PCR-RFLP method to distinguish Frankliniella occidentalis, Frankliniella intonsa, Frankliniella pallida and Frankliniella tenuicornis

Przybylska, Arnika; Fiedler, Z; Obrępalska‐Stęplowska, Aleksandra

doi:10.1515/jppr-2016-0009

Cited by 11 publications

(11 citation statements)

References 13 publications

(22 reference statements)

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“…There are methods for the molecular identification of T. palmi based on real-time polymerase chain reaction (PCR) detection (Kox et al , 2005), sequence-characterized amplified region marker-based real-time PCR detection using a TaqMan probe (Walsh et al , 2005), PCR–restriction fragment length polymorphism (RFLP) detection (Brunner et al , 2002; Toda & Komazaki, 2002), and loop-mediated isothermal amplification assays (Przybylska et al , 2015). There are also protocols for identifying F. occidentalis using a real-time PCR assay with a TaqMan probe (Huang et al , 2010), PCR–RFLP (Brunner et al , 2002; Toda & Komazaki, 2002; Mainali et al , 2008; Przybylska et al , 2016), and a cytochrome oxidase I mitochondrial DNA marker (Zhang et al , 2012) with a duplex PCR (Zhang et al , 2014). Methods are also available describing a multiplex PCR assay for F. occidentalis, Frankliniella intonsa, T. tabaci , and T. palmi (Nakahara & Minoura, 2015) as well as a multiplex PCR assay for F. occidentalis, F. intonsa, T. tabaci , and Thrips hawaiiensis (Yeh et al , 2014).…”

Section: Introductionmentioning

confidence: 99%

Real-time PCR assay for distinguishingFrankliniella occidentalisandThrips palmiArnika Przybylska, Żaneta Fiedler, Aleksandra Obrępalska-Stęplowska

Przybylska

Fiedler

Frąckowiak

et al. 2017

Bull. Entomol. Res.

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Thrips palmi and Frankliniella occidentalis (order Thysanoptera) are thrips species that represent major plant pests. They are polyphagous insects capable of adversely affecting crop production. As such, in the European Union, these thrips species should be regulated as quarantine organisms. T. palmi and F. occidentalis can cause considerable damage to susceptible plants by feeding on them and transmitting several viruses responsible for serious plant diseases. Successful pest control strategies are based on an early, fast, and reliable diagnosis, which precedes the selection of appropriate steps to limit the effects of harmful organisms. We herein describe a novel diagnostic approach that enables the sensitive and species-specific detection (and differentiation) of these pests in a duplex polymerase chain reaction assay, which was adapted for both standard and real-time quantitative assays. Our method is based on the amplification of a 5.8S-internal transcribed spacer 2 ribosomal DNA fragment that is conserved between T. palmi and F. occidentalis.

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Section: Introductionmentioning

confidence: 99%

Real-time PCR assay for distinguishingFrankliniella occidentalisandThrips palmiArnika Przybylska, Żaneta Fiedler, Aleksandra Obrępalska-Stęplowska

Przybylska

Fiedler

Frąckowiak

et al. 2017

Bull. Entomol. Res.

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“…The first PCR-based characterization of thrips species was undertaken during the late 1990s ( Crespi et al, 1998 ). Subsequently, RAPD, RFLP, AFLP, SSR, SCAR, and qPCR have been reported for discrimination of thrips species ( Fang et al, 2005 ; Meena et al, 2005 ; XiangQin et al, 2010 ; Przybylska et al, 2016 ; Przybylska et al, 2018 ; Cao et al, 2019 ; Ghosh et al, 2021a ). Besides, the molecular techniques aid in identifying cryptic diversity, biotypes, host races, genetic structure, gene flow, reproductive isolation, and resolving species ambiguities in thrips which are otherwise impossible using morphological keys ( Ghosh et al, 2021a ).…”

Section: Discussionmentioning

confidence: 99%

Development of a Polymerase Spiral Reaction-Based Isothermal Assay for Rapid Identification of Thrips palmi

Jangra

Ghosh

Mukherjee

et al. 2022

Front. Mol. Biosci.

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Thrips cause considerable economic losses to a wide range of food, feed, and forest crops. They also transmit several plant viruses. Being cryptic, it is often difficult to distinguish thrips species in crops and large consignments by conventional methods. Melon thrips (Thrips palmi Karny, Thysanoptera: Thripidae) is an invasive insect pest of vegetables, legumes, and ornamentals besides being vector to several viruses. It poses a threat to domestic and international plant biosecurity and can invade and establish in new areas. Here, we report a polymerase spiral reaction (PSR)-based isothermal assay for rapid, sensitive, specific, low-cost, and on-site detection of T. palmi. To the best of our knowledge, this is the first application of PSR in the identification of any insect species. A primer pair designed based on 3′-polymorphism of mtCOIII region can specifically identify T. palmi without any cross-reactivity with predominant thrips species. The assay uses crude lysate of a single thrips saving time and reagents involved in nucleic acid extraction. The presence of T. palmi is visualized by the appearance of bright fluorescence under ultraviolet light or a change in reaction color thus avoiding gel electrophoresis steps. The entire process can be completed in 70 min on-site using only an ordinary water bath. The assay is sensitive to detecting as little as 50 attograms of T. palmi template. The assay was validated with known thrips specimens and found to be efficient in diagnosing T. palmi under natural conditions. The described method will be useful for non-expert personnel to detect an early infestation, accidental introduction to a new area, restrict the spread of diseases and formulate appropriate management strategies.

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“…Morphometric key-based identification of thrips species is timetaking, labor-intensive, requires expertise, and dependent on developmental stage as only adults can be identified. The speed, reproducibility, and accuracy of molecular techniques have made them a valuable tool for identification of thrips vectors [13,[29][30][31][32][33][34][35][36]. The simultaneous identification of several species using multiplex PCR has become popular for its added advantages of saving time, money, and effort over other methods of diagnosis [37,38].…”

Section: Discussionmentioning

confidence: 99%

A multiplex PCR assay for rapid identification of major tospovirus vectors reported in India

et al. 2020

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Background: To date, four thrips vectors have been reported to transmit five different tospoviruses in India. Their identification at an early stage is crucial in formulating appropriate pest management strategies. Since morphometric key-based thrips identification based on the adult stage is time-consuming, there is a need to develop diagnostic tools which are rapid, accurate, and independent of developmental stages. Here, we report a multiplex PCR assay to identify four major thrips vectors viz. Thrips palmi, T. tabaci, Scirtothrips dorsalis, and Frankliniella schultzei present in India. Results: Cytochrome oxidase subunit III and internal transcribed spacer region 2 were utilized to design speciesspecific primers. Of 38 pairs of primers tested, primer pairs AG35F-AG36R, AG47F-AG48R, AG87F-AG88R, and AG79F-AG80R amplified 568 bp, 713 bp, 388 bp, and 200 bp products from the DNA templates of T. palmi, S. dorsalis, T. tabaci, and F. schultzei, respectively at same PCR conditions. The specificity of the primer pairs was validated with a large number of known specimens and no cross-reactivity was observed with other thrips species. The multiplex PCR assay with a cocktail of all the four primer pairs detected four thrips vectors efficiently and could discriminate all of them concurrently in a single reaction. Conclusion: The multiplex PCR reported in this study could identify the major thrips vectors reported in India. The assay will be useful in ascertaining distribution profile of major thrips vectors, disease epidemiology, screening large samples, and quarantine.

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PCR-RFLP method to distinguish Frankliniella occidentalis, Frankliniella intonsa, Frankliniella pallida and Frankliniella tenuicornis

Cited by 11 publications

References 13 publications

Real-time PCR assay for distinguishingFrankliniella occidentalisandThrips palmiArnika Przybylska, Żaneta Fiedler, Aleksandra Obrępalska-Stęplowska

Real-time PCR assay for distinguishingFrankliniella occidentalisandThrips palmiArnika Przybylska, Żaneta Fiedler, Aleksandra Obrępalska-Stęplowska

Development of a Polymerase Spiral Reaction-Based Isothermal Assay for Rapid Identification of Thrips palmi

A multiplex PCR assay for rapid identification of major tospovirus vectors reported in India

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