Ppe.XapF: High Throughput KASP Assays to Identify Fruit Response to Xanthomonas arboricola pv. pruni (Xap) in Peach

Bacterial spot, caused by Xanthomonas arboricola pv. pruni (Xap), is a serious peach disease with symptoms that traverse severe defoliation and black surface pitting, cracking or blemishes on peach fruit with global economic impacts. A management option for control and meeting consumer demand for chemical-free, environmentally friendly fruit production is the development of resistant or tolerant cultivars. We developed simple, accurate, and efficient DNA assays (Ppe.XapF) based on SNP genotyping with KASP technology to quickly test for bacterial spot resistance alleles in peach fruit that allows breeders to cull seedlings at the greenhouse stage. The objective of this research was to validate newly developed DNA tests that target the two major QTLs for fruit resistance in peach with diagnostic utility in predicting fruit response to bacterial spot infection. Our study confirms that only two Ppe.XapF DNA tests, Ppe.XapF1-1 and Ppe.XapF6-2, are needed to distinguish between susceptible and resistant alleles. Use of these efficient and accurate Ppe.XapF KASP tests resulted in 44% reduction in seedling planting rate in the Clemson University peach breeding program.

Ppe.XapF: High Throughput KASP Assays to Identify Fruit Response to Xanthomonas Arboricola pv. pruni (Xap) in Peach

Background Bacterial spot, caused by Xanthomonas arboricola pv. pruni (Xap), is a serious peach disease with symptoms that traverse severe defoliation and black surface pitting, cracking or blemishes on peach fruit with global economic impacts. A management option for control and meeting consumer demand for chemical-free, environmentally friendly fruit production is the development of resistant or tolerant cultivars. Results We developed simple, accurate, and efficient Ppe.XapF DNA tests based on SNP genotyping with KASP technology to quickly test for bacterial spot resistance alleles in peach fruit and cull at the greenhouse stage. The objective of this research was to validate newly developed Ppe.XapF DNA tests that target the two major QTLs for fruit resistance in peach with diagnostic utility in predicting fruit response to bacterial spot infection. Conclusion Our study confirms that only two Ppe.XapF DNA tests, Ppe.XapF1-1 and Ppe.XapF6-2, are needed to distinguish between susceptible and resistant haplotypes. Use of these efficient and accurate Ppe.XapF KASP tests resulted in 44% reduction in seedling planting rate in the Clemson University peach breeding program.

Progress in Developing Bacterial Spot Resistance in Tomato

Bacterial spot (BS), caused by four species of Xanthomonas: X. euvesicatoria, X. vesicatoria, X. perforans and X. gardneri in tomato (Solanum lycopersicum L.) results in severe loss in yield and quality by defoliation and the appearance of lesions on fruits, respectively. The combined industry standard for BS control (foliar applications Actigard® rotated with copper plus mancozeb) does not offer sufficient protection, especially when weather conditions favor disease spread. Development of tomato cultivars with BS resistance is thus an important measure to minimize losses. Hypersensitive and non-hypersensitive resistance has been identified in different wild accessions and cultivated tomato relatives and has been transferred to cultivated tomato. However, complete resistance is yet to be obtained. With the advent of next generation sequencing and precise genome editing tools, the genetic regions that confer resistance to bacterial spot can be targeted and enriched through gene pyramiding in a new commercial cultivar which may confer higher degree of horizontal resistance to multiple strains of Xanthomonas causing bacterial spot in tomato.

The Type III Effector AvrBsT Enhances Xanthomonas perforans Fitness in Field-Grown Tomato

Type III secretion system effectors contribute to pathogenicity through various mechanisms. Recent surveys showed an increasing prevalence of the type III secretion effector avrBsT among Xanthomonas perforans strains. We hypothesized that the acquisition of avrBsT has a fitness advantage for the pathogen. The contribution of avrBsT to fitness on tomato was evaluated based on disease severity, in planta growth, competition, and recovery rates of wild-type (WT) and avrBsT mutant strains in greenhouse and field plants. GEV872 and GEV1001, representative strains of two phylogenomic groups of X. perforans, were selected for generating avrBsT mutants. Disease severity was higher for WT strains compared with the avrBsT mutant strains. X. perforans WT and avrBsT mutant strains did not differ following leaf infiltration of greenhouse plants in direct competition and in planta growth assays. The effect of avrBsT on pathogen fitness was noticeable under field conditions. Differences in strain recovery were significant, with WT being recovered two to eight times more than avrBsT mutant strains in the case of both strains GEV872 and GEV1001. WT strains were capable of spreading longer distances across field plots compared with avrBsT mutant strains. Findings suggest that the functional AvrBsT affects the fitness of X. perforans under field conditions, making it an ideal candidate for bacterial spot resistance breeding efforts in tomato.

Protecting vegetable cultivars in Brazil: a chili pepper case-study research

ABSTRACT: In all countries, members of the UPOV (Union Internationale pour la Protection des Obtentions Végétales), of which Brazil is a signatory, to protect a new plant variety is necessary to carry out DUS (Distinctness, Uniformity and Stability) tests. DUS testing forCapsicum spp. is based on 48 descriptors, which involve qualitative and quantitative traits, observed from germination to fruit harvest. This paper describes the performance of DUS tests on lines of chili peppers, which are candidates for protection; we discuss the main aspects related to operational difficulties, the relevance of some descriptors for the protection process and highlight the importance of protecting new plant varieties as national intellectual property. Four C. annuum var.annuum recombinant inbred lines were tested. The Capsicum Breeding Program of the Universidade Estadual do Norte Fluminense Darcy Ribeiro developed the lines, which are resistant to bacterial spot. They were tested under greenhouse conditions from June to November 2013; and from January to July 2014 in Campos dos Goytacazes, Rio de Janeiro State, Brazil. The 'Jalapeño M' commercial genotype was used as control and treatments were arranged in a randomized block design with seven plants per plot with five replications. Besides the 48 descriptors, stipulated by legislation for Capsicum DUS testing, we included a descriptor for bacterial spot resistance. The descriptors that enabled distinction varied with each line. Although being distinctive for some descriptors, L1 and L2 lines were neither homogeneous nor stable. L6 and L8 recombined lines were characterized by homogeneity and stability. Fruit shape, capsaicin presence, number of days for flowering and bacterial spot resistance were descriptors that allowed for differentiation between treatments. In conclusion, the L6 and L8 recombined lines met the DUS test requirements; therefore, they may be subjected to the protection process with the Brazilian Ministry of Agriculture, Livestock and Food Supply.

Association Analysis for Bacterial Spot Resistance in a Directionally Selected Complex Breeding Population of Tomato

Bacterial spot of tomato is caused by at least four species of Xanthomonas with multiple physiological races. We developed a complex breeding population for simultaneous discovery of marker-trait linkage, validation of existing quantitative trait loci (QTL), and pyramiding of resistance. Six advanced accessions with resistance from distinct sources were crossed in all combinations and their F1 hybrids were intercrossed. Over 1,100 segregating progeny were evaluated in the field following inoculation with X. euvesicatoria race T1 strains. We selected 5% of the most resistant and 5% of the most susceptible progeny for evaluation as plots in two subsequent replicated field trials inoculated with T1 and T3 (X. perforans) strains. The estimated heritability of T1 resistance was 0.32. In order to detect previously reported resistance genes, as well as novel QTL, we explored methods to correct for population structure and analysis based on single markers or haplotypes. Both single-point and haplotype analyses identified strong associations in the genomic regions known to carry Rx-3 (chromosome 5) and Rx-4/Xv3 (chromosome 11). Accounting for kinship and structure generally improved the fit of statistical models. Detection of known loci was improved by adding kinship or a combination of kinship and structure using a Q matrix from model-based clustering. Additional QTL were detected on chromosomes 1, 4, 6, and 7 for T1 resistance and chromosomes 2, 4, and 6 for T3 resistance (P < 0.01). Haplotype analysis improved our ability to trace the origin of positive alleles. These results demonstrate that both known and novel associations can be identified using complex breeding populations that have experienced directional selection.