Insecticide Resistance Management Programs Research Articles

Genome-wide analysis of detoxification genes conferring diamide insecticide resistance in Spodoptera exigua identifies CYP9A40

For over a decade, diamide insecticides have been effective against lepidopteran pests like beet armyworm, Spodoptera exigua (Hübner, 1808). However, the evolution of resistance poses a challenge to their sustainable use. We identified an I4790M mutation in the S. exigua ryanodine receptor (RyR) gene, but its correlation with resistance varied across the field-collected Korean populations of S. exigua. RNA sequencing and differential gene expression analysis were performed to investigate other resistance mechanisms. Diamide-resistant and susceptible strains and F1 hybrids were compared by mapping RNA-seq reads to the S. exigua reference genome. CYP9A40 was identified as a critical gene in diamide resistance due to its high expression in the resistant strains. Synergist bioassays with piperonyl butoxide supported the role of P450s in diamide metabolic resistance in S. exigua. A strong positive correlation between CYP9A40 over-expression levels (up to 80-fold) and diamide LC50 values was obtained for field-collected populations uniformly showing a 100% frequency of the RyR I4790M target-site resistance allele. To validate the function of CYP9A40 in diamide detoxification, we recombinantly expressed the gene and tested its ability to bind and degrade chlorantraniliprole as a substrate. The results confirmed its catalytic role in diamide metabolism. CYP9A40 has been identified and validated to confer metabolic resistance in Korean S. exigua populations. It works alongside the RyR target-site I4790M mutation to enhance diamide resistance. These mechanisms offer insights for resistance monitoring and support insecticide resistance management programs to improve control strategies for S. exigua.

Realized heritability and fitness costs of flupyradifurone resistance in the cowpea aphid, Aphis craccivora Koch

The cowpea aphid, Aphis craccivora Koch (Homoptera: Aphididae) is a serious insect pest that infests legume crops. Flupyradifurone (FPF) is a butenolide insecticide that selectively targets sucking insects including aphids and whiteflies. However, the realized heritability (h2) and fitness costs associated with resistance to FPF have not yet been assessed with A. craccivora. In this study, A. craccivora showed the potential to develop resistance to FPF under continuous selection, with an overall mean estimated h2 of resistance of 0.21. Results of fitness costs showed no differences in the developmental time (except for N2 and N3), longevity, fecundity, adult pre-reproductive period (APRP), total pre-reproductive period (TPRP), and oviposition days (Od) of the FPF-R strain compared to the S-strain. In contrast, all population growth parameters, including the growth reproductive rate (GRR), net reproductive rate (R0), intrinsic rate of increase (r), finite rate of increase (λ), mean generation time (T), and doubling time (DT), were significantly lower in the FPF-R strain than in the S-strain. Fitness cost studies revealed that resistance to FPF decreased the relative fitness of the FPF-R strain to 0.76 compared with that of the S-strain. The estimation of realized heritable h2 and identification of resistance fitness costs provide a basis for designing insecticide resistance management (IRM) programs to sustain susceptibility in insect populations.

Evaluating foliar insecticides and economic thresholds for Tychius picirostris (Coleoptera: Curculionidae) management in Oregon white clover seed production.

The clover seed weevil, Tychius picirostris Fabricius (Coleoptera: Curculionidae), is a major pest in Oregon white clover seed crops. Reliance on synthetic pyrethroid insecticides and limited availability of diverse modes of action (MoAs) has increased insecticide resistance selection in regional T. picirostris populations, emphasizing the need to evaluate novel chemistries and rotational strategies for effective insecticide resistance management (IRM). The efficacy of 8 foliar insecticide formulations for managing T. picirostris adult and larval life stages was determined in small and large-plot field trials across 2 crop years. In both years, bifenthrin (Brigade 2EC), the grower's standard, showed negligible adult and larval suppression. Insecticide formulations with isocycloseram and cyantraniliprole active ingredients reduced adult and larval populations when applied at BBCH 59-60 (prebloom) and BBCH 65-66 (full bloom) growth stages, respectively. While differences in T. picirostris abundance were observed among insecticide treatments, seed yield differences were not detected in large-plot trials. Larval abundance was correlated with reduced seed yield, and an economic threshold of ≥3 larvae per 30 inflorescences was determined as a conservative larval threshold to justify foliar applications of diamide insecticides. Additional commercial white clover seed fields were surveyed to compare larval scouting techniques, including a standard Berlese funnel and a grower's do-it-yourself funnel. Both larval extraction techniques were correlated and provided similar estimates of larval abundance. These findings demonstrate new MoAs, optimal insecticide application timing, and larval monitoring methods that can be incorporated into an effective T. picirostris IRM program in white clover seed crops.

Monitoring the susceptibility of Bemisia tabaci Middle East-Asia Minor 1 (Hemiptera: Aleyrodidae) to afidopyropen, cyantraniliprole, dinotefuran, and flupyradifurone in south Florida vegetable fields.

Bemisia tabaci Middle East-Asia Minor 1 (MEAM1) is a significant pest that damages a wide range of high-value vegetable crops in south Florida. This pest has demonstrated the ability to develop resistance to various insecticide groups worldwide. Monitoring the resistance levels of MEAM1 populations and maintaining baseline susceptibility data are crucial for the long-term effectiveness of insecticide management strategies. We conducted serial dilution bioassays on 15 field populations of MEAM1 collected in south Florida to assess their resistance to 4 key insecticides: afidopyropen, cyantraniliprole, dinotefuran, and flupyradifurone. To quantify resistance levels, resistance ratios (RR) were generated by comparing the LC50 values of field populations to those of a known susceptible MEAM1 colony reared in the laboratory. Our findings reveal that all field-collected populations were susceptible to dinotefuran (RR 1-8) and flupyradifurone (RR 2-8). While over 80% of the populations tested were susceptible to afidopyropen (RR 1-9), 2 populations exhibited low (RR 38) and moderate resistance (RR 51), respectively. In contrast, most of the populations (57%) showed low to moderate resistance to cyantraniliprole (RR 21-78), and the remaining populations were susceptible (RR 3-10). The 2 populations with resistance to afidopyropen also exhibited moderate resistance to cyantraniliprole. Further research in this direction can aid in refining insecticide resistance management programs in Florida and other regions where B. tabaci MEAM1 is a major pest. Exploring the implications of these findings will be essential for insecticide use and integrated pest management strategies in south Florida.

Soybean aphid management and perception of insecticide resistance by farmers

Abstract In the Midwest United States, soybean aphids can cause significant economic losses in soybean production. The repeated use of insecticides within the same family has led to the development of insecticide resistance in soybean aphids. Implementing integrated pest management (IPM) and insecticide-resistance management (IRM) strategies can help farmers reduce the frequency of insecticide applications and delay the evolution of soybean aphid resistance while reducing production costs, maintaining yields, and protecting the environment. Farmers play a crucial role in implementing such practices; therefore, to understand farmers’ practices for management of soybean aphids and their perceptions of insecticide resistance, a survey of soybean farmers in Minnesota and North Dakota was conducted in 2021. Overall, the reported decision-making sources were similar for both farmers who changed and those who did not change their aphid management due to insecticide resistance; these were crop consultants and agricultural retailers. The threshold used by farmers in both groups to determine whether to use a foliar insecticide was lower than the research-based threshold for soybean aphids, and farmers were aware that this could contribute to insecticide resistance. In addition, farmers who changed soybean aphid management were more concerned than farmers who did not change about insecticide resistance and were implementing good practices such as scouting, using labeled application rates, and rotating insecticide modes of action. The results of this research can inform the refinement of IPM and IRM programs as well as extension education efforts.

RATIONAL MANAGEMENT OF INSECTICIDES IN MONOCULTURE AREAS

In Mexico, some monocultures occupy large areas, and organo-synthetic insecticides are the dominant method of pest control. The conventional chemical strategy is heavily reliant on new insecticides being developed in other countries. This paper discusses the characteristics of traditional/conventional and rational insecticide management. In traditional management, companies compete to position their products in the market once authorized. When pests develop resistance, the farmer must increase the dose and frequency of applications. These actions seriously threaten the environment, human health, and crop profitability. The hope is that another insecticide will enter the market to repeat the cycle of abuse and its consequences. The alternative is a rational strategy based on scientific evidence for chemical pest control. Farmers must adhere to an insecticide resistance management program that keeps resistance gene expression and intensity at acceptable levels. We propose the creation of public and private laboratories to systematically estimate the dynamics of insecticide resistance through bioassays. This will enable decisions to be made based on scientific rather than commercial information. Changing the current paradigm for the use of these agrochemicals will result in significant improvements in the time it takes to maintain acceptable biological efficacy, economic benefits to the producer, and significant reductions in adverse effects on human health and the environment.

Evaluation of Phytochemical Compounds in Senna siamea stembark Powders for Insect Pest Control

A laboratory screening was conducted to evaluate the phytochemical compounds in Senna siamea stembark powders and their insecticidal activity on Callosobruchus subinnotatus PIC on stored Bambara groundnut through untargeted Gas Chromatography – Mass Spectrometry (GC- MS) using three different extraction solvents (ethanol, methanol and distilled water). To mention but few, the ethanol extracts revealed Octanoic acid, 4,6-dimethyl-, methyl ester, (4S,6S)-(+)- (C11H22O2); cis-Vaccenic acid (C19H36O2); Z,Z-10,12-Hexadecadien-1-ol acetate (C18H32O2); 2-Methyl-Z,Z-3,13-octadecadienol (C19H36O). Methanol extracts showed Oleic Acid (C18H34O2); Cyclononasiloxane, octadecamethyl- (C18H54O9Si9); Cyclodecasiloxane, eicosamethyl- (C20H60O10Si10); 2-Methyl-Z,Z-3,13-octadecadienol (C19H36O). Distilled water extraction revealed cis-Vaccenic acid (C19H36O2); 1,2-Benzisothiazole,3-(hexahydro- 1H-azepin-1-yl)-, 1,1-dioxide (C13H16N2O2S); Oleic Acid (C18H34O2); 11-Octadecenoic acid, methyl ester (C19H36O2)as compounds with the highest percentage area. Phytochemicals have been used for many years to control insect pest damage in agricultural crops. Pyretre, Nicotine, Rotenone and tobacco have long been used as control agents against insects in some regions of sub-Saharan Africa. Unlike synthetic chemical insecticides that kill both pests and non-target organisms, botanicals pesticides are relatively target specific. They are also biodegradable, environmentally friendly, and can also be used in insecticide resistance management programs. Hence, could serve as good alternatives to chemical insecticides. Further research on the bio insecticidal activity of these compounds is highly advocated.

PRELIMINARY DETERMINATION OF PHYTOCHEMICAL COMPOUNDS IN SENNA SIAMEA LEAVES POWDERS FOR INSECT PEST CONTROL

Laboratory evaluation of phytochemical compounds and insecticidal/repellent effect of Senna siamea L. leaves was conducted. Ethanol, methanol and distilled water were used as extraction solvents for the conduct of untargeted Gas Chromatography and Mass Spectrometry (GC-MS) analysis of the leave extracts. To mention but few, the ethanol extracts revealed the presence of 2-[2-(2-Ethoxyethoxy) ethoxy]ethyl acetate (C9H18O5); 3-Methylmannoside (C7H14O6); 7H-Chromen-7-one, 8-hydroxy-2,4-dimethyl (C11H10O3); 3,5-Dimethylanisole C9H12O as compound with the highest percentage area.Methanol extracts showed Thiophene, tetrahydro-2-methyl- (C5H10S); 4,6-Di-O-methyl-.alpha.-d-galactose (C8H16O6); 2-Furancarboxaldehyde, 5-[(5-methyl-2-furanyl)methyl]- (C11H10O3); 5,9-Undecadien-2-one, 6,10-dimethyl- (C13H22O) as compound with the highest percentage area. Distilled water extraction revealed Phosphoric acid, dimethyl(4-methoxy-3-t-butylphenyl) ester (C13H21O5P); Vitamin E (C29H50O2); Naphtalene-1,3-dicarbonitrile, 5,6,7,8-tetrahydro-2-hydroxy-4-phenyl (C18H14N2O); Lanosterol (C30H50O) as compound with the highest percentage area. Phytochemicals have been used for many years to control insect pest damage in agricultural crops. Pyretre, Nicotine, Rotenone and tobacco have long been used as control agents against insects in some regions of sub-Saharan Africa. Unlike synthetic chemical insecticides that kill both pests and non-target organisms, botanicals pesticides are relatively target specific. They are also biodegradable, environmentally friendly, and can also be used in insecticide resistance management programs. Hence, could serve as good alternatives to chemical insecticides.

Exploring lethal and sublethal effects of conventional insecticides and insect growth regulators on a picture-winged fly (Diptera: Ulidiidae) pest of sweet corn

Picture-winged flies (Diptera: Ulidiidae) known as corn silk flies are intensively managed with pyrethroids in fresh market sweet corn, Zea mays L., grown in Florida. The identification of alternatives to pyrethroids is needed to increase the efficacy of insecticidal management and to allow the rotation of modes of action for insecticide resistance management (IRM). This laboratory study determined the lethal and sublethal effects of six pyrethroids, five neonicotinoids, four insect growth regulators, two diamides, one spinosyn, one avermectin, and one oxadiazine on adults of the corn silk fly, Euxesta eluta Loew, following topical or ingestion exposure. For topical exposure, pyrethroids caused the highest mortality (≥89%), followed by spinetoram (50%), whereas the remaining insecticides caused <15% mortality. For ingestion exposure, spinetoram caused 95% mortality, followed by dinotefuran, clothianidin, and thiamethoxam, which caused ≥74% mortality. The further evaluation of insecticides causing <15% mortality showed that topical exposure to abamectin reduced by 61.5% the fertility of E. eluta females expressed as the number of larvae per cumulative female-day. None of the insecticides causing <15% mortality following ingestion exposure affected E. eluta fertility. This study confirms that pyrethroids represent the main commercial insecticide option for effective foliar applications targeting adult corn silk flies. Potential alternatives to pyrethroids were highly effective only through ingestion exposure, suggesting the need for the development of management tactics promoting ingestion of insecticides. This study complements pyrethroid susceptibility monitoring for the development of an IRM program in Florida sweet corn.

Alpha-Cypermethrin Resistance in Musca domestica: Resistance Instability, Realized Heritability, Risk Assessment, and Insecticide Cross-Resistance.

Musca domestica L., the common house fly, is a cosmopolitan carrier of human and livestock disease pathogens. The species exhibits resistance to many insecticides; therefore, effective M. domestica insecticide resistance management programs are required worldwide. In the present study, the development of alpha-cypermethrin resistance, realized heritability (h2), instability of resistance trait (DR), and cross-resistance (CR) was investigated in an alpha-cypermethrin-selected M. domestica strain (Alpha-Sel) across 24 generations (Gs). Compared with an alpha-cypermethrin-unselected strain (Alpha-Unsel), resistance to alpha-cypermethrin increased from 46.4-fold (G5) to 474.2-fold (G24) in Alpha-Sel females and 41.0-fold (G5) to 253.2-fold (G24) in Alpha-Sel males. Alpha-cypermethrin resistance declined by between -0.10 (G5) and -0.05 (G24) in both M. domestica sexes without insecticide exposure for 24 generations. The h2 of alpha-cypermethrin resistance was 0.17 and 0.18 for males and females, respectively, in G1-G24. With selection intensities of 10-90%, the G values required for a tenfold increase in the LC50 of alpha-cypermethrin were 6.3-53.7, 4.1-33.8, and 3.0-24.7, given h2 values of 0.17, 0.27, and 0.37, respectively, and a constant slope of 2.1 for males and h2 values of 0.18, 0.28, and 0.38, respectively, and a constant slope of 2.0 for females. Compared with Alpha-Unsel, Alpha-Sel M. domestica exhibited moderate CR to bifenthrin (15.5-fold), deltamethrin (28.4-fold), and cyfluthrin (16.8-fold), low CR to two pyrethroids and five organophosphates, and no CR to insect growth regulators. The instability of resistance trait, low h2, and absent or low CR associated with alpha-cypermethrin resistance in M. domestica indicate resistance could be managed with rotational use of the insecticide.

Trends towards Lower Susceptibility of Spodoptera frugiperda (Lepidoptera: Noctuidae) to Teflubenzuron in Brazil: An Evidence for Field-Evolved Resistance

Susceptibility monitoring to insecticides is a key component to implementing insecticide resistance management (IRM) programs. In this research, the susceptibility to teflubenzuron in Spodoptera frugiperda (J.E Smith) was monitored in more than 200 field-collected populations from major corn-growing regions of Brazil, from 2004 to 2020. Initially, we defined a diagnostic concentration of 10 µg mL-1 of teflubenzuron using a diet-overlay bioassay for monitoring the susceptibility. A variation in the susceptibility to teflubenzuron in S. frugiperda was detected among populations from different locations. We also detected a significant reduction in the susceptibility to teflubenzuron throughout time in all the populations of S. frugiperda evaluated, with larval survival at diagnostic concentration varying from values of <5% in 2004 to up 80% in 2020. Thus, this research provides evidence of field-evolved resistance of S. frugiperda to teflubenzuron and reinforces that IRM practices are urgently needed to be implemented in Brazil.

Sublethal effects of growth-regulating insecticides of synthetic and botanical origins on the biological parameters of Spodoptera frugiperda (Lepidoptera: Noctuidae).

The objective of this study was to evaluate the effects of growth-regulating insecticides of synthetic (e.g., Certero 480 SC, Intrepid 240 SC, Match EC and Mimic 240 SC) and botanical origins (e.g., Azamax 1.2 EC, Agroneem 850 EC, Azact 2.4 EC and Fitoneem 850 EC) on the biological parameters and fertility life table of Spodoptera frugiperda (J.E. Smith) under laboratory conditions. Larvae were fed insecticides that were incorporated into artificial diets. To develop the fertility life table, the following biological parameters were evaluated: survival at 7 days after infestation (d.a.i) and survivorship at adult eclosion, duration of the neonate-to-adult eclosion period, larval and pupal weights and total fecundity (number of total eggs per female). The results indicated that S. frugiperda neonates surviving LC25 or LC50 concentrations of the evaluated insecticides showed longer larval and egg-to-adult periods, lower larval and pupal weights and reduced fecundity, when compared to the control treatment. Larvae exposed to Azamax at LC25 or LC50 concentrations showed the greatest increase in generation duration (75 d). In addition, S. frugiperda adults emerged from pupae when larvae reared on an artificial diet containing growth regulating insecticides of synthetic and botanical origins produced fewer females per female per generation (Ro). As well as, lower rates of natural population increase per day (rm) compared to insects fed the control diet. Our findings indicated that, neem-derived products and growth-regulating insecticides of synthetic origin may be employed within integrated management strategies that aim to keep populations of S. frugiperda below levels that cause economic damage. Similarly, they offer alternatives for insecticide resistance management programs.

Insecticidal effects of some selected plant extracts against Anopheles stephensi (Culicidae: Diptera)

BackgroundThe use of synthetic insecticides against mosquitoes may lead to resistance development and potential health hazards in humans and the environment. Consequently, a paradigm needs to shift towards the alternative use of botanical insecticides that could strengthen an insecticide resistance management programme. This study aimed to assess the insecticidal effects aqueous, hexane, and methanol crude leaf extracts of Calpurnia aurea, Momordica foetida, and Zehneria scabra on an insectary colony of Anopheles stephensi larvae and adults.MethodsFresh leaves of C. aurea, M. foetida and Z. scabra were collected and dried, then separately ground to powder. Powdered leaves of test plants were extracted using sonication with aqueous, hexane, and methanol solvents. The extracts were concentrated, and a stock solution was prepared. For comparison, Temephos (Abate®) and control solutions (a mixture of water and emulsifier) were used as the positive and negative controls, respectively. Different test concentrations for the larvae and the adults were prepared and tested according to WHO (2005) and CDC (2010) guidelines to determine lethal concentration (LC) values. Mortality was observed after 24 h exposure. The statistical analyses were performed using Statistical Package for the Social Sciences (SPSS) software (Kruskal-Wallis test) and R software (a generalized linear model was used to determine LC50 and LC90 values of the extracts).ResultsThe lowest LC50 values were observed in aqueous extracts of M. foetida followed by Z. scabra extract and C. aurea leaves at 34.61, 35.85, and 38.69 ppm, respectively, against the larvae. Larval mortality was not observed from the hexane extracts and negative control, while the standard larvicide (temephos) achieved 100% mortality. Further, the adulticidal efficacy was greatest for aqueous extract of Z. scabra with LC50 = 176.20 ppm followed by aqueous extract of C. aurea (LC50 = 297.75 ppm).ConclusionThe results suggest that the leaf extracts of the three test plants have the potential of being used for the control of vector An. stephensi larvae and adult instead of synthetic mosquitocides. Further studies need to be conducted to identify the active ingredients and their mode of action.

Pyrethroid Susceptibility in Field Populations of Picture-Winged Flies (Diptera: Ulidiidae) Infesting Fresh Market Sweet Corn in Florida

Picture-winged flies (Diptera: Ulidiidae) are the most damaging insect pests of sweet corn (Zea mays L.) produced in Florida for the fresh market. Management of these pests, referred to as corn silk flies, relies on frequent pyrethroid applications targeting adults. In response to the need for an insecticide resistance management (IRM) program for corn silk flies in this highly intensive crop system, glass vial bioassays were conducted to determine the susceptibility of 12 corn silk fly populations to the pyrethroid beta-cyfluthrin. Two Euxesta eluta Loew and nine Euxesta stigmatias Loew populations were obtained by collecting infested ears in commercial and experimental fields in 2020 and 2021. One E. eluta laboratory colony was used as a susceptible reference population. The E. eluta reference colony was the most susceptible population, with an LC50 value of 0.01 µg/vial. The E. stigmatias field populations were generally less susceptible to beta-cyfluthrin than the E. eluta field populations, with the highest LC50 values attaining 3.51 µg/vial and 0.19 µg/vial, respectively. In addition, the five E. stigmatias populations from commercial sweet corn fields were as much as 17.6 times less susceptible than the four E. stigmatias populations from nontreated fields. Results suggest that E. stigmatias is less susceptible to pyrethroids than E. eluta. Results also suggest that corn silk flies in commercial sweet corn fields are selected for reduced pyrethroid susceptibility throughout the growing season. This study successfully used the glass vial bioassay method for corn silk flies, providing a new tool to initiate an IRM program.

Impact of the nicotinic acetylcholine receptor mutation R81T on the response of European Myzus persicae populations to imidacloprid and sulfoxaflor in laboratory and in the field

Sulfoxaflor (Isoclast™ active) is a sulfoximine insecticide that is active on a broad range of sap-feeding insects, including species that exhibit reduced susceptibility to currently available insecticides. Colonies of Myzus persicae (green peach aphid) were established from aphids collected in the field from peach (Prunus persica) and nectarine (Prunus persica var. nucipersica) orchards in France, Italy and Spain. The presence of the nicotinic acetylcholine receptor (nAChR) point mutation R81T was determined for all the colonies. Eight of the 35 colonies collected were susceptible relative to R81T (i.e., R81T absent), three of the colonies were found to be homozygous for R81T while 24 colonies had R81T present in some proportion (heterozygous). Sulfoxaflor and imidacloprid were tested in the laboratory against these M. persicae field colonies, which exhibited a wide range of susceptibilities (sulfoxaflor RR = 0.6 to 61, imidacloprid RR = 0.7 to 986) (resistance ratios, RR) to both insecticides. Although sulfoxaflor was consistently more active than imidacloprid against these field collected M. persicae, there was a statistically significant correlation across all colonies between the RRs for imidacloprid and sulfoxaflor (Pearson's r = 0.939, p < 0.0001). However, when a larger group of the colonies from Spain possessing R81T were analyzed, there was no correlation observed for the RRs between imidacloprid and sulfoxaflor (r = 0.2901, p = 0.3604). Thus, consistent with prior studies, the presence of R81T by itself is not well correlated with altered susceptibility to sulfoxaflor. In field trials, sulfoxaflor (24 and 36 gai/ha) was highly effective (~avg. 88–96% control) against M. persicae, demonstrating similar levels of efficacy as flonicamid (60–70 gai/ha) and spirotetramat (100–180 gai/ha) at 13–15 days after application, in contrast to imidacloprid (110–190 gai/ha) and acetamiprid (50–75 gai/ha) with lower levels of efficacy (~avg. 62–67% control). Consequently, sulfoxaflor is an effective tool for use in insect pest management programs for M. persicae. However, it is recommended that sulfoxaflor be used in the context of an insecticide resistance management program as advocated by the Insecticide Resistance Action Committee involving rotation with insecticides possessing other modes of action (i.e., avoiding rotation with other Group 4 insecticides) to minimize the chances for resistance development and to extend its future utility.

Mosquito population structure, pathogen surveillance and insecticide resistance monitoring in urban regions of Crete, Greece.

BackgroundIn Greece vector borne diseases (VBD) and foremost West Nile virus (WNV) pose an important threat to public health and the tourist industry, the primary sector of contribution to the national economy. The island of Crete, is one of Greece’s major tourist destinations receiving annually over 5 million tourists making regional VBD control both a public health and economic priority.MethodologyUnder the auspices of the Region of Crete, a systematic integrative surveillance network targeting mosquitoes and associated pathogens was established in Crete for the years 2018–2020. Using conventional and molecular diagnostic tools we investigated the mosquito species composition and population dynamics, pathogen infection occurrences in vector populations and in sentinel chickens, and the insecticide resistance status of the major vector species.Principal findingsImportant disease vectors were recorded across the island including Culex pipiens, Aedes albopictus, and Anopheles superpictus. Over 75% of the sampled specimens were collected in the western prefectures potentially attributed to the local precipitation patterns, with Cx. pipiens being the most dominant species. Although no pathogens (flaviviruses) were detected in the analysed mosquito specimens, chicken blood serum analyses recorded a 1.7% WNV antibody detection rate in the 2018 samples. Notably detection of the first WNV positive chicken preceded human WNV occurrence in the same region by approximately two weeks. The chitin synthase mutation I1043F (associated with high diflubenzuron resistance) was recorded at an 8% allelic frequency in Lasithi prefecture Cx. pipiens mosquitoes (sampled in 2020) for the first time in Greece. Markedly, Cx. pipiens populations in all four prefectures were found harboring the kdr mutations L1014F/C/S (associated with pyrethroid resistance) at a close to fixation rate, with mutation L1014C being the most commonly found allele (≥74% representation). Voltage gated sodium channel analyses in Ae. albopictus revealed the presence of the kdr mutations F1534C and I1532T (associated with putative mild pyrethroid resistance phenotypes) yet absence of V1016G. Allele F1534C was recorded in all prefectures (at an allelic frequency range of 25–46.6%) while I1532T was detected in populations from Chania, Rethymnon and Heraklion (at frequencies below 7.1%). Finally, no kdr mutations were detected in the Anopheles specimens included in the analyses.Conclusions/SignificanceThe findings of our study are of major concern for VBD control in Crete, highlighting (i) the necessity for establishing seasonal integrated entomological/pathogen surveillance programs, supporting the design of targeted vector control responses and; ii) the need for establishing appropriate insecticide resistance management programs ensuring the efficacy and sustainable use of DFB and pyrethroid based products in vector control.

In vivo functional validation of the V402L voltage gated sodium channel mutation in the malaria vector An. gambiae.

Pyrethroids are the most widely used insecticides for the control of malaria transmitting Anopheles gambiae mosquitoes and rapid increase in resistance to this insecticide class is of major concern. Pyrethroids target the Voltage Gated Sodium Channels (VGSCs), that have a key role in the normal function of the mosquitoes' nervous system. VGSC mutations L995F and L995S have long been associated with pyrethroid resistance and screening for their presence is routine in insecticide resistance management programs. Recently, a VGSC haplotype containing two amino acid substitutions associated with resistance in other species, V402L and I1527T, was identified. These two VGSC mutations are found in tight linkage and are mutually exclusive to the classical L995F/S mutations. We identify the presence of the V402L-I1527T haplotype in resistant An. coluzzii colonized strains and in field populations from Burkina Faso, at frequencies higher than previously reported; in some cases almost reaching fixation. Functional validation of V402L in insecticide resistance using a CRISPR/Cas9 genome modified line showed that it confers reduced mortality after exposure to all tested pyrethroids and DDT, but at lower levels compared to L995F. In contrast to L995F however, no fitness costs were identified for mosquitoes carrying V402L under laboratory conditions. The V402L substitution confers pyrethroid resistance in An. gambiae in the absence of any other VGSC substitution and/or alternative resistance mechanisms. The lower fitness cost associated with this kdr mutation may provide a selective advantage over the classical kdr in some settings and genotyping at this locus should be added in the list of resistant alleles for routine screening.

First Sprayable Double-Stranded RNA-Based Biopesticide Product Targets Proteasome Subunit Beta Type-5 in Colorado Potato Beetle (Leptinotarsa decemlineata).

Colorado potato beetle (CPB, Leptinotarsa decemlineata) is a major pest of potato and other solanaceous vegetables in the Northern Hemisphere. The insect feeds on leaves and can completely defoliate crops. Because of the repeated use of single insecticide classes without rotating active ingredients, many chemicals are no longer effective in controlling CPB. Ledprona is a sprayable double-stranded RNA biopesticide with a new mode of action that triggers the RNA interference pathway. Laboratory assays with second instar larvae fed Ledprona showed a dose–response where 25×10−6g/L of dsPSMB5 caused 90% mortality after 6days of initial exposure. We also showed that exposure to Ledprona for 6h caused larval mortality and decreased target messenger RNA (mRNA) expression. Decrease in PSMB5 protein levels was observed after 48h of larval exposure to Ledprona. Both PSMB5 mRNA and protein levels did not recover over time. Ledprona efficacy was demonstrated in a whole plant greenhouse trial and performed similarly to spinosad. Ledprona, currently pending registration at EPA, represents a new biopesticide class integrated pest management and insecticide resistance management programs directed against CPB.

Malaria Transmission, Vector Diversity, and Insecticide Resistance at a Peri-Urban Site in the Forest Zone of Ghana

IntroductionRecent surge ofAnophelesresistance to major classes of World Health Organization (WHO)-approved insecticides globally necessitates the need for information about local malaria vector populations. It is believed that insecticide efficacy loss may lead to operational failure of control interventions and an increase in malaria infection transmission. We investigated the susceptibility levels of malaria vectors to all classes of WHO-approved vector control insecticides and described the dynamics of malaria transmission in a peri-urban setting.MethodsFit 3–5-day-old adults that emerged fromAnopheleslarvae collected from several different sites in the study area were subjected to the WHO bioassay for detecting insecticide resistance. The knockdown resistance gene (kdr) mutations within the vector populations were detected using PCR. Entomological inoculation rates were determined using the human landing catch technique andPlasmodium falciparumcircumsporozoite ELISA.ResultsThe malaria vectors from the study area were resistant to all classes of insecticides tested. Out of the 284Anophelescomplex specimen assayed for the resistance study, 265 (93.30%) were identified asAnopheles gambiae s.s.The kdr gene was detected in 90% of theAnopheles gambiae s.s.assayed. In an area whereAnopheles coluzziiresistance to insecticides had never been reported, the kdr gene was detected in 78% of theAnopheles coluzziisampled. The entomological inoculation rate (EIR) for the dry season was 1.44 ib/m/n, whereas the EIR for the rainy season was 2.69 ib/m/n.ConclusionsThis study provides information on the high parasite inoculation rate and insecticide resistance of malaria vectors in a peri-urban community, which is critical in the development of an insecticide resistance management program for the community.

Baseline Response, Monitoring, and Cross-Resistance of Spodoptera frugiperda (Lepidoptera: Noctuidae) to Sodium Channel Blocker Insecticides in Brazil

Spodoptera frugiperda (J.E. Smith) is one of the key cross-crop pests in Brazilian agroecosystems. Field-evolved resistance of S. frugiperda to some conventional insecticides and Bt proteins has already been reported. Thus, the use of insecticides with new mode of action such as sodium channel blockers (indoxacarb and metaflumizone) could be an important tool in insecticide resistance management (IRM) programs. To implement a proactive IRM, we conducted baseline response and monitoring to indoxacarb and metaflumizone in 87 field populations of S. frugiperda collected from major maize-growing regions of Brazil from 2017 to 2020, estimated the frequency of resistance alleles to indoxacarb, and evaluated cross-resistance of indoxacarb and metaflumizone to some selected insecticides and Bt proteins. Low variation in susceptibility to indoxacarb (4.6-fold) and metaflumizone (2.6-fold) was detected in populations of S. frugiperda in 2017. The frequency of the resistance allele to indoxacarb was 0.0452 (0.0382-0.0527 CI 95%), by using F2 screen method. The mean survival at diagnostic concentration, based on CL99, varied from 0.2 to 12.2% for indoxacarb and from 0.0 to 12.7% for metaflumizone, confirming high susceptibility of S. frugiperda to these insecticides in Brazil. No cross-resistance was detected between sodium channel blocker insecticides and other insecticides (organophosphate, pyrethroid, benzoylurea, spinosyn, and diamide) and Bt proteins. These findings showed that sodium channel blocker insecticides are important candidates to be exploited in IRM strategies of S. frugiperda in Brazil.