Identification of mutations in the housefly para -type sodium channel gene associated with knockdown resistance ( kdr ) to pyrethroid insecticides

Abstract

We report the isolation of cDNA clones containing the full 6.3-kb coding sequence of the para-type sodium channel gene of the housefly, Musca domestica. This gene has been implicated as the site of knockdown resistance (kdr), an important resistance mechanism that confers nerve insensitivity to DDT and pyrethroid insecticides. The cDNAs predict a polypeptide of 2108 amino acids with close sequence homology (92% identity) to the Drosophila para sodium channel, and around 50% homology to vertebrate sodium channels, Only one major splice form of the housefly sodium channel was detected, in contrast to the Drosophila para transcript which has been reported to undergo extensive alternative splicing. Comparative sequence analysis of housefly strains carrying kdr or the more potent super-kdr factor revealed two amino acid mutations that correlate with these resistance phenotypes. Both mutations are located in domain II of the sodium channel. A leucine to phenylalanine replacement in the hydro-phobic IIS6 transmembrane segment was found in two independent kdr strains and six super-kdr strains of diverse geographic origin, while an additional methionine to threonine replacement within the intracellular IIS4-S5 loop was found only in the super-kdr strains. Neither mutation was present in five pyrethroid-sensitive strains. The mutations suggest a binding site for pyrethroids at the intracellular mouth of the channel pore in a region known to be important for channel inactivation.

Primer reporte de la mutación F1534C, asociada con resistencia cruzada a DDT y piretroides, en Aedes aegypti de Colombia

“…La comparación de las secuencias de los nueve mosquitos de Sincelejo, con una secuencia homó-loga de A. aegypti sensible a insecticidas (aislamiento 1534Phe, número de acceso KF527415.1) y una resistente a permetrina (aislamiento 1534Cys, número de acceso KF527414.1), puso en evidencia que el cambio de T por G (figura 2) afectaba específicamente el codón UUC, el cual cambió a UGC y codificó para el residuo 1534 del dominio IIIS6 del canal de sodio dependiente de voltaje, al igual que en la secuencia homóloga de referencia del también díptero Musca domestica (número de acceso: NM_001286885) (23). Esta sustitución no sinónima condujo al cambio de fenilalanina por cisteína en la proteína.…”

Sequence variation in the cadherin gene of Ostrinia nubilalis: a tool for field monitoring

“…Most of the documented instances of insect resistance involve a small number of genes (Roush and McKenzie, 1987). Insecticide resistance occurred by a single amino acid change in acetylcholinesterase, the target of organophosphate and carbamate insecticides in house fly (Williamson et al, 1996), and the sodium channel peptide targeted by pyrethroids in cockroach (Miyazaki et al, 1996). Similarly, cyclodiene resistance occurs by a single nonsynonymous point mutation in the Drosophila gaminobutyric acid (GABA) receptor (ffrench-Constant et al, 1993).…”

Synergistic interaction between two cockroach sodium channel mutations and a tobacco budworm sodium channel mutation in reducing channel sensitivity to a pyrethroid insecticide

“…One type of resistance mechanism is knockdown resistance (kdr), which results from reduced nerve sensitivity to pyrethroids and also to DDT (Soderlund and Bloomquist, 1990). Although first reported in 1951 in the house fly (Busvine, 1951), our understanding of the molecular basis of kdr and kdr-type resistance was limited until the recent cloning of sodium channel genes (homologous to the Drosophila para gene) from several kdr and kdr-type insects Miyazaki et al, 1996;Williamson et al, 1996;Dong, 1997).…”

“…A single leucine (L) to phenylalanine (F) substitution (L1014F in the house fly, and L993F in the German cockroach, see Fig. 1) in segment 6 of domain II (IIS6) is associated with kdr and kdr-type resistance (Miyazaki et al, 1996;Williamson et al, 1996;Dong, 1997). Subsequently, the same mutation was found in other important insect pests, including diamondback moth, Plutella xylostella (Martinez-Torres et al, 1997), horn fly, Haematobia irritans (Guerrero et al, 1997), mosquitoes, Anopheles gambiae and Culex pipiens (Martinez-Torres et al, 1998), aphids, Myzus persicae (Martinez-Torres et al, 1999), and Colorado potato beetle, Leptinotarsa decemlineata (Lee et al, 1999a).…”

“…Interestingly, higher levels of pyrethroid resistance are found if the L to F mutation is combined with additional sodium channel mutations. In the house fly, for example, a methionine to threonine mutation (M918T) in the linker region between IIS4 and IIS5 together with L1014F confers a much higher level of kdr (super-kdr) and sodium channel insensitivity (Williamson et al, 1996;Lee et al, 1999c;Vais et al, 2000). The German cockroach has additional four mutations, D57G, E434K, C764R and P1880L that are present in several highly pyrethroid-resistant German cockroach populations (Liu et al, 2000).…”