Mosquito Mortality Research Articles

Modified World Health Organization (WHO) Tunnel Test for Higher Throughput Evaluation of Insecticide-Treated Nets (ITNs) Considering the Effect of Alternative Hosts, Exposure Time, and Mosquito Density.

Simple SummaryMembrane feeding assays have been widely used in malaria transmission research and insectary colony maintenance. Here, we investigate whether a membrane feeder can replace animal baits for evaluating insecticide-treated nets (ITNs) bio-efficacy in the World Health Organization (WHO) tunnel test. The effect of (1) alternative baits, (2) exposure time, and (3) mosquito density on the endpoints of mosquito mortality and feeding inhibition or feeding success was investigated. Our results show that similar mortality at 24-h (M24) or 72-h (M72) is estimated using either a membrane feeder or a rabbit bait with an overnight (12 h) exposure. However, the membrane measured higher blood feeding inhibition than the rabbit, likely due to the absence of host cues, notably carbon dioxide. Therefore, the membrane feeder may be used instead of an animal bait to test mortality endpoints in WHO tunnel tests and blood feeding rates need to be improved. Experimental results demonstrated that using 50 or 100 mosquitoes per replicate measure the same for mortality and feeding inhibition endpoints with an animal bait. Therefore, WHO tunnel tests may be run with lower mosquito densities. This will reduce strain on insectaries to produce sufficient mosquitoes to meet the large sample sizes needed for bio-efficacy durability monitoring of chlorfenapyr ITNs that must be evaluated in “free-flying” bioassays.The standard World Health Organization (WHO) tunnel test is a reliable laboratory bioassay used for “free-flying” testing of insecticide-treated nets (ITNs) bio-efficacy where mosquitoes pass through a ITN sample to reach a live animal bait. Multiple parameters (i.e., bait, exposure time, and mosquito density) may affect the outcomes measured in tunnel tests. Therefore, a comparison was conducted of alternative hosts, exposure time, and lower mosquito density against the current gold standard test (100 mosquitoes, animal bait, and 12-h exposure) as outlined in the WHO ITN evaluation guideline. This was done with the aim to make the tunnel test cheaper and with higher throughput to meet the large sample sizes needed for bio-efficacy durability monitoring of chlorfenapyr ITNs that must be evaluated in “free-flying” bioassays. Methods: A series of experiments were conducted in the WHO tunnel test to evaluate the impact of the following factors on bio-efficacy endpoints of mosquito mortality at 24-h (M24) and 72-h (M72) and blood-feeding success (BFS): (1) baits (rabbit, membrane, human arm); (2) exposure time in the tunnel (1 h vs. 12 h); and (3) mosquito density (50 vs. 100). Finally, an alternative bioassay using a membrane with 50 mosquitoes (membrane-50) was compared to the gold standard bioassay (rabbit with 100 mosquitoes, rabbit-100). Pyrethroid-resistant Anopheles arabiensis and pyrethroid susceptible Anopheles gambiae were used to evaluate Interceptor® and Interceptor® G2 ITNs. Results: Using a human arm as bait gave a very different BFS, which impacted measurements of M24 and M72. The same trends in M24, M72 and BFS were observed for both Interceptor® ITN and Interceptor® G2 unwashed and washed 20 times measured using the gold standard WHO tunnel test (rabbit-100) or rabbit with 50 mosquitoes (rabbit-50). M24, M72 and BFS were not statistically different when either 50 or 100 mosquitoes were used with rabbit bait in the tunnel bioassay for either the susceptible or resistant strains. No systematic difference was observed between rabbit-50 and rabbit-100 in the agreement by the Bland and Altman method (B&A). The mean difference was 4.54% (−22.54–31.62) in BFS and 1.71% (−28.71–32.12) in M72 for rabbit-50 versus rabbit-100. Similar M24, M72 and lower BFS was measured by membrane-50 compared to rabbit-100. No systematic difference was observed in the agreement between membrane-50 and rabbit-100, by B&A. The mean difference was 9.06% (−11.42–29.64) for BSF and −5.44% (−50.3–39.45) for M72. Both membrane-50, rabbit-50 and rabbit-100 predicted the superiority of Interceptor® G2 over Interceptor® ITN for the resistant strain on M72. Conclusion: These results demonstrate that WHO tunnel tests using rabbit bait may be run with 50 mosquitoes to increase sample sizes needed for bio-efficacy durability monitoring of ITNs in “free-flying” bioassays. Using a membrane feeder with 50 mosquitoes is a potential replacement for the WHO tunnel bioassay with animal bait if control blood feeding rates can be improved to 50% because blood feeding impacts mosquito survival after exposure to insecticides.

Diet and Oviposition Deprivation Effects on Survivorship, Gonotrophic Dissociation, and Mortality of Anopheles gambiae s.s.

Diet quality is of paramount importance for egg batch size, longevity, and mortality of vector mosquitoes. Oviposition site presence and absence assumed to be dry season means a lot to the survivorship and mortality of most anthropophilic malaria vectors in sub-Saharan Africa. This study has assessed the effect of different diets and oviposition-site deprivation (OSD) on survivorship, longevity, and mortality of An. gambiae s.s. To determine the effect of diet and OSD on mortality, gonotrophic dissociation rates, longevity, and survivorship, six treatments were employed: Blood Fed with Oviposition (BFO), Blood Fed without oviposition (BF), Blood and Sugar Fed with Oviposition (BSFO), Blood and Sugar Fed without oviposition (BSF), Sugar Fed with Oviposition (SFO), and Sugar Fed without oviposition (SF). Mortality and gonotrophic dissociation were monitored daily. This study found that female mosquitoes offered blood meals with sugar solution and oviposition deprivation survived longer than those deprived of oviposition deprivation. Similarly, female mosquitoes fed on blood and provided with oviposition deprivation lived longer than those without oviposition deprivation. The gonotrophic dissociation rates were found to be lower in groups provided with oviposition deprivation. Our results show that OSD has a direct impact on the survivorship, gonotrophic dissociation rate, and longevity of the malaria anthropophilic vector, An. gambiae s.s., regardless of the diet.

Mathematics of a single-locus model for assessing the impacts of pyrethroid resistance and temperature on population abundance of malaria mosquitoes

This study presents a genetic-ecology modeling framework for assessing the combined impacts of insecticide resistance, temperature variability, and insecticide-based interventions on the population abundance and control of malaria mosquitoes by genotype. Rigorous analyses of the model we developed reveal that the boundary equilibrium with only mosquitoes of homozygous sensitive (resistant) genotype is locally-asymptotically stable whenever a certain ecological threshold, denoted by , is less than one. Furthermore, genotype i drives genotype j to extinction whenever and (where i, j = SS or RR, with i ≠ j). The model exhibits the phenomenon of bistability when both thresholds are less than one. In such a bistable situation, convergence to any of the two boundary equilibria depends on the initial allele distribution in the state variables of the model. Furthermore, in this bistable case, where , the basin of attraction of the boundary equilibrium of the mosquito genotype with lower value of the ecological threshold is larger. Specifically, the basin of attraction of the boundary equilibrium for genotype i is larger than that of genotype j if . When both ecological thresholds exceed one , the two boundary equilibria lose their stability, and a coexistence equilibrium (where all three mosquito genotypes coexist) becomes locally-asymptotically stable. Global sensitivity analysis shows that the key parameters that greatly influence the dynamics and population abundance of resistant mosquitoes include the proportion of new adult mosquitoes that are females, the insecticide-induced mortality rate of adult female mosquitoes, the coverage level and efficacy of adulticides used in the community, the oviposition rates for eggs of heterozygous and homozygous resistant genotypes, and the modification parameter accounting for the reduction in insecticide-induced mortality due to resistance. Numerical simulations show that the adult mosquito population increases with increasing temperature until a peak is reached at 31 °C, and declines thereafter. Simulating the model for moderate and high adulticide coverage, together with varying fitness costs of resistance, shows a switch in the dominant genotype at equilibrium as temperature is varied. In other words, this study shows that, for certain combinations of adulticide coverage and fitness costs of insecticide resistance, increases in temperature could result in effective management of resistance (by causing the switch from a stable resistant-only boundary equilibrium (at 18 °C) to a stable sensitive-only boundary equilibrium (at 25 °C)). Finally, this study shows that, for moderate fitness costs of resistance, density-dependent larval mortality suppresses the total population of adult mosquitoes with the resistant allele for all temperature values in the range [18 °C–36 °C].

Experimental evaluation of a metofluthrin passive emanator against Aedes albopictus.

With the recent occurrence of locally transmitted Aedes-borne viruses in the continental United States and Europe, and a lack of effective vaccines, new approaches to control Aedes aegypti and Aedes albopictus are needed. In sub-tropical urban settings in the US, Ae. albopictus is a dominant nuisance and arbovirus vector species. Unfortunately, the vector control toolbox against Ae. albopictus is not as well developed as for Ae. aegypti. Here, we evaluate the efficacy, longevity, and range of protectiveness of a novel passive metofluthrin emanator (10% active ingredient in a polyethylene mesh) against Ae. albopictus indoors and outdoors. Four studies were conducted comparing the presence of the metofluthrin emanator to a control lacking emanator with interest in quantifying efficacy by human landing counts. Studies evaluated the effect of an emanator at varying distances from one or more human volunteers indoors and outdoors. Efficacy of emanators over time since activation was also evaluated. Mixed-effects models determined that sitting in close proximity to an emanator reduced landings by 89.5% outdoors and by 74.6% indoors. The emanator was determined protective when located immediately next to a human volunteer outdoors but not uniformly protective when located further away. The emanator was protective at all tested distances from the device indoors. Mortality of mosquitoes exposed to metofluthrin emanators was ~2x higher than those who were not exposed in indoor conditions. Finally, a Generalized Additive Model determined that emanators used continuously outdoors lost their effect after 2.5 weeks and stopped inducing paralysis in mosquitoes after 3.8 weeks of use. We show strong and lasting efficacy of 10% metofluthrin emanators against field Ae. albopictus both in indoor and outdoor conditions. Metofluthrin emanators can protect people from Ae. albopictus bites, representing a viable option for reducing human-mosquito contacts at home and beyond.

Effects of ivermectin treatment of backyard chickens on mosquito dynamics and West Nile virus transmission.

BackgroundVector control strategies typically rely on pesticides to target mosquitoes involved in enzootic and zoonotic transmission of West Nile virus (WNV). Nevertheless, increasing insecticide resistance and a desire to reduce pesticide usage provide the impetus for developing alternative strategies. Ivermectin (IVM), an antiparasitic drug which is widely used in human and veterinary medicine, is a potential alternative for targeted control because Culex mosquitoes experience increased mortality following ingestion of IVM in bloodmeals.Methodology/Principal findingsWe conducted a randomized field trial to investigate the impact of treating backyard chicken flocks with IVM in urban neighborhoods across Davis, California on mosquito populations and WNV transmission dynamics. We observed a significant reduction in WNV seroconversions in treated vs. untreated chickens, suggesting a reduction in WNV transmission intensity around treated flocks. We also detected a reduction in parity rates of Cx. tarsalis near treated vs. untreated flocks and increased mortality in wild mosquitoes following a bloodmeal on treated chickens (IVM serum concentration > 5ng/mL) vs. chickens with IVM serum concentrations < 5 ng/mL. However, we did not find a significant difference in abundance or infection prevalence in mosquitoes between treatment groups associated with the reductions in seroconversions. Mosquito immigration from surrounding larval habitat, relatively low WNV activity in the study area, and variable IVM serum concentrations likely contributed to uncertainty about the impact.Conclusions/SignificanceTaken together, our results point to a reduction in WNV transmission due to the impact of IVM on Culex mosquito populations and support the ongoing investigation of oral administration of IVM to wild birds for local control of WNV transmission, although further work is needed to optimize dosing and understand effects on entomological endpoints.

Development of Plant-Based Larvicide and Herbal Mosquito Repellent Fast Card with Reference to Identification of the Functional Bioactive Compounds Effective Against Culex Mosquito.

Warm and humid climate creates ideal conditions for mosquito breeding. The ability of these vectors to spread a number of diseases to humans causes millions of deaths every year. Indiscriminate use of synthetic insecticides leads to the development of resistance in vector mosquitoes and along with this, these pesticides cause biological magnification of toxic components and affects adversely the non-target organisms including human being. Commercially available, chemically manufactured mosquito repellent fast cards are convenient to use and quite effective but burning of such card generates a lot of smoke and might be hazardous to human health in the long run. Thus, alternative approaches are to be adopted to control the population load of vector mosquito. Like that, the present study also reveals the larvicidal effect of Duranta leaf extract against Culex mosquito. In the present study, mosquito repellent fast card has been developed by Duranta-algal mixture which has shown better result than commercially available fast card on the basis of mosquito mortality as well as the amount of gases emitted. Again, the ethanolic crude extract of Duranta leaves leads to 100% mortality of all instars (Culex pipiens) larvae at both 1000ppm and 500ppm concentration. Therefore, the active component of Duranta has also been investigated. In Duranta, highest area percentage and peak have been shown by propionic acid in the retention time 18.086 by GC-MS. So, it can be confirmed that the major active ingredient is propionic acid in Duranta which is responsible for the mosquitocidal properties. Occurrence of propionic acid in Duranta has also been confirmed by the HPLC analysis.

Comparative efficacy of two pyrethroid-piperonyl butoxide nets (Olyset Plus and PermaNet 3.0) against pyrethroid resistant malaria vectors: a non-inferiority assessment

BackgroundPyrethroid-PBO nets were conditionally recommended for control of malaria transmitted by mosquitoes with oxidase-based pyrethroid-resistance based on epidemiological evidence of additional protective effect with Olyset Plus compared to a pyrethroid-only net (Olyset Net). Entomological studies can be used to assess the comparative performance of other brands of pyrethroid-PBO ITNs to Olyset Plus.MethodsAn experimental hut trial was performed in Cové, Benin to compare PermaNet 3.0 (deltamethrin plus PBO on roof panel only) to Olyset Plus (permethrin plus PBO on all panels) against wild pyrethroid-resistant Anopheles gambiae sensu lato (s.l.) following World Health Organization (WHO) guidelines. Both nets were tested unwashed and after 20 standardized washes compared to Olyset Net. Laboratory bioassays were also performed to help explain findings in the experimental huts.ResultsWith unwashed nets, mosquito mortality was higher in huts with PermaNet 3.0 compared to Olyset Plus (41% vs. 28%, P < 0.001). After 20 washes, mortality declined significantly with PermaNet 3.0 (41% unwashed vs. 17% after washing P < 0.001), but not with Olyset Plus (28% unwashed vs. 24% after washing P = 0.433); Olyset Plus induced significantly higher mortality than PermaNet 3.0 and Olyset Net after 20 washes. PermaNet 3.0 showed a higher wash retention of PBO compared to Olyset Plus. A non-inferiority analysis performed with data from unwashed and washed nets together using a margin recommended by the WHO, showed that PermaNet 3.0 was non-inferior to Olyset Plus in terms of mosquito mortality (25% with Olyset Plus vs. 27% with PermaNet 3.0, OR = 1.528, 95%CI = 1.02–2.29) but not in reducing mosquito feeding (25% with Olyset Plus vs. 30% with PermaNet 3.0, OR = 1.192, 95%CI = 0.77–1.84). Both pyrethroid-PBO nets were superior to Olyset Net.ConclusionOlyset Plus outperformed PermaNet 3.0 in terms of its ability to cause greater margins of improved mosquito mortality compared to a standard pyrethroid net, after multiple standardized washes. However, using a margin of non-inferiority defined by the WHO, PermaNet 3.0 was non-inferior to Olyset Plus in inducing mosquito mortality. Considering the low levels of mortality observed and increasing pyrethroid-resistance in West Africa, it is unclear whether either of these nets would demonstrate the same epidemiological impact observed in community trials in East Africa.

High-content phenotypic screening identifies novel chemistries that disrupt mosquito activity and development

New classes of chemistries are needed to control insecticide resistant populations of mosquitoes and prevent transmission of vector-borne diseases (VBDs). Organismal screens of chemical collections have played an important role in the search for new vector insecticides and the identification of active ingredients (AIs) that cause rapid mortality of mosquitoes. Advances in image-based screening offer an opportunity to identify chemistries that operate via novel biochemical modes and investigate the range of phenotypes exhibited by mosquitoes following exposure to lethal and sub-lethal chemical dose. An automated, high throughput phenotypic screen (HTS) employing high-content imaging of first instar (L1) Aedes aegypti larvae was developed to identify chemistries associated with mortality and atypical morphological phenotypes. A pilot screen of the Library of Pharmacologically Active Compounds (LOPAC1280) identified 92 chemistries that disrupted larval activity and development, including conventional insecticides and chemistries known to modulate G protein-coupled receptors (GPCRs) and other molecular targets in mammalian systems. Secondary assay series were used to evaluate a selection of chemistries for impacts on mosquito activity, survival and development. Ritodrine hydrochloride reduced mobility of larvae but had no observable effect on survival and development of mosquitoes. High doses of metergoline suppressed larval activity and sub-lethal dose resulted in pupal mortality. Assay data support the utility of phenotypic screening and diverse entomological end-points for discovery of novel insecticidal chemical scaffolds. The insecticide discovery process must consider how multi-modal efficacy spectra contribute to vector and VBD control.

Deltamethrin interacts with Culex quinquefasciatus odorant-binding protein: a novel potential resistance mechanism

BackgroundOdorant-binding proteins (OBPs) play important roles in many physiological processes of mosquitoes. Previous high-throughput sequencing studies have revealed that some OBPs of Culex quinquefasciatus might be involved in the development of resistance to insecticides.MethodsBased on the results of sequencing analyses, the OBP28 gene was selected for evaluation in this study. Three laboratory strains of Cx. quinquefasciatus [susceptible strain (SS), deltamethrin-resistant strain 1 (HN) and deltamethrin-resistant strain 2 (RR)] were first examined by using the Centers for Disease Control and Prevention bottle bioassay, after which the expression level of the OBP28 gene in the susceptible and deltamethrin-resistant strains was determined by real-time quantitative polymerase chain reaction. The OBP28 gene in deltamethrin-resistant strain RR was silenced using RNA interference technology. The expression level of OBP28 and the resistance level were tested in the silenced strain and control strain after microinjection of double-stranded RNA for a 48-h interference period. Four field-collected strains (henceforth ‘field strains’) of Cx. quinquefasciatus were also examined for their resistance to deltamethrin and levels of OBP28 expression. Finally, a correlation analysis between deltamethrin resistance and gene expression was carried out for all seven strains, i.e. the four field strains and the three laboratory strains.ResultsIn the bioassay, the mortality of SS, HN and RR was 100%, 21.33% and 1.67%, respectively. The relative expression levels of OBP28 in strains HN and RR were 6.30- and 6.86-fold higher, respectively, than that of strain SS. After silencing of the OBP28 gene, the mortality of strain RR was 72.20% and that of the control strain 26.32%. The mortality of strain RR increased significantly after interference compared to that of the control strain. There was a negative correlation between OBP28 gene expression and mortality in adult mosquitoes after exposure to deltamethrin.ConclusionsTo our knowledge, this study shows for the first time a correlation between the expression of a gene coding for OBP and insecticide resistance in mosquitoes. The potential resistance mechanism that was elucidated provides a new target gene for the surveillance of resistance in mosquitoes.Graphical abstract

Laboratory evaluation of the efficacy of boric acid containing toxic sugar baits against Anopheles culicifacies, An. stephensi and Aedes aegypti mosquitoes.

Attractive toxic sugar baits (ATSB) is a novel tool which employs mosquito sugar feeding behaviour to kill them. The potential of ATSB against mosquito vectors has been demonstrated in limited scope around the world including Israel, some of the African countries and USA. But their efficacy against mosquito vectors of India is yet to be ascertained. Therefore, current study was planned to evaluate the efficacy of TSB (without attractant) against two important malaria vectors Anopheles culicifacies and An. stephensi along with major dengue vector Aedes aegypti. TSB solution was prepared by dissolving different concentrations of boric acid in glucose and tested against each of the mosquito species. Another experiment was done by spraying this boric acid sugar solution on to Calendula officinalis plant. It served as a sole source of mosquito feed and mortality of mosquitoes was counted after 24 hours. The TSB resulted in 100% mortality of Ae. aegypti and An. stephensi at 4% concentration of boric acid while in An. culicifacies 100% mortality was achieved at 3% concentration TSB solution. TSB solution with 2% boric acid, resulted in 99.1% mortality in An. culicifacies while ~95 % mortality of An. stephensi and Ae. aegypti. When TSB solution with 2% boric acid was sprayed on plant, ~89-94% mortality was observed in both An. stephensi and An. culicifacies. Based on the results, boric acid based toxic sugar bait solution could be a promising tool for vector control. Further studies are needed to find out its toxicity of TSB against non-target organisms and residual efficacy in field trials in different ecotypes.

Effective residual life of bendiocarb insecticide on sprayed surfaces

Effective residual life of insecticides on sprayed surfaces determines the number of spraying cycles required to cover malaria transmission season. However, residual efficacy of insecticides varies with spray surface type. This study assessed effective residual life of bendiocarb on cow dung plastered and mud plastered wall surface types of human dwellings against An. arabiensis. Wall cone bioassays were carried out following the WHO (World Health Organization) standard operating procedures. Percentage mortality was calculated according to WHO bioassay protocol and Poisson regression model was fitted to show the association of mosquito mortality with explanatory variables using SAS (SAS Institute Inc. 2016. SAS® 9.4 Language Reference: Concepts, Sixth Edition. Cary, NC: SAS Institute Inc.). Efficacy of bendiocarb against An. arabiensis was 100% for the first 35 days on both cow dung and mud wall surface types. However, its efficacy deteriorated earlier on non-plastered wall surfaces after 35 days. The relationship between mosquito mortality and explanatory variables was: Mortality =3.637542, Sprayed = + 0.282841, Plastering= - 0.0094612 35-Days after spray, - 0.600159 65-Days after spray. Because coefficients for cone position were not significant, they were excluded from the regression equation. Generally, the effective residual life of bendiocarb on mud plastered wall surface was too short to cover malaria transmission season. Therefore, alternative insecticide with longer residual effective life is required in areas having more than two months of malaria transmission season.

BASELINE SURVEY ON THE INSECTICIDE RESISTANCE PROFILE OF ANOPHELES GAMBIAE (DIPTERA: CULICIDAE) TO SELECTED INSECTICIDES IN IGBOKODA, SOUTHWESTERN-NIGERIA

Malaria is a growing concern in African countries, including Nigeria. Among the multiple malaria control approaches, insecticides are of key importance, and this main practice is threatened by the emergence of resistance in malaria vectors. Studies on the susceptibility of Anopheles towards insecticides are lacking in Igobokoda Southwestern-Nigeria, and this study investigated the susceptibility status of Anopheles gambiae s.l using five selected insecticides including permethrin (0.75%), lambda cyhalothrin (0.05%), propoxur (1.0%), bendiocarb (0.1%) and pirimiphos-methyl (0.1%), which are frequently used as control against the vector population. Immature stages of Anopheles mosquitoes were collected using 350mL deep ladles and scooping spoons and bred in the laboratory following the WHO standard protocols. Mosquitoes aged between 2-5 days were exposed to the selected insecticides using the WHO standard test procedure. Knock down time (KDT) for 50 and 95% was determined using probit analysis. Mortality of mosquitoes exposed to bendiocarb was 100%, followed by propoxur (98%), pirimiphos-methyl and permethrin (95%) and least was observed for lambda-cyhalothrin (67%). Percentage knockdown was highest in bendiocarb compared to other insecticides and the difference was significant (F=5.2, P&lt;0.0001). KDT50 and KDT95 ranged between 12.07 to 54.12min and 28.18 to 91.38min, respectively, and was lowest for bendiocarb (12.07 and 28.18min, respectively). This study therefore imperatively suggests the continuous use of permethrin, adoption of bendiocarb and combined insecticides on the treated long lasting bed nets as well as indoor coating of walls or spraying.

HMGB1-Like Dorsal Switch Protein 1 Triggers a Damage Signal in Mosquito Gut to Activate Dual Oxidase via Eicosanoids

Several mosquitoes transmit human pathogens by blood feeding, with the gut being the main entrance for the pathogens. Thus, the gut epithelium defends the pathogens by eliciting potent immune responses. However, it was unclear how the mosquito gut discriminates pathogens among various microflora in the lumen. This study proposed a hypothesis that a damage signal might be specifically induced by pathogens in the gut. The Asian tiger mosquito, Aedes albopictus, encodes dorsal switch protein 1 (Aa-DSP1) as a putative damage-associated molecular pattern (DAMP). Aa-DSP1 was localized in the nucleus of the midgut epithelium in naïve larvae. Upon infection by a pathogenic bacterium, Serratia marcescens, Aa-DSP1 was released to hemocoel and activated phospholipase A2 (PLA2). The activated PLA2 increased the level of prostaglandin E2 (PGE2) in the gut and subsequently increased Ca2+ signal to produce reactive oxygen species (ROS) via dual oxidase (Duox). Inhibition of Aa-DSP1 via RNA interference or specific inhibitor treatment failed to increase PGE2/Ca2+ signal upon the bacterial infection. Thus, the inhibitors specifically targeting eicosanoid biosynthesis significantly prevented the upregulation of ROS production in the gut and enhanced mosquito mortality after the bacterial infection. However, such inhibitory effects were rescued by adding PGE2. These suggest that Aa-DSP1 plays an important role in immune response of the mosquito gut as a DAMP during pathogen infection by triggering a signaling pathway, DSP1/PLA2/Ca2+/Duox.

Toxicity of Legiayu incense as Insecticide and Larvicide Against Aedes aegypti Mosquitoes Mortality

Community-based DHF vector control has been implemented in Indonesia but has not yet obtained optimal results. Thus, in the community choosing synthetic insecticides to control disease vectors. However, irregular and excessive use of insecticides has a toxic effect and resistance to mosquitoes. Burning mosquito coils and incense containing synthetic dyes and fragrances have the potential to reduce environmental quality. Therefore, this research was conducted to analyze the toxicity of Legiayu incense as an insecticide and larvicide against Aedes aegypti mosquito mortality. The research design is experimental with a completely randomized design. Testing was conducted by providing exposure to smoke and ash of Legiayu incense five times on twenty-five Aedes aegypti mosquitoes. Statistical analysis used one-way ANOVA, LSD, and probit test. The test result as insecticide value (p=0.000) effective exposure for 20 minutes with a durability of 6 hours. The test result as larvicide value (p=0.000) effective exposure for 24 hours. Thus, exposure toismoke and ash of iLegiayuiincense has a very noticeable effect on the mortality of Aedes aegypti mosquitoes. Exposureito Legiayu incense smoke obtained an LT50 value of 0,9012 ≤ 5 (super toxic category) with a time of 15 minutes 39 seconds, coefficient determination of 99.24%, and correlation coefficient of 99.62% while exposure to the ash of Legiayu incense obtained LT50 value of 0,05896 ≤ 5 (super toxic category) with time 19 hours 15 minutes 34 seconds, coefficient determination and correlation coefficient of 100%. Histopathological test results showed that Legiayu incense smoke did not cause tissue degeneration, necrosis, hyperplasia, and metaplasia in the lung tissue of mice (mus musculus) within a period of 12 weeks. Thus, Legiayu incense is effective as insecticides and larvicides against Aedes aegypti mosquitoes. Legiayu incense has potential substitute for mosquito repellent coils, temephos, and synthetic incense circulating in the market.

Comparative Insecticidal Effects of Dry Ocimum Gratissium (Scent Leaves) and Rambo™ Paper on Mosquitoes in Jos, Nigeria

Insecticides are chemicals or biological substances that are used to kill or disable insects. Blood feeding mosquitoes are responsible for the intolerable biting nuisance and transmission of large number of diseases such as malaria, yellow fever, dengue fever, filarias is amongst others, causing serious health problems to humans and obstacles to socioeconomic development of developing nations like Nigeria. The insecticidal effect of scent leaves (Ocimum gratissum) and Rambo™ paper on mosquitoes was investigated. The study is aimed at comparing the insecticidal effects of Ocimum gratissimum and Rambo™ paper on mosquitoes in Jos. 100 mosquitoes were exposed to dried Ocimum gratissum and Rambo™ paper at different time intervals of 5, 10 and 12 min. Results obtained showed a time dependent insecticidal effect on mosquitoes, which was 54.2%, 54.0% and 55.6% total mortality of mosquitoes at respective time intervals on exposures to both Rambo™ paper insecticide and Osimum gratissum, indicating that there was a significant difference in the lethal effect of Rambo™ paper insecticide and scent leave on mosquitoes at (p&lt;0.05). In comparing the lethal effect of Rambo™ paper insecticide and Ocimum grasstisimum on mosquitoes at differnt locations in Jos, at Dogon Karfe, after 10 min of treatment, Rambo™ paper had the highest lethal effect of 21 (84.0%) compared to scent leaves 6 (24.0%) and at Abattoir Jos, after 12 min of treatment, Rambo™ paper had the highest lethal effect of 17 (94.4%) compared to Ocimum grasstisimum 3 (16.7%). These comparisons were significant at p 0.05. This study provides evidence that Ocimum grasstisimum has a mosquitocidal effect. However, the Rambo™ paper gave a better mosquitocidal effect than Ocimum grasstisimum. There is a need to discover better additive or extract options that could give Ocimum grasstisimum a better effect as a natural product available in Africa towards the malaria eradication programme.

EFEKTIVITAS KELAMBU BERINSEKTISIDA DALAM PENGENDALIAN VEKTOR MALARIA DI INDONESIA

Malaria is still a health problem in several parts of Indonesia. National malaria elimination is targeted at 2030. One of the programs carried out by the government in malaria control to achieve elimination is the mass distribution and use of insecticide-treated mosquito nets. The use of insecticide-treated mosquito nets is one of the efforts to reduce malaria cases through vector control. Insecticide mosquito nets are recommended as a strategic step to break the chain of malaria transmission because the insecticide content in the treated fibers can kill mosquitoes. The data used in this paper is data on the percentage of mosquito mortality from the efficacy test of insecticide-treated mosquito nets to see the killing power of mosquito nets which is then used to assess the effectiveness of mosquito nets in vector control. The data were obtained from scientific studies of articles published in scientific journals. From the data collected, it is known that some insecticide-treated mosquito nets used by the community are effective in vector control, but some are no longer effective. The best period for using mosquito nets is less than 6 months by paying attention to the proper washing method so that insecticide-treated mosquito nets are still effective.

Multiple insecticide resistance mechanisms in urban population of Anopheles coluzzii (Diptera: culicidae) from Lagos, South-West Nigeria

Malaria is a major public health challenge in Africa with Nigeria accounting for the highest burden of the disease in the world. Vector control has proved to be a highly effective component of malaria control, however, the development and spread of insecticide resistance in major vectors of malaria have been a major challenge. This study assessed resistance mechanisms in Anopheles coluzzii populations from Kosofe, Lagos mainland and Ojo Local Government Areas in Lagos, Nigeria where An. gambiae s.l is resistant to DDT and Permethrin. WHO susceptibility bioassay test was used in determining resistance status of An. coluzzii to discriminating doses of DDT and Permethrin while synergist assay was used to assess the involvement of monooxygenases in resistance development. Sub-species of An. gambiae s.l (An. gambiae and An. coluzzii) were identified using polymerase chain reaction (PCR) and Restriction Fragment Length Polymorphism (PCR-RFLP) while Allele-Specific Polymerase Chain Reaction (AS-PCR) assay was used to detect knockdown mutation (kdr-West; L1014F). Biochemical assays were used in determining the activities of metabolic enzymes. High DDT resistance was recorded in An. coluzzii populations from the three sites. Mortality rate of mosquitoes exposed confirmed Permethrin resistance in Kosofe (50%) and Lagos mainland (48%) but resistance was suspected in Ojo (96%). All specimens tested were confirmed as An. coluzzii with low kdr frequency; 11.6%, 16.4% and 6.7% in Kosofe, Lagos mainland and Ojo respectively. Pre-exposure to synergist (PBO) before exposure to Permethrin led to increased mortality in all populations. Esterase activity was insignificantly overexpressed in Kosofe (p = 0.849) and Lagos mainland (p = 0.229) populations. In contrast, GST activity was significantly lower in populations from Lagos mainland (63.650 ± 9.861; p = 0.007) and Ojo (91.765 ± 4.959; p = 0.042) than Kisumu susceptible strains (120.250 ± 13.972). Monooxygenase activity was higher in Lagos mainland (2.371 ± 0.261) and Ojo (1.361 ± 0.067) populations, albeit significantly in Lagos mainland (p = 0.007) only. Presence of target-site mutation in all populations, increased mortality with pre-exposure to PBO and elevated monooxygenase in Lagos mainland population were confirmed. Multiple resistance mechanisms in some urban populations of An. coluzzii from Lagos, Nigeria calls for appropriate resistance management strategies.

Assessing transfluthrin mortality against Aedes aegypti and Culex quinquefasciatus inside and outside US military tents in a northern Florida environment

Mortality caused by passive resin transfluthrin diffusers (∼5 mg AI per 24 h release rate) suspended in small 2-person tents was measured for colony-reared sentinel pyrethroid susceptible Aedes aegypti and Culex quinquefasciatus female mosquitoes, as well as a pyrethroid-resistant strain of Aedes aegypti, in a USA military field camp scenario. Mortality effects were investigated for impact by factors such as sentinel cage location (inside tent, tent doorway and outside tent), exposure time (15, 30, 45 and 60 min), and environmental temperature (°C), all of which were examined over an 8-week period. Analyses determined there was a significant interaction between mosquito strain and transfluthrin susceptibility, with the two susceptible strains experiencing significantly greater mean mortality than the resistant Ae. aegypti strain. Significant differences were likewise observed between the mosquito strains over the 8-week study period, where study week and temperature were both positively correlated with an increase in observed mean mosquito mortality. Mosquito proximity to the transfluthrin diffusers was also influenced by week and showed that sentinel cage placement in the environment demonstrates different mortality measurements, depending on the environmental conditions. The length of exposure to transfluthrin, however, was determined to not significantly impact transfluthrin efficacy on the examined mosquito strains, although increased exposure did result in increased susceptible strain mortality. These results suggest that transfluthrin is highly effective in causing mortality against susceptible Ae. aegypti and Cx. quinquefasciatus mosquitoes under field conditions but is minimally effective against pyrethroid-resistant Ae. aegypti mosquitoes. Transfluthrin-infused devices are influenced by environmental factors that can combine to impact mosquito mortality in the field.

The Effect of Insect Repellent Exposure on Leukocyte Profile and Histopathologic Findings in Lungs

Organophosphate compounds in insect repellent have a role in contributing to mosquito mortality but have toxic effects for humans when exposed for a long time. The research is aimed to analyze the effect of insect repellent exposure in blood leukocyte profile and histopathologic findings in lungs. The study used thirty males Rattus novergicus, which were divided into three groups, such as electric liquid insect repellent (P1) with contain 0.031% dimefluthrin, anti-mosquito coils (P2) with 0.014% dimefluthrin, and electric mat mosquito repellent with 0.566% dimefluthrin for 8 hours in 20 days respectively. Leukocyte profiles were determined by using the blood smear method, and the lung’s health was identified by histopathologic findings. Based on the results study showed mosquito coils exposure increase the lymphocytes count. Meanwhile, the electric liquid-repellent increased the basophil’s numbers. The electric mat exposure had more eosinophils, neutrophils stab, neutrophils segment, and monocytes in the blood. The leukocyte profile of each group showed there were no statistically significant differences (P-value &gt; 0.05). Based on histopathology, lung findings showed that the electric mat exposure contributed to cells degeneration 7.5% and pleural thickening 30%. The higher dimefluthrin concentrations in insect repellents could affect leukocyte profile and lungs health.

Knockout of Anopheles stephensi immune gene LRIM1 by CRISPR-Cas9 reveals its unexpected role in reproduction and vector competence.

PfSPZ Vaccine against malaria is composed of Plasmodium falciparum (Pf) sporozoites (SPZ) manufactured using aseptically reared Anopheles stephensi mosquitoes. Immune response genes of Anopheles mosquitoes such as Leucin-Rich protein (LRIM1), inhibit Plasmodium SPZ development (sporogony) in mosquitoes by supporting melanization and phagocytosis of ookinetes. With the aim of increasing PfSPZ infection intensities, we generated an A. stephensi LRIM1 knockout line, Δaslrim1, by embryonic genome editing using CRISPR-Cas9. Δaslrim1 mosquitoes had a significantly increased midgut bacterial load and an altered microbiome composition, including elimination of commensal acetic acid bacteria. The alterations in the microbiome caused increased mosquito mortality and unexpectedly, significantly reduced sporogony. The survival rate of Δaslrim1 mosquitoes and their ability to support PfSPZ development, were partially restored by antibiotic treatment of the mosquitoes, and fully restored to baseline when Δaslrim1 mosquitoes were produced aseptically. Deletion of LRIM1 also affected reproductive capacity: oviposition, fecundity and male fertility were significantly compromised. Attenuation in fecundity was not associated with the altered microbiome. This work demonstrates that LRIM1’s regulation of the microbiome has a major impact on vector competence and longevity of A. stephensi. Additionally, LRIM1 deletion identified an unexpected role for this gene in fecundity and reduction of sperm transfer by males.