Resistance tests to diseases of tomato varieties for listing in Common Catalogue Loredana Sigillo, Romana Bravi Ente Nazionale Sementi Elette l.sigillo@ense.it
3rd International DUS Distinctness Uniformity Stability According to the official criteria, a vegetal variety can be listed in National and European Catalogue of Varieties if it is distinct, uniform and stable in fact According to the Acts of the UPOV Convention, protection can only be granted in respect of a new plant variety after examination of the variety has shown that is distinct (D) from any other variety whose existence is a matter of common knowledge and that it is sufficiently uniform (U) and stable (S), or DUS in short (UPOV, TG/1/3). Article 7 of the 1961/1972 and 1978 Acts and Article 12 of the 1991 Act For DUS tests, a variety is descripted by phenotipical characterization
Basic requirements that a characteristic used for description should fulfill The characteristic - results from a given genotype or combination of genotypes - is sufficiently consistent and repeteable in a particular environment - exhibits sufficient variation between varieties to be able to establish distinctness - is capable of precise definition and recognition - allows uniformity and stability requirements to be fulfilled For some species, disease resistance is used as a characteristic in DUS examination
Criteria for use of disease resistance characteristic - Knowing of which genes are responsable for resistance (one or more genes) - Standardization of the test resistance method - As a result of a resistance test, we should have sufficient variation between varieties to establish distincteness - The use of well defined reference materials The procedures to perfom DUS tests are reported in CPVO document.
CPVO guidelines Resistance assays are carried out according Community Plant Variety Office (CPVO) technical protocols. The protocol to be used in tomato variety characterization is the TP/44/3
CPVO protocols compulsory parameters to be respected The C.P.V.O. protocols set out the most important parameters to be respected in the trials as: - Sample size (number of plant to be tested) - Plant stage for inoculation - Inoculum density - Temperature for seedling growth and disease development - Growing method (in climatic room, in glasshouse) - Duration of the test - Symptom notation (diseased plant, health plant or plant with intermediate level of disease) The C.P.V.O. protocols show varieties and pathogen strains to be used as reference materials. For the proper pathogen denomination American Phytopathological Society and International Committee fot Taxonomy of Viruses are accepted
Resistance characters evaluated in tomato variety description Resistance to Meloydogine incognita Resistance to Fusarium oxysporum f. sp. lycopersici race 0 (ex 1) (FOL 0) Resistance to Fusarium oxysporum f. sp. lycopersici race 1 (ex 2) (FOL 1) Resistance to Verticillium dahlie race 0 Resistance to Tomato mosaic virus (ToMV) race 0, 1, 2 and 2a In 2003, only resistance against fungi were compulsory. From 2009, among virus races, the only resistance characteristic to be evaluated was the resistance against ToMV race 0
The role of ENSE laboratories in tomato variety Registration in National and European Catalogue The operative unit of Battipaglia (SA, Italy) works in phenotipical characterization of tomato varieties The Laboratory of Phytopathological Analyses performs resistance assays to evaluate the resistance of tomato candidate varieties against the most important pathogens
ENSE Internal Protocols ENSE laboratories produced internal resistance test protocols in harmonisation with the international guidelines according to CPVO and UPOV principles Basically - Resistance tests against fungi are performed by dipping root inoculation - Resistance tests against ToMV are performed by rubbing of the cotyledons - Resistance tests against nematodes are performed by inoculation of seeds with eggs
ENSE Reference Collections V. dahliae strain Vert 1 FOL 1 strain ATCC 16605 Low temperature collection Seed collection ENSE has a collection of reference materials: pathogen strains and varieties
Reference strains and varieties used in tomato tests Pathogen Reference strain Reference varieries M. incognita Mis strain (NL) Susceptible: Gianna (I) Resistant: Bolsena (I) Intermediate resistant: evaluating FOL 0 ATCC 16417 Susceptible: Gianna (I) Resistant: Revenge (I), Pressing (I) FOL 1 ATCC 16605 Susceptible: Gianna (I), Revenge Resistant: Pressing (I) V. dahliae Vert 5 (I) Susceptible: Gianna (I) Resistant: Bolsena (I) ToMV race 0 GdK (NL) Susceptible: Gianna (I) Resistant: Momor (I) ToMV race 1 SPS (NL) Susceptible: Gianna (I) Resistant: Momor (I) ToMV race 2 GeR0 (NL) Susceptible: Gianna (I) Resistant: Momor (I) ToMV race 2a GM6S (NL) Susceptible: Gianna (I)
Resistance tests carried out from 2003 to 2009 in Italy (ENSE laboratories) In the last seven years we tested 201 tomato varieties. The tests are repeated at least in two independent cycles. Totally more than 400 samples in 1400 resistance assays
Resistant varieties against the different pathogens from 2003 to 2009 In general, from 2003 to 2009, the number of resistant varieties to the different pathogens remained stable 100 90 80 70 60 50 40 30 20 10 0 M. incognita FOL 0 FOL 1 V. dahliae ToMV 0 ToMV 1 ToMV 2 ToMV 2a 2003 2004 2005 2006 2007 2008 2009 Pathogens
Resistant varieties FOL 0: about 95% FOL 1: about 60% ToMV: from 41% to 63% V. dahliae from 55% to 75% M. incognita from 13 to 56%.
Critical control points CCP in a standardized protocols - use reference strains characterized by reaction on differential sets of cultivar - use a known inoculum concentration - control of growth temperature - correct interpretation of symptoms - use the correct scale to note the disease symptoms (disease present/absent or 1-3 scale or 1-9 scale)
3 rd International
Future perspective It is generally assumed that resistant in vivo test needs good experience by the technicians. In vivo tests are time consuming and easly affected by external factors The use of genetical markers togheter with phenotypical descriptions is hoped UPOV working groups (Biochemical Molecular Techniques) are working on the realization of a molecular marker database and on harmonization of procedures. Some documents are proposed for maize. The items of molecular analyses is to allow the integrated statistical elaboration of genetical and morphological data distances. In tomato, the elaboration of protocols to detect resistance genes for M. incognita and ToMV resistances is in progress The definition of a procedures to test resistance against TSWV is needed
Thanks to Veronica Senape, Giovanna Serratore, Vincenzo Spina for technical collaboration to Liliana Barra and Marco Faina for slide background to all the working group of DUS testing Thank you for the attention!