Novel Tetraplex Quantitative PCR Assays for Simultaneous Detection and Identification of Xylella fastidiosa Subspecies in Plant Tissues
Identifieur interne : 000444 ( Main/Exploration ); précédent : 000443; suivant : 000445Novel Tetraplex Quantitative PCR Assays for Simultaneous Detection and Identification of Xylella fastidiosa Subspecies in Plant Tissues
Auteurs : Enora Dupas [France] ; Martial Briand [France] ; Marie-Agnès Jacques [France] ; Sophie Cesbron [France]Source :
- Frontiers in Plant Science [ 1664-462X ] ; 2019.
Abstract
Url:
DOI: 10.3389/fpls.2019.01732
PubMed: 31956326
PubMed Central: 6951419
Affiliations:
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strict</wicri:regionArea></affiliation></author><author><name sortKey="Cesbron, Sophie" sort="Cesbron, Sophie" uniqKey="Cesbron S" first="Sophie" last="Cesbron">Sophie Cesbron</name><affiliation wicri:level="1"><nlm:aff id="aff1"><institution>IRHS, Agrocampus-Ouest, INRA, University of Angers, SFR 4207 QuaSaV</institution>,<addr-line>Beaucouzé</addr-line>,<country>France</country></nlm:aff><country xml:lang="fr">France</country><wicri:regionArea># see nlm:aff country strict</wicri:regionArea></affiliation></author></titleStmt><publicationStmt><idno type="wicri:source">PMC</idno><idno type="pmid">31956326</idno><idno type="pmc">6951419</idno><idno type="url">http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6951419</idno><idno type="RBID">PMC:6951419</idno><idno type="doi">10.3389/fpls.2019.01732</idno><date when="2019">2019</date><idno type="wicri:Area/Pmc/Corpus">000C87</idno><idno type="wicri:explorRef" wicri:stream="Pmc" wicri:step="Corpus" wicri:corpus="PMC">000C87</idno><idno 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type="wicri:Area/Main/Curation">000444</idno><idno type="wicri:Area/Main/Exploration">000444</idno></publicationStmt><sourceDesc><biblStruct><analytic><title xml:lang="en" level="a" type="main">Novel Tetraplex Quantitative PCR Assays for Simultaneous Detection and Identification of <italic>Xylella fastidiosa</italic> Subspecies in Plant Tissues</title><author><name sortKey="Dupas, Enora" sort="Dupas, Enora" uniqKey="Dupas E" first="Enora" last="Dupas">Enora Dupas</name><affiliation wicri:level="1"><nlm:aff id="aff1"><institution>IRHS, Agrocampus-Ouest, INRA, University of Angers, SFR 4207 QuaSaV</institution>,<addr-line>Beaucouzé</addr-line>,<country>France</country></nlm:aff><country xml:lang="fr">France</country><wicri:regionArea># see nlm:aff country strict</wicri:regionArea></affiliation><affiliation wicri:level="1"><nlm:aff id="aff2"><institution>French Agency for Food, Environmental and Occupational Health & Safety, Plant Health Laboratory</institution>,<addr-line>Angers</addr-line>,<country>France</country></nlm:aff><country xml:lang="fr">France</country><wicri:regionArea># see nlm:aff country strict</wicri:regionArea></affiliation></author><author><name sortKey="Briand, Martial" sort="Briand, Martial" uniqKey="Briand M" first="Martial" last="Briand">Martial Briand</name><affiliation wicri:level="1"><nlm:aff id="aff1"><institution>IRHS, Agrocampus-Ouest, INRA, University of Angers, SFR 4207 QuaSaV</institution>,<addr-line>Beaucouzé</addr-line>,<country>France</country></nlm:aff><country xml:lang="fr">France</country><wicri:regionArea># see nlm:aff country strict</wicri:regionArea></affiliation></author><author><name sortKey="Jacques, Marie Agnes" sort="Jacques, Marie Agnes" uniqKey="Jacques M" first="Marie-Agnès" last="Jacques">Marie-Agnès Jacques</name><affiliation wicri:level="1"><nlm:aff id="aff1"><institution>IRHS, Agrocampus-Ouest, INRA, University of Angers, SFR 4207 QuaSaV</institution>,<addr-line>Beaucouzé</addr-line>,<country>France</country></nlm:aff><country xml:lang="fr">France</country><wicri:regionArea># see nlm:aff country strict</wicri:regionArea></affiliation></author><author><name sortKey="Cesbron, Sophie" sort="Cesbron, Sophie" uniqKey="Cesbron S" first="Sophie" last="Cesbron">Sophie Cesbron</name><affiliation wicri:level="1"><nlm:aff id="aff1"><institution>IRHS, Agrocampus-Ouest, INRA, University of Angers, SFR 4207 QuaSaV</institution>,<addr-line>Beaucouzé</addr-line>,<country>France</country></nlm:aff><country xml:lang="fr">France</country><wicri:regionArea># see nlm:aff country strict</wicri:regionArea></affiliation></author></analytic><series><title level="j">Frontiers in Plant Science</title><idno type="eISSN">1664-462X</idno><imprint><date when="2019">2019</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p><italic>Xylella fastidiosa</italic> (<italic>Xf</italic>) is an insect-borne bacterium confined to the xylem vessels of plants. This plant pathogen has a broad host range estimated to 560 plant species. Five subspecies of the pathogen with different but overlapping host ranges have been described, but only three subspecies are widely accepted, namely subspecies <italic>fastidiosa</italic>, <italic>multiplex</italic>, and <italic>pauca</italic>. Initially limited to the Americas, <italic>Xf</italic> has been detected in Europe since 2013. As management of <italic>X. fastidiosa</italic> outbreaks in Europe depends on the identification of the subspecies, accurate determination of the subspecies in infected plants as early as possible is of major interest. Thus, we developed various tetraplex and triplex quantitative PCR (qPCR) assays for <italic>X. fastidiosa</italic> detection and subspecies identification <italic>in planta</italic> in a single reaction. We designed primers and probes using SkIf, a bioinformatics tool based on <italic>k</italic>-mers, to detect specific signatures of the species and subspecies from a data set of 58 genome sequences representative of <italic>X. fastidiosa</italic> diversity. We tested the qPCR assays on 39 target and 30 non-target strains, as well as on 13 different plant species spiked with strains of the different subspecies of <italic>X. fastidiosa</italic>, and on samples from various environmental and inoculated host plants. Sensitivity of simplex assays was equal or slightly better than the reference protocol on purified DNA. Tetraplex qPCR assays had the same sensitivity than the reference protocol and allowed <italic>X. fastidiosa</italic> detection in all spiked matrices up to 10<sup>3</sup> cells.ml<sup>−1</sup>. Moreover, mix infections of two to three subspecies could be detected in the same sample with tetraplex assays. In environmental plant samples, the tetraplex qPCR assays allowed subspecies identification when the current method based on multilocus sequence typing failed. The qPCR assays described here are robust and modular tools that are efficient for differentiating <italic>X. fastidiosa</italic> subspecies directly in plant samples.</p></div></front><back><div1 type="bibliography"><listBibl><biblStruct><analytic><author><name sortKey="Abraham, N D" uniqKey="Abraham N">N. D. Abraham</name></author><author><name sortKey="Chitrampal, P" uniqKey="Chitrampal P">P. Chitrampal</name></author><author><name sortKey="Kerio, S" uniqKey="Kerio S">S. Keriö</name></author><author><name sortKey="Leboldus, J M" uniqKey="Leboldus J">J. M. LeBoldus</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Bergsma Vlami, M" uniqKey="Bergsma Vlami M">M. Bergsma-Vlami</name></author><author><name sortKey="Van De Bilt, J L J" uniqKey="Van De Bilt J">J. L. 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Bromley</name></author><author><name sortKey="Spring Pearson, S" uniqKey="Spring Pearson S">S. Spring-Pearson</name></author><author><name sortKey="Stouthamer, R" uniqKey="Stouthamer R">R. Stouthamer</name></author><author><name sortKey="Nunney, L" uniqKey="Nunney L">L. Nunney</name></author></analytic></biblStruct><biblStruct><analytic><author><name sortKey="Zitnick Anderson, K" uniqKey="Zitnick Anderson K">K. Zitnick-Anderson</name></author><author><name sortKey="Simons, K" uniqKey="Simons K">K. Simons</name></author><author><name sortKey="Pasche, J S" uniqKey="Pasche J">J. S. Pasche</name></author></analytic></biblStruct></listBibl></div1></back></TEI><affiliations><list><country><li>France</li></country></list><tree><country name="France"><noRegion><name sortKey="Dupas, Enora" sort="Dupas, Enora" uniqKey="Dupas E" first="Enora" last="Dupas">Enora Dupas</name></noRegion><name sortKey="Briand, Martial" sort="Briand, Martial" uniqKey="Briand M" first="Martial" last="Briand">Martial Briand</name><name sortKey="Cesbron, Sophie" sort="Cesbron, Sophie" uniqKey="Cesbron S" first="Sophie" last="Cesbron">Sophie Cesbron</name><name sortKey="Dupas, Enora" sort="Dupas, Enora" uniqKey="Dupas E" first="Enora" last="Dupas">Enora Dupas</name><name sortKey="Jacques, Marie Agnes" sort="Jacques, Marie Agnes" uniqKey="Jacques M" first="Marie-Agnès" last="Jacques">Marie-Agnès Jacques</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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