Euphytica 96: 339–344, 1997. 339c 1997 Kluwer Academic Publishers. Printed in the Netherlands.

Transfer of resistance to potato cyst nematode (Globodera pallida) intocultivated potato Solanum tuberosum through first division restitution 2npollen

Rodomiro Ortiz1;*, Javier Franco2 & Masaru Iwanaga2

1 The Royal Veterinary & Agricultural University, 40 Thorvaldsensvej, DK 1871, Frederiksberg C, Copenhagen,Denmark; 2 Centro Internacional de la Papa, Apartado Postal 1558, Lima, Peru; (* author for correspondence)

Received 24 September 1996; accepted 8 April 1997

Key words: Solanum tuberosum, 4x-2x crosses, breeding values, pest resistance, tuber-bearing Solanums

Summary

Genetic resistance to potato cyst nematode is considered as one of the most effective means of increasing yield andreducing nematode infestation levels in potato fields. In this study, resistance to this nematode was successfullytransferred from diploid tuber-bearing Solanums to the tetraploid gene pool using a 4x-2x breeding approach. Morespecifically, resistance from Solanum vernei, S. sparsipilum and haploids of S. tuberosum group Andigena wasintrogressed into conventional tetraploid clones, using first division restitution (FDR) 2n gametes. Furthermore,some of the FDR diploid parents had similar breeding values as advanced resistant tetraploid clones which weredeveloped only after several cycles of selection against the potato cyst nematode.

Introduction

Potato cyst nematode (Globodera pallida Stone Chez.)is an important constraint to potato production in tem-perate areas worldwide. While chemical treatmentmay increase yield, it is expensive and highly toxic,and does not effectively control nematode populations.Cultural control methods, such as crop rotation, maybe effective against nematodes but they require time.The use of resistant cultivars is the most appealingcontrol strategy, because it reduces both yield loss andnematode populations in infested fields.

Transfer of resistance to potato cyst nematodehas been the subject of intensive worldwide research(Brodie et al., 1991; Chavez et al., 1988; Dellaert etal., 1988; Phillips et al., 1979a; Phillips & Dale, 1982).Genetic variability has been reported for the potato cystnematode which makes resistance breeding more dif-ficult. Furthermore, the resistance to this pest seems tobe complex: on one or two dominant genes but inter-acting with minor genes (Dellaert et al., 1988). Dale& Phillips (1985) suggested that different non-specificfield resistance genes should be combined to increase

the durability of the resistance when breeding cultivarswith a high level of resistance to this pest.

Potato cyst nematode populations P4A and P5A areamong the most aggressive pathotypes. At least twogenes with epistatic effects control resistance to P4A(Gonzalez, 1981). Resistance to P5A is determined byan independent and more complex system. Evaluationof haploid progeny from resistant parents indicatedthat field resistance to potato cyst nematode was quan-titative with predominantly additive gene effects (DeMaine, 1978). Artzen & Dellaert (1987) also suggestedthat resistance to this nematode was under polygeniccontrol and could be durable.

A breeding strategy, using haploids (extracted fromresistant parents) producing 2n gametes has been sug-gested by De Maine et al. (1986) to transfer resistanceto potato cyst nematode across ploidy levels in potato.Dellaert (1987) advocated a slightly different strategyto accumulate resistance genes. This author indicatedthat haploids from the cultivated tetraploid (S. tubero-sum group Tuberosum) or diploid (e.g. S. tuberosumgroup Phureja) should be crossed with resistant tuber-bearing diploid Solanums (Chavez et al., 1988; Del-laert & Hoekstra, 1987) to produce resistant progenies.

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Table 1. Parental materials of families evaluated for potato cyst nematode resistance

Clone Genetic background Ploidy Reaction to

for resistance level P4A P5A

381348.7 Solanum sparsipilum diploid Resistant (R) R

84.28.58 S. vernei diploid R R

84.122.13 S. tuberosum gp. Andigena diploid Moderately (M) R Susceptible (S)

84.122.45 S. tuberosum gp. Andigena diploid MR R

H8.5 – diploid S S

14XY.7 – tetraploid S S

275186.13 S. tuberosum gp. Andigena tetraploid R MR

280240.11 S. tuberosum gp. Andigena, tetraploid Highly (H) R R

S. vernei?

281414.6 S. tuberosum gp. Andigena, tetraploid Highly (H) R HR

S. vernei?

CUP-199 – tetraploid S S

G-82138.2 S. tuberosum gp. Andigena, tetraploid Highly (H) R R

S. vernei?

P-3 – tetraploid S S

Combining resistance genes with incomplete domi-nance from wild species with additive resistance genesfrom the cultivated gene pool could result in a highlevel of resistance (Dellaert et al., 1988). Subsequentgermplasm enhancement would be done at the diploidlevel prior to transfer of resistance to the tetraploid genepool. This transfer would be done by 4x-2x crosses inwhich 2n gametes occur due to first division restitu-tion (FDR). This approach would minimize the lossof resistance factors. Furthermore, the proper manage-ment of minor and major resistance genes will increasethe durability of the resistance.

This research, therefore, aimed to determine thefeasibility of the 4x-2x approach in potato breeding,with respect to transfer of resistance to potato cystnematode from the wild diploid species S. sparsipilumand S. vernei to the tetraploid potato gene pool usingFDR 2n pollen.

Materials and methods

Seven tetraploid clones with different levels of resis-tance to potato cyst nematode isolates P4A fromHuancayo and P5A from Otuzco (Scurrah & Fran-co, 1987) were selected as female parents for thisstudy. They were crossed to three diploid clones withFDR 2n pollen (Watanabe et al., 1994) and threetetraploids (Table 1). A total of 35 tetraploid fami-lies was obtained. Triploid hybrids from 4x-2x crosses

are very rare due to a strong triploid block in potato(Johnston & Hanneman, 1995; Ortiz & Ehlendfeldt,1992). Also, five diploid clones, three of which wereused as parents in 4x-2x crosses, were intermated toproduce six diploid families.

Plant materials were grown in the greenhouse ofthe International Potato Center (CIP) Highland Stationin Huancayo (Peru) during 1987/1988. Forty seedlingswere grown in a tray and inoculated with isolates P4Aor P5A of G. pallida at the rate of 50 eggs per gram ofsoil. The seedling mass screening procedure of Fran-co & Gonzalez (1986) was then used to determine cystnematode resistance. The number of immature femalesin the tray was evaluated on 10 week old seedlingssince the fertility of the female influences the penetra-tion, establishment and development of virulent potatocyst nematode populations (Dellaert, 1987). A seedlingfamily with less than 56 immature females, counted in16 random open areas in the tray or around its bor-ders, was considered as resistant. This was determinedbecause of the significant correlation between the num-ber of immature females and the rate of multiplicationof potato cyst nematode (Franco & Gonzalez, 1986).Number of seedlings per tray, seedling height, rootdevelopment and number of tubers were recorded foreach family.

Breeding values for each parent were determinedby progeny testing. Phenotypic correlations were cal-culated between the number of immature females inthe seedlings and number of seedlings, seedling height,

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Table 2. Number of immature females per tray in tetraploid and diploid families evaluated for two races of potato cystnematode, using the greenhouse’s seedling mass test

Family P4A Rating1 P5A Rating

4x-2x

14XY.7 � 84.28.58 0 Resistant (R) 395 Susceptible (S)

275186.12 � 84.28.58 0 R 106 S

281414.6 � 84.28.58 80 S 0 R

CUP-199 � 84.28.58 140 S 766 S

G82138.2 � 84.28.58 1 R 0 R

P-3 � 84.28.58 0 R 1 R

14XY.7 � 84.122.45 202 S 0 R

281414.6 � 84.122.45 0? S2 515 S

CUP-199 � 84.122.45 0 R 0 R

G82.138.2 � 84.122.45 0 R

P-3 � 84.122.45 322 S 0 R

14XY.7 � 381348.7 1 R 10 R

CUP-199 � 381348.7 0 R 0 R

275186.13 � 381348.7 0 R 0 R

280240.11 � 281348.7 30 R 78 S

G82138.2 � 381348.7 1 R 0 R

P-3 � 381348.7 0 R 0 R

4x-4x

275186.13 � 14XY.7 0 R 8 R

280240.11 � 14XY.7 0 R 0 R

281414.6 � 14XY.7 0 R 0 R

CUP-199 � 14XY.7 182 S 171 S

G82138.2 � 14XY.7 0 R 1 R

P-3 � 14XY.7 68 S 0 ?3

14XY.7 � G82138.2 152 S 0 R

275186.13 � G82138.2 188 S 0 R

280240.11 � G82138.12 0 R 0 R

281414.6 � G82138.2 0 R 133 S

CUP-199 � G82138.2 0 R 0 R

14XY.7 selfed 163 S

2x-2x

381348.7 � 84.122.45 3 R 0 R

381348.7 � 84.122.13 320 S 0 R

84.28.58 � 381348.7 0 R 0 R

84.28.58 � 84.122.13 320 S 0 R

84.122.13 � 381348.7 341 S 0 R

H8.5 � 381348.7 190 S 0 R

Susceptible control to P5A 406 S

1 Resistance and susceptibility were measured by the number of Globodera pallida females in seedling mass testing.2 Many immature females in tray borders.3 Poor root development which did not allow to score for potato cyst nematode resistance, using seedling mass testing.

root development and number of tubers. All statisticalanalysis were performed with MSTAT-C (Anonymous,1989).

Results

Significantly different resistance reactions wereobserved between families for both pathotypes. More

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Table 3. Potato cyst nematode resistance at the tetraploid level, as measured by seedling masstesting, according to type of mating

Type of mating Percentage of resistant progenies to

P4A P5A P4A + P5A

Resistant (R) tetraploid � R diploid 75 57 43

Susceptible (S) tetraploid � R diploid 67 77 55

R tetraploid � R tetraploid 67 67 33

S tetraploid � R tetraploid (or reciprocal) 100 87 87

S tetraploid � S tetraploid 0 0 0

specifically, only a few families displayed resistance toboth P5A and P4A isolates (Table 2). Families derivedfrom crosses between susceptible parents were sus-ceptible to both pathotypes. Two families (281414.6�84.122.45 and CUP-199� 14XY.7) were rated as sus-ceptible to the isolate P4A because many immaturefemales were found on the borders of the tray.The reac-tion to the isolate P5A in P-3� 14XY.7 was not deter-mined due to poor root development of the seedlings.

All diploid families were resistant to isolate P5Awhile only 33% of them were resistant to isolate P4A.Resistance to P4A was observed when both parentswere also resistant to this pathotype, e.g. 381348.7 �84.122.45 or 84.28.58� 381348.7.

Resistance was successfully transferred from thediploid to the tetraploid level using FDR 2n pollenin 4x-2x crosses (Table 3). Interestingly, resistant 4x-2x families were equally in number or more frequentthan resistant 4x-4x families. It should be noted thatresistant clones were developed after several cyclesof selection. Also, the highest frequencies of resis-tant progeny were observed in families derived fromcrossing a susceptible tetraploid female with a resistanttetraploid male.

Based on breeding values (Table 4), the bestmale parent for potato cyst nematode resistance was381348.7. This is a FDR diploid clone with potato cystnematode resistance genes from S. sparsipilum. Theother two FDR clones with resistance alleles from S.vernei (84.28.58) and S. tuberosum group Andigena(84.122.45) have similar breeding values as the resis-tant tetraploid male parents with similar sources ofresistance. The best tetraploid female parent for pota-to cyst nematode resistance was G82138.2. However,some susceptible tetraploid female parents had a high-er or equal percentage of resistant families than theresistant tetraploid female parent 281414.6.

Table 4. Breeding value of tetraploid and diploid parent for potatocyst nematode resistance, as measured by percentage of resistantfamilies in seedling mass testing in 4x-2x and 4x-4x crosses

Parent Percentage of resistant

progenies to

P4A P5A P4A + P5A

Resistant diploid male parents

381348.7 100 83 83

84.28.58 67 50 33

84.122.45 40 75 0

Resistant tetraploid male parents

280240.11 83 60 40

G82138.2 60 80 40

Susceptible tetraploid male parent1

14XY.7 57 67 67

Resistant tetraploid female parents

275186.13 80 60 40

280240.11 100 67 67

281414.6 40 60 20

G82138.2 100 100 100

Susceptible tetraploid female parents

14XY.7 50 80 40

CUP-199 67 60 40

P-3 60 80 40

1 Resistant families were obtained only in crosses with resistanttetraploid females or with either diploid or tetraploid resistant males.

Discussion

The seedling mass screening technique was general-ly efficient in detecting different levels of resistanceamong the families evaluated. However, poor rootdevelopment prevented sometimes detection of sus-ceptible reactions that were based on immature femalecount. This was revealed by the phenotypic correla-tions between cyst nematode resistance and some mor-phological traits (Table 5).

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Table 5. Phenotypic correlations between number of immatureGlobodera pallida females in the seedling mass testing andother characteristics

Number of immature

females per tray

P4A P5A

Number of seedlings per tray 0.28 NS 0.32 *

Seedling height - 0.44 ** - 0.01 NS

Root development 0.36 * 0.40 **

Number of tubers 0.20 NS 0.27 NS

NS, * and ** indicate non-significance and significance at the5% and 1%, respectively.

The differences in breeding value suggested that theparents were segregating for resistance genes. Also,these differences between breeding values of resis-tant and susceptible clones indicated that progeny test-ing should be considered for parental selection. Thisconfirms previous reports on the contribution of non-resistant parents to resistance (Balandras et al., 1987;Dale & Phillips, 1985). Phillips et al. (1979b) foundthat specific combining ability effects were similar orgreater than general combining ability effects in cross-es among resistant parents, while the general com-bining ability effects predominate in crosses betweenresistant and susceptible parents. This breeding valueof susceptible parents could result from minor resis-tance genes which could be expressed in specific com-binations, thereby increasing the level of potato cystnematode resistance (Artzen & Dellaert, 1987). There-fore, specific combining ability for potato cyst nema-tode resistance should be determined in order to capi-talize on non-allelic gene interaction (Balandras et al.,1987; De Maine et al., 1986; Dellaert et al., 1988;Gonzalez, 1981).

The breeding value of the FDR diploid clones wasequal to, or higher than, those of resistant tetraploidclones selected after several cycles. This result indi-cates that the 4x-2x approach is appropriate for resis-tance transfer from wild or cultivated diploid tuber-bearing Solanums to the cultivated tetraploid genepool.

Acknowledgements

We thank Dr. Maria Scurrah (CIP) for providingtetraploid potato cyst nematode resistant parents andDr. Abdou Tenkouano (IITA) for his critical review.

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