Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios
Almonds and pistachios are fed upon by a diverse assemblage of lepidopteran insects, several of which are economically important pests. Unfortunately, identification of these pests can be difficult, as specimens are frequently damaged during collection, occur in traps with non-target species, and ar...
Main Authors: | , , |
---|---|
Format: | Article |
Language: | English |
Published: |
MDPI AG
2021-06-01
|
Series: | Insects |
Subjects: | |
Online Access: | https://www.mdpi.com/2075-4450/12/6/553 |
id |
doaj-699e2098848e4c60a5588d4f143628cd |
---|---|
record_format |
Article |
spelling |
doaj-699e2098848e4c60a5588d4f143628cd2021-07-01T00:10:55ZengMDPI AGInsects2075-44502021-06-011255355310.3390/insects12060553Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and PistachiosRohith Vulchi0Kent M. Daane1Jacob A. Wenger2Department of Plant Science, California State University, Fresno, 2415 E. San Ramon Ave, Fresno, CA 93740-8033, USADepartment of Environmental Science, Policy, & Management, University of California Berkeley, Berkeley, CA 94720-3114, USADepartment of Plant Science, California State University, Fresno, 2415 E. San Ramon Ave, Fresno, CA 93740-8033, USAAlmonds and pistachios are fed upon by a diverse assemblage of lepidopteran insects, several of which are economically important pests. Unfortunately, identification of these pests can be difficult, as specimens are frequently damaged during collection, occur in traps with non-target species, and are morphologically similar up to their third instar. Here, we present a quantitative PCR based melt curve analysis for simple, rapid, and accurate identification of six lepidopteran pests of almonds and pistachios: navel orangeworm (<i>Amyelois transitella</i>), peach twig borer (<i>Anarsia lineatella</i>), oriental fruit moth (<i>Grapholita molesta)</i>, obliquebanded leafroller (<i>Choristoneura rosaceana)</i>, raisin moth (<i>Cadra figulilella</i>), and Indian meal moth (<i>Plodia interpunctella</i>). In this approach, the dissociation (melt) temperature(s) of a 658 bp section of cytochrome c oxidase subunit 1 was determined using quantitative PCR (qPCR). Within these six species, the distribution and the number of melt peak temperatures provide an unambiguous species level identification that is reproducible when unsheared DNA can be extracted. The test is robust across a variety of sampling approaches including insects removed from sticky card traps, museum specimens, and samples that were left in the field for up to 7 days. The melt curve’s simplicity allows it to be performed in any basic molecular biology laboratory with a quantitative PCR.https://www.mdpi.com/2075-4450/12/6/553quantitative PCRreal-time PCRpest identificationorchardsdiagnostic protocolmelt curve profile |
collection |
DOAJ |
language |
English |
format |
Article |
sources |
DOAJ |
author |
Rohith Vulchi Kent M. Daane Jacob A. Wenger |
spellingShingle |
Rohith Vulchi Kent M. Daane Jacob A. Wenger Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios Insects quantitative PCR real-time PCR pest identification orchards diagnostic protocol melt curve profile |
author_facet |
Rohith Vulchi Kent M. Daane Jacob A. Wenger |
author_sort |
Rohith Vulchi |
title |
Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios |
title_short |
Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios |
title_full |
Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios |
title_fullStr |
Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios |
title_full_unstemmed |
Development of DNA Melt Curve Analysis for the Identification of Lepidopteran Pests in Almonds and Pistachios |
title_sort |
development of dna melt curve analysis for the identification of lepidopteran pests in almonds and pistachios |
publisher |
MDPI AG |
series |
Insects |
issn |
2075-4450 |
publishDate |
2021-06-01 |
description |
Almonds and pistachios are fed upon by a diverse assemblage of lepidopteran insects, several of which are economically important pests. Unfortunately, identification of these pests can be difficult, as specimens are frequently damaged during collection, occur in traps with non-target species, and are morphologically similar up to their third instar. Here, we present a quantitative PCR based melt curve analysis for simple, rapid, and accurate identification of six lepidopteran pests of almonds and pistachios: navel orangeworm (<i>Amyelois transitella</i>), peach twig borer (<i>Anarsia lineatella</i>), oriental fruit moth (<i>Grapholita molesta)</i>, obliquebanded leafroller (<i>Choristoneura rosaceana)</i>, raisin moth (<i>Cadra figulilella</i>), and Indian meal moth (<i>Plodia interpunctella</i>). In this approach, the dissociation (melt) temperature(s) of a 658 bp section of cytochrome c oxidase subunit 1 was determined using quantitative PCR (qPCR). Within these six species, the distribution and the number of melt peak temperatures provide an unambiguous species level identification that is reproducible when unsheared DNA can be extracted. The test is robust across a variety of sampling approaches including insects removed from sticky card traps, museum specimens, and samples that were left in the field for up to 7 days. The melt curve’s simplicity allows it to be performed in any basic molecular biology laboratory with a quantitative PCR. |
topic |
quantitative PCR real-time PCR pest identification orchards diagnostic protocol melt curve profile |
url |
https://www.mdpi.com/2075-4450/12/6/553 |
work_keys_str_mv |
AT rohithvulchi developmentofdnameltcurveanalysisfortheidentificationoflepidopteranpestsinalmondsandpistachios AT kentmdaane developmentofdnameltcurveanalysisfortheidentificationoflepidopteranpestsinalmondsandpistachios AT jacobawenger developmentofdnameltcurveanalysisfortheidentificationoflepidopteranpestsinalmondsandpistachios |
_version_ |
1721349332813217792 |