Nested-quantitative PCR approach with improved sensitivity for the detection of low titer levels of Candidatus Liberibacter asiaticus in the Asian citrus psyllid, Diaphorina citri Kuwayama
Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal agent of citrus greening disease. The current method of detection for CLas within plant and insect samples is by a presence/absence qPCR assay...
Saved in:
Published in | Journal of microbiological methods Vol. 102; pp. 15 - 22 |
---|---|
Main Authors | , , , , , |
Format | Journal Article |
Language | English |
Published |
Netherlands
Elsevier B.V
01.07.2014
|
Subjects | |
Online Access | Get full text |
Cover
Loading…
Abstract | Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal agent of citrus greening disease. The current method of detection for CLas within plant and insect samples is by a presence/absence qPCR assay using the CLas 16S rDNA gene target. Although qPCR is highly sensitive, low bacterial titers or suboptimal qPCR conditions can result in false-negatives. Using a nested qPCR assay, we determined the false-negative rate of the 16S presence/absence qPCR assay was greater than 50%. Studies to determine the performance parameters of the qPCR assays for CLas 16S and Wingless (Wg), the D. citri endogenous gene, using plasmid and psyllid DNA, revealed suboptimal and variable performance of the 16S assay in psyllid samples. Average efficiencies and sensitivity limits of the plasmid assays were 99.0% and 2.7 copies of template for Wg, respectively, and 98.5% and 2.2-22.1 copies for 16S, respectively. Variability in efficiency was significantly greater in psyllid samples for both gene targets compared to the corresponding plasmid assays, and efficiencies as low as 76% were obtained for 16S. A secondary structure analysis revealed the formation of two stem-loop structures that block the forward and probe binding sites in the 16S template, which could hinder amplification. In summary, our results suggest that suboptimal qPCR efficiency is not uncommon for the 16S presence/absence qPCR assay, which combined with lowCLas titers in some samples, could contribute significantly to the under-reporting of CLas infection in psyllid and plant samples.
•A nested-qPCR assay to detect Candidatus Liberibacter asiaticus is described.•The false-negative rate using the current qPCR method is greater than 50%.•Performance of the qPCR gene targets used in the current method is reported.•The 16S target used in the current method is suboptimal and highly variable.•Low abundance and secondary structure of 16S probably contribute to poor performance. |
---|---|
AbstractList | Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal agent of citrus greening disease. The current method of detection for CLas within plant and insect samples is by a presence/absence qPCR assay using the CLas 16S rDNA gene target. Although qPCR is highly sensitive, low bacterial titers or suboptimal qPCR conditions can result in false-negatives. Using a nested qPCR assay, we determined the false-negative rate of the 16S presence/absence qPCR assay was greater than 50%. Studies to determine the performance parameters of the qPCR assays for CLas 16S and Wingless (Wg), the D. citri endogenous gene, using plasmid and psyllid DNA, revealed suboptimal and variable performance of the 16S assay in psyllid samples. Average efficiencies and sensitivity limits of the plasmid assays were 99.0% and 2.7 copies of template for Wg, respectively, and 98.5% and 2.2-22.1 copies for 16S, respectively. Variability in efficiency was significantly greater in psyllid samples for both gene targets compared to the corresponding plasmid assays, and efficiencies as low as 76% were obtained for 16S. A secondary structure analysis revealed the formation of two stem-loop structures that block the forward and probe binding sites in the 16S template, which could hinder amplification. In summary, our results suggest that suboptimal qPCR efficiency is not uncommon for the 16S presence/absence qPCR assay, which combined with lowCLas titers in some samples, could contribute significantly to the under-reporting of CLas infection in psyllid and plant samples.
•A nested-qPCR assay to detect Candidatus Liberibacter asiaticus is described.•The false-negative rate using the current qPCR method is greater than 50%.•Performance of the qPCR gene targets used in the current method is reported.•The 16S target used in the current method is suboptimal and highly variable.•Low abundance and secondary structure of 16S probably contribute to poor performance. Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal agent of citrus greening disease. The current method of detection for CLas within plant and insect samples is by a presence/absence qPCR assay using the CLas 16S rDNA gene target. Although qPCR is highly sensitive, low bacterial titers or suboptimal qPCR conditions can result in false-negatives. Using a nested qPCR assay, we determined the false-negative rate of the 16S presence/absence qPCR assay was greater than 50%. Studies to determine the performance parameters of the qPCR assays for CLas 16S and Wingless (Wg), the D. citri endogenous gene, using plasmid and psyllid DNA, revealed suboptimal and variable performance of the 16S assay in psyllid samples. Average efficiencies and sensitivity limits of the plasmid assays were 99.0% and 2.7 copies of template for Wg, respectively, and 98.5% and 2.2-22.1 copies for 16S, respectively. Variability in efficiency was significantly greater in psyllid samples for both gene targets compared to the corresponding plasmid assays, and efficiencies as low as 76% were obtained for 16S. A secondary structure analysis revealed the formation of two stem-loop structures that block the forward and probe binding sites in the 16S template, which could hinder amplification. In summary, our results suggest that suboptimal qPCR efficiency is not uncommon for the 16S presence/absence qPCR assay, which combined with lowCLas titers in some samples, could contribute significantly to the under-reporting of CLas infection in psyllid and plant samples. |
Author | Pelz-Stelinski, K.S. Kuhns, E.H. Hoffmann, M. Stelinski, L.L. Kingdom Gibbard, H.N. Coy, M.R. |
Author_xml | – sequence: 1 givenname: M.R. surname: Coy fullname: Coy, M.R. email: mrcoy@ufl.edu – sequence: 2 givenname: M. surname: Hoffmann fullname: Hoffmann, M. – sequence: 3 givenname: H.N. surname: Kingdom Gibbard fullname: Kingdom Gibbard, H.N. – sequence: 4 givenname: E.H. surname: Kuhns fullname: Kuhns, E.H. – sequence: 5 givenname: K.S. surname: Pelz-Stelinski fullname: Pelz-Stelinski, K.S. – sequence: 6 givenname: L.L. surname: Stelinski fullname: Stelinski, L.L. |
BackLink | https://www.ncbi.nlm.nih.gov/pubmed/24769405$$D View this record in MEDLINE/PubMed |
BookMark | eNqFkd2O0zAQhS20iO0uPAES8iUXpNjOn3PBxar8igoQgmvLsSfqVImTtZ1WfTceDqdduARpJMtnvjkjzbkhV250QMhzztac8er1fj3gAHEtGC_WLBWrH5EVl7XIZF42V2SVqDqrGRfX5CaEPWO8zAv5hFyLoq6agpUr8usLhAg2u5-1ixh1xAPQb5vvVE-TH7XZ0SPGHcUh_Q5gaQAXMEEYT7QbPY07oBYimIijo2NH-_FIkxF42sMB-rBoG-0sWh3nQLfYgsdWm4XQAdNCk2R0Z6e7JDhqMPqkTeHU92hf0beop93o0elzC-nn-ahPetBPyeNO9wGePby35Of7dz82H7Pt1w-fNnfbzOSNiFkr26YCnleizrktKylLyTpuhRSGJ0RCxURVNRW3DGwlWC26vC1LKKQxBS_yW_Ly4puOcD-ng6kBg4G-1w7GOSienBtZS8b-j5a5KBZQJjS_oMaPIXjo1ORx0P6kOFNLwmqvzgmrJWHFUrE6Tb14WDC3A9i_M38iTcCbC5CODwcEr4JBcAYs-hSTsiP-c8FvwFK9Lg |
CitedBy_id | crossref_primary_10_1007_s00248_023_02197_8 crossref_primary_10_1111_jam_13302 crossref_primary_10_1016_j_bios_2019_111631 crossref_primary_10_1007_s10658_016_0921_y crossref_primary_10_3390_microorganisms8010092 crossref_primary_10_1038_s41598_019_39164_8 crossref_primary_10_1093_jee_toac080 crossref_primary_10_3389_fphys_2021_777908 crossref_primary_10_1128_spectrum_03614_22 crossref_primary_10_1094_PHYTOFR_04_22_0046_FI crossref_primary_10_1007_s11356_021_17211_0 crossref_primary_10_1007_s42161_022_01155_6 crossref_primary_10_1038_s42003_021_02851_2 crossref_primary_10_1021_acs_jafc_7b03490 crossref_primary_10_1016_j_pmpp_2016_03_001 crossref_primary_10_1089_fpd_2023_0003 crossref_primary_10_3390_insects10050141 crossref_primary_10_1007_s10658_017_1372_9 crossref_primary_10_3389_fmars_2022_988918 crossref_primary_10_4081_ijfs_2023_11134 crossref_primary_10_1093_jee_toz197 crossref_primary_10_3389_fevo_2018_00185 crossref_primary_10_1111_een_12439 crossref_primary_10_1016_j_jip_2023_107959 crossref_primary_10_1007_s00248_016_0733_9 crossref_primary_10_1016_j_ijfoodmicro_2024_110752 crossref_primary_10_1093_jee_tov038 crossref_primary_10_3389_fmicb_2019_01948 crossref_primary_10_1094_PDIS_09_23_1994_SR crossref_primary_10_3389_fagro_2021_675247 crossref_primary_10_1111_eea_12268 crossref_primary_10_1094_PDIS_11_17_1854_RE crossref_primary_10_1094_PHYTO_06_21_0240_FI crossref_primary_10_3390_agronomy11101956 crossref_primary_10_1016_j_cropro_2018_08_004 crossref_primary_10_1016_j_agee_2015_06_027 crossref_primary_10_1111_jen_12466 crossref_primary_10_1016_j_chemosphere_2022_133773 crossref_primary_10_1094_PDIS_08_22_1839_SR crossref_primary_10_1007_s11356_021_12419_6 crossref_primary_10_1653_024_098_0157 crossref_primary_10_1007_s41348_018_0160_x crossref_primary_10_3390_agronomy11010010 crossref_primary_10_1371_journal_pone_0129373 crossref_primary_10_1021_acs_jafc_8b01015 crossref_primary_10_1111_ppl_13662 |
Cites_doi | 10.1094/PDIS-12-12-1174-RE 10.1073/pnas.92.9.3814 10.1016/j.mimet.2005.10.018 10.1093/bioinformatics/btm404 10.1146/annurev-phyto-073009-114418 10.1016/j.jviromet.2011.06.012 10.1093/nar/gks596 10.1603/EC10123 10.1016/j.jviromet.2011.09.017 10.1094/PDIS-11-12-1007-RE 10.1093/nar/gki591 10.1590/S1519-566X2004000500001 10.1016/j.mimet.2008.11.002 10.1016/j.soilbio.2013.01.007 10.1093/nar/26.14.3340 10.1128/JCM.44.3.1029-1039.2006 10.1111/ppa.12193 10.1146/annurev-ento-120811-153542 10.1038/sj.leu.2401811 |
ContentType | Journal Article |
Copyright | 2014 Elsevier B.V. Copyright © 2014 Elsevier B.V. All rights reserved. |
Copyright_xml | – notice: 2014 Elsevier B.V. – notice: Copyright © 2014 Elsevier B.V. All rights reserved. |
DBID | CGR CUY CVF ECM EIF NPM AAYXX CITATION 7X8 7SS 7T7 8FD C1K FR3 P64 |
DOI | 10.1016/j.mimet.2014.04.007 |
DatabaseName | Medline MEDLINE MEDLINE (Ovid) MEDLINE MEDLINE PubMed CrossRef MEDLINE - Academic Entomology Abstracts (Full archive) Industrial and Applied Microbiology Abstracts (Microbiology A) Technology Research Database Environmental Sciences and Pollution Management Engineering Research Database Biotechnology and BioEngineering Abstracts |
DatabaseTitle | MEDLINE Medline Complete MEDLINE with Full Text PubMed MEDLINE (Ovid) CrossRef MEDLINE - Academic Entomology Abstracts Engineering Research Database Technology Research Database Industrial and Applied Microbiology Abstracts (Microbiology A) Biotechnology and BioEngineering Abstracts Environmental Sciences and Pollution Management |
DatabaseTitleList | MEDLINE Entomology Abstracts MEDLINE - Academic |
Database_xml | – sequence: 1 dbid: NPM name: PubMed url: https://proxy.k.utb.cz/login?url=http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed sourceTypes: Index Database – sequence: 2 dbid: EIF name: MEDLINE url: https://proxy.k.utb.cz/login?url=https://www.webofscience.com/wos/medline/basic-search sourceTypes: Index Database |
DeliveryMethod | fulltext_linktorsrc |
Discipline | Biology |
EISSN | 1872-8359 |
EndPage | 22 |
ExternalDocumentID | 10_1016_j_mimet_2014_04_007 24769405 S0167701214001079 |
Genre | Research Support, Non-U.S. Gov't Journal Article |
GroupedDBID | --- --K --M -~X .1- .~1 0R~ 1B1 1P~ 1RT 1~. 1~5 29L 3O- 4.4 457 4G. 53G 5GY 5VS 7-5 71M 8P~ 9JM AAAJQ AABNK AACTN AAEDT AAEDW AAIAV AAIKJ AAKOC AALRI AAOAW AAQFI AAQXK AARKO AAXUO ABFNM ABFRF ABGSF ABJNI ABMAC ABUDA ABXDB ABYKQ ACDAQ ACGFO ACGFS ACIUM ACRLP ADBBV ADEZE ADMUD ADUVX AEBSH AEFWE AEHWI AEKER AENEX AFCTW AFKWA AFTJW AFXIZ AGEKW AGHFR AGRDE AGUBO AGYEJ AHHHB AIEXJ AIKHN AITUG AJBFU AJOXV AJUYK ALMA_UNASSIGNED_HOLDINGS AMFUW AMRAJ ASPBG AVWKF AXJTR AZFZN BKOJK BLXMC CJTIS CNWQP CS3 DOVZS DU5 EBS EFJIC EFLBG EJD EO8 EO9 EP2 EP3 F5P FDB FEDTE FGOYB FIRID FNPLU FYGXN G-2 G-Q G8K GBLVA HMG HVGLF HZ~ IHE J1W KOM LUGTX M41 MO0 N9A O-L O9- OAUVE OZT P-8 P-9 P2P PC. Q38 R2- RIG ROL RPZ SCC SDF SDG SDP SES SEW SIN SPCBC SSI SSU SSZ T5K WH7 WUQ Y6R Z5R ~02 ~G- ~KM AKRWK CGR CUY CVF ECM EIF NPM AAHBH AAXKI AAYXX CITATION 7X8 7SS 7T7 8FD C1K FR3 P64 |
ID | FETCH-LOGICAL-c392t-b8b96e1362731d5688580f1d282c1c398e60266961d0ed62072f3b55e48cc4143 |
IEDL.DBID | .~1 |
ISSN | 0167-7012 |
IngestDate | Fri Aug 16 10:10:18 EDT 2024 Fri Aug 16 11:18:25 EDT 2024 Thu Sep 12 17:48:54 EDT 2024 Thu May 23 23:19:50 EDT 2024 Fri Feb 23 02:22:56 EST 2024 |
IsPeerReviewed | true |
IsScholarly | true |
Keywords | Template secondary structure Huanglongbing Microbial detection methods Plant pathogen False-negatives Low-abundance template |
Language | English |
License | Copyright © 2014 Elsevier B.V. All rights reserved. |
LinkModel | DirectLink |
MergedId | FETCHMERGED-LOGICAL-c392t-b8b96e1362731d5688580f1d282c1c398e60266961d0ed62072f3b55e48cc4143 |
Notes | ObjectType-Article-1 SourceType-Scholarly Journals-1 ObjectType-Feature-2 content type line 23 |
PMID | 24769405 |
PQID | 1532480038 |
PQPubID | 23479 |
PageCount | 8 |
ParticipantIDs | proquest_miscellaneous_1627987800 proquest_miscellaneous_1532480038 crossref_primary_10_1016_j_mimet_2014_04_007 pubmed_primary_24769405 elsevier_sciencedirect_doi_10_1016_j_mimet_2014_04_007 |
PublicationCentury | 2000 |
PublicationDate | 2014-07-01 |
PublicationDateYYYYMMDD | 2014-07-01 |
PublicationDate_xml | – month: 07 year: 2014 text: 2014-07-01 day: 01 |
PublicationDecade | 2010 |
PublicationPlace | Netherlands |
PublicationPlace_xml | – name: Netherlands |
PublicationTitle | Journal of microbiological methods |
PublicationTitleAlternate | J Microbiol Methods |
PublicationYear | 2014 |
Publisher | Elsevier B.V |
Publisher_xml | – name: Elsevier B.V |
References | Li, Hartung, Levy (bb0065) 2006; 66 Larkin, Blackshields, Brown, Chenna, McGettigan, McWilliam, Valentin, Wallace, Wilm, Lopez, Thompson, Gibson, Higgins (bb0060) 2007; 23 Christoforou, Pantelides, Kanetis, Ioannou, Tsaltas (bb0020) 2014 Santos, Reis, dos Santos, Damas, Silva, Viana, Ferraz, Carnauba, El-Far, Serra, Diaz (bb0090) 2012; 179 Sontakke, Cadenas, Maggi, Diniz, Breitschwerdt (bb0095) 2009; 76 VSN International (bb0135) 2012 Ananthakrishnan, Choudhary, Roy, Sengoda, Hartung, Stone, Damsteegt, Schneider, Munyaneza, Brlansky (bb0005) 2013; 97 Grafton-Cardwell, Stelinski, Stansly (bb0035) 2013; 58 Hargreaves, Roberto, Hofmockel (bb0040) 2013; 59 Karrer, Lincoln, Hagenhout, Bennett, Bostock, Martineau, Lucas, Gilchrist, Alexander (bb0050) 1995; 92 White, Quake, Curr (bb0125) 2012; 179 Hochhaus, Weisser, La Rose'e, Emig, Müller, Saußele, Reiter, Kuhn, Berger, Hehlmann, Cross (bb0130) 2000; 14 U.S. Department of Agriculture (bb0115) 2012 Untergrasser, Cutcutache, Koressaar, Ye, Faircloth, Remm, Rozen (bb0110) 2012; 40 McDowell, Burns, Parkes (bb0075) 1998; 26 Bové (bb0010) 2006; 88 Engelbrecht, Duong, van den Berg (bb0025) 2013; 97 Markham, Zuker (bb0070) 2005; 33 Pelz-Stelinski, Brlansky, Ebert, Rogers (bb0085) 2010; 103 Takahashi, Nakayama (bb0100) 2006; 44 Gottwald (bb0030) 2010; 48 Mitchell (bb0080) 2004; 33 Li W., Duan Y., Brlansky R.H., Twieg E., & Levy L., "Incidence and population of 'Candidatus Liberibacter asiaticus' in Asian citrus psyllids (Diaphorina citri) on citrus plants affected by huanglongbing in Florida", Proceedings International Research Conference on Huanglongbing (bb9005) 2008, Dec. Bustin, Nolan (bb0015) 2004; 15 Grafton-Cardwell (10.1016/j.mimet.2014.04.007_bb0035) 2013; 58 Markham (10.1016/j.mimet.2014.04.007_bb0070) 2005; 33 Larkin (10.1016/j.mimet.2014.04.007_bb0060) 2007; 23 McDowell (10.1016/j.mimet.2014.04.007_bb0075) 1998; 26 Mitchell (10.1016/j.mimet.2014.04.007_bb0080) 2004; 33 White (10.1016/j.mimet.2014.04.007_bb0125) 2012; 179 Karrer (10.1016/j.mimet.2014.04.007_bb0050) 1995; 92 Santos (10.1016/j.mimet.2014.04.007_bb0090) 2012; 179 Gottwald (10.1016/j.mimet.2014.04.007_bb0030) 2010; 48 Li W., Duan Y., Brlansky R.H., Twieg E., & Levy L., "Incidence and population of 'Candidatus Liberibacter asiaticus' in Asian citrus psyllids (Diaphorina citri) on citrus plants affected by huanglongbing in Florida", Proceedings International Research Conference on Huanglongbing (10.1016/j.mimet.2014.04.007_bb9005) 2008 Sontakke (10.1016/j.mimet.2014.04.007_bb0095) 2009; 76 Bové (10.1016/j.mimet.2014.04.007_bb0010) 2006; 88 Christoforou (10.1016/j.mimet.2014.04.007_bb0020) 2014 Bustin (10.1016/j.mimet.2014.04.007_bb0015) 2004; 15 Takahashi (10.1016/j.mimet.2014.04.007_bb0100) 2006; 44 Engelbrecht (10.1016/j.mimet.2014.04.007_bb0025) 2013; 97 Pelz-Stelinski (10.1016/j.mimet.2014.04.007_bb0085) 2010; 103 Hargreaves (10.1016/j.mimet.2014.04.007_bb0040) 2013; 59 Hochhaus (10.1016/j.mimet.2014.04.007_bb0130) 2000; 14 U.S. Department of Agriculture (10.1016/j.mimet.2014.04.007_bb0115) Untergrasser (10.1016/j.mimet.2014.04.007_bb0110) 2012; 40 VSN International (10.1016/j.mimet.2014.04.007_bb0135) 2012 Li (10.1016/j.mimet.2014.04.007_bb0065) 2006; 66 Ananthakrishnan (10.1016/j.mimet.2014.04.007_bb0005) 2013; 97 |
References_xml | – volume: 14 start-page: 998 year: 2000 end-page: 1005 ident: bb0130 article-title: Detection and quantification of residual disease in chronic myelogenous leukemia publication-title: Leukemia contributor: fullname: Cross – volume: 76 start-page: 217 year: 2009 end-page: 225 ident: bb0095 article-title: Use of broad range 16S rDNA PCR in clinical microbiology publication-title: J. Microbiol. Methods contributor: fullname: Breitschwerdt – year: 2014 ident: bb0020 article-title: Rapid detection and quantification of viable potato cyst nematodes using qPCR in combination with propidium monoazide publication-title: Plant Pathol contributor: fullname: Tsaltas – volume: 48 start-page: 119 year: 2010 end-page: 139 ident: bb0030 article-title: Current epidemiological understanding of citrus huanglongbing publication-title: Annu. Rev. Phytopathol. contributor: fullname: Gottwald – volume: 58 start-page: 413 year: 2013 end-page: 432 ident: bb0035 article-title: Biology and management of Asian citrus psyllid, vector of the Huanglongbing pathogens publication-title: Annu. Rev. Entomol. contributor: fullname: Stansly – volume: 26 start-page: 3340 year: 1998 end-page: 3347 ident: bb0075 article-title: Localised sequence regions possession high melting temperatures prevent the amplification of a DNA mimic in competitive PCR publication-title: Nucleic Acids Res. contributor: fullname: Parkes – year: 2008, Dec. ident: bb9005 article-title: 1–5, Orlando, Florida; Plant Management Network contributor: fullname: Li W., Duan Y., Brlansky R.H., Twieg E., & Levy L., "Incidence and population of 'Candidatus Liberibacter asiaticus' in Asian citrus psyllids (Diaphorina citri) on citrus plants affected by huanglongbing in Florida", Proceedings International Research Conference on Huanglongbing – volume: 97 start-page: 1235 year: 2013 end-page: 1243 ident: bb0005 article-title: Development of primers and probes for genus and species specific detection of ‘Candidatus Liberibacter species’ by real-time PCR publication-title: Plant Dis. contributor: fullname: Brlansky – volume: 40 start-page: e115 year: 2012 ident: bb0110 article-title: Primer3 — new capabilities and interfaces publication-title: Nucleic Acids Res. contributor: fullname: Rozen – volume: 66 start-page: 104 year: 2006 end-page: 115 ident: bb0065 article-title: Quantitative real-time PCR for detection and identification of Candidatus Liberibacter species associated with citrus huanglongbing publication-title: J. Microbiol. Methods contributor: fullname: Levy – volume: 97 start-page: 1012 year: 2013 end-page: 1017 ident: bb0025 article-title: Development of a nested quantitative real-time PCR for detecting publication-title: Plant Dis. contributor: fullname: van den Berg – year: 2012 ident: bb0115 article-title: New pest response guidelines citrus greening disease contributor: fullname: U.S. Department of Agriculture – volume: 103 start-page: 1531 year: 2010 end-page: 1541 ident: bb0085 article-title: Transmission parameters for Candidatus Liberibacter asiaticus by Asian citrus psyllid (Hemiptera: Psyllidae) publication-title: J. Econ. Entomol. contributor: fullname: Rogers – volume: 179 start-page: 45 year: 2012 end-page: 50 ident: bb0125 article-title: Digital PCR provides absolute quantification of viral load for occult RNA virus publication-title: J. Virol. Methods contributor: fullname: Curr – volume: 88 start-page: 7 year: 2006 end-page: 37 ident: bb0010 article-title: Huanglongbing: a destructive, newly-emerging, century-old disease of citrus publication-title: J. Plant Pathol. contributor: fullname: Bové – volume: 33 start-page: W577 year: 2005 end-page: W581 ident: bb0070 article-title: DINAMelt web server for nucleic acid melting prediction publication-title: Nucleic Acids Res. contributor: fullname: Zuker – year: 2012 ident: bb0135 article-title: GenStat for Windows 15th Edition contributor: fullname: VSN International – volume: 44 start-page: 1029 year: 2006 end-page: 1039 ident: bb0100 article-title: Novel technique of quantitative nested real-time PCR assay for publication-title: J. Clin. Microbiol. contributor: fullname: Nakayama – volume: 59 start-page: 89 year: 2013 end-page: 97 ident: bb0040 article-title: Reaction-and sample-specific inhibition affect standardization of qPCR assays of soil bacterial communities publication-title: Soil Biol. Biochem. contributor: fullname: Hofmockel – volume: 23 start-page: 2947 year: 2007 end-page: 2948 ident: bb0060 article-title: Clustal W and Clustal X version 2.0 publication-title: Bioinformatics contributor: fullname: Higgins – volume: 15 start-page: 155 year: 2004 end-page: 166 ident: bb0015 article-title: Pitfalls of quantitative real-time reverse-transcription polymerase chain reaction publication-title: J. Biomol. Tech. contributor: fullname: Nolan – volume: 33 start-page: 519 year: 2004 end-page: 545 ident: bb0080 article-title: Heteroptera as vectors of plant pathogens publication-title: Neotrop. Entomol. contributor: fullname: Mitchell – volume: 179 start-page: 17 year: 2012 end-page: 20 ident: bb0090 article-title: The use of real-time PCR to detect hepatitis C virus RNA in dried blood spots from Brazilian patients infected chronically publication-title: J. Virol. Methods contributor: fullname: Diaz – volume: 92 start-page: 3814 year: 1995 end-page: 3818 ident: bb0050 article-title: In situ isolation of mRNA from individual plant cells: creation of cell-specific cDNA libraries publication-title: PNAS contributor: fullname: Alexander – year: 2008 ident: 10.1016/j.mimet.2014.04.007_bb9005 contributor: fullname: Li W., Duan Y., Brlansky R.H., Twieg E., & Levy L., "Incidence and population of 'Candidatus Liberibacter asiaticus' in Asian citrus psyllids (Diaphorina citri) on citrus plants affected by huanglongbing in Florida", Proceedings International Research Conference on Huanglongbing – volume: 97 start-page: 1235 year: 2013 ident: 10.1016/j.mimet.2014.04.007_bb0005 article-title: Development of primers and probes for genus and species specific detection of ‘Candidatus Liberibacter species’ by real-time PCR publication-title: Plant Dis. doi: 10.1094/PDIS-12-12-1174-RE contributor: fullname: Ananthakrishnan – volume: 92 start-page: 3814 year: 1995 ident: 10.1016/j.mimet.2014.04.007_bb0050 article-title: In situ isolation of mRNA from individual plant cells: creation of cell-specific cDNA libraries publication-title: PNAS doi: 10.1073/pnas.92.9.3814 contributor: fullname: Karrer – volume: 15 start-page: 155 year: 2004 ident: 10.1016/j.mimet.2014.04.007_bb0015 article-title: Pitfalls of quantitative real-time reverse-transcription polymerase chain reaction publication-title: J. Biomol. Tech. contributor: fullname: Bustin – volume: 66 start-page: 104 year: 2006 ident: 10.1016/j.mimet.2014.04.007_bb0065 article-title: Quantitative real-time PCR for detection and identification of Candidatus Liberibacter species associated with citrus huanglongbing publication-title: J. Microbiol. Methods doi: 10.1016/j.mimet.2005.10.018 contributor: fullname: Li – volume: 23 start-page: 2947 year: 2007 ident: 10.1016/j.mimet.2014.04.007_bb0060 article-title: Clustal W and Clustal X version 2.0 publication-title: Bioinformatics doi: 10.1093/bioinformatics/btm404 contributor: fullname: Larkin – volume: 48 start-page: 119 year: 2010 ident: 10.1016/j.mimet.2014.04.007_bb0030 article-title: Current epidemiological understanding of citrus huanglongbing publication-title: Annu. Rev. Phytopathol. doi: 10.1146/annurev-phyto-073009-114418 contributor: fullname: Gottwald – volume: 179 start-page: 17 year: 2012 ident: 10.1016/j.mimet.2014.04.007_bb0090 article-title: The use of real-time PCR to detect hepatitis C virus RNA in dried blood spots from Brazilian patients infected chronically publication-title: J. Virol. Methods doi: 10.1016/j.jviromet.2011.06.012 contributor: fullname: Santos – volume: 40 start-page: e115 year: 2012 ident: 10.1016/j.mimet.2014.04.007_bb0110 article-title: Primer3 — new capabilities and interfaces publication-title: Nucleic Acids Res. doi: 10.1093/nar/gks596 contributor: fullname: Untergrasser – volume: 103 start-page: 1531 year: 2010 ident: 10.1016/j.mimet.2014.04.007_bb0085 article-title: Transmission parameters for Candidatus Liberibacter asiaticus by Asian citrus psyllid (Hemiptera: Psyllidae) publication-title: J. Econ. Entomol. doi: 10.1603/EC10123 contributor: fullname: Pelz-Stelinski – volume: 179 start-page: 45 year: 2012 ident: 10.1016/j.mimet.2014.04.007_bb0125 article-title: Digital PCR provides absolute quantification of viral load for occult RNA virus publication-title: J. Virol. Methods doi: 10.1016/j.jviromet.2011.09.017 contributor: fullname: White – volume: 88 start-page: 7 year: 2006 ident: 10.1016/j.mimet.2014.04.007_bb0010 article-title: Huanglongbing: a destructive, newly-emerging, century-old disease of citrus publication-title: J. Plant Pathol. contributor: fullname: Bové – ident: 10.1016/j.mimet.2014.04.007_bb0115 contributor: fullname: U.S. Department of Agriculture – year: 2012 ident: 10.1016/j.mimet.2014.04.007_bb0135 contributor: fullname: VSN International – volume: 97 start-page: 1012 year: 2013 ident: 10.1016/j.mimet.2014.04.007_bb0025 article-title: Development of a nested quantitative real-time PCR for detecting Phytophthora cinnamomi in Persea americana rootstocks publication-title: Plant Dis. doi: 10.1094/PDIS-11-12-1007-RE contributor: fullname: Engelbrecht – volume: 33 start-page: W577 year: 2005 ident: 10.1016/j.mimet.2014.04.007_bb0070 article-title: DINAMelt web server for nucleic acid melting prediction publication-title: Nucleic Acids Res. doi: 10.1093/nar/gki591 contributor: fullname: Markham – volume: 33 start-page: 519 year: 2004 ident: 10.1016/j.mimet.2014.04.007_bb0080 article-title: Heteroptera as vectors of plant pathogens publication-title: Neotrop. Entomol. doi: 10.1590/S1519-566X2004000500001 contributor: fullname: Mitchell – volume: 76 start-page: 217 year: 2009 ident: 10.1016/j.mimet.2014.04.007_bb0095 article-title: Use of broad range 16S rDNA PCR in clinical microbiology publication-title: J. Microbiol. Methods doi: 10.1016/j.mimet.2008.11.002 contributor: fullname: Sontakke – volume: 59 start-page: 89 year: 2013 ident: 10.1016/j.mimet.2014.04.007_bb0040 article-title: Reaction-and sample-specific inhibition affect standardization of qPCR assays of soil bacterial communities publication-title: Soil Biol. Biochem. doi: 10.1016/j.soilbio.2013.01.007 contributor: fullname: Hargreaves – volume: 26 start-page: 3340 year: 1998 ident: 10.1016/j.mimet.2014.04.007_bb0075 article-title: Localised sequence regions possession high melting temperatures prevent the amplification of a DNA mimic in competitive PCR publication-title: Nucleic Acids Res. doi: 10.1093/nar/26.14.3340 contributor: fullname: McDowell – volume: 44 start-page: 1029 year: 2006 ident: 10.1016/j.mimet.2014.04.007_bb0100 article-title: Novel technique of quantitative nested real-time PCR assay for Mycobacterium tuberculosis DNA publication-title: J. Clin. Microbiol. doi: 10.1128/JCM.44.3.1029-1039.2006 contributor: fullname: Takahashi – year: 2014 ident: 10.1016/j.mimet.2014.04.007_bb0020 article-title: Rapid detection and quantification of viable potato cyst nematodes using qPCR in combination with propidium monoazide publication-title: Plant Pathol doi: 10.1111/ppa.12193 contributor: fullname: Christoforou – volume: 58 start-page: 413 year: 2013 ident: 10.1016/j.mimet.2014.04.007_bb0035 article-title: Biology and management of Asian citrus psyllid, vector of the Huanglongbing pathogens publication-title: Annu. Rev. Entomol. doi: 10.1146/annurev-ento-120811-153542 contributor: fullname: Grafton-Cardwell – volume: 14 start-page: 998 year: 2000 ident: 10.1016/j.mimet.2014.04.007_bb0130 article-title: Detection and quantification of residual disease in chronic myelogenous leukemia publication-title: Leukemia doi: 10.1038/sj.leu.2401811 contributor: fullname: Hochhaus |
SSID | ssj0015348 |
Score | 2.3583765 |
Snippet | Candidatus Liberibacter asiaticus (CLas) is a phloem-limited bacterium transmitted by the Asian citrus psyllid, Diaphorina citri, and the presumptive causal... |
SourceID | proquest crossref pubmed elsevier |
SourceType | Aggregation Database Index Database Publisher |
StartPage | 15 |
SubjectTerms | Animals Citrus Citrus - microbiology Diaphorina citri DNA, Bacterial - genetics DNA, Ribosomal - genetics False-negatives Hemiptera - microbiology Huanglongbing Humans Insecta - microbiology Kuwayama Low-abundance template Microbial detection methods Plant pathogen Plasmids Polymerase Chain Reaction - methods Real-Time Polymerase Chain Reaction - methods Rhizobiaceae - genetics Rhizobiaceae - isolation & purification RNA, Ribosomal, 16S - genetics Sensitivity and Specificity Template secondary structure |
Title | Nested-quantitative PCR approach with improved sensitivity for the detection of low titer levels of Candidatus Liberibacter asiaticus in the Asian citrus psyllid, Diaphorina citri Kuwayama |
URI | https://dx.doi.org/10.1016/j.mimet.2014.04.007 https://www.ncbi.nlm.nih.gov/pubmed/24769405 https://search.proquest.com/docview/1532480038 https://search.proquest.com/docview/1627987800 |
Volume | 102 |
hasFullText | 1 |
inHoldings | 1 |
isFullTextHit | |
isPrint | |
link | http://utb.summon.serialssolutions.com/2.0.0/link/0/eLvHCXMwnV1Na9wwEBUhpdBLSb-3ScMUeoyztixb8nHZNmwbupS2gdyMbMnUYde7xbuEveSX5cdlZmwHekgOBV9syUZoxppn-c0bIT5J6RCFhyZICl8FSoYuyLyLAl9heLOyLDJOF_s-T2cX6ttlcrknpkMuDNEq-7W_W9N5te6vjPvZHK_revyLCPSaJMkUfdhoSuJTGIzQp09v7mke-EJzBS3W96beg_IQc7yW9dIToTJSp52K9kPR6SH0yVHo7EA87-EjTLoRvhB7vnkpnnYFJXevxO2cty-Dv1vbcPYYrmXwY_oTBulwoH1XqHknwTtoib7e1Y8ARK-AaBCc3zA9q4FVBYvVNbDyMiyIXdTStSllwji72bbAfJO6YMVnoHxMkvJooW74SRNK0ISS8zpg3e4Wi9qdwOfarv8Q7c9yUw3n22u7s0v7Wlycffk9nQV9fYagRFS1CQqDlvQRhkAdRy5JjUlMWEUOv-LKCLsYT_WtUrS2C71LZahlFRdJ4pUpS4VA7Y3Yb1aNfydAJqXVNsVQGReqoCPWMi0zZzwiOhWPxMlgl3zdyXDkAz_tKmcz5mTGPMQj1CORDrbL__GmHAPF4zd-HCyNjS39PLGNX23bHB1JKkM_Uh_pgzORGY29RuJt5yb3o5VKpxmi4_f_O7RD8YzOOqrwkdhH2_kPCIg2xTF7_LF4Mvl6PpvfATDqDOA |
link.rule.ids | 315,786,790,4521,24144,27957,27958,45620,45714 |
linkProvider | Elsevier |
linkToHtml | http://utb.summon.serialssolutions.com/2.0.0/link/0/eLvHCXMwnV1da9swFBVdythexr6Xbd3uYI_1YsuyJT-GtCVd2jC2FvpmZEumGomT4YSS_7Yft3tlu7CH9qHgJ30YoSt0j6Rzz2XsK-cGUXiogqSwVSB4aILMmiiwFbo3zcsi8-Fi5_N0eim-XyVXe2zSx8IQrbLb-9s93e_WXcmom83R2rnRLyLQS5IkE3Swkdkjti8SGYkB2x-fzqbz28eEJPZJtLzEN3XoxYc8zWvplpY4lZH41gpp3-Wg7gKg3hGdPGfPOgQJ43aQL9ierV-yx21Oyd0r9nfubzCDP1td-wAy3M7gx-Qn9OrhQFev4PxlgjXQEIO9TSEBCGABASEYu_EMrRpWFSxWN-DFl2FBBKOGyiYUDGP0ZtuAp5y4wos-A4VkkppHA672fxpTjCaUPrQD1s1usXDmEI6cXl8T80_7Kgez7Y3e6aV-zS5Pji8m06BL0RCUCKw2QaHQmDZCLyjjyCSpUokKq8jgQa6MsImylOIqRYOb0JqUh5JXcZEkVqiyFIjV3rBBvartOwY8KbXUKXrLuBAFfbHkaZkZZRHUiXjIDnu75OtWiSPvKWq_c2_GnMyYh_iFcsjS3nb5fwsqR19xf8cvvaWxsqH3E13b1bbJcSFxoegt9Z42OBOZkthqyN62y-R2tFzINEOA_P6hQ_vMnkwvzs_ys9P57AN7SjUtc_gjG6Ad7QHio03xqVv__wBllQ-W |
openUrl | ctx_ver=Z39.88-2004&ctx_enc=info%3Aofi%2Fenc%3AUTF-8&rfr_id=info%3Asid%2Fsummon.serialssolutions.com&rft_val_fmt=info%3Aofi%2Ffmt%3Akev%3Amtx%3Ajournal&rft.genre=article&rft.atitle=Nested-quantitative+PCR+approach+with+improved+sensitivity+for+the+detection+of+low+titer+levels+of+Candidatus+Liberibacter+asiaticus+in+the+Asian+citrus+psyllid%2C+Diaphorina+citri+Kuwayama&rft.jtitle=Journal+of+microbiological+methods&rft.au=Coy%2C+M.R.&rft.au=Hoffmann%2C+M.&rft.au=Kingdom+Gibbard%2C+H.N.&rft.au=Kuhns%2C+E.H.&rft.date=2014-07-01&rft.pub=Elsevier+B.V&rft.issn=0167-7012&rft.eissn=1872-8359&rft.volume=102&rft.spage=15&rft.epage=22&rft_id=info:doi/10.1016%2Fj.mimet.2014.04.007&rft.externalDocID=S0167701214001079 |
thumbnail_l | http://covers-cdn.summon.serialssolutions.com/index.aspx?isbn=/lc.gif&issn=0167-7012&client=summon |
thumbnail_m | http://covers-cdn.summon.serialssolutions.com/index.aspx?isbn=/mc.gif&issn=0167-7012&client=summon |
thumbnail_s | http://covers-cdn.summon.serialssolutions.com/index.aspx?isbn=/sc.gif&issn=0167-7012&client=summon |