Genetic markers associated with insecticide resistance and resting behaviour in Anopheles gambiae mosquitoes in selected sites in Kenya

Abstract Background Molecular diagnostic tools have been incorporated in insecticide resistance monitoring programmes to identify underlying genetic basis of resistance and develop early warning systems of vector control failure. Identifying genetic markers of insecticide resistance is crucial in enhancing the ability to mitigate potential effects of resistance. The knockdown resistance (kdr) mutation associated with resistance to DDT and pyrethroids, the acetylcholinesterase-1 (ace-1R) mutation associated with resistance to organophosphates and carbamates and 2La chromosomal inversion associated with indoor resting behaviour, were investigated in the present study. Methods Anopheles mosquitoes sampled from different sites in Kenya and collected within the context of malaria vector surveillance were analysed. Mosquitoes were collected indoors using light traps, pyrethrum spray and hand catches between August 2016 and November 2017. Mosquitoes were identified using morphological keys and Anopheles gambiae sensu lato (s.l.) mosquitoes further identified into sibling species by the polymerase chain reaction method following DNA extraction by alcohol precipitation. Anopheles gambiae and Anopheles arabiensis were analysed for the presence of the kdr and ace-1R mutations, while 2La inversion was only screened for in An. gambiae where it is polymorphic. Chi-square statistics were used to determine correlation between the 2La inversion karyotype and kdr-east mutation. Results The kdr-east mutation occurred at frequencies ranging from 0.5 to 65.6% between sites. The kdr-west mutation was only found in Migori at a total frequency of 5.3% (n = 124). No kdr mutants were detected in Tana River. The ace-1R mutation was absent in all populations. The 2La chromosomal inversion screened in An. gambiae occurred at frequencies of 87% (n = 30), 80% (n = 10) and 52% (n = 50) in Baringo, Tana River and Migori, respectively. A significant association between the 2La chromosomal inversion and the kdr-east mutation was found. Conclusion The significant association between the 2La inversion karyotype and kdr-east mutation suggests that pyrethroid resistant An. gambiae continue to rest indoors regardless of the presence of treated bed nets and residual sprays, a persistence further substantiated by studies documenting continued mosquito abundance indoors. Behavioural resistance by which Anopheles vectors prefer not to rest indoors may, therefore, not be a factor of concern in this study’s malaria vector populations.

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Status of Insecticide Resistance and Its Mechanisms in Anopheles gambiae and Anopheles coluzzii Populations from Forest Settings in South Cameroon

Genes ◽

10.3390/genes10100741 ◽

2019 ◽

Vol 10 (10) ◽

pp. 741 ◽

Cited By ~ 6

Author(s):

Roland Bamou ◽

Nadège Sonhafouo-Chiana ◽

Konstantinos Mavridis ◽

Timoléon Tchuinkam ◽

Charles S. Wondji ◽

...

Keyword(s):

Insecticide Resistance ◽

Anopheles Gambiae ◽

Expression Profiles ◽

Rapid Expansion ◽

Molecular Diagnostic ◽

Diagnostic Tools ◽

Anopheles Coluzzii ◽

The South ◽

Potential Implication ◽

Forest Region

A key factor affecting malaria vector control efforts in Cameroon is the rapid expansion of insecticide resistance in Anopheles gambiae s.l (An. gambiae) populations; however, mechanisms involved in insecticide resistance in forest mosquito populations are still not well documented yet. The present study was conducted to screen molecular mechanisms conferring insecticide resistance in An. gambiae s.l. populations from the South Cameroon forest region. WHO bioassays were conducted with F0 An. gambiae females aged three to four days from forest (Sangmelima, Nyabessan, and Mbandjock) and urban sites (Yaoundé (Bastos and Nkolondom)), against pyrethroids (permethrin 0.75% and deltamethrin 0.05%) and carbamates (bendiocarb 0.1%). Members of the An. Gambiae s.l. species complex were identified using molecular diagnostic tools. TaqMan assays were used to screen for target site mutations. The expression profiles of eight genes implicated in insecticide resistance were assessed using RT-qPCR. Cuticle hydrocarbon lipids were measured to assess their potential implication in insecticide resistance. Both An. Gambiae and An. coluzzii were detected. An. gambiae was highly prevalent in Sangmelima, Nyabessan, Mbandjock, and Nkolondom. An. coluzzii was the only species found in the Yaoundé city center (Bastos). Low mortality rate to both pyrethroids and bendiocarb was recorded in all sites. High frequency of L1014F allele (75.32–95.82%) and low frequencies of L1014S (1.71–23.05%) and N1575Y (5.28–12.87%) were recorded. The G119S mutation (14.22–35.5%) was detected for the first time in An. gambiae populations from Cameroon. This mutation was rather absent from An. coluzzii populations. The detoxification genes Cyp6m2, Cyp9k1, Cyp6p4, Cyp6z1, as well as Cyp4g16 which catalyzes epicuticular hydrocarbon biosynthesis, were found to be overexpressed in at least one population. The total cuticular hydrocarvbon content, a proxy of cuticular resistance, did not show a pattern associated with pyrethroid resistance in these populations. The rapid emergence of multiple resistance mechanisms in An. Gambiae s.l. population from the South Cameroon forest region is of big concern and could deeply affect the sustainability of insecticide-based interventions strategies in this region.

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The new Internal Transcribed Spacer 2 diagnostic tool clarifies the taxonomic position and geographic distribution of the North American malaria vector Anopheles punctipennis

Malaria Journal ◽

10.1186/s12936-021-03676-4 ◽

2021 ◽

Vol 20 (1) ◽

Author(s):

James M. Hodge ◽

Andrey A. Yurchenko ◽

Dmitriy A. Karagodin ◽

Reem A. Masri ◽

Ryan C. Smith ◽

...

Keyword(s):

Malaria Vector ◽

Internal Transcribed Spacer ◽

Single Species ◽

Its2 Sequence ◽

Pathogen Transmission ◽

Molecular Diagnostic ◽

Diagnostic Tools ◽

Sequence Length ◽

Phylogenetic Position ◽

Internal Transcribed Spacer 2

Abstract Background The malaria mosquito Anopheles punctipennis, a widely distributed species in North America, is capable of transmitting human malaria and is actively involved in the transmission of the ungulate malaria parasite Plasmodium odocoilei. However, molecular diagnostic tools based on Internal Transcribed Spacer 2 (ITS2) of ribosomal DNA are lacking for this species. Anopheles punctipennis is a former member of the Anopheles maculipennis complex but its systematic position remains unclear. Methods In this study, ITS2 sequences were obtained from 276 An. punctipennis specimens collected in the eastern and midwestern United States and a simple and robust Restriction Fragment Length Polymorphism approach for species identification was developed. The maximum-likelihood phylogenetic tree was constructed based on ITS2 sequences available through this study and from GenBank for 20 species of Anopheles. Results The analysis demonstrated a consistent ITS2 sequence length and showed no indications of intragenomic variation among the samples based on ITS2, suggesting that An. punctipennis represents a single species in the studied geographic locations. In this study, An. punctipennis was found in urban, rural, and forest settings, suggesting its potential broad role in pathogen transmission. Phylogeny based on ITS2 sequence comparison demonstrated the close relationship of this species with other members of the Maculipennis group. Conclusions This study developed molecular tools based on ITS2 sequences for the malaria vector An. punctipennis and clarified the phylogenetic position of the species within the Maculipennis group.

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Structure of the G119S Mutant Acetylcholinesterase of the Malaria Vector Anopheles gambiae Reveals Basis of Insecticide Resistance

Structure ◽

10.1016/j.str.2017.11.021 ◽

2018 ◽

Vol 26 (1) ◽

pp. 130-136.e2 ◽

Cited By ~ 15

Author(s):

Jonah Cheung ◽

Arshad Mahmood ◽

Ravi Kalathur ◽

Lixuan Liu ◽

Paul R. Carlier

Keyword(s):

Insecticide Resistance ◽

Anopheles Gambiae ◽

Malaria Vector

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Major effect genes or loose confederations? The development of insecticide resistance in the malaria vector Anopheles gambiae

Parasites & Vectors ◽

10.1186/1756-3305-3-74 ◽

2010 ◽

Vol 3 (1) ◽

pp. 74 ◽

Cited By ~ 18

Author(s):

Basil D Brooke ◽

Lizette L Koekemoer

Keyword(s):

Insecticide Resistance ◽

Anopheles Gambiae ◽

Malaria Vector ◽

Major Effect

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Insecticide resistance in Bemisia tabaci Gennadius (Homoptera: Aleyrodidae) and Anopheles gambiae Giles (Diptera: Culicidae) could compromise the sustainability of malaria vector control strategies in West Africa

Acta Tropica ◽

10.1016/j.actatropica.2013.06.004 ◽

2013 ◽

Vol 128 (1) ◽

pp. 7-17 ◽

Cited By ~ 22

Author(s):

Olivier Gnankiné ◽

Imael H.N. Bassolé ◽

Fabrice Chandre ◽

Isabelle Glitho ◽

Martin Akogbeto ◽

...

Keyword(s):

West Africa ◽

Insecticide Resistance ◽

Vector Control ◽

Anopheles Gambiae ◽

Bemisia Tabaci ◽

Malaria Vector ◽

Control Strategies ◽

Malaria Vector Control ◽

Anopheles Gambiae Giles

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Dynamics of insecticide resistance in the malaria vector Anopheles gambiae s.l. from an area of extensive cotton cultivation in Northern Cameroon

Tropical Medicine & International Health ◽

10.1111/j.1365-3156.2008.02025.x ◽

2008 ◽

Vol 13 (4) ◽

pp. 476-486 ◽

Cited By ~ 71

Author(s):

M. Chouaïbou ◽

J. Etang ◽

T. Brévault ◽

P. Nwane ◽

C. K. Hinzoumbé ◽

...

Keyword(s):

Insecticide Resistance ◽

Anopheles Gambiae ◽

Malaria Vector ◽

Northern Cameroon ◽

Anopheles Gambiae S.L

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Resting behaviour of malaria vectors in a highland and a lowland site of western Kenya: Implication on malaria vector control measures

10.1101/815175 ◽

2019 ◽

Author(s):

Maxwell G. Machani ◽

Eric Ochomo ◽

Fred Amimo ◽

Jackline Kosgei ◽

Stephen Munga ◽

...

Keyword(s):

Insecticide Resistance ◽

Vector Control ◽

Malaria Vector ◽

Control Measures ◽

Malaria Vectors ◽

Western Kenya ◽

Sporozoite Rate ◽

Resting Behaviour ◽

Outdoor Resting ◽

Indoor And Outdoor

AbstractBackgroundUnderstanding the interactions between increased insecticide resistance in field malaria vector populations and the subsequent resting behaviour patterns is important for planning adequate vector control measures in a specific context and sustaining the current vector interventions. The aim of this study was to investigate the resting behavior, host preference and infection with Plasmodium falciparum sporozoites by malaria vectors in different ecological settings of western Kenya with different levels of insecticide resistance.MethodsIndoor and outdoor resting Anopheline mosquitoes were sampled during the dry and rainy seasons in Kisian (lowland site) and Bungoma (highland site), both in western Kenya. WHO tube bioassay was used to determine levels of phenotypic resistance of first generation offspring (F1 progeny) of malaria vectors resting indoors and outdoors to deltamethrin. PCR-based molecular diagnostics were used for mosquito speciation, genotype for resistance mutations and to determine specific host blood meal origins. Enzyme-linked Immunosorbent Assay (ELISA) was used to determine mosquito sporozoite infections.ResultsOverall, 3,566 female Anopheles mosquitoes were collected with Anopheles gambiae s.l [In Bungoma, An. gambiae s.s (90.9%), An arabiensis (7.6%) and in Kisian, An. gambiae s.s (38.9%), An. arabiensis (60.2%)] being the most abundant species (74.7%) followed by An. funestus s.l (25.3%). The majority of An. gambiae s.l (85.4 and 58%) and An. funestus (96.6 and 91.1%) were caught resting indoors in Bungoma and Kisian respectively.Vgsc-1014S was observed at a slightly higher frequency in An. gambiae s.s hereafter(An. gambiae) resting indoor than outdoor (89.7 vs 84.6% and 71.5 vs 61.1%) in Bungoma and Kisian respectively. For An. arabiensis, Vgsc-1014S was 18.2% indoor and outdoor (17.9%) in Kisian. In Bungoma, the Vgsc-1014S was only detected in An. arabiensis resting indoors with a frequency of 10%. The Vgsc-1014F mutation was only present in An. gambiae resting indoors from both sites, but at very low frequencies in Kisian compared to Bungoma (0.8 and 9.2% respectively. In Bungoma, the sporozoite rates for An. funestus, An. gambiae, and An. arabiensis resting indoors were 10.9, 7.6 and 3.4 % respectively. For outdoor resting, An. gambiae and An. arabiensis in Bungoma, the sporozoite rates were 4.7 and 2.9 % respectively.Overall, in Bungoma, the sporozoite rate for indoor resting mosquitoes was 8.6% and 4.2% for outdoors. In Kisian the sporozoite rate was 0.9% for indoor resting An. gambiae. None of the outdoor collected mosquitoes in Kisian tested positive for sporozoite infections.ConclusionThe study reports high densities of insecticide-resistant An. gambiae and An. funestus resting indoors and the persistence of malaria transmission indoors with high entomological inoculation rates (EIR) regardless of the use of Long-lasting insecticidal nets (LLINs). These findings underline the difficulties of controlling malaria vectors resting and biting indoors using the current interventions. Supplemental vector control tools and implementation of sustainable insecticide resistance management strategies are needed in western Kenya.

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Insecticide resistance mechanisms associated with different environments in the malaria vector Anopheles gambiae: a case study in Tanzania

Malaria Journal ◽

10.1186/1475-2875-13-28 ◽

2014 ◽

Vol 13 (1) ◽

pp. 28 ◽

Cited By ~ 80

Author(s):

Theresia E Nkya ◽

Idir Akhouayri ◽

Rodolphe Poupardin ◽

Bernard Batengana ◽

Franklin Mosha ◽

...

Keyword(s):

Insecticide Resistance ◽

Anopheles Gambiae ◽

Malaria Vector ◽

Resistance Mechanisms

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Mosquitoes cloak their legs to resist insecticides

Proceedings of The Royal Society B Biological Sciences ◽

10.1098/rspb.2019.1091 ◽

2019 ◽

Vol 286 (1907) ◽

pp. 20191091 ◽

Cited By ~ 12

Author(s):

Vasileia Balabanidou ◽

Mary Kefi ◽

Michalis Aivaliotis ◽

Venetia Koidou ◽

Juan R. Girotti ◽

...

Keyword(s):

Insecticide Resistance ◽

Anopheles Gambiae ◽

Malaria Vector ◽

Malaria Incidence ◽

Detoxification Enzymes ◽

Female Mosquito ◽

Cuticular Proteins ◽

Year 2000 ◽

Major Malaria Vector

Malaria incidence has halved since the year 2000, with 80% of the reduction attributable to the use of insecticides. However, insecticide resistance is now widespread, is rapidly increasing in spectrum and intensity across Africa, and may be contributing to the increase of malaria incidence in 2018. The role of detoxification enzymes and target site mutations has been documented in the major malaria vector Anopheles gambiae ; however, the emergence of striking resistant phenotypes suggests the occurrence of additional mechanisms. By comparing legs, the most relevant insect tissue for insecticide uptake, we show that resistant mosquitoes largely remodel their leg cuticles via enhanced deposition of cuticular proteins and chitin, corroborating a leg-thickening phenotype. Moreover, we show that resistant female mosquitoes seal their leg cuticles with higher total and different relative amounts of cuticular hydrocarbons, compared with susceptible ones. The structural and functional alterations in Anopheles female mosquito legs are associated with a reduced uptake of insecticides, substantially contributing to the resistance phenotype.

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