Acta Univ. Agric. Silvic. Mendelianae Brun. 2017, 65(1), 205-210 | DOI: 10.11118/actaun201765010205

Detecting Pathogens of Verticillium Wilt in Winter Oilseed Rape Using ELISA and PCR - Comparison of the Two Methods and With Visual Stand Evaluation

Tomáš Spitzer, Pavel Matušinsky
Agrotest fyto, Ltd, Kromeriz, Czech Republic

During 2013-2015, oilseed rape samples from various locations in the Czech Republic were analysed for the presence of Verticillium wilt. Samples were evaluated in the lab using ELISA and PCR as well as by visual evaluation during the samples collection. A comparison of detection match in individual methods also was made. ELISA and PCR matched in detecting Verticillium wilt in 60 % of cases. For practical use, a higher match rate would be necessary in cases where samples were to be analysed in various laboratories using only one of these techniques. The possibility was demonstrated to use kits as well as primers not targeted specifically to just Verticillium longisporum but recording both main species on oilseed rape (V. longisporum and V. dahliae). The match rates of lab analyses with visual evaluation of stand infection were surprisingly high at 62 % for ELISA and 77 % for PCR. All three stand evaluation methods matched in 56 % of cases.

Keywords: Verticillium longisporum, Verticillium dahliae
Grants and funding:

This publication was made possible by institutional support provided for the long-term development of a research organization under Decision of the Ministry of Agriculture of the Czech Republic No. RO1116 from 28 February 2011 and was supported by the Ministry of Agriculture of the Czech Republic, Project No. QJ1310227.

Prepublished online: February 28, 2017; Published: March 1, 2017  Show citation

ACS AIP APA ASA Harvard Chicago IEEE ISO690 MLA NLM Turabian Vancouver
Spitzer, T., & Matušinsky, P. (2017). Detecting Pathogens of Verticillium Wilt in Winter Oilseed Rape Using ELISA and PCR - Comparison of the Two Methods and With Visual Stand Evaluation. Acta Universitatis Agriculturae et Silviculturae Mendelianae Brunensis, 65(1), 205-210. doi: 10.11118/actaun201765010205
Share...
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)
    • Open full article

    References

    1. BOKOR, P., BEČKA, D. and TÓTHOVÁ, M. 2014. Occurrence of white rot and verticillium wilt in Winter Rapeseed fields in Slovakia during 2014 [in Slovak]. In: Proceedings from Prosperous Oil Crops 2014 [in Czech: Prosperující olejniny]. Czech University of Life Sciences Prague, 11-12 December 2014. Prague: Czech University of Life Sciences Prague, 82-85.
    2. BREITENMOSER, S., DUBUIS, N., GRILLOT, L., BRODARD, J. and BALMELLI, C. 2011. Viral diseases of oilseed rape in Switzerland [in French: Les viroses du colza en Suisse]. Recherche Agronomique Suisse, 2: 388 -395.
    3. CARDER, J., MORTON, A., TABRETT, A. M. and BARBARA, D. J. 1994. Detection and differentiation by PCR of subspecific groups within two Verticillium species causing vascular wilts in herbaceous hosts. In: SCHOTS, A., DEWEY, F. M. and OLIVER, R. (eds.), Modern assays for plant pathogenic fungi. Oxford: CAB International, 91-97.
    4. COULES, A. and ROSSALL, S. 2003. Disease control and the consequences of timing on the yield of oilseed rape. In: Proceedings of the BCPC International Congress: Crop Science and Technology, Volumes 1 and 2. Scottish Exhibition and Conference Centre, 10-12. November 2003. Alton, Hampshire, UK: British Crop Production Council, 659-662.
    5. DEBODE, J., VAN POUCKE, K., FRANÇA, S. C., MAES, M., HÖFTE, M. and HEUNGENS, K. 2011. Detection of multiple Verticillium species in soil using density flotation and real-time polymerase chain reaction. Plant Disease, 95: 1571-1580. DOI: 10.1094/PDIS-04-11-0267 Go to original source...
    6. DUNKER, S., KEUNECKE, H., STEINBACH P. and VON TIEDEMANN, A. 2008. Impact of Verticillium longisporum on yield and morphology of winter oilseed rape (Brassica napus) in relation to systemic spread in the plant. Journal of Phytopathology, 156: 698-707. DOI: 10.1111/j.1439-0434.2008.01429.x Go to original source...
    7. FORTNAGEL, B. and SCHLOSSER, E. 1996. Polyclonal antibodies against Verticillium dahliae Kleb from oil-seed rape via immunized hens. Journal of Plant Diseases and Protection, 103(6): 582 - 589.
    8. GAO, W., NAN, T., TAN, G., ZHAO, H., WANG, B., LI, Q. X. and MENG, F. 2012. Development of a sensitive monoclonal antibody-based enzyme-linked immunosorbent assay for the analysis of cadmium ions in water, soil and rape samples. Food and Agricultural Immunology, 23: 27-39. DOI: 10.1080/09540105.2011.589045 Go to original source...
    9. INDERBITZIN, P., DAVIS, R. M., BOSTOCK, R. M. and SUBBARAO, K. V. Identification and differentiation of Verticillium species and V. longisporum lineages by simplex and multiplex PCR assays. PLoS One, 8(6): e65990. DOI: 10.1371/journal.pone.0065990
    10. JOHANSSON, A. 2006. Verticillium longisporum, infection, host range, prevalence and plant defence responses. Licentiate thesis. Uppsala: Swedish University of Agricultural Sciences.
    11. JOHANSSON, A., GOUD, J-K. C. and DIXELIUS, C. 2006. Plant host range of Verticillium longisporum and microsclerotia density in Swedish soils. European Journal of Plant Pathology, 114: 139-149. DOI: 10.1007/s10658-005-2333-2 Go to original source...
    12. KARAPAPA, V. K, BAINBRIDGE, B. W. and HEALE, J. B. 1997. Morphological and molecular characterization of Verticillium longisporum comb. nov., pathogenic to oilseed rape. Mycological Research, 101: 1281-1294. DOI: 10.1017/S0953756297003985 Go to original source...
    13. KARAPAPA, V. K. and TYPAS, M. A. 2001. Molecular characterization of the host-adapted pathogen Verticillium longisporum on the basis of a group-I intron found in the nuclear SSU-rRNA gene. Current Microbiology, 42: 217-224. Go to original source...
    14. MERCADO-BLANCO, J., RODRÍGUEZ-JURADO, D. and PARRILLA-ARUAJO, S. 2003. Simultaneous detection of the defoliating and nondefoliating Verticillium dahliae pathotypes in infected olive plants by duplex, nested polymerase chain reaction. Plant Disease, 87: 1487-1494. DOI: 10.1094/PDIS.2003.87.12.1487 Go to original source...
    15. PANTOU, M. P., STRUNNIKOVA, O. K., SHAKHNAZAROVA, V. Y., VISHNEVSKAYA, N. A., PAPALOUKA, V. G. and TYPAS, M. A. 2005. Molecular and immunochemical phylogeny of Verticillium species. Mycological Research, 109: 889:902. DOI: 10.1017/S0953756205003345 Go to original source...
    16. POPOVIĆ, T., BALAŽ, J., IGNJATOV, M., MITROVIĆ, P., GAVRILOVIĆ, V. and JOŠIĆ, D. 2014. Identification and genetic characterisation of Xanthomonas campestris pv. campestris as an oilseed rape pathogen in Serbia. Journal of Plant Pathology, 96: 553-560.
    17. SPITZER, T., MATUŠINSKY, P., MIKOLÁŠOVÁ, R. and WOLF, G. 2008. Occurrence of Verticillium wilt of oilseed rape in the Czech Republic detected by PCR and ELISA. In: Proceedings of the 56. Deutsche Pflanzenschutztagung. Kiel, Germany, 22-25 September 2008. Quedlinburg, Germany: Julius Kühn-Institut.
    18. STEVENTON, L. A., FAHLESON, J., HU, Q. and DIXELIUS, C. 2002. Identification of the causal agent of Verticillium wilt of winter oilseed rape in Sweden, V. longisporum. Mycological Research, 106: 570-578. DOI: 10.1017/S0953756202005865 Go to original source...
    19. WAGNER, M.-H., DUCOURNAUA, S., LUCIANIA, A. and LÉCHAPPÉA, J. 2012. From knowledge-based research towards accurate and rapid testing of seed quality in winter rape. Seed Science Research, 22(S1): S80 - S85. DOI: 10.1017/S0960258511000316 Go to original source...
    20. YU, S., SU, T., CHEN, J., WANG, W., ZHANG, F., YU, Y., ZHANG, D., ZHAO, X. and LU, G. 2015. Real-time PCR as a diagnostic tool for evaluating the resistance of Chinese cabbage cultivars to Verticillium wilt. European Journal of Plant Pathology, 143: 543-557. DOI: 10.1007/s10658-015-0706-8 Go to original source...
    21. ZHOU, L., HU, Q., JOHANSSON A. and DIXELIUS C. 2006. Verticillium longisporum and V. dahliae: infection and disease in Brassica napus. Plant Pathology, 55: 137-144. DOI: 10.1111/j.1365-3059.2005.01311.x Go to original source...

    This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License (CC BY NC ND 4.0), which permits non-comercial use, distribution, and reproduction in any medium, provided the original publication is properly cited. No use, distribution or reproduction is permitted which does not comply with these terms.


    Return to the content