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01.12.2012 | Methodology | Ausgabe 1/2012 Open Access

Malaria Journal 1/2012

Development of an allele-specific, loop-mediated, isothermal amplification method (AS-LAMP) to detect the L1014F kdr-w mutation in Anopheles gambiae s. l.

Zeitschrift:
Malaria Journal > Ausgabe 1/2012
Autoren:
Athanase Badolo, Kyioshi Okado, Wamdaogo M Guelbeogo, Hiroka Aonuma, Hironori Bando, Shinya Fukumoto, N’Fale Sagnon, Hirotaka Kanuka
Wichtige Hinweise

Electronic supplementary material

The online version of this article (doi:10.​1186/​1475-2875-11-227) contains supplementary material, which is available to authorized users.

Competing interests

The authors declare that they have no competing interests.

Authors' contributions

AB and HK conceived the study and wrote the paper. AB designed the primers and carried out the molecular study. OK, HA and SF performed experiments in the laboratory. AB, NS, WMG, OK and HB contributed to the fieldwork related to collecting the Anopheles mosquitoes. All the authors read and approved the final manuscript.

Abstract

Background

Malaria control relies heavily on treated bed nets and indoor residual spraying with pyrethroid insecticides. Unfortunately, the resistance to pyrethroid insecticides, mainly due to the kdr mutation, is spreading in the main malaria vector Anopheles gambiae s.l., decreasing the insecticides’ efficacy. To manage the insecticide resistance rapidly and flexibly, simple and effective tools for the early detection of resistant mosquitoes are needed. This study aimed to develop an allele-specific, loop-mediated, isothermal amplification (AS-LAMP) method to detect the West African-type kdr mutation (kdr-w; L1014F) in field-collected mosquitoes.

Methods

DNA fragments of the wild-type and the mutated kdr gene were used to select the primers and develop the method. The primers were designed with the mutation at the 5’ end of the backward inner primer (BIP). The AS-LAMP method was compared to the AS-PCR method using the genomic DNA of 120 field-collected mosquitoes.

Results

The AS-LAMP method could discriminate between the wild-type homozygote, the heterozygote, and the kdr-w homozygote within 75 min. The AS-LAMP method has the advantage of being faster and at least as sensitive and specific as the AS-PCR method.

Conclusions

The AS-LAMP method can be used to detect the kdr mutation for quick decision-making, even in less well-equipped laboratories.
Zusatzmaterial
Authors’ original file for figure 1
12936_2012_2124_MOESM1_ESM.pdf
Authors’ original file for figure 2
12936_2012_2124_MOESM2_ESM.pdf
Authors’ original file for figure 3
12936_2012_2124_MOESM3_ESM.pdf
Literatur
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