Variation among Accessions of Pisum fulvum for Resistance to Pea Weevil

S. L. Clement,* D. C. Hardie, and L. R. Elberson

ABSTRACT tents reduces yield, while feeding scars and holes on The pea weevil, Bruchus pisorum (L.) (Coleoptera: Bruchidae), is testa reduce the quality and marketability of pea seed. one of the most intractable pest problems of cultivated pea, Pisum In addition, weevil-damaged seed has lower germination sativum L. The availability of resistant cultivars would give growers rates and is prone to structural weakening during har- more pest management options. Searches for plant resistance to pea vest (Brindley and Hinman, 1937; Baker, 1990). High weevil were expanded to the Pisum secondary gene pool (P. fulvum levels of weevil-infested seed have been reported in Sm.) because seed resistance had not been located in P. sativum and Australia (10.6 to 71.5%; Horne and Bailey, 1991), subspecies. The objectives of this study were to determine the extent Spain (12.2 to 25.7%; Marzo et al., 1997), and the USA of pod and seed resistance to pea weevil in P. fulvum, and to use the (up to 64%; Pesho et al., 1977; Bragg and Burns, 2000). life table format to characterize weevil stage-specific mortality and survivorship on different P. fulvum accessions. Mortality of first instar Worldwide, pea producers rely mainly on contact in- larvae on pods, mortality of all weevil stages within seed, adult emer- secticides to control adults in pea fields before females gence from seed, and seed damage levels were quantified. In two lay eggs on pods (Horne and Bailey, 1991; O’Keeffe et greenhouse trials, more larvae died (14 to 50% averages) on pods of al., 1992; Clement et al., 2000). However, timing chemi- P. fulvum accessions than on pods of ‘Alaska 81’ (6% average), and cal applications to coincide with female egg laying is mortality of first instar larvae entering seed of P. fulvum accessions difficult. More than one application may be required if averaged 83.7%. Seed damage ratings (1 ϭ feeding scar on seed testa, weevil invasions continue for 2 to 4 wk in a pea field 0-1% cotyledon tissue eaten, dead first instar larva; 5 ϭ extensive (Michael et al., 1990). The development and use of culti- Ͻ damage, live adult) averaged 3.0 for 26 P. fulvum accessions, com- vars with pod and seed resistance to B. pisorum would pared with mean ratings of 4.9 for Alaska 81. Using weevil mortality reduce control costs and provide an environmentally and survivorship values in life tables and adult emergence rates, entries were classified as susceptible (two controls and five accessions), mod- safer option than contact insecticides for adult weevil erately resistant (14 accessions), and resistant (12 accessions). Antibio- control. sis resistance was based on the death of weevil larvae on pods and Some P. sativum lines with the Np gene respond to seed testa and cotyledon tissues. The results identify sources of natural the presence of pea weevil eggs on pods by forming weevil resistance in the Pisum genome (26 moderately resistant and callus (neoplastic pod trait) that reduces larval entry resistant accessions of P. fulvum) to endow pea cultivars with pod into the pod (Hardie, 1990; Berdnikov et al., 1992; Doss and/or seed resistance to B. pisorum. et al., 2000). In a field trial, this pod-based resistance was responsible for a lower rate of weevil infested seed (62.2%) in Np plants compared with that in a susceptible nsect pests are a major problem in the worldwide line (85.4%) (Doss et al., 2000). In addition, plant bio- I production of field pea, with the pea weevil being technology has the potential to protect peas from B. one of the most destructive pests of this grain legume pisorum damage, as evidenced by the development of (Clement et al., 2000). Adult weevils leave winter hiber- transgenic P. sativum for resistance to Callosobruchus nation sites and invade pea fields in the spring to feed weevils (Shade et al., 1994) and B. pisorum (Schroeder on pea pollen and other parts of the pea flower (Brin- et al., 1995; Morton et al., 2000). This resistance is based dley, 1933; Clement, 1992). Females must feed on pollen on the insecticidal activity of the ␣-amylase inhibitor in before they can lay eggs and pea pollen is most effective seeds of bean, Phaseolus vulgaris L., which blocks the in promoting oogenesis (Pesho and Van Houten, 1982). action of the starch-digesting enzyme ␣-amylase and Eggs are laid on the surface of immature pea pods and thus prevents weevil larvae from digesting starches in hatch to produce first instar (neonate) larvae that chew seed. More research is required before weevil-resistant through the pod wall from the underside of eggs. Once transgenic peas can be released to growers (Morton et a neonate larva bores through a pod wall and reaches al., 2000). Moreover, any concerns about the biosafety the inside of a pod, it soon locates and starts to feed on of genetically engineered peas, as voiced by consumer a developing seed (Brindley, 1934). There are four larval and environmental groups for transgenic crops (Stewart instars, with second, third, and fourth instars consuming et al., 2000), must be addressed by researchers and pro- a large part of the cotyledon contents of seed (Brindley, ducer groups before transgenic peas are commercially 1933; Smith, 1990). Before pupating, a larva chews a deployed for weevil protection. circular window in the seed testa that is pushed open An alternative to genetically engineered crops is the by the emerging adult. Larval consumption of seed con- development and deployment of crop cultivars with nat- ural insect resistance from primary and secondary gene S.L. Clement and L.R. Elberson, USDA–ARS Plant Germplasm pools. In the absence of seed resistance to pea weevil Introduction and Testing Research Unit, Washington State Univ., in the Pisum primary gene pool (P. sativum and all sub- Pullman, WA 99164-6402; D.C. Hardie, Entomology Section, Dep. of Agriculture Western Australia, 3 Baron-Hay Court, South Perth, species) (Hardie, 1990; Clement et al., 1994), searches WA, 6151 Australia. Received 3 Aug. 2001. *Corresponding author for resistance were expanded to the secondary gene ([email protected]).

Published in Crop Sci. 42:2167–2173 (2002). Abbreviations: GRIN, Genetic Resources Information Network. 2167 2168 CROP SCIENCE, VOL. 42, NOVEMBER–DECEMBER 2002 pool in the late 1980s, resulting in the discovery of seed to 10 d, two or three flat or swollen pea pods from greenhouse- resistance in P. fulvum during field evaluations (Hardie grown P. sativum were placed each day in the Pullman (Alaska et al., 1995; 1999). This wild species from the eastern 81) and South Perth (‘Pennant’) cages as oviposition sub- Mediterranean and Near East areas, where pea domesti- strates. In both environments, mated females began to lay cation took place (Zohary, 1973) and where B. pisorum fertile eggs on excised pea pods after feeding on pollen for 7 to 14 d. Cages were held at 25 Ϯ 2ЊC and a 12- to 14-h is native (Clement et al., 1999), is the only species in photophase during feeding, reproduction, and egg laying on the secondary gene pool (Muehlbauer et al., 1994). Be- pea pods. cause P. fulvum is cross-compatible with P. sativum when it is used as the pollen parent (Muehlbauer et al., 1994), conventional plant breeding could potentially Experimental Protocol transfer P. fulvum resistance traits to domesticated pea. In Pullman, seeds of wild P. fulvum accessions were germi- Further consideration of P. fulvum as a potential source nated following methods described by Kaiser et al. (1997) for of weevil resistance genes for cultivar development re- wild Cicer seed. Seeds were scarified by scoring testa with quires detailed knowledge of the level and extent of seed sandpaper before they were placed in labeled cheese cloth resistance and identification of pod-based resistance in bags in 1000-mL beakers filled with distilled water that was aerated with laboratory-supplied air. Every 2 d, the water in this species. each beaker was changed and any germinated seeds were Data from life tables (Poole, 1974, p. 11–15; South- removed for planting. P. sativum ‘Alaska 81’ seeds were germi- wood, 1978, p. 366–369) may provide insight into the nated in a similar fashion except they were not scarified. genetics of plant resistance to insects if the resistance Newly germinated seeds were planted individually in 15- through high insect mortality is expressed by different cm pots containing a commercial soil mix and grown in a plant parts. Indeed, the expression of resistance by dif- greenhouse between March and July 1995 and 1996, without ferent plant parts (i.e., a pod factor and seed antibiosis) any fertilizer. No pesticides were applied. The 1995 and 1996 can be under the control of different genes (Rusoke trials evaluated 22 and 12 P. fulvum accessions, respectively. and Fatunla, 1987; Kennedy and Barbour, 1992). The However, a total of 31 accessions were evaluated because objectives of this research were (i) to determine the three accessions were included in both trials. A pea weevil- susceptible cultivar (Alaska 81) of P. sativum (Clement et al., extent of pod and seed resistance to pea weevil in P. 1996; Hardie et al., 1999) was included in each trial. Entries fulvum accessions, and (ii) to use the life table format (PI accessions and susceptible control) were arranged on two to characterize weevil stage-specific mortality and survi- greenhouse benches in a completely randomized design with vorship on P. fulvum accessions. five replications (potted plants). Both greenhouse trials were conducted under prevailing natural light/dark cycles and 15 to 21ЊC nighttime and 23 to 28ЊC daytime temperatures. MATERIALS AND METHODS Mature weevil eggs (dark brown head visible through Plants and Insects chorion) were transferred from oviposition cages with a moist- ened fine-tipped brush to surfaces of pods in the late flat and Research was conducted in the USA (Pullman, WA) and early swollen pod stages (Meichenheimer and Muehlbauer, Australia (South Perth, WA). Seed of P. fulvum accessions 1982) on potted test plants in Pullman and South Perth trials for a South Perth trial was acquired by D.C. Hardie in the using procedures in Hardie and Clement (2001). Eggs were late 1980s from several seed repositories (Hardie et al., 1995). oriented ventrally so hatching neonate larvae could burrow Seed of 29 P. fulvum accessions were acquired from several directly into the pod wall. sources (D.C. Hardie, Agriculture Western Australia, South In Pullman, each pod received two eggs placed ≈15 mm Perth, Australia; J.G. Waines, University of California, River- apart for a total of 10 eggs per plant and 50 eggs per entry (2 side, CA, USA; John Innes Institute, Norwich, UK; Nordic eggs per pod ϫ 5 pods per plant ϫ 5 plants per entry). Infested Gene Bank, Alnarp, Sweden) for Pullman trials. Before this pods were observed daily and the number of hatching eggs study, the Pisum germplasm collection at the USDA-ARS was recorded. Pods were harvested at maturity and seeds were Western Regional Plant Introduction Station, Pullman, WA, placed in labeled petri dishes (100 ϫ 15 mm) on a laboratory USA contained viable seed of only two accessions of P. fulvum bench (≈25ЊC) and observed for adult emergence every 3 to (PI accessions 343955 and 531199). The Genetic Resources 4 d. Newly emerged adults were weighed and sexed. After Information Network (GRIN) (http://www.ars-grin.gov/npgs/) 90 d, seed with feeding scars, circular windows on testa, and lists the original collection sites for only 13 of the accessions (10 with adult exit holes were split and scored for damage on a from Israel, two from Turkey, one from Jordan) in this study. scale of 1 to 5, where 1 ϭ feeding scar or puncture on testa, Field-infested pea seeds were harvested at Farmington, 0 to 1% cotyledon tissue eaten or damaged, dead first instar; WA, USA (46Њ43Ј N, 117Њ9Ј W) and York, WA, Australia 2 ϭ 2 to 5% cotyledon tissue eaten or damaged, dead first (31Њ52Ј S, 117Њ48Ј E) and stored at 4ЊC (Pullman) or 10ЊC instar; 3 ϭϾ5% cotyledon tissue eaten, dead second to fourth (South Perth), for 7 to 12 mo before greenhouse trials were instar; 4 ϭ extensive damage, dead prepupa or adult; and 5 ϭ conducted at each location. Adults emerging from infested extensive damage, live adult, or adult emergence (Hardie and peas were sexed using the dimorphic character described by Clement, 2001). The larval stage was determined by head Pesho and Van Houten (1982). Five to ten pairs were placed capsule width (Brindley, 1934). in cages and allowed to feed on pollen, which was in the form In South Perth, scarification involved scoring seed testa of pellets from commercial honey bee hives (Healthy Life with a pair of pliers, followed by soaking the seed in a 10% Carousel, Cannington, WA, Australia) in South Perth cages solution of sodium hypochlorite for 5 min and washing in (130 ϫ 200 ϫ 80 mm3 clear plastic). Fresh flowers of green- boiled water. After washing, seed were placed on moistened house-grown P. sativum ‘Alaska 81’ provided pollen in Pull- filter paper in petri dishes (100 ϫ 15 mm) on a laboratory man cages (125 ϫ 175 ϫ 45 mm3 clear plastic) at a rate of 10 bench (≈24ЊC) for germination. Seed of Pennant, a weevil- to 15 flowers every 2 to 3 d for 10 to 14 d per cage. After 7 susceptible P. sativum (Hardie et al., 1999), was not scarified CLEMENT ET AL.: VARIATION IN PISUM FULVUM FOR RESISTANCE TO PEA WEEVIL 2169 Table 1. Life table values for Bruchus pisorum from starting cohorts of 48 or 50 neonate larvae on three PI accessions of Pisum fulvum at Pullman, WA, 1995 and 1996. PI 560065 (S)‡ PI 595942 (MR)‡ PI 595940 (R)‡ x† lx§ dx¶ 100qx# 100Sx†† lx§ dx¶ 100qx# 100Sx†† lx§ dx¶ 100qx# 100Sx†† %%% Before reaching seed First instar 48 14 29.2 70.8 50 17 34.0 66.0 50 20 40.0 60.0 Within seed First instar 34 5 14.7 85.3 33 29 87.9 12.1 30 30 100.0 0 Second to fourth instar 29 0 0 100.0 4 1 25.0 75.0 – – – – Prepupa to adult 29 0 0 100.0 3 0 0 100.0 – – – – Total within seed 34 5 14.7 85.3 33 30 90.9 9.1 30 30 100.0 0 Total (first instar to adult) 48 19 39.6 60.4 50 47 94.0 6.0 50 50 100.0 0 † Life Stage ‡ S, susceptible; MR, moderately resistant; R, Resistant. § Number entering a stage. ¶ Number dying in a stage. # Percentage mortality in a stage. †† Percentage survival in a stage.

but was germinated on moistened filter paper. Five germinated weevil on each plant (replicate) of each entry were: lx, the seeds per entry were planted in a 25-cm pot containing washed number of individuals entering stage x; dx, the number of river sand and top dressed with a slow release granular fertil- individuals dying in stage x; 100qx, the percentage mortality 1 izer (Osmocote, The Scotts Company, Marysville, OH) .No in stage x; and 100Sx, the percentage surviving stage x. Instead pesticides were applied. One trial was conducted in 1993 with of presenting these values in complete life tables for all pea 23 P. fulvum accessions (also in Pullman trials) and the Pen- weevil-entry interactions, tables are presented for B. pisorum nant control. The 24 pots were randomly arranged on a green- on three PI accessions of P. fulvum in which values were house bench. The greenhouse trial was conducted under the generated from starting entry-cohorts of 48 or 50 neonate prevailing natural light/dark cycle and temperatures ranged larvae (Table 1). This approach was used to conserve space from 10 to 32ЊC. while illustrating the structure of three representative life ta- Each pod on South Perth plants received eight mature eggs, bles. To simplify the presentation of all life table results and but a variable number of pods per entry were infested (5 to to facilitate assessment of pod and seed resistance among 28 pods per entry from 1 to 6 pods per plant). Pods were entries, entry means of three parameters were statistically harvested at maturity and stored at room temperature in la- analyzed: (i) mortality [100qx] of first instar larvae before beled seed envelopes. After 60 d, when adult weevils began reaching seed; (ii) mortality [100qx] of all weevil stages within emerging from Pennant seeds, all seeds were inspected and seed; and (iii) survival [100Sx] to the adult stage (Tables 2 assessed for adult emergence. A seed was classified as infested and 3). if a live pupa or adult was found in the seed or an adult exit The Fmax test, a sample variance ratio procedure (Sokal and hole was found. Rohlf, 1981, p. 403), was used to ensure that the assumption of homogeneity of variances was met before data were analyzed. Data Analysis Nonsignificant heterogeneity of variances justified analysis of variance on all original data sets (seed weights, life table A few eggs on pods did not hatch in the Pullman trials, parameters [100qx, 100Sx], seed damage ratings, and adult wee- leaving a starting cohort of 7 to 10 on plants (range of 46 to vil weights). Entry means for life table parameters and seed 50 eggs per entry). The absence of feeding and penetration damage ratings were computed from replicate (n ϭ 5 plants) marks on testa was used to calculate the number of larvae means, whereas mean seed weights were based on 20 undam- dying en route to the outer surfaces of developing seeds inside aged seeds per entry and mean adult weevil weights were green pods. Larvae on each infested pod were categorized as calculated from all emerging adults per entry (Tables 2 and 0, 1, or 2 dead if feeding damage was recorded on 2, 1, or 0 3). General linear model procedures were used for unbalanced seeds, respectively. Within-seed mortality of B. pisorum was data and entry means were compared by Fisher’s least signifi- compartmentalized, as follows: first instars consuming 0 to cant difference (LSD0.05). Pearson’s correlation coefficients 5% of the cotyledon tissue (Damage Ratings 1 and 2); second were calculated to examine relationships between mean seed Ͼ to fourth instars consuming 5% of the cotyledon tissue weights and mean within-seed mortality rates and between (Damage Rating 3); and development to prepupa or adult mean seed weights and mean seed damage ratings of P. fulvum with extensive damage to seed contents (Damage Ratings 4 accessions (SAS Institute, 1987). Since the entries differed and 5). among years (except for four entries), data from each year The life table format (Poole, 1974, p. 11–15; Southwood, were analyzed separately. Results from the South Perth trial 1978, p. 366–369) was previously used to record numerical were expressed as percentage adult weevil emergence from changes in arthropod populations on cotton, Gossypium hirsu- seed of all egg-infested pods per entry. tum L., (Trichilo and Leigh, 1985) and cowpea, Vigna unguicu- lata (L.) Walp., cultivars (Messina, 1984) and was used to quantify pea weevil mortality and survival on P. fulvum acces- RESULTS AND DISCUSSION sions and a susceptible pea cultivar (Alaska 81) in the Pullman trials. Parameters estimated in the life table analysis for pea There was significant (P Ͻ 0.05) variation among the entries in each of the Pullman trials for larval mortality 1 Mention of a trademark or proprietary product does not constitute before reaching seed (100qx ) and within-seed (100qx ), a guarantee or warranty by the USDA and does not imply its approval seed damage rating, adult emergence (100Sx ), and over other suitable products. weight of emerging adults. As expected, Alaska 81 was 2170 CROP SCIENCE, VOL. 42, NOVEMBER–DECEMBER 2002

Table 2. Mortality and survivorship of Bruchus pisorum on Pisum sativum ‘Alaska 81’ and accessions of P. fulvum, and levels of weevil- inflicted seed damage from a greenhouse study at Pullman, WA, in 1995. Means

Larval mortality (100qx )§ Adult Seed Before reaching Within Seed damage Emergence Entry Entry† wt.‡ seed seed rating¶ (100Sx )§ Wt.# classification†† mg % % mg ‘Alaska 81’ 240.0a‡‡ 6.0a 8.9a 4.8a 86.0a 12.8a S PI 560065 77.4de 29.1bcdef 14.4a 4.5ab 60.4b 8.7b S PI 595941 78.8d 22.7abcd 37.1b 4.1bc 48.4bc 7.0cde S PI 595938 77.2de 20.9abcd 46.4bc 3.6cd 42.2bc 8.5bc S PI 595953 73.8de 18.4abc 53.8c 3.3d 38.7c 8.0bcd S PI 595943 69.1ef 22.7abcd 59.5c 3.3d 32.4c 6.4def S PI 595939 90.4b 26.0bcde 88.4d 2.4e 8.0d 8.0bcd MR PI 595952 81.7cd 24.0abcde 88.5d 2.3ef 8.0d 6.3ef MR PI 560064 60.2g 18.7abc 92.7d 2.3ef 6.0§§ 4.8f MR PI 595942 55.9gh 34.0def 90.0d 2.2efgh 6.0§§ 5.5ef MR PI 595951 58.7g 36.6cdef 92.5d 2.2ef 4.0§§ 3.9, 7.9¶¶ MR PI 595948 94.9b 14.7ab 95.0d 2.2efg 4.0§§ 4.8, 5.0¶¶ MR PI 343955 87.9bc 36.0cdef 93.1d 2.1efgh 4.0§§ 8.9, 10.7¶¶ MR PI 595937 73.7de 46.5f 97.5d 2.1efgh 2.0§§ 6.8¶¶ MR PI 560061 69.5ef 32.4bcdef 97.1d 2.0efgh 2.0§§ 5.9¶¶ MR PI 595932 60.5g 29.5bcdef 96.0d 1.8fgh 2.0§§ 3.0¶¶ MR PI 595936 76.0de 28.0bcdef 100.0d 1.9efgh 0 – MR## NGB 102148 – 32.0bcdef 100.0d 2.0efgh 0 – R PI 595950 49.4h 35.1cdef 100.0d 2.0efgh 0 – R PI 560066 55.6gh 38.7def 100.0d 2.0efgh 0 – R PI 560062 80.9cd 20.0abcd 100.0d 1.9efgh 0 – R PI 595933 59.7g 14.0ab 100.0d 1.7gh 0 – R PI 560063 63.9fg 43.1ef 100.0d 1.6h 0 – R LSD 8.59 19.17 14.21 0.54 18.26 1.67 † Cultivar, PI accession, and Nordic Gene Bank (NGB) accession. ‡ n ϭ 20 undamaged seeds per entry. § Means computed from five replicate (plant) means. ¶ Means computed from five replicate (plant) means where 1 ϭ little or no damage and 5 ϭ extensive damage. # Means computed from weights of all emerging adults from five replicates (plants). †† S, susceptible; MR, moderately resistant; R, Resistant. ‡‡ Means within a column followed by the same letter are not different at P Ͻ 0.05 on the basis of LSD. §§ Too few adults emerged and therefore not statistically analyzed. ¶¶ Weights of one or two adults. ## Moderately resistant because a few adults emerged in the Australia trial (see text). highly susceptible to B. pisorum (Tables 2 and 3). Al- sustained 88.4 to 100% within-seed mortality on 17 of though there was statistical concordance (P Ͼ 0.05) 22 P. fulvum accessions in 1995 and 85.3 to 100% on between first instar mortality rates on pods of Alaska all accessions in 1996. This high level of seed resistance 81 and pods of nine P. fulvum accessions in 1995 and in P. fulvum is also revealed by the seed damage ratings pods of two accessions in 1996, the overall results reveal in both trials, which for all accessions except PI 560065 the presence of pod-based resistance in the Pisum sec- (1995 trial) were significantly lower (P Ͻ 0.05) than the ondary gene pool. The percentage of first instar larvae average ratings of 4.8 (1995) and 5.0 (1996) for Alaska that died before reaching seeds of P. fulvum accessions 81. Seed damage ratings averaged less than 3.0 for 17 averaged 14 to 43% and 22 to 50% in 1995 and 1996, P. fulvum accessions in 1995 and for all accessions in respectively, compared with an average larval mortality 1996 (Tables 2 and 3). rate of 6% on pods of Alaska 81 in both Pullman trials Seed of P. fulvum accessions in both Pullman trials (Tables 2 and 3). We observed larvae wandering about weighed significantly less (P Ͻ 0.05) than seed of Alaska on the pod surfaces of P. fulvum before attempting to 81 (Tables 2 and 3). However, resistance in small seeded bore into the pod wall, which suggested that pods of P. fulvum accessions did not appear to be the result of this wild species may be less hospitable to neonate larvae limited food for larval development, as evidenced by than are pods of P. sativum. Ninety-nine out of 100 nonsignificant (1995: r ϭ 0.29, P ϭ 0.2105; 1996: r ϭ neonates bored directly into the pod walls of Alaska 81 0.40, P ϭ 0.1994) correlation coefficients between P. from the underside of eggs in an earlier study (Clement, fulvum seed weights and damage ratings and nonsignifi- 1992, unpublished data). This normal neonate behavior cant (1995: r ϭϪ0.25, P ϭ 0.2692) or weakly significant on P. sativum pods may protect newly hatched larvae (1996: r ϭ 0.57, P ϭ 0.0507) correlation coefficients from predators, parasitoids, and dessication, as well as between seed weights and within-seed mortality rates. shield them from contact insecticides in agricultural Moreover, 81.8% (n ϭ 768) and 87.5% (n ϭ 384) of settings. the first instar larvae that reached seed of P. fulvum On the basis of within-seed mortality rates of larvae, entries in 1995 and 1996, respectively, died before con- seeds of all P. fulvum accessions except PI 560065 (1995 suming Ͼ5% of the cotyledon tissue of seed and before trial) were significantly more (P Ͻ 0.05) resistant to developing into second instars. This is further evidence larval feeding than seeds of Alaska 81. Pea weevil larvae that starvation was not responsible for high mortality CLEMENT ET AL.: VARIATION IN PISUM FULVUM FOR RESISTANCE TO PEA WEEVIL 2171 Table 3. Mortality and survivorship of Bruchus pisorum on Pisum sativum ‘Alaska 81’ and accessions of P. fulvum, and levels of weevil- inflicted seed damage from a greenhouse study at Pullman, WA, in 1996. Means

Larval mortality (100qx )§ Adult Seed Before reaching Within Seed damage Emergence Entry Entry† wt.‡ seed seed rating¶ (100Sx )§ Wt.# classification†† mg % % mg ‘Alaska 81’ 240.4a‡‡ 6.0a 0a 5.0a 94.0a 12.3a S PI 595945 87.9b 40.0bcde 85.3b 2.9b 8.0b 8.1b MR PI 595947 83.8bc 48.0de 95.0c 1.9cde 2.0§§ 6.5¶¶ MR PI 560061 69.5de 26.0abc 97.1c 2.2c 2.0§§ 8.0¶¶ MR PI 595944 70.0de 22.0ab 100.0c 1.9de 0 – MR## PI 595946 58.5fg 28.0bcd 100.0c 2.1cd 0 – R PI 595935 66.1ef 38.0bcde 100.0c 2.1cd 0 – R PI 560067 75.6cd 28.0bcd 100.0c 1.9cde 0 – R PI 531199 82.3bc 32.0cde 100.0c 1.8e 0 – R PI 595940 64.2ef 40.0bcde 100.0c 1.8e 0 – R PI 595934 52.1g 34.0bcde 100.0c 1.7e 0 – R PI 560062 80.9bc 46.0cde 100.0c 1.7e 0 – R PI 595933 59.7fg 50.0e 100.0c 1.6e 0 – R LSD 8.45 20.19 6.81 0.28 10.31 1.27 † Cultivar and PI accession. ‡ n ϭ 20 undamaged seeds per entry. § Means computed from five replicate (plant) means. ¶ Means computed from five replicate (plant) means where 1 ϭ little or no damage and 5 ϭ extensive damage. # Means computed from weights of all emerging adults from five replicates (plants) per entry. †† S, susceptible; MR, moderately resistant; R, Resistant. ‡‡ Means within a column followed by the same letter are not different at P Ͻ 0.05 on the basis of LSD. §§ Too few adults emerged and therefore not statistically analyzed. ¶¶ Weight of one adult. ## Moderately resistant because a few adults emerged in the Australia trial (see text). of first instars on P. fulvum seeds of weights that aver- 595941, PI 595943, PI 595945, PI 595947). Adult emer- aged 49.4 to 94.9 mg in 1995 and 52.1 to 87.9 mg in 1996. gence from the susceptible P. sativum control (Pennant) We speculate that insecticidal properties are responsible averaged 85%. There was good concordance between for seed resistance, although the absence of ␣-amylase the Pullman and South Perth results. A few adults inhibitor in seed of P. fulvum (Schroeder et al., 1995) emerged (2 to 8%) from seven accessions (PI 595939, precludes associating this seed protein with weevil toxic- PI 560064, PI 595942, PI 595948, PI 560061, PI 595932, ity. Schoonhoven et al. (1983) speculated that factors PI 595937) in Pullman (Table 2), but none emerged other than seed size and weight were responsible for from these entries in South Perth, and a few adults resistance in small seeded and uncultivated bean acces- emerged (4 to 15%) from three accessions (PI 595947, sions to the bean weevil, Acanthoscelides obtectus (Say), PI 595944, PI 595936) in South Perth, but none emerged and Mexican bean weevil, Zabrotes subfasciatus (Bohe- from these entries in Pullman (Tables 2 and 3). man) (Coleoptera: Bruchidae). Any variation among weevil populations (including Mean adult emergence values (100Sx ) from P. fulvum B. pisorum) for virulence to resistant genotypes could were significantly lower (P Ͻ 0.05) than those from minimize the ability of a resistant cultivar developed in Alaska 81, which had emergence values of 86% (1995) one country to confer resistance to weevil populations and 94% (1996). However, emergence rates were rela- in other countries (Dick and Credland, 1986; Shade et tively high (32.4 to 60.4%) for five P. fulvum accessions. al., 1999). However, differences in adult emergence Adults that emerged from P. fulvum weighed signifi- rates between Pullman and South Perth trials were not cantly less (P Ͻ 0.05) than those from Alaska 81 seed of a magnitude for us to conclude that significant vari- in both Pullman trials (Tables 2 and 3), prompting us ability exists among pea weevil populations in eastern to speculate that these very small adults would have Washington and Western Australia for virulence to re- reduced survivability and reproductive fitness. No large sistant P. fulvum accessions. Thus, the resistant acces- departure from a 1&:1( sex ratio was recorded for sions in this study could be used to develop pea cultivars adults emerging from Alaska 81 in 1995 and 1996 in different geographical areas with durable pea wee- (49:51%) and from all P. fulvum accessions in both vil resistance. trials (52:48%). Three distinct patterns of B. pisorum mortality and In the South Perth trial, 15 P. fulvum accessions pro- survival on the entries in the Pullman trials emerged duced no adults (PI 595939, PI 560064, PI 595942, PI from the construction of life tables, illustrated by the 595948, PI 560061, PI 595932, PI 595937, PI 595946, PI values for the weevil on PI 560065, PI 595942, and PI 595934, PI 595935, PI 595933, PI 560062, PI 560067, PI 595940 (Table 1). The PI 560065 life table characterizes 595940, PI 560063). However, adults emerged from eight pea weevil susceptibility in P. fulvum, a pattern that accessions with values averaging between 4 and 6% for was exhibited by four other accessions (PI 595941, PI three accessions (PI 595936, PI 595944, PI 343955) and 595938, PI 595953, PI 595943) in the 1995 trial. These between 15 and 47% for five accessions (PI 595938, PI susceptible accessions had within-seed larval mortality 2172 CROP SCIENCE, VOL. 42, NOVEMBER–DECEMBER 2002 rates of 14.4 to 59.5%, seed damage ratings Ͼ3.0, and tein to provide durable protection against B. pisorum adult emergence percentages of 32.4 to 60.4% (Table in one cultivar (Schroeder et al., 1995). 2). Although the remaining accessions could not be sep- arated into resistant classes on the basis of statistically ACKNOWLEDGMENTS different mean values for within-seed mortality and seed damage ratings (Tables 2 and 3), their adult emergence We thank J. Burns, F. Muehlbauer, and E. Zakarison for values yielded two distinct patterns. First, 13 P. fulvum manuscript review, H. Collie for technical assistance, and M. Evans for statistical advice. Research was supported in part accessions from the Pullman trails and two accessions by grants from the USDA-Foreign Agricultural Service-Inter- from the South Perth trial produced a few adults, albeit national Cooperation and Development-Research and Scien- only one or two small adults or emergence percentages tific Exchanges Division (AS37) in the USA and the Grains of 8% or less (exemplified by PI 595942 in Table 1), Research and Development Corporation (GRDC) in Aus- and thus were classified as moderately resistant. Second, tralia. 14 accessions shared by the Pullman and South Perth trials produced no adults and were classified as resistant REFERENCES (exemplified by PI 595940 in Table 1) (Tables 2 and 3). A wide range in first instar mortality rates on pods Baker, G.J. 1990. 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