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Eric B. Bish, Thomas A. Bewick, and Donn G. Shilling

Laboratory experiments were conducted to evaluate the relationship between leaf area, leaf number, dry weight, and mg extract of Lycopersicon hirsutum (LA 1777) leaf washes and germination or root growth of common purslane (Portulaca oleracea) seeds. Additional experiments were conducted to determine the relationship between L. hirsutum (accessions 1777 and 1625) leaf washes and germination or root growth of common purslane seeds. Activity of separated sesquiterpenes from trichomes were compared to crude leaf washes. Results from the leaf washes of the L. hirsutum accessions (1777 and 1625) indicated that there was no significant difference between hexane leaf washes, methanol leaf washes, or crude leaf extracts when common purslane was used as the assay species. The accession 1777 was greater than 800 x more inhibitory to germination and greater than 300x more inhibitory to root growth of purslane seeds than accession 1625.

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John C. Snyder

Breeding for resistance to insects and other arthropod pests in vegetables has been a difficult endeavor. Greater public awareness of health and environmental issues requires that we as horticultural scientists reexamine why breeding for resistance has been difficult. The literature clearly suggests the potential for a genetic solution, and the literature also reveals some reasons why achievement of genetic resistance to arthropod pests has not been as successful as the achievement of resistance to pathogens. The thesis of my presentation is that the complexity of plant-arthropod interactions often prevents simple genetic approaches to breeding for resistance. Data using Lycopersicon hirsutum and its interaction with spider mites will provide examples of the these complex interactions. L. hirsutum is a wild relative of L. esculentum, the common tomato, and is nearly immune to insect attack. However, there are few or no clear examples of this taxa contributing to the insect resistance of tomato. The complexity of the interaction between mites and trichomes on L. hirsutum will be highlighted as it pertains to environment and genetics of the plant, and the development of the arthropod.

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John R. Stommel and Stephen L. Sinden

Cultured leaf explants obtained from 36 accessions of the wild tomato species, Lycopersicon hirsutum Humb. and Bonpl., were evaluated for morphogenic capacity in response to three cytokinins (zeatin, BA, and kinetin) in combination with IAA. Media containing 0.1 μm IAA plus 4.6 or 9.2 μm zeatin were optimal for shoot induction. Cotyledon explants were superior to true leaf explants for obtaining shoot formation. Morphogenic responses of L. hirsutum f. typicum and L. hirsutum f. glabratum were clearly accession-dependent and ranged from exceptional with numerous shoots produced to recalcitrant with no shoots produced. The high morphogenetic capacity of leaf explants from L. hirsutum f. typicum accession 128644 was also evident in protoplast-derived calli that readily regenerated shoots. Chemical names used (E)-2-methyl-4-(1H-purin-6-ylamino)-2-buten-1-ol (zeatin), N-(phenylmethyl) -1H-purin-6-amine (BA), N- (2-furanylmethyl) -1H- purin-6-amine (kinetin), 1H- indole-3-acetic acid (IAA).

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John R. Stommel

Cultured leaf explants obtained from 36 accessions of the wild tomato Lycopersicon hirsutum were evaluated for morphogenic capacity in response to 3 cytokinins [zeatin, benzylamino purine (BA) and kinetin] in combination with indoleacetic acid (IAA). Morphogenic responses within this wild species were accession-dependent, Cotyledon tissue, in comparison to true leaf explants, were superior for callus and shoot formation. Optimal callus induction medium varied with accession, but most often contained 13.3 μM BA plus 1.7 μM IAA. Media containing 4.6 or 9.2 μM zeatin plus 0.1 μM iaa were optimal shoot induction media. Explants of L. hirsutum f. typicum accessions 126445, 127826, 128644, and 390663 and L. hirsutum f. glabratum accessions 365904, 365905, and 365906 exhibited the highest levels of shoot formation.

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John R. Stommel

Cultured leaf explants obtained from 36 accessions of the wild tomato Lycopersicon hirsutum were evaluated for morphogenic capacity in response to 3 cytokinins [zeatin, benzylamino purine (BA) and kinetin] in combination with indoleacetic acid (IAA). Morphogenic responses within this wild species were accession-dependent, Cotyledon tissue, in comparison to true leaf explants, were superior for callus and shoot formation. Optimal callus induction medium varied with accession, but most often contained 13.3 μM BA plus 1.7 μM IAA. Media containing 4.6 or 9.2 μM zeatin plus 0.1 μM iaa were optimal shoot induction media. Explants of L. hirsutum f. typicum accessions 126445, 127826, 128644, and 390663 and L. hirsutum f. glabratum accessions 365904, 365905, and 365906 exhibited the highest levels of shoot formation.

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John C. Snyder, George Antonious, and Richard Thacker

Many accessions of Lycopersicon hirsutum are highly resistant to insects. Trichomes and their secretions have been extensively indicated as factors of resistance. One mechanism of resistance mediated by secretions is repellency, a mechanism that is consistent with the observation that few insects visit plants of L. hirsutum. Trichome secretions from certain accessions of L. hirsutum f. typicum are repellent to spider mites. However, the composition of secretions from different accessions of f. typicum are chemically diverse. Sesquiterpene hydrocarbons are prevalent in secretions, but are structurally diverse. How structure may relate to repellency is of interest but difficult to address because isolation of pure sesquiterpene hydrocarbons from hydrocarbon mixtures is difficult. To begin examining relationships between structure and activity we determined how chain length of n-alkanes related to repellency of spider mites. n-Alkanes having chain lengths from 8 to 22 carbon atoms were assayed for repellency. The C16-C18 alkanes were most repellent. Smaller and larger hydrocarbons were less repellent. The EC50 for n-hexadecane was equal to that of the most repellent natural products we have isolated from trichome secretions of L. hirsutum.

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Rebecca C. Lough and R.G. Gardner

During the last century Phytophthora infestans (Mont.) de Bary, which causes the devastating disease late blight of tomato and potato, has been controlled with pesticides. Recently, the difficulty of controlling late blight has increased due to the appearance of new strains of P. infestans that are more virulent and are resistant to metalaxyl. Numerous P. infestans resistance genes exist within the Solanaceae; however, most of these are race-specific and have the potential of being overcome. To achieve durable resistance, it may be necessary to utilize multigenic resistance or gene pyramiding. The Lycopersicon hirsutum Kunth accession LA1033 is highly resistant to P. infestans. To incorporate resistance into a useful background, the L. esculentum Miller inbred line NC215E was used as a recurrent parent in backcrossing with L. hirsutum LA1033. A population of 264 BC3F1 plants derived from 11 BC2F2 families was planted at Fletcher and Waynesville, N.C., in July 1998 in a replicated field trial. BC3F2 seed were collected from a single highly resistant BC3F1 plant. The BC3F2 population was tested for resistance using a detached leaf screen. To verify growth chamber test results, BC3F3 seeds were collected from the BC3F2 individuals and were planted in a field trial at Fletcher in July 1999. The ratio of resistant to susceptible progeny fit the expected ratio for an incompletely dominant trait controlled by two loci. To identify molecular markers linked to the resistance loci, DNA was extracted from the highly resistant and susceptible BC3F2 individuals, and bulks of DNA were constructed. The resistant and susceptible bulks were screened with AFLP (amplified fragment length polymorphism) markers. Results of the AFLP study indicate marker linkage to resistance.

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T.J. Montagno, P.S. Jourdan, and S. Z. Berry

Unilateral incompatibility has limited the direction of crossing between L. esculentum and L. hirsutum; the latter can only serve as the pollen parent. In an attempt to introduce the L. hirsutum cytoplasm into L. esculentum, thirty-three somatic hybrid plants have been regenerated following four separate fusions between leaf protoplasts of L. hirsutum PI 126445 and etiolated hypocotyl protoplasts of L. esculentum (`OH7870', `OH832', and `OH8245'). A 33% PEG solution supplemented with 10% DMSO was used as the fusogen. Selection of fusion products was based on treatment of L. hirsutum protoplasts with 1 mM iodoacetic acid and non-regenerability of the L. esculentum genotypes. Hybridity was initially confirmed by intermediate morphology, including leaf shape, type of trichomes, flower shape, stigma placement, and fruit size and color. Isozyme analysis for GOT, PGM, and 6-PDH verified hybridity. Six of the hybrids produced viable seed upon selfing. At least some of the hybrids contained chloroplast DNA from L. hirsutum, indicating that the wild species cytoplasm may be present in these plants.

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T.J. Montagno, S.Z. Berry, and P.S. Jourdan

L. hirsutum has been previously reported as recalcitrant to culture and plant regeneration. We have modified tomato protoplasm culture protocols and obtained high frequencies of plant regeneration from leaf protoplasts of L. hirsutum PI 126445, LA 94, and LA 1393, as well as from 8 interspecific hybrids of PI 126445 (male parent) with L. esculentum `Floradade', `Marglobe', `Tropic', `OH7870', `OH7983', `OH832', `OH8243', and `OH8245'. Protoplasts were isolated from 3-week old low light pretreated seedlings and cultured in modified LCM containing 1 mg/L NAA 0.5 m /L BA, and 0.5 mg/L 2,4-D. Cultures were kept in the dark at 30 C, diluted every 3 days with LCM containing only 0.75 mg/L BA and gradually moved to the light. After 2-3 weeks, colonies of 1-2 mm were transferred to solid MS medium containing 0.5 mg/L BA and 0.05 mg/L NAA. Calli containing dark green bud primordia were then placed on MS with 2% sucrose and 2 mg/L zeatin riboside for shoot production.

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John C. Snyder, Richard Thacker, Jan St. Pvrek, and Jack P. Goodman

Spider mites (Tetranychus urticae Koch) readily colonize the cultivated tomato. Lycopersicon esculentum L. However, mites have extreme difficulty colonizing the wild relative of tomato, L. hirsutum Humb. and Bonpl. When mites approach leaves of L. hirsutum, they often veer away, suggesting the presence of a deterrent or repellent. Initial experiments indicated that trichome secretions on leaflets of L. hirstum deterred mites. In vitro bioassays indicated that at least four distinct compounds present in these sequiterpenoid secretions of L. hirsutum P.I. 251303 were deterrent. At least two of the compounds were soluble in dilute NaOH. Based on mass spectra and 1H and 13C NMR the structure of two base soluble compounds were established as two related bisabolane derived carboxylic acids.