Liebman_2015_Malar.J_14_74

BACKGROUND: Resistance to multiple classes of insecticides has been detected in the malaria vector Anopheles albimanus in northwest Peru. Acetylcholinesterase (AChE) insensitivity has previously been associated with resistance to organophosphate (OP) and carbamate (CA) insecticides in arthropods. A single point mutation on the ace-1 gene (G119S) associated with resistance to OPs and CAs has been described previously in four anopheline species, but not in field-collected An. albimanus. The present study aimed to characterize the role of ace-1 in conferring resistance to both OPs and CAs in the An. albimanus population in Tumbes, Peru.
METHODS: The frequency and intensity of resistance to OPs and CAs was quantified through bioassays of female An. albimanus collected between 2012 and 2014, and the presence of insensitive AChE was confirmed using biochemical assays. A portion of the ace-1 gene flanking codon 119 was amplified and sequenced from individuals used in the bioassays and biochemical assays, as well as from historical samples collected in 2008. Statistical analyses were conducted to determine: (1) associations between genotype and AChE insensitivity; and, (2) associations between genotype and resistance phenotype.
RESULTS: After confirming high levels of resistance to fenitrothion, malathion, and bendiocarb through bioassays, two novel polymorphisms were identified at the first and second loci of codon 119, with all individuals from the 2012-2014 collections being heterozygous at the first base (G/T) and either heterozygous (G/C) or homozygous mutants (C/C) at the second base. Based on sequence data from historical samples, these mutations arose prior to 2008, but became fixed in the population between 2008 and 2012. Homozygotes at the second locus had significantly higher levels of AChE insensitivity than heterozygotes (p <0.05). Individuals phenotypically susceptible to OPs and CAs were more likely to be heterozygous at the second locus (p <0.01). Cloning identified four individuals each containing three distinct genotypes, suggesting that a duplication of the ace-1 gene may have occurred.
CONCLUSIONS: The occurrence of heterozygotes at two loci and the presence of three genotypes in four individuals suggest that balancing selection could be maintaining OP and CA resistance in this population, while minimizing associated fitness costs.