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F.A. Hammerschlag

A detached-leaf bioassay was used to evaluate peach [Prunus persica (L.) Batsch] regenerants derived from zygotic embryo callus cultures of cultivars Sunhigh (susceptible to leaf spot) and Redhaven (moderately resistant to leaf spot) for resistance to Xanthomonas campestris pv. pruni [(E.F. Sm.) Dews], the causal agent of bacterial leaf spot. Regenerants obtained from calli produced on two `Sunhigh' embryos, #61 and #156, and on three `Redhaven' embryos were evaluated. Sixty-four percent of the regenerants derived from `Sunhigh' embryo #156 and 13% of the regenerants derived from `Sunhigh' embryo #61 demonstrated significantly greater spot resistance than `Sunhigh'. Regenerants with resistance greater than `Redhaven' were also obtained from both `Sunhigh' embryos. The frequency of variation in the `Sunhigh' seedling population, with respect to the response to bacterial leaf spot, was not so great as that exhibited by the regenerants derived from `Sunhigh' embryo #156. None of the `Redhaven' seedlings or any of the regenerants derived from `Redhaven' embryos were more resistant than `Redhaven'. These studies suggest that the frequency of somaclonal variation is genetically determined and that screening for somaclonal variation may be a feasible approach to obtaining leaf spot-resistant peach plants.

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F.A. Hammerschlag

A detached-leaf bioassay was used to evaluate peach [Prunus persica (L.) Batsch] somaclone 122-1 (derived from callus produced on an immature embryo of peach cultivar Redhaven) for resistance to several virulent strains of Xanthomonas campestris pv. pruni [E.F. Sm.) Dows], causal agent of bacterial leaf spot, and to a virulent isolate of Pseudomonas syringae van Hall pv. syringae, causal agent of bacterial canker. The detached-leaf bioassay was also used to evaluate progeny of 122-1 for resistance to X. campestris pv. pruni virulent strain XP1. Somaclone 122-1 was significantly more resistant to most strains of X. campestris pv. pruni than was `Redhaven', and all of its progeny exhibited high levels of resistance to X. campestris pv. pruni strain XP1. Somaclone 122-1 exhibited significantly higher levels of resistance to Pseudomonas syringae pv. syringae than did `Redhaven' and this resistance was retained over time in the greenhouse and following a 2-year cycle of tissue culture propagation.

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B.L. Topp, W.B. Sherman, D.A. Huber, and S.B. Linda

Analysis of a five-parent diallel in a greenhouse estimated general and specific combining ability (GCA and SCA) effects for resistance of Japanese plum (Prunus salicina Lindl. and hybrids) to Xanthomonas campestris pv. pruni (Smith) Dye stem canker, as measured by length of inoculated cankers, canker appearance rating, and canker expansion rate. `Friar' and `Gulfruby' were the most susceptible parents. `Burbank', `Wilson', and' Wade' had similar GCA values for length of inoculated cankers, but' Wade' was the superior parent in transmitting canker resistance, as measured by canker appearance rating. SCA was not important in determining the performance of a cross. Canker appearance rating was the best measure of resistance in the greenhouse tests and provided a greater separation of GCA estimates and lower coefficient of variation.

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B.L. Topp, W.B. Sherman, R.E. Stall, G.V. Minsavage, and C.J. Wilcox

Four greenhouse leaf inoculation methods for screening Japanese plum (Prunus salicina L. and hybrids) for resistance to Xanthomonas campestris pv. pruni (Smith) Dye were compared for repeatability, ability to differentiate among plant genotype responses, and correlations with field ratings. Clonally propagated trees were inoculated artificially in a greenhouse by immersing leaves in 2.5 × 108 cfu/ml inoculum (DIP), rubbing the adaxial side of leaves with a slurry of 2.5 × 108 cfu/ml inoculum and Carborundum powder (CARB), infiltrating leaves with 5 × 105 cfu/ml inoculum using a needle-less syringe (INFS), and infiltrating with 5 × 106 cfu/ml inoculum (INF6). No greenhouse method was superior in all assessment categories. The CARB method was most repeatable (t = 0.78) but had a low Spearman's correlation (rs = 0.29), indicating that greenhouse rankings did not correspond closely with field rankings. The INF6 method was unsuitable because it did not differentiate between plant genotypes. The DIP method appeared most suitable, having moderate repeatability (t = 0.46) for four observations per leaf and moderate Spearman's correlation with field performance (rs = 0.56). The INF5 method may be appropriate for identifying bacterial spot resistance that is associated with resistance in the leaf mesophyll.

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John R. Clark and James N. Moore

’ is an early, midseason clingstone with non-melting flesh like that found in processing cling peaches. It has a standard acid flavor. These cultivars have good to very good resistance to bacterial spot [caused by Xanthomonas campestris pv. pruni

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Freddi A. Hammerschlag, Ghazala Hashmi, Robin Huettel, Dennis Werner, and David Ritchie

One approach for obtaining useful genetic variation is to select for somaclonal variants generated by tissue culture techniques. Increased levels of resistance to bacterial leaf spot (Xanthomonas campestris pv. pruni) have been observed in toxin-selected and unselected peach regenerants in vitro, in the greenhouse and under field conditions. Peach regenerants have also demonstrated increased levels of bacterial canker (Pseudomonas syringae pv. syringae) and root-knot nematode (Meloidogyne incognita) resistance. Random amplified polymorphic DNA (RAPD) primers have been used to study genetic variation at the DNA level among the somaclonal variants. Sixty RAPD primers (10-mers) were screened and 10 proved useful as markers to detect polymorphisms, thus establishing a genetic basis for somaclonal variation. These studies demonstrate the feasibility of using tissue culture techniques to generate fruit trees with increased levels of disease resistance.

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Gregory L. Reighard, Eldon I. Zehr, and Freddi Hammerschlag

Peach tree short life (PTSL) is a serious peach tree disease syndrome on replant orchard sites in the Southeast. Pseudomonas syringae pv. syringae is a bacterial disease often associated with tree injury and death on these PTSL sites. Rootstocks that have better tolerance to ring nematodes such as Lovell have less PTSL death. Tissue-cultured peach embryos and/or explants have shown increased resistance to Pseudomonas syringae and Xanthomonas campestris pv. pruni, another bacterial peach pathogen, in laboratory and greenhouse screenings. Tissue-cultured `Redhaven' (RH), `Redskin' (RS), and `Sunhigh' (SH) peach cultivars on their own roots were planted with SH seedlings and RH and RS budded to Lovell rootstock on a severe PTSL site in South Carolina. Treatments beside cultivar/rootstock combination included preplant fumigation vs. nonfumigation. PTSL appeared in the third year and by year 4 significant tree death occurred. Tissue-cultured RH, RS, and SH trees had 54%, 55%, and 88% PTSL death, respectively, compared to RH (17%) and RS (29%) on Lovell or the SH seedlings (25%). Fumigation significantly decreased PTSL in both RS combinations but not RH. These data suggest that the tolerance of the cultivar root system to PTSL-inducing factors such as ring nematodes was more important in PTSL than scion resistance to bacteria.

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David Mills and Freddi A. Hammerschlag

Pathogenic bacteria, such as Xanthomonas campestris pv. pruni, cause diseases of significant economical implications in the Prunus genus. Cecropins are naturally occurring bactericidal peptides found in the hemolymph of insects. Cecropins cause channel formation in membranes and lysis of bacterial cells. We are interested in engineering the gene for cecropin into peach (Prunus persica) and other fruit tree species. The objective of this study was to determine the effect of cecropin B on viability, using fluorescein diacetate staining, and on changes in transmembrane electrical potential (PD) using the fluorescing probe merocyanine-540. Protoplasts were isolated from shoot-tip cultures in a CPW13M (salts + 0.71M mannitol) solution containing 2% cellulase and 0.5% macerase, while cells were isolated in CPW15.4S (salts + 0.45M sucrose) containing 0.5% cellulase and 0.5% macerase. Cecropin B (1μM) had no effect on viability and changes in PD, while 10μM had a slight effect, and 100μM cecropin B caused significant depolarization and lysis of peach protoplasts. No effect on viability and change in PD were observed in cells when treated with 1-100μM cecropin B. These results suggest that cells and protoplasts of peach can resist cecropin B in the concentration range that causes lysis of plant pathogenic bacteria. The implication of using cecropin to increase microbial disease resistance will be discussed.

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John R. Clark and James N. Moore