Temephos resistance and esterase activity in the mosquito Aedes aegypti in Havana, Cuba increased dramatically between 2006 and 2008

Aedes aegypti (L.) (Diptera: Culicidae) control programmes in Cuba rely on the application of the organophosphate temephos for larval control. Hence, the monitoring of resistance to this insecticide is an essential component of such programmes. Here, 15 field populations from different municipalities of Havana City were assayed for resistance to temephos. High levels of resistance were detected in all strains and resistance ratios were highly correlated with esterase activity (P = 0.00001). Populations from three municipalities were tested in both 2006 and 2008; resistance and esterase activities both significantly increased during this 2-year period. Synergist studies demonstrated that neither glutathione transferases nor monooxygenases were associated with the increase in resistance to temephos in this period. The duration of the efficacy of commercial formulations of temephos in controlling Ae. aegypti populations in Havana City was reduced by the high level of temephos resistance observed; hence these data are of clear operational significance for the dengue control programme in Cuba. New integrated strategies to avoid further increases in temephos resistance in Cuba are necessary.     

Adaptive larval thermotolerance and induced cross-tolerance to propoxur insecticide in mosquitoes Anopheles stephensi and Aedes aegypti

Abstract. Fourth-instar larvae of mosquitoes Anopheles stephensi and Aedes aegypti normally died within 90 min at 43C. Pre-exposure to high but sublethal temperatures conferred adaptive thermotolerance, dependent on the temperature and the duration of pre-exposure. Adaptive cross-tolerance to propoxur (a carbamate insecticide) was also induced in larvae by pre-exposing them to sublethal temperatures. Pre-exposure to sublethal concentrations of propoxur was found to confer cross-thermotolerance to a lower extent. These results suggest that the shock proteins (e.g. heat shock proteins) induced by unrelated stress factors play an important role in the development of adaptive cross-protection (stress response) to other stress conditions.     

Chronology of sodium channel mutations associated with pyrethroid resistance in Aedes aegypti

Aedes aegypti is the primary mosquito vector of dengue, yellow fever, Zika and chikungunya. Current strategies to control Ae. aegypti rely heavily on insecticide interventions. Pyrethroids are a major class of insecticides used for mosquito control because of their fast acting, highly insecticidal activities and low mammalian toxicity. However, Ae. aegypti populations around the world have begun to develop resistance to pyrethroids. So far, more than a dozen mutations in the sodium channel gene have been reported to be associated with pyrethroid resistance in Ae. aegypti. Co-occurrence of resistance-associated mutations is common in pyrethroid-resistant Ae. aegypti populations. As global use of pyrethroids in mosquito control continues, new pyrethroid-resistant mutations keep emerging. In this microreview, we compile pyrethroid resistance-associated mutations in Ae. aegypti in a chronological order, as they were reported, and summarize findings from functional evaluation of these mutations in an in vitro sodium channel expression system. We hope that the information will be useful for tracing possible evolution of pyrethroid resistance in this important human disease vector, in addition to the development of methods for global monitoring and management of pyrethroid resistance in Ae. aegypti.     

Insecticide resistance,associated mechanisms and fitness aspects in two Brazilian Stegomyia aegypti (= Aedes aegypti) populations

In Brazil, insecticide resistance in Stegomyia aegypti (= Aedes aegypti) (Diptera: Culicidae) populations to pyrethroids and to the organophosphate (OP) temephos is disseminated. Currently, insect growth regulators (IGRs) and the OP malathion are employed against larvae and adults, respectively. Bioassays with mosquitoes from two northeast municipalities, Crato and Aracaju, revealed, in both populations, susceptibility to IGRs and malathion (RR₉₅ ≤ 2.0), confirming the effectiveness of these compounds. By contrast, temephos and deltamethrin (pyrethroid) resistance levels were high (RR₉₅ > 10), which is consistent with the use of intense chemical control. In Crato, RR₉₅ values were > 50 for both compounds. Knock‐down‐resistant (kdr) mutants in the voltage‐gated sodium channel, the pyrethroid target site, were found in 43 and 32%, respectively, of Aracaju and Crato mosquitoes. Biochemical assays revealed higher metabolic resistance activity (esterases, mixed function oxidases and glutathione‐S‐transferases) at Aracaju. With respect to fitness aspects, mating effectiveness was equivalently impaired in both populations, but Aracaju mosquitoes showed more damaging effects in terms of longer larval development, decreased bloodmeal acceptance, reduced engorgement and lower numbers of eggs laid per female. Compared with mosquitoes in Crato, Aracaju mosquitoes exhibited lower OP and pyrethroid RR₉₅, increased activity of detoxifying enzymes and greater effect on fitness. The potential relationship between insecticide resistance mechanisms and mosquito viability is discussed.     

Effect of proteolytic and detoxification enzyme inhibitors on Bacillus thuringiensis var. israelensis tolerance in the mosquito Aedes aegypti

Bacillus thuringiensis var. israelensis (Bti) is highly pathogenic to mosquito larvae and is widely used for mosquito control. Its mosquitocidal activity however is relatively low compared to many chemical insecticides. The detoxification mechanisms in the mosquito, among other things, might neutralize the Bti activity, resulting in resistance or tolerance. We tested whether or not the detoxification mechanisms against chemical insecticides might also operate against Bti, rendering it less effective. We targeted four enzymes in Aedes aegypti larvae involved in detoxification with inhibitors that have been used in resistance studies in chemical insecticides and assayed their effects on Bti toxicity. Results revealed that phenylmethanesulphonyl fluoride (PMSF), diethyl maleate, phenobarbital (PB), and piperonyl butoxide (PBO) altered Bti toxicity to various degrees. PMSF is a serine protease inhibitor that prevents Bti digestion and improves Bti activity. PB that induces several detoxifying enzymes had two different effects depending on the method of treatment. Mortality was higher when treatment with PB was discontinuous (149%) whereas with continuous treatment it was lower (101%). PBO, a typical cytochrome P450 inhibitor, increased Bti effect (159%). The combination of discontinuous pretreatment of larvae with PB followed by PBO had a synergistic effect and showed increased activity (146%). It appears that the mechanism for Bti resistance in mosquitoes is similar to that of chemical insecticides. Our studies indicate that we may be able to increase Bti activity by inhibiting some of the detoxification systems as active as broad spectrum chemical insecticides.     

Competition and resistance to starvation in larvae of container-inhabiting Aedes mosquitoes

Abstract. 1. Hypotheses about declining populations of container-inhabiting Aedes mosquitoes following the invasion by additional species were tested.
2. The larval competition hypothesis was studied experimentally in pure and mixed cultures of Aedes aegypti (L.), A.albopictus (Skuse) and A.triseriatus (Say). The experiments used decomposing leaf litter in the laboratory, as opposed to most previous research which used non-natural food.
3. Resistance to starvation is introduced as a new measure of larval performance and competitiveness. The hypothesis is that more successful larvae store larger energy reserves and resist the lack of food longer.
4. Contrary to previous research showing better performance of A.aegypti in mixed cultures, A.albopictus developed faster and had greater survival when natural food was used.
5. Resistance to starvation was greater in the better performing species (i.e. A.aegypti with non-natural food and A.albopictus with leaf litter). Oxygen consumption by starved larvae was similar in the three container species, and in the ground-water mosquito, A.taeniorhynchus (Wied.), whose resistance to starvation was comparatively very low.     

Evaluation of biological and chemical insecticide mixture against Aedes aegypti larvae and adults by thermal fogging in Singapore

To improve the operational efficiency of dengue vector control in Singapore, larvicide and adulticide were applied together by thermal fog generator (Agrofog AF40). The mixture consisted of Bacillus thuringiensis var. israelensis (Vectobac 12 AS) as biological larvicide at 1.5 L/ha and pirimiphos-methyl (Actellic 50 EC) as adulticide at 100 g ai/ha, diluted 10-fold with water. Aerosol of this mixture was evaluated against the mosquito Aedes aegypti (L.) (Diptera: Culicidae) in bioassays using cages of 10 adult females exposed at heights of 0.3-2.4 m and distances of 3-12 m from the hand-held generator. Cups containing 200 mL water were treated at ground level by exposure to the aerosol application at the same distances from the generator. Subsequent larval bioassays on days 1, 7, 14, 21 and 28 post-spray involved exposing 20 larvae/cup for 48 h. Droplets had VMD 57 microm and female mosquitoes were killed by 2 s exposure to the aerosol at 3 m. We obtained 92-100% mortality of the adult mosquitoes and 100% control of larvae at 3 m distance, but only 10-13% mortality at 12 m from the fogger. In treated cups, larvae showed high mortality (92%) when exposed for 48 h even 1 month post-treatment. Results demonstrate the practical advantage of using this mixture of Vectobac 12AS and Actellic 50 EC for simultaneous control of Aedes adults and larvae, with prolonged larvicidal efficacy in treated containers.     

不同种群棉铃虫三龄幼虫酯酶活性频率分布与抗药性的关系

选用室内饲养的棉铃虫Helicoverpa armigera偃师和湖北2个敏感品系、对辛硫磷高抗的PCP20品系、对氰戊菊酯高抗的YG45品系及1999或2000年采自山东阳谷、河北邯郸和河南安阳的田间高抗种群,江苏徐州、湖北武汉的田间中等抗性水平种群和新疆沙湾的田间敏感种群,采用酶标板酶动力学法测定了各品系（种群）的3龄幼虫个体酯酶活性频率分布和平均酯酶活性。结果表明,偃师敏感品系、湖北敏感品系和新疆沙湾田间敏感种群的酯酶活性个体频率分布相似,三个品系（种群）的平均酯酶活性相近,分别为991、1 138、1 055 mOD·min^-1·larva^-1。室内选育的PCP20抗性品系、YG45抗性品系及山东阳谷、河北邯郸 、河南安阳田间高抗种群的高酯酶活性（活性在1 800 mOD·min^-1·larva^-1以上）个体频率明显高于三个敏感品系（种群）,平均酯酶活性在1 510～2 482 mOD·min^-1·larva^-1之间。江苏徐州、湖北武汉的田间中抗水平种群高酯酶活性个体频率及平均酯酶活性都介于敏感和高抗品系（种群）之间,平均酯酶活性为1 258～1.404 mOD·min^-1·larva^-1。棉铃虫各品系（种群）平均酯酶活性与对拟除虫菊酯类杀虫剂抗性个体频率的相关性要比对有机磷类的高,相关系数分别为0.82和0.42。分析各品系（种群）高酯酶活性个体频率与棉铃虫对拟除虫菊酯类、有机磷类杀虫剂抗性个体频率的相关性,得到相似的结果。考虑到酯酶并不是棉铃虫对拟除虫菊酯抗性的主要机理,建议酯酶活性可作为棉铃虫抗药性生化检测的一个参考指标。本文还讨论了酯酶与棉铃虫对拟除虫菊酯类杀虫剂及有机磷类杀虫剂抗性的关系。     

A comparison of mosquito densities,weather and infection rates of Aedes aegypti during the first epidemics of Chikungunya (2014) and Zika (2016) in areas with and without vector control in Puerto Rico

In Puerto Rico, the first records of the transmission of Chikungunya (CHIKV) and Zika (ZIKV) viruses were confirmed in May 2014 and December 2015, respectively. Transmission of CHIKV peaked in September 2014, whereas that of ZIKV peaked in August 2016. The emergence of these mosquito‐transmitted arboviruses in the context of a lack of human population immunity allowed observations of whether the outbreaks were associated with Aedes aegypti (Diptera: Culicidae) densities and weather. Mosquito density was monitored weekly in four communities using sentinel autocidal gravid ovitraps (AGO traps) during 2016 in order to provide data to be compared with the findings of a previous study carried out during the 2014 CHIKV epidemic. Findings in two communities protected against Ae. aegypti using mass AGO trapping (three traps per house in most houses) were compared with those in two nearby communities without vector control. Mosquito pools were collected to detect viral RNA of ZIKV, CHIKV and dengue virus. In areas without vector control, mosquito densities and rates of ZIKV detection in 2016 were significantly higher, similarly to those observed for CHIKV in 2014. The density of Ae. aegypti in treated sites was less than two females/trap/week, which is similar to the putative adult female threshold for CHIKV transmission. No significant differences in mosquito density or infection rates with ZIKV and CHIKV at the same sites between years were observed. Although 2016 was significantly wetter, mosquito densities were similar.     

Heterogeneous genetic invasions of three insecticide resistance mutations in Indo‐Pacific populations of Aedes aegypti (L.)

Nations throughout the Indo‐Pacific region use pyrethroid insecticides to control Aedes aegypti, the mosquito vector of dengue, often without knowledge of pyrethroid resistance status of the pest or origin of resistance. Two mutations (V1016G + F1534C) in the sodium channel gene (Vssc) of Ae. aegypti modify ion channel function and cause target‐site resistance to pyrethroid insecticides, with a third mutation (S989P) having a potential additive effect. Of 27 possible genotypes involving these mutations, some allelic combinations are never seen whereas others predominate. Here, five allelic combinations common in Ae. aegypti from the Indo‐Pacific region are described and their geographical distributions investigated using genome‐wide SNP markers. We tested the hypothesis that resistance allele combinations evolved de novo in populations versus the alternative that dispersal of Ae. aegypti between populations facilitated genetic invasions of allele combinations. We used latent factor mixed‐models to detect SNPs throughout the genome that showed structuring in line with resistance allele combinations and compared variation at SNPs within the Vssc gene with genome‐wide variation. Mixed‐models detected an array of SNPs linked to resistance allele combinations, all located within or in close proximity to the Vssc gene. Variation at SNPs within the Vssc gene was structured by resistance profile, whereas genome‐wide SNPs were structured by population. These results demonstrate that alleles near to resistance mutations have been transferred between populations via linked selection. This indicates that genetic invasions have contributed to the widespread occurrence of Vssc allele combinations in Ae. aegypti in the Indo‐Pacific region, pointing to undocumented mosquito invasions between countries.     

Changes in malathion and pyrethroid resistance after cypermethrin selection of Culex quinquefasciatus field populations of Cuba

Abstract. Use. of the organophosphorus insecticide malathion for mosquito control in Cuba, for 7 years up to 1986, selected elevated non-specific esterase and altered acetylcholinesterase (AChE) resistance mechanisms in Culex quinquefasciatus. In central Havana space-spraying of malathion was replaced by the pyrethroid cypermethrin in 1987: alternate cycles of malathion and cypermethrin were applied in some of the more rural areas of Havana district during 1987-91. Consequently, populations of Cx quinquefasciatus in the central area of Havana developed resistance to cypermethrin, but there is no evidence of pyrethroid resistance in the outlying areas. Malathion resistance levels declined significantly after 1986, measured both by bioassay and the frequency of the elevated esterase resistance mechanism, and then stabilized with no measurable decline during 1990 in any of the populations tested. These populations had less than 10% frequency of susceptible homozygotes for both the esterase and AChE resistance mechanisms, indicating that organophosphate resistance is still prevalent in Cuban Cx quinquefasciatus. These two mechanisms appear to be in linkage equilibrium, suggesting that current selection for double resistance is not strong. In the central Havana region, pirimiphos-methyl, an organophosphorus insecticide unaffected by the two common malathion resistance mechanisms, is now being used in a resistance management strategy designed to avoid pyrethroid resistance spreading.     

昆虫钠离子通道基因突变及其与杀虫剂抗性关系的研究进展

昆虫电压门控钠离子通道(voltage-gated sodium channel)存在于所有可兴奋细胞的细胞膜上,在动作电位的产生和传导上起重要作用,是有机氯和拟除虫菊酯杀虫剂的靶标位点。在农业和医学害虫控制过程中,由于有机氯和拟除虫菊酯杀虫剂的广泛使用,抗药性问题日益突出。其中,由于钠离子通道基因突变,降低了钠离子通道对有机氯和拟除虫菊酯类杀虫剂的亲和性,从而产生击倒抗性(knock-down resistance, kdr),已成为抗性产生的重要机制之一。本文综述了昆虫钠离子通道的跨膜拓扑结构、功能、进化及其基因的克隆;更重要的是总结了已报道的40多种昆虫40个钠离子通道基因非同义突变,以及钠离子通道基因选择性mRNA剪接和编辑,以及它们与杀虫剂抗性的关系;也评述了钠离子通道基因突变引起蛋白质结构的改变,从而对杀虫剂抗性的影响机制。这些研究对于进一步鉴定与杀虫剂抗性相关的突变及抗性机制,开发有机氯和拟除虫菊酯类杀虫剂抗性分子监测方法具有重要意义。     

Surveillance of the mosquito Aedes aegypti and its biocontrol with the copepod Mesocyclops aspericornis in Australian wells and gold mines

Abstract. A survey of the dengue vector mosquito Aedes aegypti was undertaken using runnel traps to detect immature stages (larvae and pupae) in flooded disused mine shafts and wells in Charters Towers, Queensland, Northern Australia. The town has a history of dengue fever since 1885 when goldminers were the first recorded victims. During the latest dengue epidemic in 1993, 2% of the population had laboratory-confirmed dengue virus Type 2, despite source reduction of Ae.aegypti breeding-sites at ground level or above. This led to suspicions that dengue vector Ae.aegypti breeding-sites might be below ground level. When surveyed in March 1994, Ae.aegypti immatures were found in 9/10 wells and 1/6 mine shafts. The water in wells and mines had similar characteristics -except that turbidity was higher in the mines, which more often contained predators of mosquito immatures.
The copepod Mesocyclops aspericornis was collected from water in 1/10 wells and 2/6 mine shafts. Laboratory predation trials resulted in 95.5–100% predation by 25 copepods/1 on Ae.aegypti first-instar larvae up to 200 larvae/1. Five wells containing Ae.aegypti in the survey were inoculated with fifty indigenous M.aspericornis , and five wells (one positive and four negative in the survey) were left untreated as controls. Nine months later, in December 1994, Ae.aegypti had been eliminated from all five treated wells but all untreated control wells contained Ae.aegypti , except for one well that contained a natural population of M.aspericornis. The role of wells and mines as winter/ dry season refuges of Ae.aegypti in northern Australia is reviewed, and we recommend the use of M.aspericornis as a cost-effective, environmentally acceptable and persistent agent for the sustainable control of Ae.aegypti , especially in inaccessible breeding sites.     

Larval sites of the mosquito Aedes aegypti formosus in forest and domestic habitats in Africa and the potential association with oviposition evolution

Adaptations to anthropogenic domestic habitats contribute to the success of the mosquito Aedes aegypti as a major global vector of several arboviral diseases. The species inhabited African forests before expanding into domestic habitats and spreading to other continents. Despite a well‐studied evolutionary history, how this species initially moved into human settlements in Africa remains unclear. During this initial habitat transition, African Ae. aegypti switched their larval sites from natural water containers like tree holes to artificial containers like clay pots. Little is known about how these natural versus artificial containers differ in their characteristics. Filling this knowledge gap could provide valuable information for studying the evolution of Ae. aegypti associated with larval habitat changes. As an initial effort, in this study, we characterized the microenvironments of Ae. aegypti larval sites in forest and domestic habitats in two African localities: La Lopé, Gabon, and Rabai, Kenya. Specifically, we measured the physical characteristics, microbial density, bacterial composition, and volatile chemical profiles of multiple larval sites. In both localities, comparisons between natural containers in the forests and artificial containers in the villages revealed significantly different microenvironments. We next examined whether the between‐habitat differences in larval site microenvironments lead to differences in oviposition, a key behavior affecting larval distribution. Forest Ae. aegypti readily accepted the artificial containers we placed in the forests. Laboratory choice experiments also did not find distinct oviposition preferences between forest and village Ae. aegypti colonies. These results suggested that African Ae. aegypti are likely generalists in their larval site choices. This flexibility to accept various containers with a wide range of physical, microbial, and chemical conditions might allow Ae. aegypti to use human‐stored water as fallback larval sites during dry seasons, which is hypothesized to have initiated the domestic evolution of Ae. aegypti.     

Changes in emergence phenology,fatty acid composition,and xenobiotic-metabolizing enzyme expression is associated with increased insecticide resistance in the Colorado potato beetle

The Colorado potato beetle (Leptinotarsa decemlineata) is a major agricultural pest of solanaceous crops. An effective management strategy employed by agricultural producers to control this pest species is the use of systemic insecticides. Recent emphasis has been placed on the use of neonicotinoid insecticides. Despite efforts to curb resistance development through integrated pest management approaches, resistance to neonicotinoids in L. decemlineata populations continues to increase. One contributing factor may be alterations in insect fatty acids, which have multiple metabolic functions and are associated with the synthesis of xenobiotic-metabolizing enzymes to mitigate the effects of insecticide exposure. In this study, we analyzed the fatty acid composition of L. decemlineata populations collected from an organic production field and from a commercially managed field to determine if fatty acid composition varied between the two populations. We demonstrate that a population of L. decemlineata that has a history of systemic neonicotinoid exposure (commercially managed) has a different lipid composition and differential expression of known metabolic detoxification mechanisms relative to a population that has not been exposed to neonicotinoids (organically managed). The fatty acid data indicated an upregulation of Δ⁶ desaturase in the commercially managed L. decemlineata population and suggest a role for eicosanoids and associated metabolic enzymes as potential modulators of insecticide resistance. We further observed a pattern of delayed emergence within the commercially managed population compared with the organically managed population. Variations in emergence timing together with specific fatty acid regulation may significantly influence the capacity of L. decemlineata to develop insecticide resistance.     

天津地区家蝇抗药性水平及与两种解毒酶的关系

用点滴法对采自天津市7个郊区县的7个不同生境野外家蝇种群的抗药性进行了测定并与实验室内的相对敏感种群进行比较;对不同地区家蝇种群进行了羧酸酯酶（carboxylesterase, CarE）和谷胱甘肽S-转移酶（glutathione S-transferase, GSTs）活性检测。结果表明,不同家蝇种群对DDVP、高效氯氰菊酯和残杀威的抗性倍数不同,对DDVP抗性最高为18.563倍,最低为1.885倍;对高效氯氰菊酯最高为14.071倍,最低为1.071倍;对残杀威抗性倍数最高为7.499倍,最低为1.071倍。从CarE平均比活力看,室外家蝇种群CarE的比活力均高于室内相对敏感种群的比活力,CarE活性的分布在敏感种群和不同地区野外种群间具有明显的重叠现象。天津不同地区F₂代家蝇GSTs的比活力抗性种群普遍高于室内相对敏感种群,大约在2～4倍之间,不同地区间也具有比较大的差异,而抗药性高的种群GSTs活性也高,GSTs活性变化与不同种群抗性差异是相符的。     

Characterising the morphological characters and carbohydrate metabolism of oat culms and their association with lodging resistance

• Lodging resistance can be improved by enhancing the mechanical strength of culms, and culm carbohydrates could improve this mechanical strength. Culm carbohydrates can regulate development of the culm and affect its toughness.
• The present study determines the relationship between lodging and carbohydrate content in oat culms. Field experiments were conducted in alpine regions in 2017 and 2018 using three oat varieties with different lodging resistance. Lodging‐related morphological characteristics were directly determined and culm carbohydrate content and enzyme activity related to cellulose synthesis and sucrose metabolism were evaluated with ultraviolet spectrophotometry.
• Results showed that the lower the gravity height or the lower ratio of gravity height to plant height, the stronger the lodging resistance of the varieties. Higher culm nonstructural (NSC) and structural (SC) carbohydrate content contributed to the ability of culms to resist lodging, especially the content of cellulose and sucrose. PCA showed that sucrose metabolism and SC content were closely related to lodging resistance. Correlation analysis showed that the lodging index (LI) was significantly negatively correlated with NSC. Sucrose content was highly and significantly positively correlated with NSC. Additionally, the activities of sucrose phosphate synthase (SPS) and sucrose synthase (SS) were highly and significantly positively correlated with sucrose and cellulose content.
• The relationship between field characters and oat lodging, as well as the regulatory mechanism of carbohydrate content on lodging resistance of the culm are discussed.