Hostname: page-component-cd9895bd7-hc48f Total loading time: 0 Render date: 2024-12-26T07:22:18.353Z Has data issue: false hasContentIssue false
  • English
  • Français

Qualitative Sybr Green real-time detection of single nucleotide polymorphisms responsible for target-site resistance in insect pests: the example of Myzus persicae and Musca domestica

Published online by Cambridge University Press:  22 July 2016

V. Puggioni ,
O. Chiesa ,
M. Panini  and
E. Mazzoni
V. Puggioni
Affiliation:
Department of Sustainable Crop Production, Section Sustainable Crop and Food Protection, Università Cattolica del Sacro Cuore, Via Emilia Parmense, 84, I-29122 Piacenza, Italy
O. Chiesa
Affiliation:
Department of Sustainable Crop Production, Section Sustainable Crop and Food Protection, Università Cattolica del Sacro Cuore, Via Emilia Parmense, 84, I-29122 Piacenza, Italy
M. Panini
Affiliation:
Department of Sustainable Crop Production, Section Sustainable Crop and Food Protection, Università Cattolica del Sacro Cuore, Via Emilia Parmense, 84, I-29122 Piacenza, Italy
E. Mazzoni*
Affiliation:
Department of Sustainable Crop Production, Section Sustainable Crop and Food Protection, Università Cattolica del Sacro Cuore, Via Emilia Parmense, 84, I-29122 Piacenza, Italy
*
*Address for correspondence Fax: +39 0523 599268 Phone: +39 0523 599237 E-mail: emanuele.mazzoni@unicatt.it
Get access Rights & Permissions [Opens in a new window]

Abstract

Chemical insecticides have been widely used to control insect pests, leading to the selection of resistant populations. To date, several single nucleotide polymorphisms (SNPs) have already been associated with insecticide resistance, causing reduced sensitivity to many classes of products. Monitoring and detection of target-site resistance is currently one of the most important factors for insect pest management strategies. Several methods are available for this purpose: automated and high-throughput techniques (i.e. TaqMan or pyrosequencing) are very costly; cheaper alternatives (i.e. RFLP or PASA–PCRs) are time-consuming and limited by the necessity of a final visualization step. This work presents a new approach (QSGG, Qualitative Sybr Green Genotyping) which combines the specificity of PASA–PCR with the rapidity of real-time PCR analysis. The specific real-time detection of Cq values of wild-type or mutant alleles (amplified used allele-specific primers) allows the calculation of ΔCqW–M values and the consequent identification of the genotypes of unknown samples, on the basis of ranges previously defined with reference clones. The methodology is applied here to characterize mutations described in Myzus persicae and Musca domestica and we demonstrate it represents a valid, rapid and cost-effective technique that can be adopted for monitoring target-site resistance in field populations of these and other insect species.

Keywords

SNPs genotypingqualitative real-time PCRM. persicaeM. domesticaR81Tkdrs-kdrkdr-hisM918LMACE
Type
Research Papers
Information
Bulletin of Entomological Research , Volume 107 , Issue 1 , February 2017 , pp. 96 - 105
Check for updates
Copyright
Copyright © Cambridge University Press 2016 

Access options

Get access to the full version of this content by using one of the access options below. (Log in options will check for institutional or personal access. Content may require purchase if you do not have access.)

References

Anstead, J.A., Williamson, M.S., Eleftherianos, I. & Denholm, I. (2004) High-throughput detection of knockdown resistance in Myzus persicae using allelic discriminating quantitative PCR. Insect Biochemistry and Molecular Biology 34(8), 871–877.CrossRefGoogle ScholarPubMed
Anstead, J.A., Williamson, M.S. & Denholm, I. (2008) New methods for the detection of insecticide resistant Myzus persicae in the UK suction trap network. Agricultural and Forest Entomology 10(3), 291–295.Google Scholar
Bai, L., Zhu, G.D., Zhou, H.Y., Tang, J.X., Li, J.L., Xu, S., Zhang, M.H., Yao, L.N., Huang, G.Q., Wang, Y.B., Zhang, H.W., Wang, S.B., Cao, J. & Gao, Q. (2014) Development and application of an AllGlo probe-based qPCR assay for detecting knockdown resistance (kdr) mutations in Anopheles sinensis . Malaria Journal 13(1), 1.CrossRefGoogle ScholarPubMed
Bass, C., Nikou, D., Donnelly, M.J., Williamson, M.S., Ranson, H., Ball, A., Vontas, J. & Field, L.M. (2007) Detection of knockdown resistance (kdr) mutations in Anopheles gambiae: a comparison of two new high-throughput assays with existing methods. Malaria Journal 6(1), 111.CrossRefGoogle ScholarPubMed
Bass, C., Puinean, A.M., Andrews, M., Cutler, P., Daniels, M., Elias, J., Paul, V.L., Crossthwaite, A.J., Denholm, I., Field, L.M., Foster, S.P., Lind, R., Williamson, M.S. & Slater, R. (2011) Mutation of a nicotinic acetylcholine receptor β subunit is associated with resistance to neonicotinoid insecticides in the aphid Myzus persicae . BMC Neuroscience 12(1), 51.CrossRefGoogle ScholarPubMed
Black, I.V. & Vontas, J.G. (2007) Affordable assays for genotyping single nucleotide polymorphisms in insects. Insect Molecular Biology 16(4), 377–387.CrossRefGoogle ScholarPubMed
Cassanelli, S., Cerchiari, B., Giannini, S., Bizzaro, D., Mazzoni, E. & Manicardi, G.C. (2005) Use of the RFLP-PCR diagnostic test for characterizing MACE and kdr insecticide resistance in the peach potato aphid Myzus persicae . Pest Management Science 61(1), 91–96.Google Scholar
Dall'Ozzo, S., Andres, C., Bardos, P., Watier, H. & Thibault, G. (2003) Rapid single-step FCGR3A genotyping based on SYBR Green I fluorescence in real-time multiplex allele-specific PCR. Journal of Immunological Methods 277(1), 185–192.CrossRefGoogle ScholarPubMed
Dhas, D.B.B., Ashmi, A.H., Bhat, B.V., Parija, S.C. & Banupriya, N. (2015) Modified low cost SNP genotyping technique using cycle threshold (Ct) & melting temperature (Tm) values in allele specific real-time PCR. Indian Journal of Medical Research 142(5), 555.Google Scholar
Eleftherianos, I., Foster, S.P., Williamson, M.S. & Denholm, I. (2008) Characterization of the M918T sodium channel gene mutation associated with strong resistance to pyrethroid insecticides in the peach-potato aphid, Myzus persicae (Sulzer). Bulletin of Entomological Research 98(02), 183–191.CrossRefGoogle ScholarPubMed
Fenton, B., Margaritopoulos, J.T., Malloch, G.L. & Foster, S.P. (2010) Micro-evolutionary change in relation to insecticide resistance in the peach–potato aphid, Myzus persicae . Ecological Entomology 35(s1), 131–146.CrossRefGoogle Scholar
Feyereisen, R., Dermauw, W. & Van Leeuwen, T. (2015) Genotype to phenotype, the molecular and physiological dimensions of resistance in arthropods. Pesticide Biochemistry and Physiology 121, 61–77.CrossRefGoogle ScholarPubMed
Fontaine, S., Caddoux, L., Brazier, C., Bertho, C., Bertolla, P., Micoud, A. & Roy, L. (2011) Uncommon associations in target resistance among French populations of Myzus persicae from oilseed rape crops. Pest Management Science 67(8), 881–885.CrossRefGoogle ScholarPubMed
Fontaine, S., Caddoux, L. & Micoud, A. (2013) Methods for characterising resistance to carbamates, pyrethroids and neonicotinoids in Myzus persicae . Euro Reference 9, 19–23.Google Scholar
Fraaije, B.A., Butters, J.A., Coelho, J.M., Jones, D.R. & Hollomon, D.W. (2002) Following the dynamics of strobilurin resistance in Blumeria graminis f. sp. tritici using quantitative allele-specific real-time PCR measurements with the fluorescent dye SYBR Green I. Plant Pathology 51(1), 45–54.CrossRefGoogle Scholar
Hardstone, M.C. & Scott, J.G. (2010) A review of the interactions between multiple insecticide resistance loci. Pesticide Biochemistry and Physiology 97(2), 123–128.CrossRefGoogle Scholar
Huang, J., Kristensen, M., Qiao, C.L. & Jespersen, J.B. (2004) Frequency of kdr gene in house fly field populations: correlation of pyrethroid resistance and kdr frequency. Journal of Economic Entomology 97(3), 1036–1041.CrossRefGoogle ScholarPubMed
Kwok, P.Y. (2001) Methods for genotyping single nucleotide polymorphisms. Annual Review of Genomics and Human Genetics 2(1), 235–258.CrossRefGoogle ScholarPubMed
Liu, N. & Pridgeon, J.W. (2002) Metabolic detoxication and the kdr mutation in pyrethroid resistant house flies, Musca domestica (L.). Pesticide Biochemistry and Physiology 73(3), 157–163.CrossRefGoogle Scholar
Mazzoni, E. & Cravedi, P. (2002) Analysis of insecticide-resistant Myzus persicae (Sulzer) populations collected in Italian peach orchards. Pest Management Science 58(9), 975–980.Google Scholar
Mazzoni, E., Chiesa, O., Puggioni, V., Panini, M., Manicardi, G.C. & Bizzaro, D. (2015) Presence of kdr and s-kdr resistance in Musca domestica populations collected in Piacenza province (Northern Italy). Bulletin of Insectology 68(1), 65–72.Google Scholar
Nabeshima, T., Kozaki, T., Tomita, T. & Kono, Y. (2003) An amino acid substitution on the second acetylcholinesterase in the pirimicarb-resistant strains of the peach potato aphid, Myzus persicae . Biochemical and Biophysical Research Communications 307(1), 15–22.Google Scholar
Panini, M., Dradi, D., Marani, G., Butturini, A. & Mazzoni, E. (2014) Detecting the presence of target-site resistance to neonicotinoids and pyrethroids in Italian populations of Myzus persicae . Pest Management Science 70(6), 931–938.Google Scholar
Panini, M., Anaclerio, M., Puggioni, V., Stagnati, L., Nauen, R. & Mazzoni, E. (2015) Presence and impact of allelic variations of two alternative s -kdr mutations, M918T and M918L, in the voltage-gated sodium channel of the green peach aphid Myzus persicae . Pest Management Science 71(6), 878–884.CrossRefGoogle ScholarPubMed
Papp, A.C., Pinsonneault, J.K., Cooke, G. & Sadée, W. (2003) Single nucleotide polymorphism genotyping using allele-specific PCR and fluorescence melting curves. Biotechniques 34(5), 1068–1073.CrossRefGoogle ScholarPubMed
Puinean, A.M., Elias, J., Slater, R., Warren, A., Field, L.M., Williamson, M.S. & Bass, C. (2013) Development of a high-throughput real-time PCR assay for the detection of the R81T mutation in the nicotinic acetylcholine receptor of neonicotinoid-resistant Myzus persicae . Pest Management Science 69(2), 195–199.Google Scholar
Qiu, X., Pan, J., Li, M. & Li, Y. (2012) PCR–RFLP methods for detection of insecticide resistance-associated mutations in the house fly (Musca domestica). Pesticide Biochemistry and Physiology 104(3), 201–205.Google Scholar
Rinkevich, F.D., Hedtke, S.M., Leichter, C.A., Harris, S.A., Su, C., Brady, S.G., Taskin, V., Qiu, X. & Scott, J.G. (2012) Multiple origins of kdr-type resistance in the house fly, Musca domestica . PLoS ONE 7(12), e52761.CrossRefGoogle ScholarPubMed
Rinkevich, F.D., Du, Y. & Dong, K. (2013) Diversity and convergence of sodium channel mutations involved in resistance to pyrethroids. Pesticide Biochemistry and Physiology 106(3), 93–100.Google Scholar
Roy, L., Fontaine, S., Caddoux, L., Micoud, A. & Simon, J.C. (2013) Dramatic changes in the genotypic frequencies of target insecticide resistance in French populations of Myzus persicae (Hemiptera: Aphididae) over the last decade. Journal of Economic Entomology 106(4), 1838–1847.CrossRefGoogle ScholarPubMed
Soderlund, D.M. & Knipple, D.C. (2003) The molecular biology of knockdown resistance to pyrethroid insecticides. Insect Biochemistry and Molecular Biology 33(6), 563–577.Google Scholar
Tsuchihashi, Z. & Dracopoli, N.C. (2002) Progress in high throughput SNP genotyping methods. The Pharmacogenomics Journal 2(2), 103–110.Google Scholar
Voudouris, C.C., Kati, A.N., Sadikoglou, E., Williamson, M., Skouras, P.J., Dimotsiou, O., Georgiou, S., Fenton, B., Skavdis, G. & Margaritopoulos, J.T. (2016) Insecticide resistance status of Myzus persicae in Greece: long-term surveys and new diagnostics for resistance mechanisms. Pest Management Science 72(4), 671–683.Google Scholar
Whalon, M.E., Mota-Sanchez, D. & Hollingworth, R.M. (2008) Analysis of global pesticide resistance in arthropods. pp. 5–31 in Whalon, M.E., Mota-Sanchez, D. & Hollingworth, R.M. (Eds) Global Pesticide Resistance in Arthropods. Wallingford, UK, CABI.Google Scholar
Yu, D.J., Chen, Z.L., Zhang, R.J. & Yin, W.Y. (2005) Real-time qualitative PCR for the inspection and identification of Bactrocera philippinensis and Bactrocera occipitalis (Diptera: Tephritidae) using SYBR Green assay. Raffles Bulletin of Zoology 53(1), 73–78.Google Scholar
Supplementary material: PDF

Puggioni supplementary material S1

Supplementary Figure

Download Puggioni supplementary material S1(PDF)
PDF 928 KB
Supplementary material: PDF

Puggioni supplementary material S2

Supplementary Figure

Download Puggioni supplementary material S2(PDF)
PDF 652.7 KB
Supplementary material: PDF

Puggioni supplementary material S3

Supplementary Figure

Download Puggioni supplementary material S3(PDF)
PDF 1.4 MB
Supplementary material: PDF

Puggioni supplementary material S4

Supplementary Table

Download Puggioni supplementary material S4(PDF)
PDF 337.6 KB