Breeding for Resistance to Leafminer in Lettuce

Leafminer (Liriomyza langei Frick) is a major pest that causes considerable damage to a wide variety of vegetable crops including lettuce, and resistant cultivars remain the most economic means of insect control. Eighty-four lettuce cultivars and introduction lines were grown in an insect cage with eight replications for resistance screening. Leafminer flies were released in the cage to feed on the plants. Significant genetic variation for leafminer stings per unit leaf area was observed among genotypes tested. Resistant lines with fewer leafminer stings were found in Lactuca sativa, L. saligna, L. serriola, and L. virosa, and the resistance was confirmed in a field experiment. Crosses were made to combine leafminer resistance and superior horticultural traits in crisphead, green leaf, red leaf, romaine, and butterhead lettuces. Leaf miner resistant plants were selected in F2 progenies of such crosses, and were backcrossed to restore horticultural type. INTRODUCTION Leafminers are major insect pests of many important agricultural crops (Parrella, 1987). They attack a wide range of plants including aster, baby’s breath, bean, broccoli, cabbage, cauliflower, celery, chrysanthemum, cucumber, lettuce, muskmelon, onion, pea, potato, spinach, summer squash, Swiss chard, tomato, watermelon, and many weeds. The principal leafminer species affecting vegetables include Liriomyza brassicae, L. sativae, L. trifolii, L. huidobrensis, and L. langei. In the major lettuce production areas in central California, the predominant species was believed to be L. huidobrensis (pea leafminer) until recently (Morgan et al., 2000). Scheffer and co-workers (Scheffer et al, 2001) identified the leafminers in central California to be the morphologically cryptic species L. langei by using polymerase chain reaction (PCR) amplification of mitochondrial DNA. Leafminer adults are small, shiny, black flies with a bright yellow triangular spot on the upper thorax between the wings. Damage occurs when adult flies puncture leaves to feed on plant sap and females lay white, oval eggs within the leaf tissue. Adult “stings” appear as holes or bumps on the leaves. Adult feeding on cotyledons may stunt seedling growth. Larvae hatch from eggs and feed between upper and lower leaf surfaces. The winding, whitish tunnels or mines they create are initially narrow, but they increase in width as the larvae grow. Larvae emerge from the mines after completing three instars and pupate in cracks in the soil or on the leaf surface. Adult flies come out of pupae in about 8 to 11 days. The entire life cycle can be completed in less than three weeks in warm weather. Many generations are produced each year in California. Adult sting and larval mining of leaves reduce photosynthetic capacity, render leaves unmarketable, and provide an entrance for disease organisms (University of California, 1992; LeStrange et al., 1999). Few studies on leafminer resistance in vegetables have been reported in the literature. Larval (L. trifolii) antibiosis was found in four interspecific hybrids of Lycopersicon pennellii, L. cheesmanii, and L. hirsutum, and adult antibiosis and antixenosis for feeding was partially a result of the tomato plant’s trichome exudates (Erb et al., 1993). No resistance to leafminers (L. trifolii) has been observed in cultivated celery, but Proc. XXVI IHC – Advances in Vegetable Breeding Eds. J.D. McCreight and E.J. Ryder Acta Hort. 637, ISHS 2004 Publication supported by Can. Int. Dev. Agency (CIDA) 58 an accession from a wild species, Apium prostratum, was found to be practically immune (Trumble and Quiros, 1988). No feeding or oviposition was observed in this species. It has been used in a backcross program in an attempt to develop leafminer-resistant celery lines (Quiros, 1993). In addition, an accession from another wild species, A. nodiflorum, demonstrated substantial insect toxicity; few mines were observed and no larvae survived to the pupal stage (Trumble et al., 1990). In lettuce, significant differences among four romaine cultivars were found in the number of pupae produced and in the total numbers of stipples when exposed to leafminers (L. trifolii), but these differences resulted from differences in adult survival. Female leafminers survived significantly longer and produced more pupae on the cultivar Tall Guzmaine than on three other cultivars (Nuessly and Nagata, 1994; Nagata et al., 1998). The genetic variation of leafminer resistance in the lettuce germplasm including the wild species has not been fully explored. The purposes of the present study were to evaluate differences in leafminer resistance among lettuce genotypes and to incorporate the resistance into elite lettuce cultivars. MATERIALS AND METHODS The experiments were conducted at the Agricultural Research Station of the U.S. Department of Agriculture, Salinas, CA in 2001. Eighty-four genotypes from the lettuce germplasm collection maintained at the station were used in this study. They include crisphead, leaf, romaine, butterhead, stem, Latin, Batavia, and primitive forms of cultivated lettuce (Lactuca sativa), and the wild species L. serriola, L. saligna, and L. virosa from different geographic areas of the world. Three weeks after planting, 8 plants of each genotype were transplanted individually into plastic pots (10 x 10 x 10-cm) with soil. Plants were placed in field cages (2 m high by 4 m wide by 4 m deep) made of polypropylene shade cloth and were arranged in a randomized complete block with a single plant as the experimental unit and 8 replications. Lettuce leaves with leafminer mines were collected from newly harvested fields around Salinas and leafminer larvae were allowed to emerge from the leaves and pupate. Pupae were collected, weighed, and put in plastic containers to allow the adult flies to emerge. The weight of a random sample of 200 pupae was used to estimate the number of pupae collected. The sampled pupae were also reared to adults to determine percentage of viable pupae. The number of flies was estimated based on weight and emergence of sampled pupae. About 2,500 flies were released in the field cage. The leaf with the most leafminer stings on each plant was counted for the number of stings per 20-cm leaf area with the aid of an optical glass binocular magnifier (OptiVisor, Donegan Optical Co., Lenexa, Kans., USA) 7 days after the introduction of leafminer flies in the cage. The genotypes were also transplanted in a field on the station in summer. Each plot consisted of six plants, with 30 cm between plants and 35 cm between rows on double-row beds of 1-m center. The experimental design was a randomized complete block with 8 replications. The leaf with the most leafminer stings on each plant was counted for the number of stings per 20-cm leaf area when plants reached maturity. Data were analyzed statistically by using the Analysis of Variance (ANOVA) functions of the Microsoft Excel (Office 2001, Microsoft Co., Redmond, Wash., USA). Means of different lettuce genotypes were compared with t tests (Petersen, 1985). RESULTS AND DISCUSSION Significant differences among genotypes were found for the number of stings per unit leaf area, and the 11 genotypes with fewest stings are shown in Fig. 1. Two L. saligna lines (PI 509525 and PI 490999) had least leafminer stings among genotypes tested. Among the cultivated lettuces (L. sativa), PI 187238, PI 491212, Merlot, Lolla Rossa, and Salad Bowl showed fewer leafminer stings per unit leaf area than other lines. Da Ye Wo Sun, a stem lettuce from China, registered the most sting damage among the genotypes tested. These results demonstrate the existence of high levels of leafminer resistance in