950","identity":"rs-2803950","version":["v1"]},"buildId":"cBFmMYwuxLRRLfASyISRj","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}