A Primer-Introduced Restriction Analysis-Polymerase Chain Reaction Method to Detect Knockdown Resistance Mutations in Anopheles gambiae

In the major malaria vector Anopheles gambiae Giles, two point mutations at the voltage-gated sodium channel have been associated with knockdown resistance (kdr) to DDT and pyrethroid insecticides. Simple allele-specific polymerase chain reaction (PCR) assays to detect these single-nucleotide polymo...

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Bibliographic Details
Published in:Journal of medical entomology 2008-03, Vol.45 (2), p.237-241
Main Authors: Janeira, F., Vicente, J. L., Kanganje, Y., Moreno, M., do Rosário, V. E., Cravo, P., Pinto, J.
Format: Article
Language:English
Subjects:
Amino Acid Substitution
Animals
Anopheles - genetics
Anopheles gambiae
Biological and medical sciences
detection
DNA Mutational Analysis - methods
DNA Primers
Female
Fundamental and applied biological sciences. Psychology
genetic resistance
genotype
insecticide resistance
Insecticide Resistance - genetics
Insecticides
kdr mutations
Medically important nuisances and vectors, pests of stored products and materials: population survey and control
Molecular Biology/Genomics
new methods
PIRA-PCR
point mutation
Polymerase Chain Reaction
Pyrethrins
resistance
restriction endonucleases
sequence analysis
Sodium Channels - genetics
Vectors. Intermediate hosts
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Summary:In the major malaria vector Anopheles gambiae Giles, two point mutations at the voltage-gated sodium channel have been associated with knockdown resistance (kdr) to DDT and pyrethroid insecticides. Simple allele-specific polymerase chain reaction (PCR) assays to detect these single-nucleotide polymorphisms are prone to lack of specificity and therefore alternative techniques have been proposed. However, these may not be easily implemented in many laboratories from malaria endemic regions. Here, we describe a primer-introduced restriction analysis (PIRA)-PCR method to detect kdr mutations in An. gambiae. This method unambiguously identified all six genotypes for the kdr locus in a sample of 113 field-collected mosquitoes for which kdr genotypes had been confirmed by DNA sequencing. Co-occurrence of both kdr alleles was found in sites from Equatorial Guinea and Gabon and the L1014F mutation was detected in M-form individuals from Angola. The PIRA-PCR proved to be a reliable, robust, and simpler alternative for the detection of kdr mutations in this malaria vector.
ISSN:0022-2585
1938-2928
DOI:10.1603/0022-2585%282008%2945%5B237%3AAPRACR%5D2.0.CO%3B2