Kdr-type Resistance And Molecular Detection Techniques Of Resistance To Pyrethroids In Plutella Xylostella L.

This paper is mainly study on the toxicological, molecular characterization of knockdown resistanceto pyrethroid insecticides and molecular techniques for the detection of resistance in the diamondbackmoth, Plutella xylostella (L.). A convenient and actable rearing technique of diamondback moth wasdiscussed by a comparison of two different crop rearing diamondback moth. Four field populations ofBeijing, Anhui, Guangxi and Hainan of diamondback moth were collected in 2003. Compared with asusceptible strain, the four field populations have developmed very high level of resistance todeltamethrin. The highest resistance ratios (RR) was＞3570-fold found in Haina P xylostella strain.Resistance to deltamethrin were mass selected for 8 generations in laboratory developed＞3500-foldresistance in diamondback moth. Susceptibility to deltamethrin was sigle pair selected for 1 generationin laboratory with 2.6-fold increase of LC₅₀s in diamondback moth. Deltamethrin selected strain caused＞1500-fold cross-resistance to Alpha-cypermetkrin. There was a positive cross-resistance to DDT indiamondback moth strain. But little cross-resistance to methomyl and phoxim were found indiamondback moth strain. Synergism of by piperonyl butoxide (PB) and DEF in deltamethrin-resistantstrain of diamondback moth was investigated The PB increased the toxicity of deltametlu-in to RRstrain, but there was still a verb, high level of resistance in presence of the microsomal monooxygenaseinhibitor PB. DEF were even less effective as synergists on deltamethrin, alpha-cypermethrin and DDT.It suggested that the high level of resistance to deltamethrin in diamondback moth strain is correlated toMFO, but it is not only resistant mechanism. The genetics of deltamethrin resistance was studied indeltamethrin-selected strain of diamondback moth. It indicated incompletely recessive autosomalinheritance of the resistance trait. The high conserved domainâ…¡region of the para-typevoltage-sensitive sodium channel (the primary target site of pyrethroids) gene from both resistant andsusceptible diamondback moth strains was cloned and sequenced by RT-PCR. A fragment of 314 bpwas amplified from resistant strain, there is very, high homology between nucleotide sequences of theRR and SS strains (98.7ï¼…). Four amino acid changes in part of the para-type voltage-sensitive sodiumehannel of the resistant strain was detected. Two of the changes are exactly the same previously beenshown to correlate with kdr in the house fly, Musa domestica, and German cockroach, Blattellagermanica: leucine (CTT, Susceptible) to phenylalanine (TTT, Resistant) replacement (L1014F) in theIIS6 transmembrane segment and methionine (ATG, Susceptible) to threonine(ACG, Resistant)substitution (Mg18T) in the IIS4-5 transmembrane segment. The other two change, however, is differentto that in house fly strains, causing a threonine (ACC, Susceptible) to isoleucine (ATC, Resistant)substitution (T929I) at the beginning of the I1S5 segment and phenylalanine (TTC, Susceptible) toleucine (TTA, Resistant) replacement (F1020L) in the IIS6 transmembrane segment . The design ofthree PCR-based monitoring techniques for the genotyping of deltamethrin-resistant diamondback mothwere studied. These techniques rely upon designing an allele-specific primer which has basecorresponding to a specific resistance-associated mutation. PCR amplication of specific alleles(one-tube PCR: PASA, double-tube PCR: Bi-BASA) and single-stranded conformational polymorphismanalysis of PCR-amplified DNA (SSCP) all can be performed to monitor specific mutations. Individualinsects can be genotyped as SS, RS or RR. The relative advantages of these genotyping techniquesagainst conventional assay in monitoring for insecticide resistance are discussed.