Double-Stranded RNA Uptake through Topical Application, Mediates Silencing of Five CYP4 Genes and Suppresses Insecticide Resistance in Diaphorina citri
Identifieur interne : 000213 ( Pmc/Curation ); précédent : 000212; suivant : 000214Double-Stranded RNA Uptake through Topical Application, Mediates Silencing of Five CYP4 Genes and Suppresses Insecticide Resistance in Diaphorina citri
Auteurs : Nabil Killiny [États-Unis] ; Subhas Hajeri [États-Unis] ; Siddharth Tiwari [États-Unis] ; Siddarame Gowda [États-Unis] ; Lukasz L. Stelinski [États-Unis]Source :
- PLoS ONE [ 1932-6203 ] ; 2014.
Abstract
Silencing of genes through RNA interference (RNAi) in insects has gained momentum during the past few years. RNAi has been used to cause insect mortality, inhibit insect growth, increase insecticide susceptibility, and prevent the development of insecticide resistance. We investigated the efficacy of topically applied dsRNA to induce RNAi for five Cytochrome P450 genes family 4 (
Url:
DOI: 10.1371/journal.pone.0110536
PubMed: 25330026
PubMed Central: 4203802
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<record><TEI><teiHeader><fileDesc><titleStmt><title xml:lang="en">Double-Stranded RNA Uptake through Topical Application, Mediates Silencing of Five CYP4 Genes and Suppresses Insecticide Resistance in <italic>Diaphorina citri</italic></title><author><name sortKey="Killiny, Nabil" sort="Killiny, Nabil" uniqKey="Killiny N" first="Nabil" last="Killiny">Nabil Killiny</name><affiliation wicri:level="1"><nlm:aff id="aff1"><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Hajeri, Subhas" sort="Hajeri, Subhas" uniqKey="Hajeri S" first="Subhas" last="Hajeri">Subhas Hajeri</name><affiliation wicri:level="1"><nlm:aff id="aff2"><addr-line>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Tiwari, Siddharth" sort="Tiwari, Siddharth" uniqKey="Tiwari S" first="Siddharth" last="Tiwari">Siddharth Tiwari</name><affiliation wicri:level="1"><nlm:aff id="aff1"><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Gowda, Siddarame" sort="Gowda, Siddarame" uniqKey="Gowda S" first="Siddarame" last="Gowda">Siddarame Gowda</name><affiliation wicri:level="1"><nlm:aff id="aff2"><addr-line>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Stelinski, Lukasz L" sort="Stelinski, Lukasz L" uniqKey="Stelinski L" first="Lukasz L." last="Stelinski">Lukasz L. Stelinski</name><affiliation wicri:level="1"><nlm:aff id="aff1"><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">25330026</idno><idno type="pmc">4203802</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4203802</idno><idno type="RBID">PMC:4203802</idno><idno type="doi">10.1371/journal.pone.0110536</idno><date when="2014">2014</date><idno type="wicri:Area/Pmc/Corpus">000213</idno><idno type="wicri:Area/Pmc/Curation">000213</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Double-Stranded RNA Uptake through Topical Application, Mediates Silencing of Five CYP4 Genes and Suppresses Insecticide Resistance in <italic>Diaphorina citri</italic></title><author><name sortKey="Killiny, Nabil" sort="Killiny, Nabil" uniqKey="Killiny N" first="Nabil" last="Killiny">Nabil Killiny</name><affiliation wicri:level="1"><nlm:aff id="aff1"><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Hajeri, Subhas" sort="Hajeri, Subhas" uniqKey="Hajeri S" first="Subhas" last="Hajeri">Subhas Hajeri</name><affiliation wicri:level="1"><nlm:aff id="aff2"><addr-line>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Tiwari, Siddharth" sort="Tiwari, Siddharth" uniqKey="Tiwari S" first="Siddharth" last="Tiwari">Siddharth Tiwari</name><affiliation wicri:level="1"><nlm:aff id="aff1"><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Gowda, Siddarame" sort="Gowda, Siddarame" uniqKey="Gowda S" first="Siddarame" last="Gowda">Siddarame Gowda</name><affiliation wicri:level="1"><nlm:aff id="aff2"><addr-line>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author><author><name sortKey="Stelinski, Lukasz L" sort="Stelinski, Lukasz L" uniqKey="Stelinski L" first="Lukasz L." last="Stelinski">Lukasz L. Stelinski</name><affiliation wicri:level="1"><nlm:aff id="aff1"><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></nlm:aff><country xml:lang="fr">États-Unis</country><wicri:regionArea>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida</wicri:regionArea></affiliation></author></analytic><series><title level="j">PLoS ONE</title><idno type="eISSN">1932-6203</idno><imprint><date when="2014">2014</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p>Silencing of genes through RNA interference (RNAi) in insects has gained momentum during the past few years. RNAi has been used to cause insect mortality, inhibit insect growth, increase insecticide susceptibility, and prevent the development of insecticide resistance. We investigated the efficacy of topically applied dsRNA to induce RNAi for five Cytochrome P<sub>450</sub> genes family 4 (<italic>CYP4</italic>) in <italic>Diaphorina citri</italic>. We previously reported that these <italic>CYP4</italic> genes are associated with the development of insecticide resistance in <italic>D. citri</italic>. We targeted five <italic>CYP4</italic> genes that share a consensus sequence with one dsRNA construct. Quantitative PCR confirmed suppressed expression of the five <italic>CYP4</italic> genes as a result of dsRNA topically applied to the thoracic region of <italic>D. citri</italic> when compared to the expression levels in a control group. Western blot analysis indicated a reduced signal of cytochrome P<sub>450</sub> proteins (45 kDa) in adult <italic>D. citri</italic> treated with the dsRNA. In addition, oxidase activity and insecticide resistance were reduced for <italic>D. citri</italic> treated with dsRNA that targeted specific <italic>CYP4</italic> genes. Mortality was significantly higher in adults treated with dsRNA than in adults treated with water. Our results indicate that topically applied dsRNA can penetrate the cuticle of <italic>D. citri</italic> and induce RNAi. These results broaden the scope of RNAi as a mechanism to manage pests by targeting a broad range of genes. The results also support the application of RNAi as a viable tool to overcome insecticide resistance development in <italic>D. citri</italic> populations. However, further research is needed to develop grower-friendly delivery systems for the application of dsRNA under field conditions. Considering the high specificity of dsRNA, this tool can also be used for management of <italic>D. citri</italic> by targeting physiologically critical genes involved in growth and development.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct></biblStruct><biblStruct><analytic><author><name sortKey="Huvenne, H" uniqKey="Huvenne H">H Huvenne</name></author><author><name sortKey="Smagghe, G" uniqKey="Smagghe G">G Smagghe</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Izant, Jg" uniqKey="Izant J">JG Izant</name></author><author><name sortKey="Weintraub, H" uniqKey="Weintraub H">H Weintraub</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Fire, A" uniqKey="Fire A">A Fire</name></author><author><name sortKey="Xu, S" uniqKey="Xu S">S Xu</name></author><author><name sortKey="Montgomery, Mk" uniqKey="Montgomery M">MK Montgomery</name></author><author><name sortKey="Kostas, Sa" uniqKey="Kostas S">SA 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<front><journal-meta><journal-id journal-id-type="nlm-ta">PLoS One</journal-id><journal-id journal-id-type="iso-abbrev">PLoS ONE</journal-id><journal-id journal-id-type="publisher-id">plos</journal-id><journal-id journal-id-type="pmc">plosone</journal-id><journal-title-group><journal-title>PLoS ONE</journal-title></journal-title-group><issn pub-type="epub">1932-6203</issn><publisher><publisher-name>Public Library of Science</publisher-name><publisher-loc>San Francisco, USA</publisher-loc></publisher></journal-meta><article-meta><article-id pub-id-type="pmid">25330026</article-id><article-id pub-id-type="pmc">4203802</article-id><article-id pub-id-type="publisher-id">PONE-D-14-33137</article-id><article-id pub-id-type="doi">10.1371/journal.pone.0110536</article-id><article-categories><subj-group subj-group-type="heading"><subject>Research Article</subject></subj-group><subj-group subj-group-type="Discipline-v2"><subject>Biology and Life Sciences</subject><subj-group><subject>Agriculture</subject><subj-group><subject>Pest Control</subject></subj-group></subj-group><subj-group><subject>Biochemistry</subject></subj-group><subj-group><subject>Biotechnology</subject></subj-group><subj-group><subject>Genetics</subject></subj-group></subj-group></article-categories><title-group><article-title>Double-Stranded RNA Uptake through Topical Application, Mediates Silencing of Five CYP4 Genes and Suppresses Insecticide Resistance in <italic>Diaphorina citri</italic></article-title><alt-title alt-title-type="running-head">Silencing of Cytochrome P<sub>450</sub> in <italic>Diaphorina citri</italic></alt-title></title-group><contrib-group><contrib contrib-type="author"><name><surname>Killiny</surname><given-names>Nabil</given-names></name><xref ref-type="aff" rid="aff1"><sup>1</sup></xref><xref ref-type="corresp" rid="cor1"><sup>*</sup></xref></contrib><contrib contrib-type="author"><name><surname>Hajeri</surname><given-names>Subhas</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref></contrib><contrib contrib-type="author"><name><surname>Tiwari</surname><given-names>Siddharth</given-names></name><xref ref-type="aff" rid="aff1"><sup>1</sup></xref></contrib><contrib contrib-type="author"><name><surname>Gowda</surname><given-names>Siddarame</given-names></name><xref ref-type="aff" rid="aff2"><sup>2</sup></xref></contrib><contrib contrib-type="author"><name><surname>Stelinski</surname><given-names>Lukasz L.</given-names></name><xref ref-type="aff" rid="aff1"><sup>1</sup></xref></contrib></contrib-group><aff id="aff1"><label>1</label><addr-line>Department of Entomology and Nematology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></aff><aff id="aff2"><label>2</label><addr-line>Department of Plant Pathology, Citrus Research and Education Center, IFAS, University of Florida, Lake Alfred, Florida, United States of America</addr-line></aff><contrib-group><contrib contrib-type="editor"><name><surname>Hansen</surname><given-names>Immo A.</given-names></name><role>Editor</role><xref ref-type="aff" rid="edit1"></xref></contrib></contrib-group><aff id="edit1"><addr-line>New Mexico State University, United States of America</addr-line></aff><author-notes><corresp id="cor1">* E-mail: <email>nabikilliny@ufl.edu</email></corresp><fn fn-type="conflict"><p><bold>Competing Interests: </bold>The authors have declared that no competing interests exist.</p></fn><fn fn-type="con"><p>Conceived and designed the experiments: NK SH ST SG LLS. Performed the experiments: NK SH SG. Analyzed the data: NK SH ST SG LLS. Contributed reagents/materials/analysis tools: NK SH ST SG LLS. Contributed to the writing of the manuscript: NK SH ST SG LLS.</p></fn></author-notes><pub-date pub-type="collection"><year>2014</year></pub-date><pub-date pub-type="epub"><day>20</day><month>10</month><year>2014</year></pub-date><volume>9</volume><issue>10</issue><elocation-id>e110536</elocation-id><history><date date-type="received"><day>1</day><month>8</month><year>2014</year></date><date date-type="accepted"><day>16</day><month>9</month><year>2014</year></date></history><permissions><copyright-year>2014</copyright-year><copyright-holder>Killiny et al</copyright-holder><license><license-p>This is an open-access article distributed under the terms of the <ext-link ext-link-type="uri" xlink:href="http://creativecommons.org/licenses/by/4.0/">Creative Commons Attribution License</ext-link>, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.</license-p></license></permissions><abstract><p>Silencing of genes through RNA interference (RNAi) in insects has gained momentum during the past few years. RNAi has been used to cause insect mortality, inhibit insect growth, increase insecticide susceptibility, and prevent the development of insecticide resistance. We investigated the efficacy of topically applied dsRNA to induce RNAi for five Cytochrome P<sub>450</sub> genes family 4 (<italic>CYP4</italic>) in <italic>Diaphorina citri</italic>. We previously reported that these <italic>CYP4</italic> genes are associated with the development of insecticide resistance in <italic>D. citri</italic>. We targeted five <italic>CYP4</italic> genes that share a consensus sequence with one dsRNA construct. Quantitative PCR confirmed suppressed expression of the five <italic>CYP4</italic> genes as a result of dsRNA topically applied to the thoracic region of <italic>D. citri</italic> when compared to the expression levels in a control group. Western blot analysis indicated a reduced signal of cytochrome P<sub>450</sub> proteins (45 kDa) in adult <italic>D. citri</italic> treated with the dsRNA. In addition, oxidase activity and insecticide resistance were reduced for <italic>D. citri</italic> treated with dsRNA that targeted specific <italic>CYP4</italic> genes. Mortality was significantly higher in adults treated with dsRNA than in adults treated with water. Our results indicate that topically applied dsRNA can penetrate the cuticle of <italic>D. citri</italic> and induce RNAi. These results broaden the scope of RNAi as a mechanism to manage pests by targeting a broad range of genes. The results also support the application of RNAi as a viable tool to overcome insecticide resistance development in <italic>D. citri</italic> populations. However, further research is needed to develop grower-friendly delivery systems for the application of dsRNA under field conditions. Considering the high specificity of dsRNA, this tool can also be used for management of <italic>D. citri</italic> by targeting physiologically critical genes involved in growth and development.</p></abstract><funding-group><funding-statement>This project was supported by a grant to LS from the Citrus Research and Development Foundation. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.</funding-statement></funding-group><counts><page-count count="8"></page-count></counts><custom-meta-group><custom-meta id="data-availability"><meta-name>Data Availability</meta-name><meta-value>The authors confirm that all data underlying the findings are fully available without restriction. All relevant data are within the paper.</meta-value></custom-meta></custom-meta-group></article-meta><notes><title>Data Availability</title><p>The authors confirm that all data underlying the findings are fully available without restriction. All relevant data are within the paper.</p></notes></front></pmc></record>Pour manipuler ce document sous Unix (Dilib)
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