Characterization of Sources of Resistance to the Watermelon Strain of Papaya Ringspot Potyvirus in Cucumber: Allelism and Cosegregation with other Potyvirus Resistances

R. Grumet, E. Kabelka, S. McQueen, and R. Humphrey

Department of Horticulture, Michigan State University, East Lansing MI 48824

Additional index words. Cucumis sativus, zucchini yellow mosaic virus, virus resistance

Abstract. At least three sources of resistance to the watermelon strain of papaya ringspot potyvirus (PRSV-W) have been identified in cucumber (Cucumis sativus L.) germplasm: 'Suinam', 'TMG-1', and 'Dina-1'. 'TMG-1', and 'Dina-1' also are resistant to several other potyviruses including zucchini yellow mosaic virus (ZYMV), watermelon mosaic virus and the Moroccan watermelon mosaic virus. We sought to understand inheritance of multiple potyvirus resistance in cucumber by studying the allelic and dominance relationships among the three sources of PRSV-W resistance and by examining the relationship between resistance to PRSV-W and the other potyvirus resistances in 'TMG-1' and 'Dina-1'. Inheritance studies indicated that resistance to PRSV-W in 'Dina-1' is conferred by a single recessive or incompletely dominant gene and that the resistances in all three genotypes are at the same locus. 'Surinam' is not resistant to the other potyviruses, and progeny of 'Surinam' x 'TMG-1', which were all resistant to PRSV-W, segregated for resistance to ZYMV. Prior or concurrent inoculation of 'Surinam' with ZYMV and PRSV-W did not influence the response to either virus. Resistance to ZYMV and PRSV-W in 'TMG-1', however, completely cosegregated in sequential inoculation experiments with F2 individuals and in F3 families. Collectively these results provide evidence for a series of alleles at the zym locus with varying viral specificities and resistance responses, or for a cluster of tightly linked potyvirus resistance genes in cucumber.

We thank Sue Hammar for technical assistance and Holly Kuhnle, Pam Weckworth, and Kurt Kuhnle for assistance in the greenhouse. This work was in part supported by the Office of USAID Cairo Agr A under Cooperative Agreement No. 263-0152-A-00-3036-00 and by the Michigan Agriculture Experiment Station. S. McQueen was supported by an MSU Undergraduate Professorial Assistantship.

The watermelon strain of papaya ring-spot virus (PRSV-W) is one of several potyviruses causing severe damage to production of cucurbit crops worldwide (Purcifull et al., 1984a). At least three sources of resistance to this virus have been identified in cucumber (Cucumis sativus L.). These include 'Surinam', a cultivar from South American (Wang et al., 1984), 'TMG-1', an inbred line derived from a single plant selection from the Taiwanese cultivar 'Taichung Mou Gua' (Provvidenti, 1985), and 'Dina-1', an inbred line derived from the Dutch hybrid, 'Dina' (Kabelka et al., 1997). Resistance in 'Surinam' is conferred by a single recessive gene (Wang et al., 1984) while resistance in 'TMG-1' appears to be conditioned by a single dominant or incompletely

dominant gene (Wai and Grumet, 1995a). The inheritance of PRSV-W resistance in 'Dina-1' and relationships among the three PRSV-W resistances is not known.

In addition to PRSV-W, there are several other potyviruses that infect cucurbit crops including zucchini yellow mosaic virus (ZYMV), watermelon mosaic virus (WMV), the Moroccan watermelon mosaic virus (MWMV) and zucchini yellow fleck virus (ZYFV) (Gilbert-Albertini et al., 1995; Lisa and Lecoq, 1984; McKern et al., 1993; Purcifull et al., 1984b). 'Surinam' is only resistant to PRSV-W, but 'TMG-1' and 'Dina-1' also are resistant to ZYMV, WMV, and MWMV (Albertini et al., 1995, Kabelka et al., 1997; Provvidenti, 1985). In 'Dina-1' and 'TMG-1', resistance to ZYMV is conferred by single genes (Abul-Hayja and Al-Shahwan, 1991; Provvidenti, 1987) that are alleles of the same locus (Kabelka et al., 1997). The zymDina allele, which allows for viral spread and distinct veinal chlorosis that is limited to one leaf, is

 

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dominant to the zymTMG allele which appears to restrict virus accumulation more rapidly; both are recessive to susceptibility. Resistance to ZYFV (Gilbert-Albertini et al., 1995) and MWMV (Kabelka and Grumet, 1997) also are controlled by single recessive genes in 'TMG-1' and 'Dina-1', while resistance to WMV in 'TMG-1' appears to be more complex (Wai and Grumet, 1995b). The gene conferring resistance to ZYMV in 'TMG-1' appears to be the same as, or tightly linked to, genes conferring resistance to WMV and MWMV (Kabelka and Grumet, 1997; Wai and Grumet, 1995b); similarly the resistances to ZYMV and MWMV completely cosegregate in 'Dina-1' (Kabelka et al., 1997).

In this investigation we sought to further understand inheritance of multiple potyvirus resistance in cucumber by studying the allelic and dominance relationships among the three sources of PRSV-W resistance, and by examining the relationship between resistance to PRSV-W and the other potyvirus resistances in 'TMG-1' and 'Dina-1'. Collectively, these results examining the various potyvirus resistances provide evidence for a cluster of tightly linked potyvirus resistance genes in cucumber.

Materials and methods

Cucumber genotypes. The inbred potyvirus resistant cucumber (Cucumis sativus L.) lines 'TMG-1' and 'Surinam' were originally provided by J. Staub (USDA, University of Wisconsin, Madison) and R. Provvidenti (Cornell University, Geneva N.Y.), respectively. Self-pollinated progeny of the Dutch hybrid 'Dina', true breeding for resistance to ZYMV ('Dina-1') were initially provided by K. Owens (Seminis Seed Co., Woodland, Calif.). All lines were increased by self- or sib-pollinations in the greenhouse. The two susceptible parental genotypes were 'Wisconsin-2757' [WI-2757 (Peterson, 1982); provided by J. Staub] and 'Straight-8' (W. Atlee Burpee and Co., Warminster, Pa.). F1, F2 and backcross progeny of the crosses among all genotypes were produced in the greenhouse. F3 families derived from ZYMV-resistant 'TMG-1' x 'WI-2757' F2 individuals were produced in the field as described in Kabelka and Grumet (Kabelka and Grumet, 1997).

Virus inocula, inoculation procedures, experimental designs, and symptom scoring. PRSV-W (ATCC PV-380), ZYMV (Connecticut strain), WMV (ATCC PV379), and MWMV (originally provided by Dr. D. Purcifull, University of Florida, Gainesville) were propagated in Cucurbita pepo L. 'Black Beauty' (SeedWay Inc., Elizabethtown, Pa.) or 'Midas' (Willhite Seed, Poolville, Texas) as described in Kabelka and Grumet (Kabelka and Grumet, 1997). Purity of the virus sources was verified by ELISA and by the use of the differential host, Phaseolus vulgaris cv. Black Turtle 2 (Provvidenti, 1984). Preparation of inocula, rub-inoculations, germination, planting, fertilization, greenhouse growth conditions, and experimental designs were as described in Kabelka and Grumet (Kabelka and Grumet, 1997).

Plants were visually scored as either resistant (symptom-free) or susceptible when symptom development was optimal on the susceptible genotypes [approximately 14 days post inoculation (dpi) with PRSV-W and seven dpi with ZYMV]. In some cases a symptom 0­4 severity scale was used with 0 = no symptoms; 1­2 = mild symptoms and an absence of symptoms in the youngest leaves; 3­4 = moderate to severe symptoms including the emerging leaves.

Results

Inheritance of prsv-w resistance in 'dina'. Segregation for PRSV-W symptom development in the progeny of 'Dina-1' x 'Straight 8' (susceptible) indicates that resistance in 'Dina-1' is conferred by a single recessive or incompletely dominant gene (Table 1). Although 3/4 of the F2, half of the backcross to 'Dina-1', and all of the backcross to 'Straight 8' developed symptoms (Table 1), an intermediate phenotype of reduced symptom severity and rate of spread could be observed in the F1 and segregating progeny. When scored on a symptom severity scale of 0­4 (Table 1, Expts. 3 and 4), the majority of the F1 progeny fell into a reduced symptom class with ratings of 1­2 (symptoms were mild and were only present on older leaves but not young emerging leaves). The number of individuals exhibiting the intermediate phenotype in the F2 and backcross generations was consistent with the numbers expected for the

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Table 1. Inheritance of PRSV-W resistance in 'Dina-1'.

Parent Susceptible

or Resistantz Intermediate Moderate/severe

progeny Expt. (0)y (1­2) (3) c2

Dina (R) 1,2 20 --- 0

3,4 30 0 0

Straight 8 (S) 1,2 0 --- 20

3,4 0 0 30

F1 1,2 2 --- 19

3,4 2 24 8

F2 1 32 --- 87 0.28ns (1:3)

3 23 35 31 5.49ns (1:2:1)

1,3 pooledx 55 --- 154 0.093ns (1:3)

BC (F1 x R) 1,2 52 --- 49 0.23ns (Expt. 1); 0ns (Expt. 2) (1:1)

3 65 68 4 0.28ns (Expt. 3); 0.01ns (Expt. 4)(1:1)

1­4 pooled 117 --- 121 0.04ns (1:1)

BC (F1 x S) 1 0 --- 100

3 1 67 69 0.16ns (Expt. 3); 0.05ns (Expt. 4) (1:1)

1­4 pooled 1 --- 236

zIn Expts. 1 and 2, plants were only scored as resistant or susceptible.

yIn Expts. 3 and 4, plants were rated on a 0­4 scale: 0 = no symptoms; 1­2 = mild symptoms and an absence of symptoms in the youngest leaves; 3­4 = moderate to severe symptoms including the emerging leaves.

xPooled data were classified as either resistant or susceptible.

nsNonsignificant.

heterozygous classes. These results suggest that a single copy of the resistance allele delayed and reduced, but did not prevent symptom development.

Relationship between resistance to prsv-w in 'tmg-1', 'dina-1', and 'surinam'. To determine if the resistance genes for PRSV in 'TMG-1', 'Dina-1' and 'Surinam' are at the same locus, segregating progeny of 'TMG-1' x 'Dina-1' and 'TMG-1' x 'Surinam' were inoculated with PRSV-W. Although the susceptible control genotype developed systemic rugosity of the foliage approximately 14 to 21 days postinoculation, the resistant parents, 'TMG-1', 'Dina-1', and 'Surinam', and all individuals of the various F1, F2, and backcross generations remained symptom free (Table 2).

Comparison of performance of the prsv-w resistance alleles. The 'Surinam' and 'Dina-1' resistance alleles both complemented the allele from 'TMG-1', indicating that all three alleles are at the same locus; however, the allele from 'Surinam' is reported to be recessive (Wang et al., 1984), while the alleles from 'Dina-1' (Table 1), and 'TMG-1' (Wai and Grumet, 1995a) appeared to be incompletely dominant or dominant. To clarify their relative performances, 'TMG-1' and 'Surinam' were each crossed to a common susceptible genotype, 'Straight 8', and their progeny were tested concurrently.

Consistent with the previous studies, the F1 progeny of 'Straight 8' x 'Surinam' exhibited PRSV-W symptoms throughout the plant that were as severe as the susceptible parent (Table 3). The F2 progeny segregated in a 1:3 (R:S) ratio, the backcross to the resistant parent segregated 1:1 (R:S) and progeny of the backcross to the susceptible parent were all susceptible.

The F1 progeny of 'TMG-1' crossed with 'WI-2757' or 'Straight 8', however, exhibited an intermediate phenotype where the young leaves and growing points were symptom free, but the older

Table 2. Tests for allelism among PRSV-W resistances in 'TMG-1', 'Surinam', and 'Dina'.z

Parent or progeny Resistant Susceptible

TMG 56 0

Surinam 16 0

Dina 40 0

Straight 8 0 36

F1 TMG x Surinam 16z 0

F2 TMG x Surinam 320 0

BC1 F1 x TMG 80 0

BC2 F1 x Surinam 200 0

F1 TMG x Dina 30y 0

F2 TMG x Dina 318 0

BC1 F1 x TMG 120 0

BC2 F1 x Dina 159 0

zData are pooled from two experiments.

yNumber of resistant or susceptible plants.

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Table 3. Response of 'TMG-1', 'Surinam, 'Straight-8', 'WI-2757', and their progenyz to inoculation with PRSV-W.

Susceptible

Parent or progeny Resistanty Intermediatex Moderate/severew Expected c2

TMG-1 15 0 0

Surinam 15 0 0

Straight 8 0 0 15

WI-2757 0 0 15

F1 Straight 8 x Surinam 0 0 40

F2 Straight 8 x Surinam 33 0 86 1:3 0.34ns

BC1 F1 x Surinam 32 0 28 1:1 0.15ns

BC2 F1 x Straight 8 0 0 60

F1 Straight 8 x TMG 0 20 0

F2 Straight 8 x TMG 33 55 32 1:2:1 0.85ns

BC1 F1 x TMG 28 32 0 1:1 0.15ns

BC2 F1 x Straight 8 0 29 31 1:1 0.02ns

F1 WI-2757 x TMG 0 20 0

F2 WI-2757 x TMG 14 29 15 1:2:1 0.03ns

BC1 F1 x TMG 21 19 0 1:1 0.03ns

BC2 F1 x WI-2757 0 17 23 1:1 0.63ns

zNumber of plants.

yPlants are symptom-free, vigorous and healthy.

xYoung leaves and growing points remain symptom free, but older leaves exhibit rugosity and/or silver banding.

wSystemic rugosity and/or silver banding throughout the plant, similar to that exhibited by either susceptible parent.

nsNonsignificant.

leaves exhibited rugosity and/or silver banding. The F2 progeny segregated in a 1:2:1 (resistant: intermediate: susceptible) ratio. Progeny of the backcross to the resistant parent segregated in a 1:1 (R:I) ratio while progeny of the backcross to the susceptible parent segregated 1:1 (I:S).

Relationship between resistances to prsv-w and zymv. As has been mentioned, 'TMG-1' and 'Dina-1' are resistant to several potyviruses including PRSV-W, ZYMV, WMV, and MWMV. Previous studies have shown that the alleles for ZYMV, WMV, and MWMV resistance in 'TMG-1', and the alleles for ZYMV and MWMV resis

tance in 'Dina-1' appear to be at the same locus or very tightly linked loci (<1 cM apart) (Kabelka et al., 1997; Wai and Grumet, 1995b). The relationship of PRSV-W resistance to the other potyvirus resistances, however, is not known. Unlike 'TMG-1' and 'Dina-1', 'Surinam' is susceptible to ZYMV, WMV and MWMV.

Relationship between response to prsv-w and zymv in 'surinam'. In addition to the genetic separation between ZYMV and PRSV-W resistance in 'Surinam', there also does not appear to be a physiological relationship. Prior inoculation with PRSV-W does not induce a subsequent resistance

Table 4. Sequential inoculation of Surinam plants with PRSV-W and ZYMV.z

Plants with symptoms/

total inoculated

Treatment Genotype PRSV ZYMV

ZYMV, 7 days Straight 8 --- 9/9

Surinam --- 9/9

ZYMV, 21 days Straight 8 --- 9/16

Surinam --- 10/16

PRSV-W, 7days Straight 8 16/16 ---

Surinam 0/16 ---

PRSV-W + ZYMV, 7 days Straight 8 9/9 9/9

Surinam 0/9 9/9

PRSV-W, 7 days; ZYMV 21 days Straight 8 9/9 ---

Surinam 0/16 12/16

zData are pooled from two experiments.

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Table 5. Response of progeny of 'TMG-1' and 'Surinam' to inoculation with ZYMV.z

Susceptible

Parent or progeny Resistant Intermediate Moderate/severe c2

TMG-1 59 0 0

Surinam 0 0 56

F1 0 56 0

F2 139 223 111 4.93ns (1:2:1)

F1 x TMG 117 112 0 0.07ns (1:1)

F1 x Surinam 0 122 100 1.99ns (1:1)

zNumber of plants; data are pooled from three experiments.

to ZYMV infection, nor does coinoculation with ZYMV and PRSV-W result in resistance to ZYMV or susceptibility to PRSV-W (Table 4). When cotyledons of 7-day-old 'Surinam' plants, or true leaves of 21-day-old plants, were inoculated with ZYMV systemic mosaic symptoms were observed; the plants inoculated with PRSV-W remained symptom free, as expected. Similarly, when cotyledons were inoculated with both viruses, the systemic leaves only exhibited ZYMV symptoms that occurred at the same time as, and were as severe as, those inoculated with ZYMV alone. Finally, when true leaves of the symptom-free PRSV-W cotyledon-inoculated plants were inoculated two weeks later with ZYMV (at a time when PRSV-W symptoms were clearly visible on the susceptible 'Straight 8' plants), the 'Surinam' plants developed obvious ZYMV symptoms. It should be noted that the early symptoms of ZYMV and

PRSV-W are clearly distinguishable both in sequential and mixed inoculation experiments; PRSV-W induces rugosity, ZYMV induces yellowing and mosaic.

The genetic separation between ZYMV and PRSV-W resistance in 'Surinam' is also evident in segregating progeny of 'TMG-1' x 'Surinam'. The F1 progeny, are resistant to PRSV-W (Table 2), but are susceptible to ZYMV (Table 5). However, unlike recessive inheritance of ZYMV resistance in crosses between 'TMG-1' and the potyvirus susceptible 'Straight 8' or 'WI-2757' lines, the F1 progeny of 'TMG-1' x 'Surinam' exhibit a distinct intermediate phenotype of delayed symptom development and reduced symptom severity.

Relationship between response to prsv-w and zymv in 'tmg-1'. Despite the separation of resistances to ZYMV and PRSV-W in 'Surinam' and its progeny, resistances to ZYMV and PRSV-W did

Table 6. Response of 'TMG-1', 'Straight 8', F1 and backcross progeny to cotyledon inoculation with PRSV-W or ZYMV and sequential true leaf inoculation of resistant individuals with ZYMV or PRSV-W.

Cotyledon True leaf

Genotype Total plants Resistant Intermediate Susceptible Resistant Susceptible

PRSV-W cotyledon, ZYMV true leaf

TMG-1 10 10 0 0 10 0

Straight 8z 10 0 0 10 --- ---

Straight 8y 5 --- --- --- 0 5

F1z 10 0 10 0 --- ---

F1y 5 --- --- --- 0 5

BC (F1 x TMG)x 160 77 83 0 77 0

ZYMV cotyledon, PRSV-W true leaf

TMG-1 10 10 0 10 0 0

Straight 8 10 0 10 --- --- ---

Straight 8 5 --- --- 0 0 5

F1 10 0 10 --- --- ---

F1 5 --- --- 0 5 0

BC (F1 x TMG) 160 78 82 78 0 0

zControl plants to verify successful inoculation at cotyledon stage.

yControl plants to verify successful inoculation at true leaf stage.

xData fit the predicted segregation ratios based on resistance to PRSV-W conferred by an incompletely dominant gene (c2 = 0.16ns) and ZYMV by a single recessive gene (c2 = 0.06ns).

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Table 7. Reaction (resistant vs. susceptible/segregating) to PRSV-W and ZYMV inoculation of F3 families derived from 'TMG-1' x 'WI-2757' F2 individuals selected for resistance to ZYMV (i.e., homozygous, recessive at the zym locus).z

Susceptible or

Virus Resistant segregating

PRSV-W 51 0

ZYMV 51 0

zTen individuals/family were tested with each virus.

F3 families screened. These F3 families also previously had been screened for response to MWMV and all were resistant (Kabelka and Grumet, 1997).

Discussion

Inheritance, allelism and performance analyses of prsv-w resistances. Inheritance studies of PRSV-W resistance in 'Dina-1' indicated, that like the resistances in 'TMG-1' and 'Surinam', resistance in 'Dina-1' is controlled by a single gene. Crosses among the three resistant genotypes, indicated that the PRSV-W resistances are at the same locus, however, resistance from 'Surinam' appeared to be recessive, whereas the resistances from 'TMG-1' and 'Dina-1' appeared to be incompletely dominant (i.e., the heterozygous progeny exhibit an intermediate phenotype of reduced symptom severity and rate of development).

The differences in performance may be due to different alleles at the PRSV-W locus, or alternatively, genetic background and/or environmental factors may influence the relative effectiveness of the resistance genes. Although environment has been observed to play a role in full expression of the PRSV-W resistance in TMG-1 (Wai and Grumet, 1995a), when compared in the same environment and crossed with the same susceptible parent, 'TMG-1' and 'Surinam' still performed differently. This suggests a genetic component that may either be due to the allele itself and/or other modifying factors, including possible association with other tightly linked potyvirus resistance loci in 'TMG-1'.

Relationship of prsv-w and zymv resistances. Earlier studies have determined that 'TMG-1' and 'Dina-1' are resistant to multiple potyviruses including PRSV-W, ZYMV, WMW, and MWMV and that the alleles for ZYMV, WMV and MWMV completely cosegregate. In this study we also have identified a tight association between resistance to PRSV-W and ZYMV in 'TMG-1'. Sequential inoculation of ZYMV resistant backcross progeny with PRSV-W (or PRSV resistant backcross progeny with ZYMV) and analysis of F3 families derived from F2 individuals selected for resistance to ZYMV, indicate that both resistances are conferred by the same gene, or tightly linked genes (<1 cM; product-ratio method). These results are

not separate in progeny of 'TMG-1' crossed with a susceptible genotype. Two approaches were take to examine the relationship between ZYMV and PRSV-W resistances in 'TMG-1'. The first was sequential inoculation of backcross progeny of ('Straight 8' x 'TMG-1') x 'TMG-1'. This procedure consisted of cotyledon inoculation with PRSV-W followed by true leaf inoculation of the resistant individuals with ZYMV (Table 6). In a separate experiment, this procedure was reversed the cotyledons were inoculated with ZYMV and the true leaves of the resistant individuals were inoculated with PRSV-W (Table 6). In all experiments, additional control plants were included to verify successful inoculation at the true leaf stage. Symptoms of ZYMV or PRSV-W developed on the susceptible genotype 'Straight 8' and the F1 progeny following either cotyledon or true-leaf inoculation. The resistant genotype, 'TMG-1', remained free of symptoms of both PRSV-W and ZYMV. In all cases, those backcross individuals that were resistant to cotyledon inoculation with the first virus remained symptom free upon true leaf inoculation with the second virus.

The second approach to evaluate whether a relationship exists between the resistance to ZYMV and resistance to PRSV-W was to screen F3 families that were produced by self pollination of F2 individuals selected for resistance to ZYMV. Progeny from each ZYMV-resistant F2 individual were inoculated with either PRSV-W or ZYMV (Table 7). As expected, all families were resistant to ZYMV. If the two resistances had been segregating independently, 9/16 of the F3 families should be susceptible or segregating for susceptibility to PRSV-W. There was, however, no segregation for susceptibility to PRSV-W either within or among the

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also consistent with analyses indicating that both PRSV-W and ZYMV resistances are linked to the bi locus for bitterfree cotyledons (Wai et al., 1997; Wang et al., 1987), and observations that have been made when working with these resistances in cucumber breeding programs (Kyle and Provvidenti, 1993).

The existence of simply inherited genes, or clusters of separate tightly linked genes that confer resistance to two or more distinct potyviruses has been described previously in several species. In Cucurbita moschata a single dominant gene confers resistance to both ZYMV and WMV (Gilbert-Albertini, 1993), while in Phaseolus vulgaris the possibility of a single gene, or cluster of tightly linked genes cosegregating as a unit with the I gene conditions resistance and/or lethal necrosis to a set of nine potyviruses (Fisher and Kyle, 1994; Kyle and Dickson, 1988). In Pisum sativum, well defined clusters of tightly linked loci conferring resistance to a total of 11 potyviruses are located on two chromosomes (Provvidenti and Hampton, 1993; Provvidenti and Niblett, 1994). Chromosome 2 contains resistance to seven potyviruses, while chromosome 6 includes resistance to three more potyviruses, two of which overlap with those on chromosome 2. Pea lines with differential resistances to the various viruses have been identified, and only one of the genes, mo, which confers resistance to bean yellow mosiac virus and WMV resistance, exhibits temperature sensitivity in the heterozygous state (reviewed in Kyle and Provvidenti, 1993).

Despite the inability to break the linkage among the resistances to PRSV-W, ZYMV, WMV, and MWMV in 'TMG-1', 'Surinam' is only resistant to PRSV-W and prior inoculation of 'Surinam' plants with PRSV-W did not induce resistance to ZYMV. Similarly, presence of PRSV-W in mixed inoculations did not induce resistance to ZYMV, nor did infection by ZYMV overcome resistance to PRSV-W. This separation of resistances also was observed in progeny of 'Surinam' x 'TMG-1'. Although all progeny are resistant to PRSV-W, they segregated for susceptibility to ZYMV.

In summary, resistance to at least four different potyviruses, ZYMV, WMV, MWMV, and

PRSV-W completely cosegregated in 'TMG-1' progeny indicating that the multiple resistances are either due to a single gene with a broad range of specificities, or alternatively, the resistances may be due to a cluster of tightly linked resistance genes. Although we cannot genetically distinguish between these possibilities at this time, four lines of evidence indicating differential performance of the different virus resistances suggest that the resistances may not all be the same, and therefore may be conferred by a gene cluster. First, resistance to PRSV-W in 'TMG-1' appears incompletely dominant while resistance to ZYMV is recessive. It is possible that these differences may be due to varying effectiveness of one gene against different potyviruses, but it is also possible that they are due to two different genes. Second, the mechanism and/or response rate of resistance to PRSV and ZYMV in 'TMG-1' differs. When 'TMG-1' was inoculated with PRSV-W, high virus levels were detected in young leaves while little or no ZYMV was detected in equivalent leaves (Wai and Grumet, 1995a). Again, this could be due to different effectiveness of one gene against different viruses, or to different genes. Third, 'Dina-1', which is resistant to ZYMV, PRSV-W and MWMV shows different responses to ZYMV than to PRSV-W or MWMV inoculation. A distinct veinal chlorosis phenotype limited to the first and second true leaves was observed with ZYMV cotyledon inoculation whereas no symptoms were observed with PRSV-W or MWMV inoculation (Kabelka et al., 1997). Fourth, although 'Surinam' is resistant to PRSV-W, it is susceptible to ZYMV, WMV and MWMV. This may be due to different responses of one gene to different potyviruses, or it may be that 'Surinam' only possesses one member of a gene cluster.

In conclusion, although we have not been able to break the linkage associations among the resistances to PRSV-W, ZYMV, WMV, and MWMV in 'TMG-1', varying responses to the different viruses with regard to dominance relationships, resistance mechanisms, symptom expression, and which viruses are protected against, support the possibility that multiple potyvirus resistance in cucumber is conferred by a tightly linked cluster of resistance genes.

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