Biology subjects

Ibrahim, S. S.

Publications and source records attributed to Ibrahim, S. S..

9 recordsLinked to original sources

bioRxiv2026

Generational selection, transcriptomics and functional characterization reveal the impact of environmental pollutants on the evolution of insecticide resistance in malaria vectors

Insecticide resistance is threatening malaria control. While the evolution and spread of resistance has been linked to scale-up in the distribution of public health insecticides, the role of environmental pollutants such as the polyaromatic hydrocarbons (PAHs) from industrial and agricultural use remains largely uncharacterized. The PAHs are potent ligands of the aryl hydrocarbon receptor (Ahr) transcription factors involved in the regulation of xenobiotic metabolizing enzymes, and potentially involved in insecticide resistance. Here, using field insecticide-resistant (Auyo) An. coluzzii and a laboratory-susceptible colony (Ngousso), we conducted a multi-generational selection experiment using naphthalene, fluorene and a mixture of both PAHs. After ten generations, the changes in susceptibility to insecticides were monitored using WHO bioassays and whole-transcriptome analysis (RNASeq) was conducted. Compared with the non-selected colony lines, PAH exposures significantly reduced pyrethroid and DDT resistance in the field population, suggesting fitness cost associated with established resistance. In contrast, Ngousso showed a significant increase in DDT resistance (p = 0.01) at the tenth generation. A significant increase in permethrin resistance was also observed at the seventh generation (p = 0.03). Several candidate genes from the major detoxification classes were overexpressed in the selected lines (including GSTe2, CYP6Z1, and CYP6P4); the most consistent were CYP6M4 and CYP4C27, as well as those from the Ahr pathway. Heterologous expression of CYP6M4 revealed its ability to metabolise pyrethroids, including permethrin, deltamethrin, and -cypermethrin, as well as PAHs (naphthalene and fluorene). These findings establish the role of environmental pollutants as additional drivers of metabolic insecticide resistance in An, coluzzii.

ecology↗

bioRxiv2026

Metabolomic changes underpinning permethrin resistance in the Anopheles gambiae malaria vector from Cameroon

Insecticide resistance reduces the effectiveness of current malaria vector control interventions. To understand the mechanisms underlying permethrin resistance in field-derived Anopheles gambiae vectors collected from Cameroon, we used Ultra-High Performance Liquid Chromatography coupled with High-Resolution Tandem Mass Spectrometry (UHPLC-HRMS/MS) to comprehensively analyze metabolic changes in resistant and susceptible samples to gain insight into mechanisms driven permethrin resistance. Resistant mosquitoes exhibited upregulated levels of inosine, nicotinic acid, dipeptides, amino acids, fumarate, uracil, and aldopentose. These data suggest that altered metabolic-based detoxification, as well as target site and metabolic shifts, and enhanced energy production contribute to permethrin resistance. Conversely, susceptible mosquitoes showed increased levels of N-acetyl-aspartic acid, xanthurenic acid, 2-hydroxyglutarate, 3-hydroxykynurenine, propanoylcarnitine, and L-pipecolic acid. These metabolites are associated with neurotoxicity, energy disruption, as well as tryptophan and lysine catabolism. These findings elucidate the metabolic pathways of permethrin-resistance and underscore the mechanisms that could lead to the emergence of pyrethroid cross-resistance.

systems biology↗

bioRxiv2025

A panel of CYP6Z genes drives broad-spectrum cross-resistance to public health insecticides in the major malaria vector An. gambiae s.s.

The spread of multiple and intense insecticide resistance in major African malaria vectors is jeopardising control efforts. Mitigating this threat requires deciphering its underlying molecular mechanisms. Here, by integrating transcriptomic profiling, analysis of genetic diversity and functional genomics approaches, we established the role of the CYP6Z genes in conferring multiple- and cross-resistance to insecticides. Investigation of CYP6Z gene expression and genetic diversity analyses indicate that resistance is primarily associated with transcriptional upregulation rather than fixed coding mutations since no predominant haplotype was selected. Structural characterisation reveals a flexible, promiscuous active site in CYP6Z enzymes, enabling binding of multiple insecticide classes. In vitro functional validation confirms that recombinant CYP6Z3 efficiently metabolizes deltamethrin (percentage depletion of 59%), permethrin (52%), -cypermethrin (37%), pirimiphos-methyl (36%), fenitrothion (25%), propoxur (25%), and possibly bendiocarb (17%). RNAi-mediated knockdown of CYP6Z genes in field-collected An. gambiae s.s. restores susceptibility to several insecticides including clothianidin: dsCYP6Z1 (mortality = 50.29%; p < 0.01), dsCYP6Z2 (47.28%; p < 0.01) and dsCYP6Z3 (39.12%; p < 0.05) compared to the control (mortality = 27.67%). Furthermore, transgenic expression in Drosophila melanogaster flies revealed that expression of CYP6Z genes alone confer cross-resistance to pyrethroids, organophosphates, carbamates and neonicotinoids, but increases susceptibility to the pro-insecticide chlorfenapyr: CYP6Z1 (mortality = 100%; p < 0.01), CYP6Z3 (98.18%; p < 0.01), and CYP6Z4 (98.01%; p < 0.01) vs. control (88.03%). This study establishes An. gambiae CYP6Z genes operate in concert to drive a broad-spectrum metabolic resistance, towards several classes of public health insecticides, while in contrast bioactivating chlorfenapyr to its more insecticidally active metabolite.

molecular biology↗

bioRxiv2024

A single mutation G454A in P450 CYP9K1 drives pyrethroid resistance in the major malaria vector Anopheles funestus reducing bed net efficacy

Metabolic resistance to pyrethroids is jeopardizing the effectiveness of insecticide-based interventions against malaria. The complexity of the Africa-wide spatio-temporal evolution of the molecular basis of this resistance, the major genetic drivers should be detected to improve resistance management. Here, we demonstrated that a single amino acid change G454A in the cytochrome P450 CYP9K1 drives pyrethroid resistance in Anopheles funestus vector in East and Central Africa. Polymorphism analysis revealed drastic reduction of diversity of the CYP9K1 gene in Uganda (2014) with the selection of a predominant haplotype (90%), exhibited a G454A mutation. However, 6 years later (2020) the Ugandan 454A-CYP9K1 haplotype was also predominant in Cameroon (84.6%), but absent in Malawi (Southern Africa) and Ghana (West Africa). In vitro comparative heterologous metabolism assays revealed that the mutant-type 454A-CYP9K1 (R) allele metabolises type II pyrethroid (deltamethrin) better than the wild-type G454-CYP9K1 (S) allele. Transgenic Drosophila melanogaster flies expressing the mutant-type 454A-CYP9K1 allele were significantly more resistant to both type I and II pyrethroids than the flies expressing the wild-type G454-CYP9K1 allele. Genotyping with a newly designed DNA-based diagnostic assay targeting the G454A replacement revealed that this mutation is strongly associated with pyrethroid resistance as mosquitoes surviving pyrethroid exposure were significantly more homozygote resistant (Odds ratio = 567, P<0.0001). Furthermore, Cone test and experimental hut trials showed that 454A-CYP9K1 reduces the efficacy of LLINs. The resistant allele (454A) is under directional selection in Eastern and Central Africa, present but not strongly selected in Southern Africa and at very low frequency in West Africa. This study reveals the rapid spread of P450-based metabolic pyrethroid resistance driven by CYP9K1, greatly reducing the efficacy of pyrethroid-based control tools. The new DNA-based assay designed here will add to the toolbox to monitor resistance in the field and improve resistance management strategies. Author SummaryThe complex molecular basis and genetic drivers of metabolic resistance in malaria vectors should be detected to improve resistance management. Here, we established that allelic variation by a single mutation G454A in P450 CYP9K1 enzyme drives pyrethroid resistance in Anopheles funestus. Drastic reduction of diversity was noted in Ugandan female An. funestus samples collected in 2014, with a major haplotype (454A) already fixed but absent in other African regions. However, this Ugandan 454A-CYP9K1 haplotype was highly selected within 6 years in An. funestus samples from Cameroon (Central Africa), but still absent in Ghana (West Africa) and Malawi (Southern Africa). Metabolism assays revealed that the 454A-resistant allele metabolized pyrethroid better than the susceptible G454 allele and driving higher pyrethroid resistance in transgenic Drosophila melanogaster flies. DNA-based diagnostics designed around the G454A-CYP9K1 marker strongly correlates with pyrethroid resistance, reducing bed net efficacy indicating that this assay should be added to the toolbox to monitor this 454A-CYP9K1 resistance which is rapidly spreading in An. funestus mosquito populations from Eastern and Central Africa.

genetics↗

bioRxiv2024

Two highly selected mutations in the tandemly duplicated CYP6P4a and CYP6P4b drive pyrethroid resistance in Anopheles funestus

Gaining a comprehensive understanding of the genetic mechanisms underlying insecticide resistance in malaria vectors is crucial for optimising the effectiveness of insecticide-based vector control methods and developing diagnostic tools for resistance management. Considering the heterogeneity of metabolic resistance in major malaria vectors, the implementation of tailored resistance management strategies is essential for successful vector control. In this study, we provide evidence demonstrating that two highly selected mutations in the tandemly duplicated cytochrome P450 genes namely CYP6P4a and CYP6P4b, are driving pyrethroid insecticide resistance in the major malaria vector Anopheles funestus, in West Africa. Through a continent-wide polymorphism survey, we observed heightened indications of directional selection in both genes between 2014 and 2021. By conducting in vitro insecticide metabolism assays with recombinant enzymes expressed from both genes, we established that mutant alleles under selection exhibit higher metabolic efficiency compared to their wild-type counterparts. Furthermore, using the GAL4-UAS transgenic system, we demonstrated that transgenic Drosophila melanogaster flies overexpressing mutant alleles displayed an increased resistance to pyrethroids. These findings were in agreement with in silico characterisation, which highlighted changes in enzyme active site architecture that enhance the affinity of mutant alleles for type I and II pyrethroids. Furthermore, we developed two DNA-based assays capable of detecting the CYP6P4a-M220I and CYP6P4b-D284E mutations, showing their current confinement to West Africa. Genotype/phenotype correlation analyses revealed that these markers are strongly associated with resistance to types I and II pyrethroids and combine to drastically reduce the efficacy of pyrethroid bednets. Overall, our study makes available two field-applicable insecticide resistance molecular markers that will help in the monitoring and better management of insecticide resistance in West Africa. TeaserTwo field-applicable diagnostic tools for detecting metabolic resistance in Anopheles funestus to enhance insecticide resistance management in West Africa.

genetics↗

bioRxiv2024

A single E205D allele of a key P450 CYP6P3 is driving metabolic pyrethroid resistance in the major African malaria vector Anopheles gambiae

Deciphering the molecular drivers of insecticide resistance is paramount to extend the effectiveness of malaria vector control tools. Here, we demonstrated that the E205D amino acid change in a key metabolic resistance P450 CYP6P3 drives pyrethroid resistance in the major malaria vector, Anopheles gambiae. Spatio-temporal whole genome Poolseq analyses in Cameroon detected a major P450-linked locus on chromosome 2R beside the sodium channel locus. In vitro metabolism assays with recombinantly expressed CYP6P3 protein revealed that the catalytic efficiency of 205D was 2.5 times higher than E205 with -cypermethrin. Similar patterns were observed for permethrin. Overexpression of the 205D allele in transgenic flies confers higher more pyrethroids and carbamates resistance, compared to controls. A DNA-based assay further supported that the CYP6P3-205D variant strongly correlates with pyrethroid resistance in field populations (OR=26.4; P<0.0001) and that it reduces the efficacy of pyrethroid-only LLINs with homozygote RR genotype exhibiting significantly higher survival following PermaNet 3.0 exposure compared to the SS genotype (OR: 6.1, p = 0.0113). Furthermore, the CYP6P3-E205D combines with the kdr target-site resistance mechanisms to worsen the loss of bednet efficacy. The 205D mutation is now predominant in West and Central Africa but less abundant or absent in East and South Africa with signs of introgression with An. coluzzii in Ghana. This study highlights the importance of P450-based resistance and designs field-applicable tools to easily track the spread of metabolic resistance and assess its impact on control interventions. One Sentence Summary: The major obstacle to malaria control and elimination is the spread of parasite resistance to anti-malarial drugs, and mosquito resistance to insecticides. In this study, we identified a key point mutation E205D in the metabolic gene CYP6P3 (cytochrome P450) conferring resistance to pyrethroids by enhancing the breakdown of insecticides used for bednets impregnation. DNA-based assays were then designed and used to determine the spread of the resistance across Africa and demonstrate that the CYP6P3-205D allele works together with the knockdown resistance in the voltage-gated Sodium channel to reduce the efficacy of insecticide-treated bednets.

genomics↗

bioRxiv2024

Nationwide surveillance identifies yellow fever and chikungunya viruses transmitted by various species of Aedes mosquitoes in Nigeria

BackgroundSince its reemergence in 2017, yellow fever (YF) has been active in Nigeria. The Nigeria Centre for Disease Control (NCDC) has coordinated responses to the outbreaks with the support of the World Health Organization (WHO). The National Arbovirus and Vectors Research Centre (NAVRC) handles the vector component of these responses. This study sought to identify the vectors driving YF transmission and any of the targeted arboviruses and their distribution across states. MethodsEggs, larvae and pupae as well as adult mosquitoes were collected in observational, analytical, and cross-sectional surveys conducted in sixteen YF outbreak states between 2017 and 2020. Adult mosquitoes (field-collected or reared from immature stages) were morphologically identified, and arboviruses were detected using RT-qPCR at the African Centre of Excellence for Genomics of Infectious Diseases (ACEGID). ResultsAedes mosquitoes were collected in eleven of the sixteen states surveyed and the mosquitoes in nine states were found infected with arboviruses. A total of seven Aedes species were collected from different parts of the country. Aedes aegypti was the most dominant (51%) species, whereas Aedes africanus was the least (0.2%). Yellow fever virus (YFV) was discovered in 33 ([~]26%) out of the 127 Aedes mosquito pools. In addition to YFV, the Chikungunya virus (CHIKV) was found in nine pools. Except for Ae. africanus, all the Aedes species tested positive for at least one arbovirus. YFV-positive pools were found in six (6) Aedes species while CHIKV-positive pools were only recorded in two Aedes species. Edo State had the most positive pools (16), while Nasarawa, Imo, and Anambra states had the least (1 positive pool). Breteau and house indices were higher than normal transmission thresholds in all but one state. ConclusionIn Nigeria, there is a substantial risk of arbovirus transmission by Aedes mosquitoes, with YFV posing the largest threat at the moment. This risk is heightened by the fact that YFV and CHIKV have been detected in vectors across outbreak locations. Hence, there is an urgent need to step up arbovirus surveillance and control activities in the country. Author summaryEpidemics of arboviral infections are fast becoming the norm across Africa, especially in Nigeria, where sporadic epidemics of Aedes-borne Yellow Fever (YF) have been on the rise since 2017. A nationwide surveillance was conducted to detect the arboviruses and the vectors transmitting them during large YF outbreaks across Nigeria. Aedes mosquitoes collected from 57 local government areas/outbreak locations, covering 16 out of the 36 states in Nigeria, were reared to adulthood, morphologically identified to species level and assayed using qRT-PCR to detect six epidemiologically important arboviruses (Dengue, Chikungunya (CHIKV), Zika, YF, West Nile and Onyong nyong). Seven Aedes species (including Ae. aegypti, Ae. albopictus, Ae. circumluteolus, Ae. vittatus, Ae. simpsoni complex, Ae. luteocephalus, and Ae. africanus), were caught, with Ae. aegypti and Ae. albopictus the most widespread and predominant. Only YF and CHKV were found in mosquito pools from 9 states, with only Ae. aegypti pools positive for CHKV, while YF virus was found predominantly in this species and Ae. albopictus. This study provides a baseline for widespread distribution of the major Aedes vectors, and their arboviral transmission profiles, which can enhance evidence-based control measures, and strengthen nationwide surveillance systems to forestall future outbreaks of public health magnitude.

zoology↗

bioRxiv2022

Molecular drivers of insecticide resistance in the Sahelo-Sudanian populations of a major malaria vector

Information on common markers of metabolic resistance in malaria vectors from countries sharing similar eco-climatic characteristics can facilitate coordination of malaria control. Here, we characterized populations of the major malaria vector Anopheles coluzzii from Sahel region, spanning four sub-Saharan African countries: Nigeria, Niger, Chad and Cameroon. Genome-wide transcriptional analysis identified major genes previously implicated in pyrethroid and/or cross resistance to other insecticides, overexpressed across the Sahel, including CYP450s, glutathione S-transferases, carboxylesterases, and cuticular proteins. Several, well-known markers of insecticide resistance were found in high frequencies - including in the voltage-gated sodium channel (V402L, I940T, L995F, I1527T and N1570Y), the acetylcholinesterase-1 gene (G280S) and the CYP4J5-L43F (fixed). High frequencies of the epidemiologically important chromosomal inversions, 2La, 2Rb and 2Rc were observed ([~]80% for 2Rb and 2Rc). The 2La alternative arrangement is fixed across the Sahel. Low frequencies of these inversions (<10%) were observed in the fully insecticide susceptible laboratory colony of An. coluzzii (Ngoussou). Several of the most commonly overexpressed metabolic resistance genes sit in these three inversions. Two commonly overexpressed genes, GSTe2 and CYP6Z2 were functionally validated. Transgenic Drosophila melanogaster expressing GSTe2 exhibited extremely high DDT and permethrin resistance (mortalities < 10% in 24 h). Serial deletion of the 5 intergenic region, to identify putative nucleotide(s) associated with GSTe2 overexpression, revealed that simultaneous insertion of adenine nucleotide and a transition (T->C), between Fork-head box L1 and c-EST putative binding sites were responsible for the high overexpression of GSTe2 in the resistant mosquitoes. Transgenic flies expressing CYP6Z2 exhibited marginal resistance towards 3-phenoxybenzylalcohol (a primary product of pyrethroid hydrolysis by carboxylesterases) and a type II pyrethroid, -cypermethrin. However, significantly higher mortalities were observed in CYP6Z2 transgenic flies compared with controls, on exposure to the neonicotinoid, clothianidin. This suggests a possible bioactivation of clothianidin into a toxic intermediate, which if true make it an ideal insecticide against populations of An. coluzzii overexpressing this P450. These findings will facilitate regional collaborations within the Sahel region, and refine implementation strategies through re-focusing interventions, improving evidence-based, cross-border policy towards local and regional malaria pre-elimination.

genomics↗

bioRxiv2021

Multi-omics analysis identifies a CYP9K1 haplotype conferring pyrethroid resistance in the malaria vector Anopheles funestus in East Africa

Metabolic resistance to pyrethroids is a menace to the continued effectiveness of malaria vector controls. Its molecular basis is complex and varies geographically across Africa. Here, we used a multi-omics approach, followed-up with functional validation to show that a directionally selected haplotype of a cytochrome P450, CYP9K1 is a major driver of resistance in Anopheles funestus. A PoolSeq GWAS using mosquitoes alive and dead after permethrin exposure, from Malawi and Cameroon, detected candidate genomic regions, but lacked consistency across replicates. Targeted deep sequencing of candidate resistance genes and genomic loci detected several SNPs associated with known pyrethroid resistance QTLs. The most significant SNP was in the cytochrome P450 CYP304B1 (Cameroon), CYP315A1 (Uganda) and the ABC transporter gene ABCG4 (Malawi). However, when comparing field resistant mosquitoes to laboratory susceptible, the pyrethroid resistance locus rp1 and SNPs around the ABC transporter ABCG4 were consistently significant, except for Uganda where CYP9K1 P450 was markedly significant. In vitro heterologous metabolism assays with recombinant CYP9K1 revealed that it metabolises type II pyrethroid (deltamethrin; 64% depletion) but not type I (permethrin; 0%), while moderately metabolising DDT (17%). CYP9K1 exhibited a drastic reduction of genetic diversity in Uganda, in contrast to other locations, highlighting an extensive selective sweep. Furthermore, a glycine to alanine (G454A) amino acid mutation located between the meander and cysteine pocket of CYP9K1 was detected in all Ugandan mosquitoes. This study sheds further light on the complex evolution of metabolic resistance in a major malaria vector, by adding further resistance genes and variants that can be used to design field applicable markers to better track this resistance Africa-wide. Author SummaryMetabolic resistance to pyrethroids is a menace to the continued effectiveness of malaria vector controls. Its molecular basis is complex and varies geographically across Africa. Here, we used several DNA based approach to associate genomic differences between resistant and susceptible mosquitoes from several field and laboratory populations of the malaria vector Anopheles funestus. We followed-up our genomic analyses with functional validation of a candidate resistance gene in East Africa. This gene (CYP9K1) is a member of the cytochrome P450 gene-family that helps to metabolise, and thereby detoxify, pyrethroid insecticides. We show that this gene is a major driver of resistance to a specific sub-class of pyrethroid insecticides only, with moderate to no effects on other insecticides used against Anopheles funestus. We were able to link resistance in this gene to a mutation that changes the amino acid glycine to alanine that may impact how the protein-product of this gene binds to target insecticides. In addition to demonstrating the biochemical specificity of an evolutionary response, we have broadened the available pool of genes can be used to monitor the spread of insecticide resistance in this species.

genomics↗