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Multiple resistance of spinach (Spinacia oleracea L.) to cucumber mosaic and beet mild yellowing viruses
Zentralbl. Mikrobiol. 144 (1989), 13-18 VEB Gustav Fischer Verlag Jena
[Institut fiir Phytopathologie Aschersleben der Akademie der Landwirtschaftswissenschaften der Deutschen Demokratischen Republik und Saatzuchtstation des VEG Pflanzenproduktion Ascherslebenl
Multiple Resistance of Spinach (Spinacia oleracea L.) to Cucumber Mosaic and Beet Mild Yellowing Viruses H. E. SCHMIDT, U. MEYER, E. BRIEST, I. HAACK and K. EISBEIN With 2 Figures
Summary The spinach cultivar "Larina" proved to be highly resistant to 4 isolates of CMV and their equal mixture (= M). Additionally a moderate level of quantitative resistance of this cultivar to BMYV was confirmed serologically by use of ELISA. Deduced from significant differences of resistance levels within the cultivars "Larina" and "Matador" associated with single isolates of CMV, 2 groups of virus isolates were classified preliminarily distinguished by their virulence. The spinach isolate SoPe 28179, a mung bean isolate Mungo 9 and M belonged to the most virulent group differentiated by the cultivar "Matador".
Zusammenfassung Die Spinatsorte "Larina" erwies sich als hoch resistent gegen 4 Isolate des Gurkenmosaik-Virus (CMV) und ihr gleichteiliges Gemisch (= M). Zusiitzlich wurde maBige Resistenz dieser Sorte gegen das Milde RiibenvergilbungsVirus (BMYV) mit Hilfe des ELISA bestiitigt. Abgeleitet von signifikanten Unterschieden des Resistenzniveaus innerhalb der Sorten "Larina" und "Matador", die mit einzelnen CMV -Isolaten verbunden waren, wurden vorliiufig 2 Gruppen von Virusisolaten klassifiziert, die sich durch ihre Virulenz unterschieden. Das Spinatisolat SoPe 28179, ein Mung-Bohnenisolat Mungo 9 und M gehorten der virulentesten Gruppe an, die mit Hilfe der Sorte "Matador" differenziert wurde.
Serious yield losses of spinach induced by cucumber mosaic virus (CMV) and the increasing occurrence of beet mild yellowing virus (BMYV) in this vegetable (BRIEST et al. 1984, SCHMIDT et al. 1985) required a more intensive evaluation of resistance to these viruses.
Materials and Methods Actual genotypes of the world cultivar collection, hybrids, lines and strains of spinach were inoculated artificially with cucumber isolates GM-I, GM Dresden, the spinach isolate SoPe 28179, the mung bean isolate Mungo 9 of CMV, with an equal mixture of these isolates ( = M) as well as with the most virulent isolate Bv A of BMYV. All virus isolates used in this study originated from the GDR. Inoculation experiments were carried out under glasshouse conditions during the spring months of March and April. Temperatures ranged from 20 to 28°C. Plants inoculated with BMYV by Myzus persicae SULZ. during autumn were wintered in the field. Plants of the cultivars "Milena" and "Matador" were included in the trials as controls. Inoculation methods, serological detection of both viruses by ELISA and the calculation of degrees of resistance (= DR) according to 4 classes of disease symptom intensity (I = severe stunting of plants, 2 = prominent mosaic, 3 = weak slight mosaic, 4 = no symptoms) were applied as described (SCHMIDT and SCHUBERT 1980, BRIEST 1985, HAACK 1985, SCHMIDT et al. \985). The mathematical assay of infection rates occurred by use of a special analysis of variance. Differences of DR values corresponding with the number of plants distributed into the four classes of symptom intensity were tested by means of RIDIT-analysis (RYAN 1960, MEYER \988).
14
H. E.
SCHMIDT
et at.
Fig. 1. Reaction of spinach plants to CMV; Artificial inoculation using M of virus isolates; A - cv. "Matador", sensitive; B - cv. "Larina", resistant.
Results The 41 spinach genotypes studied were confined to a high-yielding strain released in 1987 as cultivar "Larina" , showing characters of combined resistance to CMV and BMYV. Compared with the standard cultivars "Matador" (Fig. 1 A), sensitive and "Milena", moderately resistant to CMV but susceptible to BMYV, most plants of "Larina" became not infected by CMV (Fig. IB). Individuals of this cultivar inoculated with BMYV by its vector M. persicae developed pale brightened leaves only. Contrarily, the growth of "Matador", "Milena", "Asta" and other cultivars was slightly inhibited by BMYV accompanied by a strong foliar yellowing. The resistance behaviour of "Larina" to CMV was studied in comparison with the spinach cultivars "Milena" and "Matador". Experiments were done in the course of five years. Results of artificial inoculation of these cultivars by the virus isolates SoPe 28179, GM Dresden, GM-I, Mungo 9 and their mixture M are summarized in Table 1. High infection rates of CMV isolates determined with the cultivar "Matador" (e.g., 94.4 ... 99.5 %) were representative for all trials. Under these conditions the high and stable resistance level of "Larina" to CMV was confirmed. As demonstrated in Figure 2 the analysis of variance of infection rates induced by the 5 variants of CMV indicated significant differences between the 3 cultivars. The overall mean artificial infection rates in sequence of "Larina" , "Milena" and "Matador" increased (10.6:41.9 and 96%) as the DR values decreased (93.3; 68.4 and 14.4, Table 1). The RIDIT -analysis included the original numbers of spinach plants belonging to four classes of disease symptom intensity. This mathematical assay supplied evidence of significant differences of mean DR values calculated for each cultivar (Table 2). True differences of DR depending on the virulence of distinct isolates of CMV were found within the cultivars "Larina" and "Matador" but not within cv. "Milena". The variant of DR within "Larina" corresponded with the virus isolate GM-I (Table 3). DR values of "Matador" associated with the virus isolates GM-I and GM Dresden were clearly distinct from those of the more virulent group of isolates constituted by SoPe 28179, Mungo 9 and M (Table 4).
Multiple Resistance of Spinach Table I. Reaction of spinach to isolates of CMV under glasshouse conditions Number of plants infected/inoculated
Degree of resistance 1)
Genotype
Isolate ofCMV
Larina Milena Matador
Mungo 9
33/231 1001227 2181219
91.9 65.3 9.1
Larina Milena Matador
GM-I
151233 1081214 2361243
96.3 71.8 21.3
Larina Milena Matador
GM Dresden
331203 90/206 235/241
91.1 63.8 16.9
Larina Milena Matador
SoPe 28179
23/233 631211 235/249
92.9 73.0 10.0
Larina Milena Matador
M
241227 711162 160/169
92.7 64.0 7.3
Larina Milena Matador
total
128/1127 43211020 108411121
93.3 68.4 14.4
1) Differences are significant by RIDIT-analysis (ex = 0.05).
100
pel" cent of plants infected
50
01234512345
Larina WSD= 7,9
Legend: 01 Mungo ~
2 GM-I
Milena WSD=13,2
2345
Matador WSD=4,5
134 SoPe 28/79 UIl5 Mixture M
~ 3 GM Dresden WSD wholly significant difference
Fig. 2. Infection rates of 3 spinach cultivars inoculated with 5 variants of CMV isolates.
15
16
H. E.
SCHMIDT
et al.
Table 2. Significant differences of degrees of resistance of 3 spinach cultivars to CMV (sum of 5 CMV variants)
Larina Milena Matador
+
DR
Milena
Matador
93.3 68.4 14.4
+
+ +
= significant by RIDIT-anaiysis of DR (0: = 0.05).
Table 3. Significant differences between symptom intensities of the cultivar "Larina" inoculated with 5 variants of CMV isolates Virus isolate
DR
4 2
96.3 92.9 92.7 91.9 91.1
I
5 3 I 5
2
5
3
+
= M, 2 = SoPe 28179, 3 = GM Dresden, 4 = GM-I, = Mungo 9, + = significant by RIDIT-analysis (0: = 0.05).
Table 4. Significant differences between symptom intensities of the cultivar "Matador" inoculated with 5 variants of CMV isolates Virus isolate
DR
4 3 2 5
21.3 16.9 10.0 9.1 7.3
I 5
3
2
5
+ +
+ +
+ +
= M, 2 = SoPe 28179, 3 = GM Dresden, 4 = GM-I, = Mungo 9, + = significant by RIDIT-analysis (0: = 0.05).
Artificially inoculated but symptomless plants of "Larina" and "Milena" tested by ELISA and by back inoculation onto Chenopodium quinoa WILLD. proved to be non- infected. This phenotypically extreme resistance was not brocken by the 4 isolates and M of CMV involved in the experiments. A similar high resistance was observed, if plants of "Larina" were grown in the field. In the course of 5 years the mean natural infection rates of cv. "Larina" and "Matador" averaged 0.1 and 24.5 %, respectively. One CMV isolate originating from a susceptible "Larina" plant was unable to overcome the resistance to CMV. Efforts to detect more virulent CMV strains breaking the CMV resistance in a long observation period over 15 years in the GDR garave negative results. Additionally to the CMV resistance screening of 41 spinach genotypes revealed evidence of the comparatively most decreased concentration of BMYV within cv. "Larina". For example, ranging over "Larina", "Milena" to "Matador" the preliminarily mean optical density (O.D. 405 urn) values of ELISA averaged 38.5,66.3 and 100% (P < 0.05), respectively. However, when high inoculum dosages of BMYV were used in the infection experiments these differences disappeared during overwintering of the spinach plants.
Multiple Resistance of Spinach
17
Discussion Up to the present time CMV followed by BMYV are predominant viruses of spinach in the GDR. Damages caused by them in future can be reduced by growing the resistant cultivar "Larina" mainly on localities known to have a high vector activity and virus incidence. The resistance to BMYV has quantitative character. More detailed investigations are necessary in order to classify the type of resistance to CMV and to identify its genetic base. The resistance of the spinach cultivar "Virginia Savoy" is governed by a single dominant gene (POUND and CHEO 1952). This resistance acts strain-specifically also in the cultivar "Old Dominion" and can be broken by virulent CMV strains found in the USA and Great Britain (FULTON 1950, DOOLITTLE and ZAUMEYER 1953, DOOLITTLE and WEBB 1960, WEBB et al. 1960, BAILISS and OKONKWO 1979). In consequence of plant quarantine no attempt was made to use such virulent virus strains as reference materials for our resistance tests. The CMV isolate Mungo 9 from mung bean (Vigna radiata (L.) R. WILCZEK var. radiata), infecting Phaseolus vulgaris systemically, was unable to overcome the resistance of cv. "Larina". In this property it differs from the resistance breaking strain D (FULTON 1950), the alfalfa strain (DOOLITTLE and ZAUMEYER 1953) and the Commelina strain ofCMV (DOOLITTLE and WEBB 1960) reported from the USA. These CMV strains induce systemic infections apparently comparable with isolate Mungo 9. Observations in the breeding process of several years and growing cv. "Larina" in the field hithertho revealed evidence of a good durability of CMV resistance on spinach stands of the GDR. Very low natural infection rates of CMV sometimes to be found, were not induced by resistance breaking strains. Some heterogeneity of the CMV resistance as reported by Bos (1975) may be the cause for the occurrence of single spontaneously diseased spinach plants in the field as well as after artificial inoculation experiments in the glasshouse. However, as shown by the trials, a notable variability of CMV isolates was stated. This agrees with accounts of FULTON (1950), WEBB et al. (1960). The authors mentioned that the symptom expression of a CMV sensitive spinach cultivar like "Bloomsdale" to any given CMV strain was relatively uniform, but its reaction to various virus strains proved to be highly variable. Therefore, the differences of DR values of cv. "Matador" to the CMV isolates studied in the former trials may be explained. The inability of cv. "Milena" and the reduced property of cv. "Larina", respectively, to differentiate the virulence of the CMV isolates used is not surprising, because the complete action of virulence genes may be prevented by corresponding resistance genes (ROBINSON 1976) or by the genetic background. DR values of the spinach cultivars were calculated on the base of visual classification of disease symptom intensities. Further investigations will have to distinguish more precisely DR or differences of the virulence of CMV isolates by metric characters, e.g. plant growth, yield, and O.D. values of ELISA. The level of CMV resistance in spinach may be degradated by increasing temperatures, e.g. between 24 to 32°C (POUND and CHEO 1952, WEBB et al. 1960, EISBEIN and HAACK 1985). The resistance level of cv. "Larina" was confirmed independently of this temperature effect under glasshouse conditions, characterized by high temperatures up to 28°C. During the spinach growing period such outdoor temperatures in the GDR are not typical. Since the resistance of "Larina" was maintained under field growing conditions, this cultivar can be recommended for horticulture. This is supported by its quantitative resistance to BMYV, which proved to be the highest of different spinach genotypes so far tested. In order to preserve the multiple virus resistance of cv. "Larina" and to apply this in breeding work, the selection of multiple restistant lines of this cultivar is recommended. According to CHOD (1983) also resistance to beet yellowing virus needs attention on the part of the spinach breeders. This virus is spread generally in several countries.
2 ZentralbL MikrobioL. Bd. 144
18
H. E. SCHMIDT et aI., MUltiple Resistance of Spinach
References BAILISS, K. W., OKONKWO, V. N. : Virus infection of spinach (Spinacia oleracea L.) in Britain. Jour. of Horticultural Science 54 (1979), 289-297. Bos, L.: Spinach. Cucumber mosaic virus. Ann. Rep. Inst. Phytopathol. Res., Wageningen 1974 (1975), 47. BRIEST, E.: Beitriige zur Priizisierung der Resistenzpriifung von Beta-Riiben gegeniiber Vergilbungsviren. Diss. A, Akad. Landw.-Wiss. DDR, Berlin (1984), 15I. SCHMIDT, H. E., KALININA, I., RiCHTER, J., ZOBYWALSKI, S.: Nachweis von Gemiisekulturen und Raps als natiirlich infizierte Winterwirte des Milden Riibenvergilbungs-Virus (beet mild yellowing virus). Arch. Phytopathol. u. Pflanzenschutz, Berlin 20 (1984), 177-179. CHaD, J.: Susceptibility of some spinach cultivars and hybrids to beet mosaic virus, beet yellowing virus and cucumber mosaic virus. Proc. XIVth Conf. Polish Plant Virologists, BiaZejewko, 1980. Zeszyty problemowe postep6w nauk roln. 291 (1983), 89-95, DOOLITTlE, S. P., WEBB, R. E.: A strain of cucumber mosaic virus I infectious to blight-resistant spinach. Phytopathology 50 (1960),7-9. - ZAUMEYER, W. J.: A pepper ringspot caused by strains of cucumber mosaic virus from pepper and alfalfa. Phytopathology 43 (1953), 333-337. EISBEIN, K., HAACK, I. : Veriinderungen im Resistenzverhalten von Spinat gegeniiber einem Stamm des Gurkenmosaik-Virus (cucumber mosaic virus) unter dem EinfluB hiiherer Temperaturen. Arch. Phytopathol. u. Pflanzenschutz, Berlin 21 (1985), 411-413. FULTON, J. P.: Studies on strains of cucumber mosaic virus 1 from spinach. Phytopathology 40 (1950),729-736. HAACK, 1.: Ein Beitrag zur Differenzierung und Diagnose von Vertretem der Cucumovirus-Gruppe. Diss. A, Akad. Landwirtsch.-Wiss. DDR, Berlin (1985), 176. MEYER, U.: Ein Verfahren zum Vergleich ordinalskalierter Bonitumoten. Arch. Phytopathol. u. Pflanzenschutz, Berlin 24 (1988), 349-35I. POUND, G. S., CHEO, P. C.: Studies on resistance to cucumber virus I in spinach. Phytopathology 42 (1952),
301-306. ROBINSON, R. A.: Plant pathosystems. Springer-Verlag, Berlin-Heidelberg-New York (1976). RYAN, T.: Significance tests for multiple comparison of proportions, variances and other statistics. Psychological Bull. 57 (1960),318-328. SCHMIDT, H. E., BRIEST, E., KALININA, I.: Das Milde Riibenvergilbungs-Virus (beet mild yellowing virus) als Ursache von Vergilbungskrankheiten bei Freilandgemiise in der Deutschen Demokratischen Republik. Arch. Gartenbau, Berlin 33 (1985), 247 - 257 . SCHUBERT, L.: Ergebnisse und Aufgaben der Virusresistenzziichtung bei Gemiiseerbsen (Pisum sativum L.), Spinat (Spinacia oleracea L.) und Tomate (Lycopersicon esculentum MILL.). Arch. Phytopath. u. Pflanzenschutz, Berlin 16 (1980), 77-88. WEBB, R. E., PERRY, B. A., JONES, H. A., McLEAN, D. M.: A new source of resistance to spinach blight. Phytopathology 50 (1960),54-56. Authors' addresses: Dr. sc. H. E. SCHMIDT, Dipl.-Mathem. U. MEYER, Dr. 1. HAACK. Institut flir Phytopathologie Aschersleben der AdL der DDR, Tbeodor-Roemer-Weg, Aschersleben, DDR - 4320; Dr. E. BRIEST, Dipl.-Landw. K. EISBEIN, VEG (P) Aschersleben, Ermslebener StraBe, Aschersleben, DDR - 4320.
[Institut fiir Phytopathologie Aschersleben der Akademie der Landwirtschaftswissenschaften der Deutschen Demokratischen Republik und Saatzuchtstation des VEG Pflanzenproduktion Ascherslebenl
Multiple Resistance of Spinach (Spinacia oleracea L.) to Cucumber Mosaic and Beet Mild Yellowing Viruses H. E. SCHMIDT, U. MEYER, E. BRIEST, I. HAACK and K. EISBEIN With 2 Figures
Summary The spinach cultivar "Larina" proved to be highly resistant to 4 isolates of CMV and their equal mixture (= M). Additionally a moderate level of quantitative resistance of this cultivar to BMYV was confirmed serologically by use of ELISA. Deduced from significant differences of resistance levels within the cultivars "Larina" and "Matador" associated with single isolates of CMV, 2 groups of virus isolates were classified preliminarily distinguished by their virulence. The spinach isolate SoPe 28179, a mung bean isolate Mungo 9 and M belonged to the most virulent group differentiated by the cultivar "Matador".
Zusammenfassung Die Spinatsorte "Larina" erwies sich als hoch resistent gegen 4 Isolate des Gurkenmosaik-Virus (CMV) und ihr gleichteiliges Gemisch (= M). Zusiitzlich wurde maBige Resistenz dieser Sorte gegen das Milde RiibenvergilbungsVirus (BMYV) mit Hilfe des ELISA bestiitigt. Abgeleitet von signifikanten Unterschieden des Resistenzniveaus innerhalb der Sorten "Larina" und "Matador", die mit einzelnen CMV -Isolaten verbunden waren, wurden vorliiufig 2 Gruppen von Virusisolaten klassifiziert, die sich durch ihre Virulenz unterschieden. Das Spinatisolat SoPe 28179, ein Mung-Bohnenisolat Mungo 9 und M gehorten der virulentesten Gruppe an, die mit Hilfe der Sorte "Matador" differenziert wurde.
Serious yield losses of spinach induced by cucumber mosaic virus (CMV) and the increasing occurrence of beet mild yellowing virus (BMYV) in this vegetable (BRIEST et al. 1984, SCHMIDT et al. 1985) required a more intensive evaluation of resistance to these viruses.
Materials and Methods Actual genotypes of the world cultivar collection, hybrids, lines and strains of spinach were inoculated artificially with cucumber isolates GM-I, GM Dresden, the spinach isolate SoPe 28179, the mung bean isolate Mungo 9 of CMV, with an equal mixture of these isolates ( = M) as well as with the most virulent isolate Bv A of BMYV. All virus isolates used in this study originated from the GDR. Inoculation experiments were carried out under glasshouse conditions during the spring months of March and April. Temperatures ranged from 20 to 28°C. Plants inoculated with BMYV by Myzus persicae SULZ. during autumn were wintered in the field. Plants of the cultivars "Milena" and "Matador" were included in the trials as controls. Inoculation methods, serological detection of both viruses by ELISA and the calculation of degrees of resistance (= DR) according to 4 classes of disease symptom intensity (I = severe stunting of plants, 2 = prominent mosaic, 3 = weak slight mosaic, 4 = no symptoms) were applied as described (SCHMIDT and SCHUBERT 1980, BRIEST 1985, HAACK 1985, SCHMIDT et al. \985). The mathematical assay of infection rates occurred by use of a special analysis of variance. Differences of DR values corresponding with the number of plants distributed into the four classes of symptom intensity were tested by means of RIDIT-analysis (RYAN 1960, MEYER \988).
14
H. E.
SCHMIDT
et at.
Fig. 1. Reaction of spinach plants to CMV; Artificial inoculation using M of virus isolates; A - cv. "Matador", sensitive; B - cv. "Larina", resistant.
Results The 41 spinach genotypes studied were confined to a high-yielding strain released in 1987 as cultivar "Larina" , showing characters of combined resistance to CMV and BMYV. Compared with the standard cultivars "Matador" (Fig. 1 A), sensitive and "Milena", moderately resistant to CMV but susceptible to BMYV, most plants of "Larina" became not infected by CMV (Fig. IB). Individuals of this cultivar inoculated with BMYV by its vector M. persicae developed pale brightened leaves only. Contrarily, the growth of "Matador", "Milena", "Asta" and other cultivars was slightly inhibited by BMYV accompanied by a strong foliar yellowing. The resistance behaviour of "Larina" to CMV was studied in comparison with the spinach cultivars "Milena" and "Matador". Experiments were done in the course of five years. Results of artificial inoculation of these cultivars by the virus isolates SoPe 28179, GM Dresden, GM-I, Mungo 9 and their mixture M are summarized in Table 1. High infection rates of CMV isolates determined with the cultivar "Matador" (e.g., 94.4 ... 99.5 %) were representative for all trials. Under these conditions the high and stable resistance level of "Larina" to CMV was confirmed. As demonstrated in Figure 2 the analysis of variance of infection rates induced by the 5 variants of CMV indicated significant differences between the 3 cultivars. The overall mean artificial infection rates in sequence of "Larina" , "Milena" and "Matador" increased (10.6:41.9 and 96%) as the DR values decreased (93.3; 68.4 and 14.4, Table 1). The RIDIT -analysis included the original numbers of spinach plants belonging to four classes of disease symptom intensity. This mathematical assay supplied evidence of significant differences of mean DR values calculated for each cultivar (Table 2). True differences of DR depending on the virulence of distinct isolates of CMV were found within the cultivars "Larina" and "Matador" but not within cv. "Milena". The variant of DR within "Larina" corresponded with the virus isolate GM-I (Table 3). DR values of "Matador" associated with the virus isolates GM-I and GM Dresden were clearly distinct from those of the more virulent group of isolates constituted by SoPe 28179, Mungo 9 and M (Table 4).
Multiple Resistance of Spinach Table I. Reaction of spinach to isolates of CMV under glasshouse conditions Number of plants infected/inoculated
Degree of resistance 1)
Genotype
Isolate ofCMV
Larina Milena Matador
Mungo 9
33/231 1001227 2181219
91.9 65.3 9.1
Larina Milena Matador
GM-I
151233 1081214 2361243
96.3 71.8 21.3
Larina Milena Matador
GM Dresden
331203 90/206 235/241
91.1 63.8 16.9
Larina Milena Matador
SoPe 28179
23/233 631211 235/249
92.9 73.0 10.0
Larina Milena Matador
M
241227 711162 160/169
92.7 64.0 7.3
Larina Milena Matador
total
128/1127 43211020 108411121
93.3 68.4 14.4
1) Differences are significant by RIDIT-analysis (ex = 0.05).
100
pel" cent of plants infected
50
01234512345
Larina WSD= 7,9
Legend: 01 Mungo ~
2 GM-I
Milena WSD=13,2
2345
Matador WSD=4,5
134 SoPe 28/79 UIl5 Mixture M
~ 3 GM Dresden WSD wholly significant difference
Fig. 2. Infection rates of 3 spinach cultivars inoculated with 5 variants of CMV isolates.
15
16
H. E.
SCHMIDT
et al.
Table 2. Significant differences of degrees of resistance of 3 spinach cultivars to CMV (sum of 5 CMV variants)
Larina Milena Matador
+
DR
Milena
Matador
93.3 68.4 14.4
+
+ +
= significant by RIDIT-anaiysis of DR (0: = 0.05).
Table 3. Significant differences between symptom intensities of the cultivar "Larina" inoculated with 5 variants of CMV isolates Virus isolate
DR
4 2
96.3 92.9 92.7 91.9 91.1
I
5 3 I 5
2
5
3
+
= M, 2 = SoPe 28179, 3 = GM Dresden, 4 = GM-I, = Mungo 9, + = significant by RIDIT-analysis (0: = 0.05).
Table 4. Significant differences between symptom intensities of the cultivar "Matador" inoculated with 5 variants of CMV isolates Virus isolate
DR
4 3 2 5
21.3 16.9 10.0 9.1 7.3
I 5
3
2
5
+ +
+ +
+ +
= M, 2 = SoPe 28179, 3 = GM Dresden, 4 = GM-I, = Mungo 9, + = significant by RIDIT-analysis (0: = 0.05).
Artificially inoculated but symptomless plants of "Larina" and "Milena" tested by ELISA and by back inoculation onto Chenopodium quinoa WILLD. proved to be non- infected. This phenotypically extreme resistance was not brocken by the 4 isolates and M of CMV involved in the experiments. A similar high resistance was observed, if plants of "Larina" were grown in the field. In the course of 5 years the mean natural infection rates of cv. "Larina" and "Matador" averaged 0.1 and 24.5 %, respectively. One CMV isolate originating from a susceptible "Larina" plant was unable to overcome the resistance to CMV. Efforts to detect more virulent CMV strains breaking the CMV resistance in a long observation period over 15 years in the GDR garave negative results. Additionally to the CMV resistance screening of 41 spinach genotypes revealed evidence of the comparatively most decreased concentration of BMYV within cv. "Larina". For example, ranging over "Larina", "Milena" to "Matador" the preliminarily mean optical density (O.D. 405 urn) values of ELISA averaged 38.5,66.3 and 100% (P < 0.05), respectively. However, when high inoculum dosages of BMYV were used in the infection experiments these differences disappeared during overwintering of the spinach plants.
Multiple Resistance of Spinach
17
Discussion Up to the present time CMV followed by BMYV are predominant viruses of spinach in the GDR. Damages caused by them in future can be reduced by growing the resistant cultivar "Larina" mainly on localities known to have a high vector activity and virus incidence. The resistance to BMYV has quantitative character. More detailed investigations are necessary in order to classify the type of resistance to CMV and to identify its genetic base. The resistance of the spinach cultivar "Virginia Savoy" is governed by a single dominant gene (POUND and CHEO 1952). This resistance acts strain-specifically also in the cultivar "Old Dominion" and can be broken by virulent CMV strains found in the USA and Great Britain (FULTON 1950, DOOLITTLE and ZAUMEYER 1953, DOOLITTLE and WEBB 1960, WEBB et al. 1960, BAILISS and OKONKWO 1979). In consequence of plant quarantine no attempt was made to use such virulent virus strains as reference materials for our resistance tests. The CMV isolate Mungo 9 from mung bean (Vigna radiata (L.) R. WILCZEK var. radiata), infecting Phaseolus vulgaris systemically, was unable to overcome the resistance of cv. "Larina". In this property it differs from the resistance breaking strain D (FULTON 1950), the alfalfa strain (DOOLITTLE and ZAUMEYER 1953) and the Commelina strain ofCMV (DOOLITTLE and WEBB 1960) reported from the USA. These CMV strains induce systemic infections apparently comparable with isolate Mungo 9. Observations in the breeding process of several years and growing cv. "Larina" in the field hithertho revealed evidence of a good durability of CMV resistance on spinach stands of the GDR. Very low natural infection rates of CMV sometimes to be found, were not induced by resistance breaking strains. Some heterogeneity of the CMV resistance as reported by Bos (1975) may be the cause for the occurrence of single spontaneously diseased spinach plants in the field as well as after artificial inoculation experiments in the glasshouse. However, as shown by the trials, a notable variability of CMV isolates was stated. This agrees with accounts of FULTON (1950), WEBB et al. (1960). The authors mentioned that the symptom expression of a CMV sensitive spinach cultivar like "Bloomsdale" to any given CMV strain was relatively uniform, but its reaction to various virus strains proved to be highly variable. Therefore, the differences of DR values of cv. "Matador" to the CMV isolates studied in the former trials may be explained. The inability of cv. "Milena" and the reduced property of cv. "Larina", respectively, to differentiate the virulence of the CMV isolates used is not surprising, because the complete action of virulence genes may be prevented by corresponding resistance genes (ROBINSON 1976) or by the genetic background. DR values of the spinach cultivars were calculated on the base of visual classification of disease symptom intensities. Further investigations will have to distinguish more precisely DR or differences of the virulence of CMV isolates by metric characters, e.g. plant growth, yield, and O.D. values of ELISA. The level of CMV resistance in spinach may be degradated by increasing temperatures, e.g. between 24 to 32°C (POUND and CHEO 1952, WEBB et al. 1960, EISBEIN and HAACK 1985). The resistance level of cv. "Larina" was confirmed independently of this temperature effect under glasshouse conditions, characterized by high temperatures up to 28°C. During the spinach growing period such outdoor temperatures in the GDR are not typical. Since the resistance of "Larina" was maintained under field growing conditions, this cultivar can be recommended for horticulture. This is supported by its quantitative resistance to BMYV, which proved to be the highest of different spinach genotypes so far tested. In order to preserve the multiple virus resistance of cv. "Larina" and to apply this in breeding work, the selection of multiple restistant lines of this cultivar is recommended. According to CHOD (1983) also resistance to beet yellowing virus needs attention on the part of the spinach breeders. This virus is spread generally in several countries.
2 ZentralbL MikrobioL. Bd. 144
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H. E. SCHMIDT et aI., MUltiple Resistance of Spinach
References BAILISS, K. W., OKONKWO, V. N. : Virus infection of spinach (Spinacia oleracea L.) in Britain. Jour. of Horticultural Science 54 (1979), 289-297. Bos, L.: Spinach. Cucumber mosaic virus. Ann. Rep. Inst. Phytopathol. Res., Wageningen 1974 (1975), 47. BRIEST, E.: Beitriige zur Priizisierung der Resistenzpriifung von Beta-Riiben gegeniiber Vergilbungsviren. Diss. A, Akad. Landw.-Wiss. DDR, Berlin (1984), 15I. SCHMIDT, H. E., KALININA, I., RiCHTER, J., ZOBYWALSKI, S.: Nachweis von Gemiisekulturen und Raps als natiirlich infizierte Winterwirte des Milden Riibenvergilbungs-Virus (beet mild yellowing virus). Arch. Phytopathol. u. Pflanzenschutz, Berlin 20 (1984), 177-179. CHaD, J.: Susceptibility of some spinach cultivars and hybrids to beet mosaic virus, beet yellowing virus and cucumber mosaic virus. Proc. XIVth Conf. Polish Plant Virologists, BiaZejewko, 1980. Zeszyty problemowe postep6w nauk roln. 291 (1983), 89-95, DOOLITTlE, S. P., WEBB, R. E.: A strain of cucumber mosaic virus I infectious to blight-resistant spinach. Phytopathology 50 (1960),7-9. - ZAUMEYER, W. J.: A pepper ringspot caused by strains of cucumber mosaic virus from pepper and alfalfa. Phytopathology 43 (1953), 333-337. EISBEIN, K., HAACK, I. : Veriinderungen im Resistenzverhalten von Spinat gegeniiber einem Stamm des Gurkenmosaik-Virus (cucumber mosaic virus) unter dem EinfluB hiiherer Temperaturen. Arch. Phytopathol. u. Pflanzenschutz, Berlin 21 (1985), 411-413. FULTON, J. P.: Studies on strains of cucumber mosaic virus 1 from spinach. Phytopathology 40 (1950),729-736. HAACK, 1.: Ein Beitrag zur Differenzierung und Diagnose von Vertretem der Cucumovirus-Gruppe. Diss. A, Akad. Landwirtsch.-Wiss. DDR, Berlin (1985), 176. MEYER, U.: Ein Verfahren zum Vergleich ordinalskalierter Bonitumoten. Arch. Phytopathol. u. Pflanzenschutz, Berlin 24 (1988), 349-35I. POUND, G. S., CHEO, P. C.: Studies on resistance to cucumber virus I in spinach. Phytopathology 42 (1952),
301-306. ROBINSON, R. A.: Plant pathosystems. Springer-Verlag, Berlin-Heidelberg-New York (1976). RYAN, T.: Significance tests for multiple comparison of proportions, variances and other statistics. Psychological Bull. 57 (1960),318-328. SCHMIDT, H. E., BRIEST, E., KALININA, I.: Das Milde Riibenvergilbungs-Virus (beet mild yellowing virus) als Ursache von Vergilbungskrankheiten bei Freilandgemiise in der Deutschen Demokratischen Republik. Arch. Gartenbau, Berlin 33 (1985), 247 - 257 . SCHUBERT, L.: Ergebnisse und Aufgaben der Virusresistenzziichtung bei Gemiiseerbsen (Pisum sativum L.), Spinat (Spinacia oleracea L.) und Tomate (Lycopersicon esculentum MILL.). Arch. Phytopath. u. Pflanzenschutz, Berlin 16 (1980), 77-88. WEBB, R. E., PERRY, B. A., JONES, H. A., McLEAN, D. M.: A new source of resistance to spinach blight. Phytopathology 50 (1960),54-56. Authors' addresses: Dr. sc. H. E. SCHMIDT, Dipl.-Mathem. U. MEYER, Dr. 1. HAACK. Institut flir Phytopathologie Aschersleben der AdL der DDR, Tbeodor-Roemer-Weg, Aschersleben, DDR - 4320; Dr. E. BRIEST, Dipl.-Landw. K. EISBEIN, VEG (P) Aschersleben, Ermslebener StraBe, Aschersleben, DDR - 4320.