Samuel F. Hutton

The Tomato Yellow Leaf Curl Virus resistance genes Ty-1 and Ty-3 are allelic and code for DFDGD-class RNA-dependent RNA polymerases.

Tomato Yellow Leaf Curl Virus Disease incited by Tomato yellow leaf curl virus (TYLCV) causes huge losses in tomato production worldwide and is caused by different related begomovirus species. Breeding for TYLCV resistance has been based on the introgression of multiple resistance genes originating from several wild tomato species. In this study we have fine-mapped the widely used Solanum chilense–derived Ty-1 and Ty-3 genes by screening nearly 12,000 plants for recombination events and generating recombinant inbred lines. Multiple molecular markers were developed and used in combination with disease tests to fine-map the genes to a small genomic region (approximately 70 kb). Using a Tobacco Rattle Virus–Virus Induced Gene Silencing approach, the resistance gene was identified. It is shown that Ty-1 and Ty-3 are allelic and that they code for a RNA–dependent RNA polymerase (RDR) belonging to the RDRγ type, which has an atypical DFDGD motif in the catalytic domain. In contrast to the RDRα type, characterized by a catalytic DLDGD motif, no clear function has yet been described for the RDRγ type, and thus the Ty-1/Ty-3 gene unveils a completely new class of resistance gene. Although speculative, the resistance mechanism of Ty-1/Ty-3 and its specificity towards TYLCV are discussed in light of the function of the related RDRα class in the amplification of the RNAi response in plants and transcriptional silencing of geminiviruses in plants.

High-Density SNP Genotyping of Tomato (Solanum lycopersicum L.) Reveals Patterns of Genetic Variation Due to Breeding

The effects of selection on genome variation were investigated and visualized in tomato using a high-density single nucleotide polymorphism (SNP) array. 7,720 SNPs were genotyped on a collection of 426 tomato accessions (410 inbreds and 16 hybrids) and over 97% of the markers were polymorphic in the entire collection. Principal component analysis (PCA) and pairwise estimates of F st supported that the inbred accessions represented seven sub-populations including processing, large-fruited fresh market, large-fruited vintage, cultivated cherry, landrace, wild cherry, and S. pimpinellifolium. Further divisions were found within both the contemporary processing and fresh market sub-populations. These sub-populations showed higher levels of genetic diversity relative to the vintage sub-population. The array provided a large number of polymorphic SNP markers across each sub-population, ranging from 3,159 in the vintage accessions to 6,234 in the cultivated cherry accessions. Visualization of minor allele frequency revealed regions of the genome that distinguished three representative sub-populations of cultivated tomato (processing, fresh market, and vintage), particularly on chromosomes 2, 4, 5, 6, and 11. The PCA loadings and F st outlier analysis between these three sub-populations identified a large number of candidate loci under positive selection on chromosomes 4, 5, and 11. The extent of linkage disequilibrium (LD) was examined within each chromosome for these sub-populations. LD decay varied between chromosomes and sub-populations, with large differences reflective of breeding history. For example, on chromosome 11, decay occurred over 0.8 cM for processing accessions and over 19.7 cM for fresh market accessions. The observed SNP variation and LD decay suggest that different patterns of genetic variation in cultivated tomato are due to introgression from wild species and selection for market specialization.

Chromosomal rearrangements between tomato and Solanum chilense hamper mapping and breeding of the TYLCV resistance gene Ty‐1

Tomato yellow leaf curl disease, a devastating disease of Solanum lycopersicum (tomato), is caused by a complex of begomoviruses generally referred to as Tomato yellow leaf curl virus (TYLCV). Almost all breeding for TYLCV resistance has been based on the introgression of the Ty-1 resistance locus derived from Solanum chilense LA1969. Knowledge about the exact location of Ty-1 on tomato chromosome 6 will help in understanding the genomic organization of the Ty-1 locus. In this study, we analyze the chromosomal rearrangement and recombination behavior of the chromosomal region where Ty-1 is introgressed. Nineteen markers on tomato chromosome 6 were used in F(2) populations obtained from two commercial hybrids, and showed the presence of a large introgression in both. Fluorescence in situ hybridization (FISH) analysis revealed two chromosomal rearrangements between S. lycopersicum and S. chilense LA1969 in the Ty-1 introgression. Furthermore, a large-scale recombinant screening in the two F(2) populations was performed, and 30 recombinants in the Ty-1 introgression were identified. All recombination events were located on the long arm beyond the inversions, showing that recombination in the inverted region was absent. Disease tests on progenies of informative recombinants with TYLCV mapped Ty-1 to the long arm between markers MSc05732-4 and MSc05732-14, an interval overlapping with the reported Ty-3 region, which led to the indication that Ty-1 and Ty-3 may be allelic. With this study we prove that FISH can be used as a diagnostic tool to aid in the accurate mapping of genes that were introgressed from wild species into cultivated tomato.

Identification of QTL associated with resistance to bacterial spot race T4 in tomato

Bacterial spot of tomato (Solanum lycopersicum L.), caused by several Xanthomonas sp., is a serious but difficult disease to control by chemical means. Development of resistance has been hindered by emergence of races virulent to tomato, by the quantitative inheritance of resistance, and by a low correlation between seedling assays and resistance in the field. Resistance to multiple races, including race T4, has been described in the S. lycopersicum var. cerasiformae accession PI 114490. We used molecular markers to identify associations with quantitative trait loci (QTL) in an elite inbred backcross (IBC) population derived from OH 9242, PI 114490 and Fla. 7600, a breeding line with tomato accession Hawaii 7998 (H7998) in its pedigree. Race T4 resistance has also been described in the advanced breeding lines Fla. 8233, Fla. 8517, and Fla. 8326, and a selective genotyping approach was used to identify introgressions associated with resistance in segregating progeny derived from crosses with these lines. In the IBC population, loci on chromosomes 11 and 3, respectively, explained as much as 29.4 and 4.8% of resistance variation. Both these loci were also confirmed by selective genotyping: PI 114490 and H7998 alleles on chromosome 11 each provided resistance. The PI 114490 allele on chromosome 3 was confirmed in the Fla. 8517 population, and an allele of undetermined descent was confirmed at this locus in the Fla. 8326 population. A chromosome 12 allele was associated with susceptibility in the Fla. 8517 population. Additional loci contributing minor effects were also implicated in the IBC population or by selective genotyping. Selection for the major QTL in a marker-directed phenotyping approach should significantly improve the efficiency of breeding for resistance to bacterial spot race T4, although as yet undetected QTL would be necessary to carry out strict marker assisted selection.

MutS HOMOLOG1-Derived Epigenetic Breeding Potential in Tomato

Silencing of a mutator gene homolog permits the alteration of plant growth behavior and the enhancement of growth potential using a novel plant breeding strategy. Evidence is compelling in support of a naturally occurring epigenetic influence on phenotype expression in land plants, although discerning the epigenetic contribution is difficult. Agriculturally important attributes like heterosis, inbreeding depression, phenotypic plasticity, and environmental stress response are thought to have significant epigenetic components, but unequivocal demonstration of this is often infeasible. Here, we investigate gene silencing of a single nuclear gene, MutS HOMOLOG1 (MSH1), in the tomato (Solanum lycopersicum) ‘Rutgers’ to effect developmental reprogramming of the plant. The condition is heritable in subsequent generations independent of the MSH1-RNA interference transgene. Crossing these transgene-null, developmentally altered plants to the isogenic cv Rutgers wild type results in progeny lines that show enhanced, heritable growth vigor under both greenhouse and field conditions. This boosted vigor appears to be graft transmissible and is partially reversed by treatment with the methylation inhibitor 5-azacytidine, implying the influence of mobile, epigenetic factors and DNA methylation changes. These data provide compelling evidence for the feasibility of epigenetic breeding in a crop plant.

Molecular Mapping of Hypersensitive Resistance from Tomato 'Hawaii 7981' to Xanthomonas perforans Race T3

Bacterial spot of tomato (Solanum lycopersicum) is caused by four species of Xanthomonas. The disease causes significant yield losses and a reduction in fruit quality. Physiological races have been described with tomato race 3 (T3) corresponding to strains of Xanthomonas perforans. The breeding line Hawaii 7981 (hereafter H7981) shows a hypersensitive reaction (HR) to race T3 strains conditioned by the interaction of the host resistance locus Xv3 and the bacterial effector avrXv3. The Xv3 gene is required for H7981-derived resistance to be effective under field conditions, though its expression is subject to genetic background. The segregation of HR in F(2) populations derived from H7981 crossed to processing tomato parents OH88119 and OH7870 was studied in 331 progeny, with the two independent crosses providing validation. We screened 453 simple-sequence repeat, insertion/deletion, and single-nucleotide polymorphism markers and identified 44 polymorphic markers each for the OH88119 and OH7870 populations covering 84.6 and 73.3% of the genome, respectively, within 20 centimorgans (cM). Marker-trait analysis using all polymorphic markers demonstrated that Xv3-mediated resistance maps to chromosome 11 in the two independent crosses. Allelism tests were conducted in crosses between lines carrying Xv3 derived from H7981, Rx-4 derived from plant introduction (PI) 128216, and resistance derived from PI 126932. These allelism tests suggested that the loci conditioning HR to race T3 strains are linked within 0.1 cM, are allelic, or are the same gene.

Analysis of wild-species introgressions in tomato inbreds uncovers ancestral origins

BackgroundDecades of intensive tomato breeding using wild-species germplasm have resulted in the genomes of domesticated germplasm (Solanum lycopersicum) being intertwined with introgressions from their wild relatives. Comparative analysis of genomes among cultivated tomatoes and wild species that have contributed genetic variation can help identify desirable genes, such as those conferring disease resistance. The ability to identify introgression position, borders, and contents can reveal ancestral origins and facilitate harnessing of wild variation in crop breeding.ResultsHere we present the whole-genome sequences of two tomato inbreds, Gh13 and BTI-87, both carrying the begomovirus resistance locus Ty-3 introgressed from wild tomato species. Introgressions of different sizes on chromosome 6 of Gh13 and BTI-87, both corresponding to the Ty-3 region, were identified as from a source close to the wild species S. chilense. Other introgressions were identified throughout the genomes of the inbreds and showed major differences in the breeding pedigrees of the two lines. Interestingly, additional large introgressions from the close tomato relative S. pimpinellifolium were identified in both lines. Some of the polymorphic regions were attributed to introgressions in the reference Heinz 1706 genome, indicating wild genome sequences in the reference tomato genome.ConclusionsThe methods developed in this work can be used to delineate genome introgressions, and subsequently contribute to development of molecular markers to aid phenotypic selection, fine mapping and discovery of candidate genes for important phenotypes, and for identification of novel variation for tomato improvement. These universal methods can easily be applied to other crop plants.

Mini Review: Potential Applications of Non-host Resistance for Crop Improvement

Plant breeding for disease resistance is crucial to sustain global crop production. For decades, plant breeders and researchers have extensively used host plant resistance genes (R-genes) to develop disease resistant cultivars. However, the general instability of R-genes in crop cultivars when challenged with diverse pathogen populations emphasizes the need for more stable means of resistance. Alternatively, non-host resistance is recognized as the most durable, broad-spectrum form of resistance against the majority of potential pathogens in plants and has gained great attention as an alternative target for managing resistance. While transgenic approaches have been utilized to transfer non-host resistance to host species, conventional breeding applications have been more elusive. Nevertheless, avenues for discovery and deployment of genetic loci for non-host resistance via hybridization are increasingly abundant, particularly when transferring genes among closely related species. In this mini review, we discuss current and developing applications of non-host resistance for crop improvement with a focus on the overlap between host and non-host mechanisms and the potential impacts of new technology.

Assessing the genetic variation of Ty-1 and Ty-3 alleles conferring resistance to tomato yellow leaf curl virus in a broad tomato germplasm

Tomato yellow leaf curl virus (TYLCV) hampers tomato production worldwide. Our previous studies have focussed on mapping and ultimately cloning of the TYLCV resistance genes Ty-1 and Ty-3. Both genes are derived from Solanum chilense and were shown to be allelic. They code for an RNA-dependent RNA polymerase (RDR) belonging to the RDRγ type defined by a DFDGD catalytic domain. In this study, we first fine-mapped the TYLCV resistance in S. chilense LA1932, LA1960 and LA1971. Results showed that chromosomal intervals of the causal genes in these TYLCV-resistant accessions overlap and cover the region where Ty-1/Ty-3 is located. Further, virus-induced gene silencing was used to silence Ty-1/Ty-3 in tomato lines carrying TYLCV resistance introgressed from S. chilense LA1932, LA1938 and LA1971. Results showed that silencing Ty-1/Ty-3 compromised the resistance in lines derived from S. chilense LA1932 and LA1938. The LA1971-derived material remained resistant upon silencing Ty-1/Ty-3. Further, we studied the allelic variation of the Ty-1/Ty-3 gene by examining cDNA sequences from nine S. chilense-derived lines/accessions and more than 80 tomato cultivars, landraces and accessions of related wild species. The DFDGD catalytic domain of the Ty-1/Ty-3 gene is conserved among all tomato lines and species analysed. In addition, the 12 base pair insertion at the 5-prime part of the Ty-1/Ty-3 gene was found not to be specific for the TYLCV resistance allele. However, compared with the susceptible ty-1 allele, the Ty-1/Ty-3 allele is characterized by three specific amino acids shared by seven TYLCV-resistant S. chilense accessions or derived lines. Thus, Ty-1/Ty-3-specific markers can be developed based on these polymorphisms. Elevated transcript levels were observed for all tested S. chilenseRDR alleles (both Ty-1 and ty-1 alleles), demonstrating that elevated expression level is not a good selection criterion for a functional Ty-1/Ty-3 allele.

Linkage between the I - 3 gene for resistance to Fusarium wilt race 3 and increased sensitivity to bacterial spot in tomato

Key messageThe negative association between theI-3gene and increased sensitivity to bacterial spot is due to linkage drag (not pleiotropy) and may be remedied by reducing the introgression size.AbstractFusarium wilt is one of the most serious diseases of tomato (Solanum lycopersicum L.) throughout the world. There are three races of the pathogen (races 1, 2 and 3), and the deployment of three single, dominant resistance genes corresponding to each of these has been the primary means of controlling the disease. The I-3 gene was introgressed from S. pennellii and confers resistance to race 3. Although I-3 provides effective control, it is negatively associated with several horticultural traits, including increased sensitivity to bacterial spot disease (Xanthomonas spp.). To test the hypothesis that this association is due to linkage with unfavorable alleles rather than to pleiotropy, we used a map-based approach to develop a collection of recombinant inbred lines varying for portions of I-3 introgression. Progeny of recombinants were evaluated for bacterial spot severity in the field for three seasons, and disease severities were compared between I-3 introgression haplotypes for each recombinant. Results indicated that increased sensitivity to bacterial spot is not associated with the I-3 gene, but rather with an upstream region of the introgression. A survey of public and private inbred lines and hybrids indicates that the majority of modern I-3 germplasm contains a similarly sized introgression for which the negative association with bacterial spot likely persists. In light of this, it is expected that the development and utilization of a reduced I-3 introgression will significantly improve breeding efforts for resistance to Fusarium wilt race 3.