scholarly journals First detection of the kdr mutation (L1014F) in the plague vector Xenopsylla cheopis (Siphonaptera: Pulicidae)

2019 ◽  
Vol 12 (1) ◽  
Author(s):  
Nian Liu ◽  
Xiangyang Feng ◽  
Mei Li ◽  
Xinghui Qiu
Keyword(s):  
Network Analysis ◽  
Insecticide Resistance ◽  
Dna Sequences ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Effective Control ◽  
Phenotypic Effect ◽  
Pyrethroid Insecticides ◽  
Xenopsylla Cheopis ◽  
Flea Control

Abstract Background The oriental rat flea, Xenopsylla cheopis, is the most efficient vector of the plague. Pyrethroid insecticides such as cypermethrin, cyhalothrin and deltamethrin have been often used to limit plague transmission via controlling the vector during outbreaks. However, this strategy is threatened by the development of insecticide resistance. Understanding the mechanisms underlying pyrethroid resistance is the prerequisite for successful flea control. Methods Partial DNA sequences of X. cheopis voltage gated sodium channel (VGSC) gene were amplified from a total of 111 individuals, collected from a natural plague epidemic foci in Baise City, Guangxi Zhuang Autonomous Region of China. These DNA fragments were sequenced. The frequency and distribution of kdr mutations were assessed in four X. cheopis populations. The origin of kdr mutations was investigated by phylogenetic and network analysis. Results The classical knockdown resistance (kdr) mutation (L1014F) was detected in four field populations at frequencies ranging between 0.021–0.241. The mutant homozygote was observed only in one of the four populations. Seven haplotypes were identified, with two of them carrying the resistance L1014F mutation. Phylogenetic tree and network analysis indicated that the L1014F allele was not singly originated. Based on polymerase chain reaction restriction fragment length polymorphism (PCR-RFLP) profiling, an easy-to-use and accurate molecular assay for screening individual fleas for the L1014F mutation was developed. Conclusions To our knowledge, this work represents the first report of the L1014F mutation in the plague vector X. cheopis. The incidence of the L1014F allele highlights the need of further studies on the phenotypic effect of this mutation in this plague vector. Early detection and monitoring of insecticide resistance is suggested in order to make effective control strategies in case of plague outbreaks in this region.

scholarly journals Functional genetic validation of key genes conferring insecticide resistance in the major African malaria vector, Anopheles gambiae

2019 ◽  
Author(s):  
Adriana Adolfi ◽  
Beth Poulton ◽  
Amalia Anthousi ◽  
Stephanie Macilwee ◽  
Hilary Ranson ◽  
...  
Keyword(s):  
Public Health ◽  
Insecticide Resistance ◽  
Anopheles Gambiae ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Resistance Mechanisms ◽  
Effective Control ◽  
Cross Resistance ◽  
Toxic Metabolite ◽  
Control Programs

ABSTRACTResistance in Anopheles gambiae to members of all four major classes (pyrethroids, carbamates, organochlorines and organophosphates) of public health insecticides limits effective control of malaria transmission in Africa. Increased expression of detoxifying enzymes has been associated with resistance, but direct functional validation in An. gambiae has been lacking. Here we perform transgenic analysis using the GAL4/UAS system to examine insecticide resistance phenotypes conferred by increased expression of the three genes - Cyp6m2, Cyp6p3 and Gste2 - most often found upregulated in resistant An. gambiae. We report the first evidence in An. gambiae that organophosphate and organochlorine resistance is conferred by overexpression of GSTE2 in a broad tissue profile. Pyrethroid and carbamate resistance is bestowed by similar Cyp6p3 overexpression, and Cyp6m2 confers only pyrethroid resistance when overexpressed in the same tissues. Conversely, such Cyp6m2 overexpression increases susceptibility to the organophosphate malathion, presumably due to conversion to a more toxic metabolite. No resistant phenotypes are conferred when either Cyp6 gene overexpression is restricted to the midgut or oenocytes, answering long standing questions related to the importance of these tissues in resistance to contact insecticides. Validation of genes conferring resistance provides markers to guide control strategies, and the observed negative cross-resistance due to Cyp6m2 gives credence to proposed dual insecticide strategies to overcome pyrethroid resistance. These trasnsgenic An. gambiae resistant lines are being used to test potential liabilities in new active compounds early in development.SIGNIFICANCE STATEMENTInsecticide resistance in Anopheles gambiae mosquitoes can derail malaria control programs, and to overcome it we need to discover the underlying molecular basis. Here, for the first time, we characterise three genes most often associated with insecticide resistance directly by their overproduction in genetically modified An. gambiae. We show that overexpression of each gene confers resistance to representatives of at least one insecticide class and, taken together, the three genes provide cross-resistance to all four major insecticide classes currently used in public health. These data validate the candidate genes as markers to monitor the spread of resistance in mosquito populations. The modified mosquitoes produced are also valuable tools to pre-screen new insecticides for potential liabilities to existing resistance mechanisms.

scholarly journals Functional genetic validation of key genes conferring insecticide resistance in the major African malaria vector,Anopheles gambiae

2019 ◽  
Vol 116 (51) ◽  
pp. 25764-25772 ◽  
Author(s):  
Adriana Adolfi ◽  
Beth Poulton ◽  
Amalia Anthousi ◽  
Stephanie Macilwee ◽  
Hilary Ranson ◽  
...  
Keyword(s):  
Insecticide Resistance ◽  
Anopheles Gambiae ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Effective Control ◽  
Cross Resistance ◽  
Toxic Metabolite ◽  
Detoxifying Enzymes ◽  
Resistant Lines ◽  
Key Genes

Resistance inAnopheles gambiaeto members of all 4 major classes (pyrethroids, carbamates, organochlorines, and organophosphates) of public health insecticides limits effective control of malaria transmission in Africa. Increase in expression of detoxifying enzymes has been associated with insecticide resistance, but their direct functional validation inAn. gambiaeis still lacking. Here, we perform transgenic analysis using the GAL4/UAS system to examine insecticide resistance phenotypes conferred by increased expression of the 3 genes—Cyp6m2,Cyp6p3, andGste2—most often found up-regulated in resistantAn. gambiae. We report evidence inAn. gambiaethat organophosphate and organochlorine resistance is conferred by overexpression of GSTE2 in a broad tissue profile. Pyrethroid and carbamate resistance is bestowed by similarCyp6p3overexpression, andCyp6m2confers only pyrethroid resistance when overexpressed in the same tissues. Conversely, suchCyp6m2overexpression increases susceptibility to the organophosphate malathion, presumably due to conversion to the more toxic metabolite, malaoxon. No resistant phenotypes are conferred when eitherCyp6gene overexpression is restricted to the midgut or oenocytes, indicating that neither tissue is involved in insecticide resistance mediated by the candidate P450s examined. Validation of genes conferring resistance provides markers to guide control strategies, and the observed negative cross-resistance due toCyp6m2gives credence to proposed dual-insecticide strategies to overcome pyrethroid resistance. These transgenicAn. gambiae-resistant lines are being used to test the “resistance-breaking” efficacy of active compounds early in their development.

scholarly journals Insecticide resistance status of Aedes aegypti in Bangladesh

2020 ◽  
Vol 13 (1) ◽  
Author(s):  
Hasan Mohammad Al-Amin ◽  
Fatema Tuj Johora ◽  
Seth R. Irish ◽  
Muhammad Riadul Haque Hossainey ◽  
Lucrecia Vizcaino ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Insecticide Resistance ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Resistance Mechanisms ◽  
Capital City ◽  
Detoxification Enzymes ◽  
Knockdown Resistance ◽  
Metabolic Resistance ◽  
Pyrethroid Insecticides

Abstract Background Arboviral diseases, including dengue and chikungunya, are major public health concerns in Bangladesh where there have been unprecedented levels of transmission reported in recent years. The primary approach to control these diseases is to control the vector Aedes aegypti using pyrethroid insecticides. Although chemical control has long been practiced, no comprehensive analysis of Ae. aegypti susceptibility to insecticides has been conducted to date. The aim of this study was to determine the insecticide resistance status of Ae. aegypti in Bangladesh and investigate the role of detoxification enzymes and altered target site sensitivity as resistance mechanisms. Methods Eggs of Aedes mosquitoes were collected using ovitraps from five districts across Bangladesh and in eight neighborhoods of the capital city Dhaka, from August to November 2017. CDC bottle bioassays were conducted for permethrin, deltamethrin, malathion, and bendiocarb using 3- to 5-day-old F0–F2 non-blood-fed female mosquitoes. Biochemical assays were conducted to detect metabolic resistance mechanisms, and real-time PCR was performed to determine the frequencies of the knockdown resistance (kdr) mutations Gly1016, Cys1534, and Leu410. Results High levels of resistance to permethrin were detected in all Ae. aegypti populations, with mortality ranging from 0 to 14.8% at the diagnostic dose. Substantial resistance continued to be detected against higher (2×) doses of permethrin (5.1–44.4% mortality). Susceptibility to deltamethrin and malathion varied between populations while complete susceptibility to bendiocarb was observed in all populations. Significantly higher levels of esterase and oxidase activity were detected in most of the test populations as compared to the susceptible reference Rockefeller strain. A significant association was detected between permethrin resistance and the presence of Gly1016 and Cys1534 homozygotes. The frequency of kdr (knockdown resistance) alleles varied across the Dhaka Aedes populations. Leu410 was not detected in any of the tested populations. Conclusions The detection of widespread pyrethroid resistance and multiple resistance mechanisms highlights the urgency for implementing alternate Ae. aegypti control strategies. In addition, implementing routine monitoring of insecticide resistance in Ae. aegypti in Bangladesh will lead to a greater understanding of susceptibility trends over space and time, thereby enabling the development of improved control strategies.

scholarly journals Voltage-Gated Sodium Channel (Vgsc) Mutation-Based Pyrethroid Resistance in Aedes aegypti Populations of Three Endemic Dengue Risk Areas of Sri Lanka

2021 ◽  
Vol 2021 ◽  
pp. 1-10
Author(s):  
Tharaka Ranathunge ◽  
Lahiru Udayanga ◽  
Sumudu Sarasija ◽  
Samudra Karunathilaka ◽  
Shavindhya Nawarathne ◽  
...  
Keyword(s):  
Sri Lanka ◽  
Insecticide Resistance ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Resistance Mechanisms ◽  
Allele Frequencies ◽  
Multiple Use ◽  
Pyrethroid Insecticides ◽  
Dengue Control ◽  
Mutation Allele

Background. Pyrethroid insecticides are widely used in many countries for chemical-based control of Ae. aegypti. Regardless of their efficacy, the constant use of insecticides has induced insecticide resistance mechanisms, such as knockdown resistance (kdr) in mosquitoes. Sri Lankan Vector Controlling Entities (VCE) have been using a variety of pyrethroid insecticides as the primary approach for dengue control. However, development of any resistance among the Aedes mosquitoes has been limitedly studied in the country. Therefore, the current study was conducted to evaluate the prevalence of F1534C, V1016G, and S989P mutations among Ae. aegypti mosquito populations in three dengue endemic high-risk regions of Sri Lanka. Methodology. Immature (both pupae and larvae) stages of Ae. aegypti mosquitoes were collected from Colombo, Gampaha, and Kandy districts of Sri Lanka from February 2018 to December 2019. Polymerase Chain Reaction- (PCR-) based assay for molecular genotyping of mutations was performed to identify the prevalence of kdr mutations in collected Ae. aegypti populations, separately. The frequencies of the resistant and susceptible kdr alleles were determined by using the Hardy–Weinberg equilibrium. Results. The Ae. aegypti populations from Colombo, Gampaha, and Kandy districts showed 46%, 42%, and 22% of F1534C mutation allele frequencies, along with 15%, 12%, and 6% of V1016G mutation allele frequencies, respectively. The mutation allele frequencies of S989 in Colombo, Gampaha, and Kandy districts were 9.5%, 8.5%, and 4.5%, respectively. The wild-type (PP) genotype remained predominant within all the three districts, whereas the homogenous (QQ) mutation genotype occurred only in minority. The abundance of Q allele frequency in Ae. aegypti mosquitoes was relatively higher for all the three mutations in Colombo. Conclusions. The findings clearly indicate that long-term insecticide applications and multiple use of pyrethroids have led to the acquisition of kdr mutations, leading to the development of insecticide resistance among local Ae. aegypti populations, especially in the Colombo and Gampaha districts. Therefore, evaluation of the prevalence levels of these kdr mutations highlights the necessity for shifting towards novel vector control strategies.

scholarly journals Insecticide resistance status of Aedes aegypti in Bangladesh

2020 ◽  
Author(s):  
Hasan Mohammad Al-Amin ◽  
Fatema Tuj Johora ◽  
Seth R. Irish ◽  
Muhammad Riadul Haque Hossainey ◽  
Lucrecia Vizcaino ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Insecticide Resistance ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Resistance Mechanisms ◽  
Capital City ◽  
Detoxification Enzymes ◽  
Health Concern ◽  
Metabolic Resistance ◽  
Pyrethroid Insecticides

AbstractBackgroundArboviral diseases including dengue and chikungunya are major public health concern in Bangladesh, with unprecedented levels of transmission reported in recent years. The primary approach to control these diseases is control of Aedes aegypti using pyrethroid insecticides. Although chemical control is long-practiced, no comprehensive analysis of Ae. aegypti susceptibility to insecticides has previously been conducted. This study aimed to determine the insecticide resistance status of Ae. aegypti in Bangladesh and investigate the role of detoxification enzymes and altered target site sensitivity as resistance mechanisms.MethodsAedes eggs were collected using ovitraps from five districts across the country and in eight neighborhoods of the capital city Dhaka from August to November 2017. CDC bottle bioassays were conducted for permethrin, deltamethrin, malathion, and bendiocarb using 3-5-day old F0-F2 non-blood fed female mosquitoes. Biochemical assays were conducted to detect metabolic resistance mechanisms and real-time PCR was performed to determine the frequencies of the knockdown resistance (kdr) mutations Gly1016, Cys1534, and Leu410.ResultsHigh levels of resistance to permethrin were detected in all Ae. aegypti populations, with mortality ranging from 0 – 14.8% at the diagnostic dose. Substantial resistance continued to be detected against higher (2X) doses of permethrin (5.1 – 44.4% mortality). Susceptibility to deltamethrin and malathion varied between populations while complete susceptibility to bendiocarb was observed in all populations. Significantly higher levels of esterase and oxidase activity were detected in most of the test populations as compared to the susceptible reference Rockefeller strain. A significant association was detected between permethrin resistance and the presence of Gly1016 and Cys1534 homozygotes. The frequency of kdr alleles varied across the Dhaka populations, and Leu410 was not detected in any of the tested populations.ConclusionsThe detection of widespread pyrethroid resistance and multiple mechanisms highlights the urgency for implementing alternate Ae. aegypti control strategies. In addition, implementing routine monitoring of insecticide resistance in Ae. aegypti in Bangladesh will lead to a greater understanding of susceptibility trends over space and time, thereby enabling the development of improved control strategies.

scholarly journals Frequency of kdr mutations in the voltage-sensitive sodium channel (VSSC) gene in Aedes aegypti from Yogyakarta and implications for Wolbachia-infected mosquito trials

2020 ◽  
Author(s):  
Juli R. Wuliandari ◽  
Ary A. Hoffmann ◽  
Warsito Tantowijoyo ◽  
Nancy M. Endersby-Harshman
Keyword(s):  
Aedes Aegypti ◽  
Inner City ◽  
Insecticide Resistance ◽  
Sodium Channel ◽  
Pyrethroid Resistance ◽  
Infected Mosquito ◽  
Simultaneous Occurrence ◽  
Pyrethroid Insecticides ◽  
Background Population ◽  
Homozygous Mutant

Abstract Background : In the inner city of Yogyakarta, Indonesia, insecticide resistance is expected in the main dengue vector, Aedes aegypti , because of the intensive local application of pyrethroid insecticides. However, detailed information about the nature of resistance in this species is required to assist the release of Wolbachia mosquitoes in a dengue control program so that we can ensure that insecticide resistance in the strain of Ae. aegypti being released matches that of the background population. Methods: High-resolution melt genotyping was used to screen for kdr mutations associated with pyrethroid resistance in the voltage-sensitive sodium channel (V SSC ) gene in Ae. aegypti of some areas in the inner city of Yogyakarta. Results: The results show that the V1016G mutation predominated, with individuals homozygous for the 1016G allele at a frequency of 82.1% and the mutant allele G at a frequency of 92%. Two patterns of co-occurrence of mutations were detected in this study, homozygous individuals V1016G/S989P; and heterozygous individuals V1016G/F1534C/S989P. We found the simultaneous occurrence of kdr mutations V1016G and F1534C at all collection sites, but not within individual mosquitoes. Homozygous mutants at locus 1016 were homozygous wildtype at locus 1534 and vice versa, and heterozygous V1016G were also heterozygous for F1534C. The most common tri-locus genotype co-occurrences were homozygous mutant 1016GG and homozygous wild-type FF1534, combined with homozygous mutant 989PP (GG/FF/PP) at a frequency of 38.28%. Conclusions: Given the relatively small differences in frequency of resistance alleles across the city area, locality variations in resistance should have minor implications for the success of Wolbachia mosquito trials being undertaken in the Yogyakarta area.

scholarly journals Frequency of kdr mutations in the voltage-sensitive sodium channel (VSSC) gene in Aedes aegypti from Yogyakarta and implications for Wolbachia-infected mosquito trials

2020 ◽  
Author(s):  
Juli R. Wuliandari ◽  
Ary A. Hoffmann ◽  
Warsito Tantowijoyo ◽  
Nancy M. Endersby-Harshman
Keyword(s):  
Inner City ◽  
Insecticide Resistance ◽  
Sodium Channel ◽  
Pyrethroid Resistance ◽  
Infected Mosquito ◽  
Simultaneous Occurrence ◽  
Wild Type ◽  
Pyrethroid Insecticides ◽  
Background Population ◽  
Homozygous Mutant

Abstract Background: In the inner city of Yogyakarta, Indonesia, insecticide resistance is expected in the main dengue vector, Aedes aegypti, because of the intensive local application of pyrethroid insecticides. However, detailed information about the nature of resistance in this species is required to assist the release of Wolbachia mosquitoes in a dengue control program, so that we can ensure that insecticide resistance in the strain of Ae. aegypti being released matches that of the background population.Methods: High-resolution melt genotyping was used to screen for kdr mutations associated with pyrethroid resistance in the voltage-sensitive sodium channel (VSSC) gene in Ae. aegypti of some areas in the inner city of Yogyakarta.Results: The results show that the V1016G mutation predominated, with individuals homozygous for the 1016G allele at a frequency of 82.1% and the mutant allele G at a frequency of 92%. Two patterns of co-occurrence of mutations were detected in this study, homozygous individuals V1016G/S989P; and heterozygous individuals V1016G/F1534C/S989P. We found the simultaneous occurrence of kdr mutations V1016G and F1534C at all collection sites, but not within individual mosquitoes. Homozygous mutants at locus 1016 were homozygous wild-type at locus 1534 and vice versa, and heterozygous V1016G were also heterozygous for F1534C. The most common tri-locus genotype co-occurrences were homozygous mutant 1016GG and homozygous wild-type FF1534, combined with homozygous mutant 989PP (GG/FF/PP) at a frequency of 38.28%.Conclusions: Given the relatively small differences in frequency of resistance alleles across the city area, locality variations in resistance should have minor implications for the success of Wolbachia mosquito trials being undertaken in the Yogyakarta area.

scholarly journals Mechanisms associated with pyrethroid resistance in populations of Aedes aegypti (Diptera: Culicidae) from the Caribbean coast of Colombia

2020 ◽  
Author(s):  
Paula X. Pareja-Loaiza ◽  
Liliana Santacoloma Varon ◽  
Gabriela Rey Vega ◽  
Doris Gómez-Camargo ◽  
Ronald Maestre-Serrano ◽  
...  
Keyword(s):  
Aedes Aegypti ◽  
Selection Pressure ◽  
Pyrethroid Resistance ◽  
Control Strategies ◽  
Activity Levels ◽  
Public Health Importance ◽  
Caribbean Coast ◽  
Pyrethroid Insecticides ◽  
The Caribbean ◽  
Great Public Health

AbstractAedes aegypti is the main vector of dengue, chikungunya, and Zika viruses, which are of great public health importance in Colombia. Aedes control strategies in Colombia rely heavily on the use of organophosphate and pyrethroid insecticides, providing constant selection pressure and the emergence of resistant populations. In recent years, insecticide use has increased due to the increased incidence of dengue and recent introductions of chikungunya and Zika. In the present study, pyrethroid resistance was studied across six populations of A. aegypti from the Caribbean coast of Colombia. Susceptibility to λ-cyhalothrin, deltamethrin, and permethrin was assessed, and resistance intensity was determined. Activity levels of enzymes associated with resistance were measured, and the frequencies of three kdr alleles (V1016I, F1534C, V410L) were calculated. Results showed variations in pyrethroid susceptibility across A. aegypti populations and altered enzyme activity levels were detected. The kdr alleles were detected in all populations, with high variations in frequencies: V1016I (frequency ranging from 0.15–0.70), F1534C (range 0.94–1.00), and V410L (range 0.05–0.72). In assays of phenotyped individuals, associations were observed between the presence of V1016I, F1534C, and V410L alleles and resistance to the evaluated pyrethroids, as well as between the VI1016/CC1534/VL410 tri-locus genotype and λ-cyhalothrin and permethrin resistance. The results of the present study contribute to the knowledge of the mechanisms underlying the resistance to key pyrethroids used to control A. aegypti along the Caribbean coast of Colombia.

scholarly journals Frequency of kdr mutations in the voltage-sensitive sodium channel (VSSC) gene in Aedes aegypti from Yogyakarta and implications for Wolbachia-infected mosquito trials

2020 ◽  
Author(s):  
Juli R. Wuliandari ◽  
Ary A. Hoffmann ◽  
Warsito Tantowijoyo ◽  
Nancy M. Endersby-Harshman
Keyword(s):  
Aedes Aegypti ◽  
Inner City ◽  
Insecticide Resistance ◽  
Sodium Channel ◽  
Pyrethroid Resistance ◽  
Infected Mosquito ◽  
Simultaneous Occurrence ◽  
High Resolution Melt ◽  
Pyrethroid Insecticides ◽  
Homozygous Mutant

Abstract Background: In the inner city of Yogyakarta, Indonesia, insecticide resistance is expected in the main dengue vector, Aedes aegypti, because of the intensive local application of pyrethroid insecticides. However, detailed information about the nature of resistance in this species is required to assist the release of Wolbachia mosquitoes in a dengue control program so that we can ensure that insecticide resistance in the strain of Ae. aegypti being released matches that of the background population.Methods: High-resolution melt genotyping was used to screen for kdr mutations associated with pyrethroid resistance in the voltage-sensitive sodium channel (VSSC) gene in Ae. aegypti of some areas in the inner city of Yogyakarta.Results: The results show that the V1016G mutation predominated, with individuals homozygous for the 1016G allele at a frequency of 82.1% and the mutant allele G at a frequency of 92%. Two patterns of co-occurrence of mutations were detected in this study, homozygous individuals V1016G/S989P; and heterozygous individuals V1016G/F1534C/S989P. We found the simultaneous occurrence of kdr mutations V1016G and F1534C at all collection sites, but not within individual mosquitoes. Homozygous mutants at locus 1016 were homozygous wildtype at locus 1534 and vice versa, and heterozygous V1016G were also heterozygous for F1534C. The most common tri-locus genotype co-occurrences were homozygous mutant 1016GG and homozygous wild-type FF1534, combined with homozygous mutant 989PP (GG/FF/PP) at a frequency of 38.28%.Conclusions: Given the relatively small differences in frequency of resistance alleles across the city area, locality variations in resistance should have minor implications for the success of Wolbachia mosquito trials being undertaken in the Yogyakarta area.

scholarly journals Identification of a rapidly-spreading triple mutant for high-level metabolic insecticide resistance in Anopheles gambiae provides a real-time molecular diagnostic for anti-malarial intervention deployment

2021 ◽  
Author(s):  
Harun Njoroge ◽  
Arjen van’t Hof ◽  
Ambrose Oruni ◽  
Dimitra Pipini ◽  
Sanjay C. Nagi ◽  
...  
Keyword(s):  
Insecticide Resistance ◽  
Anopheles Gambiae ◽  
Pyrethroid Resistance ◽  
Border Region ◽  
Malaria Vectors ◽  
Molecular Diagnostic ◽  
Triple Mutant ◽  
Pyrethroid Insecticides ◽  
High Level ◽  
Mutant Haplotype

AbstractInsecticide resistance provides both an increasingly pressing threat to the control of vector-borne diseases and insights into the remarkable capacity of natural populations to show rapid evolutionary responses to contemporary selection. Malaria control remains heavily dependent on deployment of pyrethroid insecticides, primarily in long lasting insecticidal nets (LLINs), but resistance in the major malaria vectors has increased over the last 15 years in concert with dramatic expansion of LLIN distributions. Identifying genetic mechanisms underlying high-level resistance in mosquitoes, which may almost entirely overcome pyrethroid efficacy, is crucial for the development and deployment of potentially resistance-breaking tools. Using the Anopheles gambiae 1000 genomes (Ag1000g) data we identified a very recent selective sweep in mosquitoes from Uganda which localized to a cluster of cytochrome P450 genes, including some commonly implicated in resistance. Further interrogation revealed a haplotype involving a trio of mutations, a nonsynonymous point mutation in Cyp6p4 (I236M), an upstream insertion of a partial Zanzibar-like transposable element (TE) and a duplication of the Cyp6aa1 gene. The mutations appear to have originated recently in An. gambiae from the Kenya-Uganda border region around Lake Victoria, with stepwise replacement of the double-mutant (Zanzibar-like TE and Cyp6p4-236M) with the triple-mutant haplotype (including Cyp6aa1 duplication), which has spread into the Democratic Republic of Congo and Tanzania. The triple-mutant haplotype is strongly associated with increased expression of genes able to metabolise pyrethroids and is strongly predictive of resistance to pyrethroids most notably deltamethrin, a commonly-used LLIN insecticide. Importantly, there was increased mortality in mosquitoes carrying the triple-mutation when exposed to nets co-treated with the synergist piperonyl butoxide (PBO). Frequencies of the triple-mutant haplotype remain spatially variable within countries, suggesting an effective marker system to guide deployment decisions for limited supplies of PBO-pyrethroid co-treated LLINs across African countries. Duplications of the Cyp6aa1 gene are common in An. gambiae across Africa and, given the enzymes metabolic activity, are likely to be a useful diagnostic for high levels of pyrethroid resistance.

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