Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species
Xanthomonas arboricola pv. pruni, the causal agent of bacterial spot disease of stone fruit, is considered a quarantine organism by the European Union and the European and Mediterranean Plant Protection Organization (EPPO). The bacterium can undergo an epiphytic phase and/or be latent and can be tra...
| Main Authors: | , , , , , |
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| Format: | Artículo |
| Language: | Inglés |
| Published: |
2017
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| Online Access: | http://hdl.handle.net/20.500.11939/4286 |
| _version_ | 1855491726498594816 |
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| author | Palacio-Bielsa, Ana Cubero, Jaime Cambra, Miguel A. Collados, Raquel Berruete, Isabel M. López, María M. |
| author_browse | Berruete, Isabel M. Cambra, Miguel A. Collados, Raquel Cubero, Jaime López, María M. Palacio-Bielsa, Ana |
| author_facet | Palacio-Bielsa, Ana Cubero, Jaime Cambra, Miguel A. Collados, Raquel Berruete, Isabel M. López, María M. |
| author_sort | Palacio-Bielsa, Ana |
| collection | ReDivia |
| description | Xanthomonas arboricola pv. pruni, the causal agent of bacterial spot disease of stone fruit, is considered a quarantine organism by the European Union and the European and Mediterranean Plant Protection Organization (EPPO). The bacterium can undergo an epiphytic phase and/or be latent and can be transmitted by plant material, but currently, only visual inspections are used to certify plants as being X. arboricola pv. pruni free. A novel and highly sensitive real-time TaqMan PCR detection protocol was designed based on a sequence of a gene for a putative protein related to an ABC transporter ATP-binding system in X. arboricola pv. pruni. Pathogen detection can be completed within a few hours with a sensitivity of 10(2) CFU ml(-1), thus surpassing the sensitivity of the existing conventional PCR. Specificity was assessed for X. arboricola pv. pruni strains from different origins as well as for closely related Xanthomonas species, non-Xanthomonas species, saprophytic bacteria, and healthy Prunus samples. The efficiency of the developed protocol was evaluated with field samples of 14 Prunus species and rootstocks. For symptomatic leaf samples, the protocol was very efficient even when washed tissues of the leaves were directly amplified without any previous DNA extraction. For samples of 117 asymptomatic leaves and 285 buds, the protocol was more efficient after a simple DNA extraction, and X. arboricola pv. pruni was detected in 9.4% and 9.1% of the 402 samples analyzed, respectively, demonstrating its frequent epiphytic or endophytic phase. This newly developed real-time PCR protocol can be used as a quantitative assay, offers a reliable and sensitive test for X. arboricola pv. pruni, and is suitable as a screening test for symptomatic as well as asymptomatic plant material. |
| format | Artículo |
| id | ReDivia4286 |
| institution | Instituto Valenciano de Investigaciones Agrarias (IVIA) |
| language | Inglés |
| publishDate | 2017 |
| publishDateRange | 2017 |
| publishDateSort | 2017 |
| record_format | dspace |
| spelling | ReDivia42862025-04-25T14:42:44Z Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species Palacio-Bielsa, Ana Cubero, Jaime Cambra, Miguel A. Collados, Raquel Berruete, Isabel M. López, María M. Xanthomonas arboricola pv. pruni, the causal agent of bacterial spot disease of stone fruit, is considered a quarantine organism by the European Union and the European and Mediterranean Plant Protection Organization (EPPO). The bacterium can undergo an epiphytic phase and/or be latent and can be transmitted by plant material, but currently, only visual inspections are used to certify plants as being X. arboricola pv. pruni free. A novel and highly sensitive real-time TaqMan PCR detection protocol was designed based on a sequence of a gene for a putative protein related to an ABC transporter ATP-binding system in X. arboricola pv. pruni. Pathogen detection can be completed within a few hours with a sensitivity of 10(2) CFU ml(-1), thus surpassing the sensitivity of the existing conventional PCR. Specificity was assessed for X. arboricola pv. pruni strains from different origins as well as for closely related Xanthomonas species, non-Xanthomonas species, saprophytic bacteria, and healthy Prunus samples. The efficiency of the developed protocol was evaluated with field samples of 14 Prunus species and rootstocks. For symptomatic leaf samples, the protocol was very efficient even when washed tissues of the leaves were directly amplified without any previous DNA extraction. For samples of 117 asymptomatic leaves and 285 buds, the protocol was more efficient after a simple DNA extraction, and X. arboricola pv. pruni was detected in 9.4% and 9.1% of the 402 samples analyzed, respectively, demonstrating its frequent epiphytic or endophytic phase. This newly developed real-time PCR protocol can be used as a quantitative assay, offers a reliable and sensitive test for X. arboricola pv. pruni, and is suitable as a screening test for symptomatic as well as asymptomatic plant material. 2017-06-01T10:09:47Z 2017-06-01T10:09:47Z 2011 JAN 2011 article Palacio-Bielsa, Ana, Cubero, Jaime, Cambra, M.A., Collados, Raquel, Berruete, I.M., Lopez, M.M. (2011). Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species. Applied and Environmental Microbiology, 77(1), 89-97. 0099-2240 http://hdl.handle.net/20.500.11939/4286 10.1128/AEM.01593-10 en openAccess Impreso |
| spellingShingle | Palacio-Bielsa, Ana Cubero, Jaime Cambra, Miguel A. Collados, Raquel Berruete, Isabel M. López, María M. Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species |
| title | Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species |
| title_full | Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species |
| title_fullStr | Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species |
| title_full_unstemmed | Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species |
| title_short | Development of an Efficient Real-Time Quantitative PCR Protocol for Detection of Xanthomonas arboricola pv. pruni in Prunus Species |
| title_sort | development of an efficient real time quantitative pcr protocol for detection of xanthomonas arboricola pv pruni in prunus species |
| url | http://hdl.handle.net/20.500.11939/4286 |
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