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

Background Pyrethroid-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. Methods An 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. Results With 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. Conclusion Olyset 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.

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.

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.

Can pyrethroid-piperonyl butoxide (PBO) nets reduce the efficacy of indoor residual spraying with pirimiphos-methyl against pyrethroid-resistant malaria vectors?

As the uptake of pyrethroid-PBO ITNs increases, their combination with IRS insecticides could become an operational reality in many malaria-endemic communities. Pirimiphos-methyl is a pro-insecticide requiring activation by mosquito cytochrome P450 enzymes to induce toxicity while PBO blocks activation of these enzymes in pyrethroid-resistant vector mosquitoes. PBO may thus antagonise the toxicity of pirimiphos-methyl IRS when combined with pyrethroid-PBO ITNs. The impact of combining two major brands of pyrethroid-PBO ITNs (Olyset Plus, PermaNet 3.0) with pirimiphos-methyl IRS (Actellic 300CS) was evaluated against pyrethroid-resistant Anopheles gambiae sl in two parallel experimental hut trials in southern Benin in comparison to bendiocarb IRS and each intervention alone. The wild vector population was resistant to pyrethroids but susceptible to pirimiphos-methyl and bendiocarb. PBO pre-exposure partially restored deltamethrin toxicity but not permethrin. Mosquito mortality in experimental huts was significantly improved in the combinations of bendiocarb IRS with Olyset Plus (33%) and PermaNet 3.0 (38%) compared to bendiocarb IRS alone (14 to 16%, p<0.001), demonstrating an additive effect. Conversely, mortality was significantly reduced in the combinations of pirimiphos-methyl IRS with Olyset Plus (59%) and PermaNet 3.0 (55%) compared to pirimiphos-methyl IRS alone (77 to 78%, p<0.001), demonstrating an antagonistic effect. Combining pirimiphos-methyl IRS with the pyrethroid-PBO ITNs provided significantly improved mosquito mortality (55 to 59%) compared to the pyrethroid-PBO ITNs alone (22 to 26%) and improved blood-feeding inhibition relative to the IRS alone. This study provided evidence of an antagonistic effect when pyrethroid-PBO ITNs were combined with pirimiphos-methyl IRS in the same household resulting in lower levels of vector mosquito mortality compared to the IRS alone. Pirimiphos-methyl IRS also showed potential to significantly enhance malaria control when deployed to complement pyrethroid-PBO ITNs in an area where PBO fails to fully restore susceptibility to pyrethroids.

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 > 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.

Exploring alternative insecticide delivery options in a “Lethal House Lure” for malaria vector control

The In2Care® EaveTube is a house modification designed to block and kill malaria mosquitoes using an electrostatic netting treated with insecticide powder. A previous study demonstrated prolonged duration of effective action of insecticide-treated electrostatic netting in a semi-field setting. As part of a cluster randomized controlled trial (CRT) of the EaveTube intervention in Cote d’Ivoire, we investigated the residual efficacy of a pyrethroid insecticide deployed in Eave Tubes under village condition of use. We also explored the scope of using existing malaria control technologies including LLINs and IRS as alternative methods to deliver insecticides in the lethal house lure. The efficacy of beta-cyfluthrin was evaluated using the “eave tube bioassay” and was found to be relatively short-lived in the field during the CRT, with mortality of pyrethroid resistant Anopheles gambiae mosquitoes declining below 80% after 4 months. The impact (mosquito mortality) of PVC tubes coated with pirimiphos methyl was similar to that of beta-cyfluthrin treated insert (66.8 vs 62.8%) in release-recapture experiments in experimental huts. Efficacy was significantly lower with all the LLINs tested; however, the roof of PermaNet 3.0 induced significantly higher mosquito mortality (50.4%) compared to Olyset Plus (25.9%) and Interceptor G2 (21.6%) LLINs. The new delivery methods showed a rapid decline in efficacy over time with mortality decreasing below 50% within 2 months in residual activity bioassays. None of the current products appeared superior to the powder treatments but as with the powders, further research and development on formulations and doses are required.

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 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.

MiR-279-3p regulates deltamethrin resistance through CYP325BB1 in Culex pipiens pallens

Background The overuse of insecticides to control insect vectors has promoted extensive insecticide resistance in mosquitoes. In this study, the functions of microRNA (miR)-279-3p and its target CYP325BB1 in the regulation of deltamethrin resistance in Culex pipiens pallens was investigated. Methods Quantitative real-time reverse transcription PCR was used to detect the expression levels of miR-279-3p and CYP325BB1. Then, the dual-luciferase reporter assay system, RNA interference, CDC bottle bioassay and Cell Counting Kit-8 (CCK-8) assay were used to explore the roles of these molecules in deltamethrin resistance both in vivo and in vitro. Results The expression patterns of miR-279-3p and CYP325BB1 were compared between deltamethrin-sensitive (DS-strain) and deltamethrin-resistant (DR-strain) mosquitoes. Luciferase activity was downregulated by miR-279-3p, the effect of which was ablated by a mutation of the putative binding site for CYP325BB1. In DR-strain mosquitoes, the expression of miR-279-3p was increased by microinjection and oral feeding of miR-279-3p agomir (mimic). CYP325BB1 mRNA levels were downregulated, which resulted in a higher mortality of the mosquitoes in miR-279-3p mimic-treated groups. In the DS-strain mosquitoes, microinjection of a miR-279-3p inhibitor decreased miR-279-3p expression, whereas the expression of CYP325BB1 was increased; the mortality of these mosquitoes decreased significantly. In addition, overexpression of pIB/V5-His-CYP325BB1 changed the sensitivity of C6/36 cells to deltamethrin in vitro. Also in DR-strain mosquitoes, downregulation of CYP325BB1 expression by microinjection of si-CYP325BB1 increased mosquito mortality in vivo. Conclusions These findings provide empirical evidence of the involvement of miRNAs in the regulation of insecticide resistance and indicate that miR-279-3p suppresses the expression of CYP325BB1, which in turn decreases deltamethrin resistance, resulting in increased mosquito mortality. Taken together, the results provide important information for use in the development of future mosquito control strategies. Graphical abstract

Omitting age-dependent mosquito mortality in malaria models underestimates the effectiveness of insecticide-treated nets

Mathematical models of vector-borne infections, including malaria, often assume age-independent mortality rates of vectors, despite evidence that many insects senesce. In this study we present survival data on insecticide-resistant Anopheles gambiae s.l. from field experiments in Côte d’Ivoire. We fit a constant mortality function and two age-dependent functions (logistic and Gompertz) to the data from mosquitoes exposed (treated) and not exposed (control) to insecticide-treated nets (ITNs), to establish biologically realistic survival functions. This enables us to explore the effects of insecticide exposure on mosquito mortality rates, and the extent to which insecticide resistance might impact the effectiveness of ITNs. We investigate this by calculating the expected number of infectious bites a mosquito will take in its lifetime, and by extension the vectorial capacity. Our results show that the predicted vectorial capacity is substantially lower in mosquitoes exposed to ITNs, despite the mosquitoes in the experiment being highly insecticide-resistant. The more realistic age-dependent functions provide a better fit to the experimental data compared to a constant mortality function and, hence, influence the predicted impact of ITNs on malaria transmission potential. In models with age-independent mortality, there is a reduction of 56.52% ( [[EQUATION]] 14.66) for the vectorial capacity under exposure compared to no exposure. However, the two age-dependent functions predicted a larger reduction due to exposure: for the logistic function the reduction is 74.38% ( [[EQUATION]] 9.93) and for the Gompertz 74.35% ( [[EQUATION]] 7.11), highlighting the impact of incorporating age in the mortality rates. These results further show that multiple exposures to ITNs had a considerable effect on the vectorial capacity. Overall, the study highlights the importance of including age dependency in mathematical models of vector-borne disease transmission and in fully understanding the impact of interventions.

EFEKTIVITAS INSEKTISIDA NABATI DAUN SALAM (SYZYGIUM POLYANTHUM) TERHADAP MORTALITAS NYAMUK AEDES AEGYPTI

Dengue Hemorrhagic Fever (DHF) is an acute disease in the form of dengue virus infection which is spread through mosquito bites. The use of synthetic insecticides actually causes negative effects on the environment, so another alternative is needed, namely the use of bay leaf vegetable insecticides. The purpose of this study was to determine the effectiveness of bay leaf insecticide (Syzygium polyanthum) as an insecticide against the mortality of Aedes Aegypti mosquitoes with the liquid electric method. This study used a quasi-experimental (quasi-experimental) design with a completely randomized design (CRD) method where the experiment was conducted on 150 mosquitoes in 5 treatments and 3 replications. The results of the percentage of bay leaf solution (Syzgium polyanthum) which has the largest number of mosquito mortality at a concentration of 80 ml/200 ml of water with a mortality percentage of 60%. The result of probit LC50 analysis of bay leaf solution was 64,315 g/ml. The higher the concentration of the solution given, the greater the number of mosquito mortality. It is necessary to modify the right tools and methods to make a solution of bay leaf (Syzgium polyanthum) so that it is more effectively applied in the community.Keywords: bioinsecticide, bay leaf, liquid electric, mortality, Aedes aegypti