Profiling grapevine trunk pathogens in planta: a case for community-targeted DNA metabarcoding.
Identifieur interne : 000209 ( Main/Corpus ); précédent : 000208; suivant : 000210Profiling grapevine trunk pathogens in planta: a case for community-targeted DNA metabarcoding.
Auteurs : Abraham Morales-Cruz ; Rosa Figueroa-Balderas ; Jadran F. García ; Eric Tran ; Philippe E. Rolshausen ; Kendra Baumgartner ; Dario CantuSource :
- BMC microbiology [ 1471-2180 ] ; 2018.
English descriptors
- KwdEn :
- Ascomycota (classification), Ascomycota (genetics), Ascomycota (isolation & purification), DNA Barcoding, Taxonomic (methods), DNA, Fungal (genetics), High-Throughput Nucleotide Sequencing (MeSH), Metagenomics (MeSH), Mycological Typing Techniques (methods), Plant Diseases (microbiology), Vitis (microbiology).
- MESH :
- chemical , genetics : DNA, Fungal.
- classification : Ascomycota.
- genetics : Ascomycota.
- isolation & purification : Ascomycota.
- methods : DNA Barcoding, Taxonomic, Mycological Typing Techniques.
- microbiology : Plant Diseases, Vitis.
- High-Throughput Nucleotide Sequencing, Metagenomics.
Abstract
BACKGROUND
DNA metabarcoding, commonly used in exploratory microbial ecology studies, is a promising method for the simultaneous in planta-detection of multiple pathogens associated with disease complexes, such as the grapevine trunk diseases. Profiling of pathogen communities associated with grapevine trunk diseases is particularly challenging, due to the presence within an individual wood lesion of multiple co-infecting trunk pathogens and other wood-colonizing fungi, which span a broad range of taxa in the fungal kingdom. As such, we designed metabarcoding primers, using as template the ribosomal internal transcribed spacer of grapevine trunk-associated ascomycete fungi (GTAA) and compared them to two universal primer widely used in microbial ecology.
RESULTS
We first performed in silico simulations and then tested the primers by high-throughput amplicon sequencing of (i) multiple combinations of mock communities, (ii) time-course experiments with controlled inoculations, and (iii) diseased field samples from vineyards under natural levels of infection. All analyses showed that GTAA had greater affinity and sensitivity, compared to those of the universal primers. Importantly, with GTAA, profiling of mock communities and comparisons with shotgun-sequencing metagenomics of field samples gave an accurate representation of genera of important trunk pathogens, namely Phaeomoniella, Phaeoacremonium, and Eutypa, the abundances of which were over- or under-estimated with universal primers.
CONCLUSIONS
Overall, our findings not only demonstrate that DNA metabarcoding gives qualitatively and quantitatively accurate results when applied to grapevine trunk diseases, but also that primer customization and testing are crucial to ensure the validity of DNA metabarcoding results.
DOI: 10.1186/s12866-018-1343-0
PubMed: 30547761
PubMed Central: PMC6295080
Links to Exploration step
pubmed:30547761Le document en format XML
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García</name><affiliation><nlm:affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA.</nlm:affiliation></affiliation></author><author><name sortKey="Tran, Eric" sort="Tran, Eric" uniqKey="Tran E" first="Eric" last="Tran">Eric Tran</name><affiliation><nlm:affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA.</nlm:affiliation></affiliation></author><author><name sortKey="Rolshausen, Philippe E" sort="Rolshausen, Philippe E" uniqKey="Rolshausen P" first="Philippe E" last="Rolshausen">Philippe E. Rolshausen</name><affiliation><nlm:affiliation>Department of Botany and Plant Sciences, University of California, Riverside, CA, 92521, USA.</nlm:affiliation></affiliation></author><author><name sortKey="Baumgartner, Kendra" sort="Baumgartner, Kendra" uniqKey="Baumgartner K" first="Kendra" last="Baumgartner">Kendra Baumgartner</name><affiliation><nlm:affiliation>United States Department of Agriculture, Agricultural Research Service, Crops Pathology and Genetics Research Unit, Davis, CA, 95616, USA.</nlm:affiliation></affiliation></author><author><name sortKey="Cantu, Dario" sort="Cantu, Dario" uniqKey="Cantu D" first="Dario" last="Cantu">Dario Cantu</name><affiliation><nlm:affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA. dacantu@ucdavis.edu.</nlm:affiliation></affiliation></author></analytic><series><title level="j">BMC microbiology</title><idno type="eISSN">1471-2180</idno><imprint><date when="2018" type="published">2018</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass><keywords scheme="KwdEn" xml:lang="en"><term>Ascomycota (classification)</term><term>Ascomycota (genetics)</term><term>Ascomycota (isolation & purification)</term><term>DNA Barcoding, Taxonomic (methods)</term><term>DNA, Fungal (genetics)</term><term>High-Throughput Nucleotide Sequencing (MeSH)</term><term>Metagenomics (MeSH)</term><term>Mycological Typing Techniques (methods)</term><term>Plant Diseases (microbiology)</term><term>Vitis (microbiology)</term></keywords><keywords scheme="MESH" type="chemical" qualifier="genetics" xml:lang="en"><term>DNA, Fungal</term></keywords><keywords scheme="MESH" qualifier="classification" xml:lang="en"><term>Ascomycota</term></keywords><keywords scheme="MESH" qualifier="genetics" xml:lang="en"><term>Ascomycota</term></keywords><keywords scheme="MESH" qualifier="isolation & purification" xml:lang="en"><term>Ascomycota</term></keywords><keywords scheme="MESH" qualifier="methods" xml:lang="en"><term>DNA Barcoding, Taxonomic</term><term>Mycological Typing Techniques</term></keywords><keywords scheme="MESH" qualifier="microbiology" xml:lang="en"><term>Plant Diseases</term><term>Vitis</term></keywords><keywords scheme="MESH" xml:lang="en"><term>High-Throughput Nucleotide Sequencing</term><term>Metagenomics</term></keywords></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en"><p><b>BACKGROUND</b></p><p>DNA metabarcoding, commonly used in exploratory microbial ecology studies, is a promising method for the simultaneous in planta-detection of multiple pathogens associated with disease complexes, such as the grapevine trunk diseases. Profiling of pathogen communities associated with grapevine trunk diseases is particularly challenging, due to the presence within an individual wood lesion of multiple co-infecting trunk pathogens and other wood-colonizing fungi, which span a broad range of taxa in the fungal kingdom. As such, we designed metabarcoding primers, using as template the ribosomal internal transcribed spacer of grapevine trunk-associated ascomycete fungi (GTAA) and compared them to two universal primer widely used in microbial ecology.</p></div><div type="abstract" xml:lang="en"><p><b>RESULTS</b></p><p>We first performed in silico simulations and then tested the primers by high-throughput amplicon sequencing of (i) multiple combinations of mock communities, (ii) time-course experiments with controlled inoculations, and (iii) diseased field samples from vineyards under natural levels of infection. All analyses showed that GTAA had greater affinity and sensitivity, compared to those of the universal primers. Importantly, with GTAA, profiling of mock communities and comparisons with shotgun-sequencing metagenomics of field samples gave an accurate representation of genera of important trunk pathogens, namely Phaeomoniella, Phaeoacremonium, and Eutypa, the abundances of which were over- or under-estimated with universal primers.</p></div><div type="abstract" xml:lang="en"><p><b>CONCLUSIONS</b></p><p>Overall, our findings not only demonstrate that DNA metabarcoding gives qualitatively and quantitatively accurate results when applied to grapevine trunk diseases, but also that primer customization and testing are crucial to ensure the validity of DNA metabarcoding results.</p></div></front></TEI><pubmed><MedlineCitation Status="MEDLINE" Owner="NLM"><PMID Version="1">30547761</PMID><DateCompleted><Year>2019</Year><Month>08</Month><Day>16</Day></DateCompleted><DateRevised><Year>2020</Year><Month>03</Month><Day>09</Day></DateRevised><Article PubModel="Electronic"><Journal><ISSN IssnType="Electronic">1471-2180</ISSN><JournalIssue CitedMedium="Internet"><Volume>18</Volume><Issue>1</Issue><PubDate><Year>2018</Year><Month>12</Month><Day>14</Day></PubDate></JournalIssue><Title>BMC microbiology</Title><ISOAbbreviation>BMC Microbiol</ISOAbbreviation></Journal><ArticleTitle>Profiling grapevine trunk pathogens in planta: a case for community-targeted DNA metabarcoding.</ArticleTitle><Pagination><MedlinePgn>214</MedlinePgn></Pagination><ELocationID EIdType="doi" ValidYN="Y">10.1186/s12866-018-1343-0</ELocationID><Abstract><AbstractText Label="BACKGROUND">DNA metabarcoding, commonly used in exploratory microbial ecology studies, is a promising method for the simultaneous in planta-detection of multiple pathogens associated with disease complexes, such as the grapevine trunk diseases. Profiling of pathogen communities associated with grapevine trunk diseases is particularly challenging, due to the presence within an individual wood lesion of multiple co-infecting trunk pathogens and other wood-colonizing fungi, which span a broad range of taxa in the fungal kingdom. As such, we designed metabarcoding primers, using as template the ribosomal internal transcribed spacer of grapevine trunk-associated ascomycete fungi (GTAA) and compared them to two universal primer widely used in microbial ecology.</AbstractText><AbstractText Label="RESULTS">We first performed in silico simulations and then tested the primers by high-throughput amplicon sequencing of (i) multiple combinations of mock communities, (ii) time-course experiments with controlled inoculations, and (iii) diseased field samples from vineyards under natural levels of infection. All analyses showed that GTAA had greater affinity and sensitivity, compared to those of the universal primers. Importantly, with GTAA, profiling of mock communities and comparisons with shotgun-sequencing metagenomics of field samples gave an accurate representation of genera of important trunk pathogens, namely Phaeomoniella, Phaeoacremonium, and Eutypa, the abundances of which were over- or under-estimated with universal primers.</AbstractText><AbstractText Label="CONCLUSIONS">Overall, our findings not only demonstrate that DNA metabarcoding gives qualitatively and quantitatively accurate results when applied to grapevine trunk diseases, but also that primer customization and testing are crucial to ensure the validity of DNA metabarcoding results.</AbstractText></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Morales-Cruz</LastName><ForeName>Abraham</ForeName><Initials>A</Initials><AffiliationInfo><Affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Figueroa-Balderas</LastName><ForeName>Rosa</ForeName><Initials>R</Initials><AffiliationInfo><Affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>García</LastName><ForeName>Jadran F</ForeName><Initials>JF</Initials><AffiliationInfo><Affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Tran</LastName><ForeName>Eric</ForeName><Initials>E</Initials><AffiliationInfo><Affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Rolshausen</LastName><ForeName>Philippe E</ForeName><Initials>PE</Initials><AffiliationInfo><Affiliation>Department of Botany and Plant Sciences, University of California, Riverside, CA, 92521, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Baumgartner</LastName><ForeName>Kendra</ForeName><Initials>K</Initials><AffiliationInfo><Affiliation>United States Department of Agriculture, Agricultural Research Service, Crops Pathology and Genetics Research Unit, Davis, CA, 95616, USA.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Cantu</LastName><ForeName>Dario</ForeName><Initials>D</Initials><Identifier Source="ORCID">0000-0002-4858-1508</Identifier><AffiliationInfo><Affiliation>Department of Viticulture and Enology, University of California Davis, One Shields Ave, Davis, CA, 95616, USA. dacantu@ucdavis.edu.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D023362">Evaluation Study</PublicationType><PublicationType UI="D016428">Journal Article</PublicationType><PublicationType UI="D013485">Research Support, 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Taxonomic</DescriptorName><QualifierName UI="Q000379" MajorTopicYN="Y">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D004271" MajorTopicYN="N">DNA, Fungal</DescriptorName><QualifierName UI="Q000235" MajorTopicYN="N">genetics</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D059014" MajorTopicYN="N">High-Throughput Nucleotide Sequencing</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D056186" MajorTopicYN="N">Metagenomics</DescriptorName></MeshHeading><MeshHeading><DescriptorName UI="D016533" MajorTopicYN="N">Mycological Typing Techniques</DescriptorName><QualifierName UI="Q000379" MajorTopicYN="Y">methods</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D010935" MajorTopicYN="N">Plant Diseases</DescriptorName><QualifierName UI="Q000382" MajorTopicYN="Y">microbiology</QualifierName></MeshHeading><MeshHeading><DescriptorName UI="D027843" MajorTopicYN="N">Vitis</DescriptorName><QualifierName UI="Q000382" MajorTopicYN="Y">microbiology</QualifierName></MeshHeading></MeshHeadingList><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="Y">Amplicon sequencing</Keyword><Keyword MajorTopicYN="Y">Botryosphaeria dieback</Keyword><Keyword MajorTopicYN="Y">Esca</Keyword><Keyword MajorTopicYN="Y">Eutypa dieback</Keyword><Keyword MajorTopicYN="Y">Grapevine trunk diseases</Keyword><Keyword MajorTopicYN="Y">High-throughput DNA sequencing</Keyword><Keyword MajorTopicYN="Y">Metagenomics</Keyword><Keyword MajorTopicYN="Y">Phomopsis dieback</Keyword></KeywordList></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2018</Year><Month>08</Month><Day>16</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2018</Year><Month>11</Month><Day>15</Day></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2018</Year><Month>12</Month><Day>15</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate 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