Genetic mapping of the Tsw locus for resistance to the Tospovirus Tomato spotted wilt virus in Capsicum spp. and its relationship to the Sw-5 gene for resistance to the same pathogen in tomato

The Tsw gene conferring dominant resistance to the Tospovirus Tomato spotted wilt virus (TSWV) in Capsicum spp. has been tagged with a random amplified polymorphic DNA marker and mapped to the distal portion of chromosome 10. No mapped homologues of Sw-5, a phenotypically similar dominant TSWV resistance gene in tomato, map to this region in C. annuum, although a number of Sw-5 homologues are found at corresponding positions in pepper and tomato. The relationship between Tsw and Sw-5 was also examined through genetic studies of TSWV. The capacity of TSWV-A to overcome the Tsw gene in pepper and the Sw-5 gene in tomato maps to different TSWV genome segments. Therefore, despite phenotypic and genetic similarities of resistance in tomato and pepper, we infer that distinct viral gene products control the outcome of infection in plants carrying Sw-5 and Tsw, and that these loci do not appear to share a recent common evolutionary ancestor.     

Identification and comparative mapping of a dominant potyvirus resistance gene cluster in Capsicum

The dominant gene Pvr7 from Capsicum chinense Jacq. ’PI159236’ confers resistance to the pepper mottle potyvirus (PepMoV) Florida (V1182) strain. This gene is tightly linked to the dominant potyvirus resistance gene Pvr4 with observed recombination frequencies of 0.012 to 0.016. A cleaved amplified polymorphic sequence (CAPS) marker linked to Pvr4 was used to localize Pvr4 and, by extension, Pvr7, to linkage group 10 on an interspecific map of pepper. Our results indicated that Pvr4, Pvr7, and Tsw, a gene conferring resistance to tomato spotted wilt virus, comprise the first identified cluster of dominant disease resistance genes in Capsicum L. This position does not correspond to the locations of dominant potyvirus resistance genes in potato or to the positions of any other mapped solanaceous resistance genes or resistance gene homologues. Received: 20 September 1999 / Accepted: 21 March 2000     

Draft Genome Sequence of Eggplant (Solanum melongena L.): the Representative Solanum Species Indigenous to the Old World

Unlike other important Solanaceae crops such as tomato, potato, chili pepper, and tobacco, all of which originated in South America and are cultivated worldwide, eggplant (Solanum melongena L.) is indigenous to the Old World and in this respect it is phylogenetically unique. To broaden our knowledge of the genomic nature of solanaceous plants further, we dissected the eggplant genome and built a draft genome dataset with 33,873 scaffolds termed SME_r2.5.1 that covers 833.1 Mb, ca. 74% of the eggplant genome. Approximately 90% of the gene space was estimated to be covered by SME_r2.5.1 and 85,446 genes were predicted in the genome. Clustering analysis of the predicted genes of eggplant along with the genes of three other solanaceous plants as well as Arabidopsis thaliana revealed that, of the 35,000 clusters generated, 4,018 were exclusively composed of eggplant genes that would perhaps confer eggplant-specific traits. Between eggplant and tomato, 16,573 pairs of genes were deduced to be orthologous, and 9,489 eggplant scaffolds could be mapped onto the tomato genome. Furthermore, 56 conserved synteny blocks were identified between the two species. The detailed comparative analysis of the eggplant and tomato genomes will facilitate our understanding of the genomic architecture of solanaceous plants, which will contribute to cultivation and further utilization of these crops.     

Complete sequencing and comparative analyses of the pepper (Capsicum annuum L.) plastome revealed high frequency of tandem repeats and large insertion/deletions on pepper plastome

Plants in the family Solanaceae are used as model systems in comparative and evolutionary genomics. The complete chloroplast genomes of seven solanaceous species have been sequenced, including tobacco, potato and tomato, but not peppers. We analyzed the complete chloroplast genome sequence of the hot pepper, Capsicum annuum. The pepper chloroplast genome was 156,781 bp in length, including a pair of inverted repeats (IR) of 25,783 bp. The content and the order of 133 genes in the pepper chloroplast genome were identical to those of other solanaceous plastomes. To characterize pepper plastome sequence, we performed comparative analysis using complete plastome sequences of pepper and seven solanaceous plastomes. Frequency and contents of large indels and tandem repeat sequences and distribution pattern of genome-wide sequence variations were investigated. In addition, a phylogenetic analysis using concatenated alignments of coding sequences was performed to determine evolutionary position of pepper in Solanaceae. Our results revealed two distinct features of pepper plastome compared to other solanaceous plastomes. Firstly, large indels, including insertions on accD and rpl20 gene sequences, were predominantly detected in the pepper plastome compared to other solanaceous plastomes. Secondly, tandem repeat sequences were particularly frequent in the pepper plastome. Taken together, our study represents unique features of evolution of pepper plastome among solanaceous plastomes.     

Recessive resistance genes against potyviruses are localized in colinear genomic regions of the tomato ( Lycopersicon spp.) and pepper ( Capsicum spp.) genomes

Resistance against both Potato virus Y (PVY) and Tobacco etch virus (TEV) was identified in the wild tomato relative Lycopersicon hirsutum PI247087. Analysis of the segregation ratio in F(2)/F(3) and BC(1) interspecific progenies indicated that a single recessive gene, or two very tightly linked recessive loci, are involved in resistance to both potyviruses. This locus was named pot-1. Using amplified fragment length polymorphism markers and a set of L. hirsutum introgression lines, pot-1 was mapped to the short arm of tomato chromosome 3, in the vicinity of the recessive py-1 locus for resistance to corky root rot. Because of the occurrence of phenotypically similar genes in pepper ( Capsicum spp.), the comparative genetics of resistance to potyviruses between tomato and pepper was investigated. Unlike most of the comparative genetic studies on resistance genes, pot-1 was tightly flanked by the same restriction fragment length polymorphism (RFLP) markers than the pvr2/pvr5 locus for resistance to PVY and TEV from pepper. These results may indicate that recessive resistance genes against potyviruses evolve less rapidly than the majority of the dominant genes cloned so far, and consequently may belong to a different family of resistance genes.     

Comparative genetics of nucleotide binding site-leucine rich repeat resistance gene homologues in the genomes of two dicotyledons: tomato and arabidopsis

The presence of a single resistance (R) gene allele can determine plant disease resistance. The protein products of such genes may act as receptors that specifically interact with pathogen-derived factors. Most functionally defined R-genes are of the nucleotide binding site-leucine rich repeat (NBS-LRR) supergene family and are present as large multigene families. The specificity of R-gene interactions together with the robustness of plant-pathogen interactions raises the question of their gene number and diversity in the genome. Genomic sequences from tomato showing significant homology to genes conferring race-specific resistance to pathogens were identified by systematically "scanning" the genome using a variety of primer pairs based on ubiquitous NBS motifs. Over 70 sequences were isolated and 10% are putative pseudogenes. Mapping of the amplified sequences on the tomato genetic map revealed their organization as mixed clusters of R-gene homologues that showed in many cases linkage to genetically characterized tomato resistance loci. Interspecific examination within Lycopersicon showed the existence of a null allele. Consideration of the tomato and potato comparative genetic maps unveiled conserved syntenic positions of R-gene homologues. Phylogenetic clustering of R-gene homologues within tomato and other Solanaceae family members was observed but not with R-gene homologues from Arabidopsis thaliana. Our data indicate remarkably rapid evolution of R-gene homologues during diversification of plant families.     

SAP-TRANSMISSIBLE MOSAIC DISEASES OF SOLANACEOUS CROPS IN TRINIDAD

Four sap-transmissible viruses were isolated from cultivated Solanaceae in Trinidad: (1) tobacco mosaic virus, from tobacco, tomato and sweet pepper; (2) cucumber mosaic virus, from tobacco and petunia; (3) 'pepper vein-banding virus', probably related to pepper mosaic viruses in Puerto Rico and Brazil, from peppers and tobacco; (4) 'egg-plant mosaic virus', possibly related to the tobacco ring-spot virus, from egg-plant and tomato. Pepper vein-banding virus causes leaf-crinkling and vein-banding in Physalis floridana , petunia, various Nicotiana spp. and most peppers; the Large Bell Hot pepper is killed; tomato and egg-plant are immune. Egg-plant mosaic virus produces mosaic, ring-spotting, or both, on different solanaceous species. It also gives local and systemic ring-spotting on Chenopodium hybridum and necrotic local lesions on the primary leaves of cowpea (var. Black-eye); cucumber is a symptomless carrier. Only cucumber mosaic virus was found naturally infecting non-solanaceous hosts, cucumber and certain common wild plants.
The thermal inactivation point of pepper vein-banding virus is 62° C, its dilution end-point 2×10^-5 and its longevity in vitro 6 day s at 23–30° C.; corresponding values for egg-plant mosaic virus are 78° C., 10^-6 and over 3 weeks. Aphisgossypii transmits cucumber mosaic and pepper vein-banding, but not egg-plant mosaic, of which Epitrix sp. is an occasional vector. Tobacco mosaic, as elsewhere, probably has no regular insect vectors in Trinidad.     

Conservation and divergence of both phosphate- and mycorrhiza-regulated physiological responses and expression patterns of phosphate transporters in solanaceous species

Here, orthologous genes of six phosphate transporter (PiT) genes, which are members of the Pht1 and Pht2 families in tomato and potato, have been cloned from the solanaceous species pepper, eggplant and tobacco. Overall, expressions of these genes in pepper, eggplant and tobacco showed similar patterns to those in tomato and potato: P-starvation enhancement in both leaves and roots for Pht1;1, P-depletion induction exclusively in roots for Pht1;2, mycorrhizal enhancement for Pht1;3, and mycorrhizal induction for both Pht1;4 and Pht1;5. In the roots of nonmycorrhizal eggplant, SmPht1;3, SmPht1;4 and SmPht1;5 were also expressed under extreme P starvation. Mycorrhizal symbiosis under high-P supply conditions reduced plant growth, with concurrent enhancement of Pht1;2 expression in the roots of pepper as well as eggplant. In addition, the mycorrhizal symbiosis down-regulated the expression of Pht2;1 genes greatly in the leaves of pepper and tobacco. The discrepancies between the evolutionary distances of the PiT genes and their expression patterns among the five species suggest greater complexity in function of PiT in plants than previously expected.     

Genome mapping in capsicum and the evolution of genome structure in the solanaceae.

We have created a genetic map of Capsicum (pepper) from an interspecific F2 population consisting of 11 large (76.2-192.3 cM) and 2 small (19.1 and 12.5 cM) linkage groups that cover a total of 1245.7 cM. Many of the markers are tomato probes that were chosen to cover the tomato genome, allowing comparison of this pepper map to the genetic map of tomato. Hybridization of all tomato-derived probes included in this study to positions throughout the pepper map suggests that no major losses have occurred during the divergence of these genomes. Comparison of the pepper and tomato genetic maps showed that 18 homeologous linkage blocks cover 98.1% of the tomato genome and 95.0% of the pepper genome. Through these maps and the potato map, we determined the number and types of rearrangements that differentiate these species and reconstructed a hypothetical progenitor genome. We conclude there have been 30 breaks as part of 5 translocations, 10 paracentric inversions, 2 pericentric inversions, and 4 disassociations or associations of genomic regions that differentiate tomato, potato, and pepper, as well as an additional reciprocal translocation, nonreciprocal translocation, and a duplication or deletion that differentiate the two pepper mapping parents.     

Comparative linkage map of the Solanum lycopersicoides and S. sitiens genomes and their differentiation from tomato.

The wild nightshades Solanum lycopersicoides and Solanum sitiens are closely affiliated with the tomatoes (Lycopersicon spp.). Intergeneric hybridization with cultivated tomato (Lycopersicon esculentum) is impeded by strong reproductive barriers including hybrid sterility and suppressed recombination. Conservation of genome structure between these nightshades and tomato was studied by construction of a genetic map from F2 S. sitiens x S. lycopersicoides and comparison with existing maps of tomato. Owing to self-incompatibility of the F1, two hybrid plants were crossed to obtain a population of 82 F2 individuals. Using 166 previously mapped RFLP markers and 5 restriction enzymes, 101 loci polymorphic in the S. sitiens x S. lycopersicoides population were identified. Analysis of linkage between the markers resulted in a map with 12 linkage groups covering 1192 cM and one unlinked marker. Recombination rates were similar to those observed in tomato; however, significant segregation distortion was observed for markers on 7 out of the 12 chromosomes. All chromosomes were colinear with the tomato map, except for chromosome 10, where a paracentric inversion on the long arm was detected. In this region, S. sitiens and S. lycopersicoides share the same chromosomal configuration previously reported for potato (S. tuberosum) and pepper (Capsicum), suggesting that of tomato is derived. The 10L inversion explains the lack of recombination detected among homeologous chromosomes of intergeneric hybrids in this region. On this basis, we recognize two principle genomes, designated L for the Lycopersicon spp., and S for S. lycopersicoides and S. sitiens, the first examples of structural differentiation between tomato and its cross-compatible wild relatives.     

Molecular properties of potato spindle tuber viroid (PSTVd) isolates of the Russian Research Institute of Phytopathology

The complete nucleotide sequences of more than 100 isolates of Potato spindle tuber viroid (PSTVd) collected from locations in the territory of Russia and the former USSR have been determined. These sequences represent 43 individual sequence variants, each containing 1–10 mutations with respect to the “intermediate” or type strain of PSTVd (GenBank Acc. No. V01465). Isolates containing 2–5 mutations were the most common, and 24 sequence variants are described here for the first time. Twenty one isolates contained a mutation found only in Russian and Ukrainian isolates of PSTVd up till now; i.e., replacement of the adenine at position 121 with cytosine (A121C). Many of these isolates contained two mutations—deletion of one of three adenine residues occupying positions 118–120 plus replacement of the adenine at position 121 with either uracil or cytosine (?A120, A121U/C). Both combinations of mutations were phenotypically neutral, i.e. symptom expression in Rutgers tomato was unaffected. Phylogenetic analysis of the sequences of different PSTVd isolates presented in work together with sequences of other naturally-occurring isolates obtained from Internet databases suggesting that known PSTVd isolates may be divided into four groups: (i) a group of isolates from potato and ornamentals where the type strain of PSTVd (PSTVd.018) may be considered to represent the ancestral sequence, (ii) a second group of isolates from potato and ornamentals where PSTVd.123 play the same role as PSTVd.018 for the first group, and iii) a group of potato isolates where PSTVd.125 is a possible ancestral sequence. The fourth and most divergent group of PSTVd isolates differs significantly from these first three groups. The majority of isolates in this group originate from New Zealand and Australia and infect different solanaceous hosts (tomato, pepper, cape gooseberry, potato, and others).     

Evolutionary meta-analysis of solanaceous resistance gene and solanum resistance gene analog sequences and a practical framework for cross-species comparisons

Cross-species comparative genomics approaches have been employed to map and clone many important disease resistance (R) genes from Solanum species-especially wild relatives of potato and tomato. These efforts will increase with the recent release of potato genome sequence and the impending release of tomato genome sequence. Most R genes belong to the prominent nucleotide binding site-leucine rich repeat (NBS-LRR) class and conserved NBS-LRR protein motifs enable survey of the R gene space of a plant genome by generation of resistance gene analogs (RGA), polymerase chain reaction fragments derived from R genes. We generated a collection of 97 RGA from the disease-resistant wild potato S. bulbocastanum, complementing smaller collections from other Solanum species. To further comparative genomics approaches, we combined all known Solanum RGA and cloned solanaceous NBS-LRR gene sequences, nearly 800 sequences in total, into a single meta-analysis. We defined R gene diversity bins that reflect both evolutionary relationships and DNA cross-hybridization results. The resulting framework is amendable and expandable, providing the research community with a common vocabulary for present and future study of R gene lineages. Through a series of sequence and hybridization experiments, we demonstrate that all tested R gene lineages are of ancient origin, are shared between Solanum species, and can be successfully accessed via comparative genomics approaches.     

High Density Molecular Linkage Maps of the Tomato and Potato Genomes

High density molecular linkage maps, comprised of more than 1000 markers with an average spacing between markers of approximately 1.2 cM (ca. 900 kb), have been constructed for the tomato and potato genomes. As the two maps are based on a common set of probes, it was possible to determine, with a high degree of precision, the breakpoints corresponding to 5 chromosomal inversions that differentiate the tomato and potato genomes. All of the inversions appear to have resulted from single breakpoints at or near the centromeres of the affected chromosomes, the result being the inversion of entire chromosome arms. While the crossing over rate among chromosomes appears to be uniformly distributed with respect to chromosome size, there is tremendous heterogeneity of crossing over within chromosomes. Regions of the map corresponding to centromeres and centromeric heterochromatin, and in some instances telomeres, experience up to 10-fold less recombination than other areas of the genome. Overall, 28% of the mapped loci reside in areas of putatively suppressed recombination. This includes loci corresponding to both random, single copy genomic clones and transcribed genes (detected with cDNA probes). The extreme heterogeneity of crossing over within chromosomes has both practical and evolutionary implications. Currently tomato and potato are among the most thoroughly mapped eukaryotic species and the availability of high density molecular linkage maps should facilitate chromosome walking, quantitative trait mapping, marker-assisted breeding and evolutionary studies in these two important and well studied crop species.     

Linkage of the <Emphasis Type="Italic">A</Emphasis> locus for the presence of anthocyanin and <Emphasis Type="Italic">fs10.1</Emphasis>, a major fruit-shape QTL in pepper

The purple color of the foliage, flower and immature fruit of pepper ( Capsicum spp.) is a result of the accumulation of anthocyanin pigments in these tissues. The expression of anthocyanins is controlled by the incompletely dominant gene A. We have mapped A to pepper chromosome 10 in a Capsicum annuum (5226) x Capsicum chinense (PI 159234) F(2) population to a genomic region that also controls anthocyanin expression in two other Solanaceous species, tomato and potato, suggesting that variation for tissue-specific expression of anthocyanin pigments in these plants is controlled by an orthologous gene(s). We mapped an additional locus, Fc, for the purple anther filament in an F(2) population from a cross of IL 579, a C. chinense introgression line and its recurrent parent 100/63, to the same position as A, suggesting that the two loci are allelic. The two anthocyanin loci were linked to a major quantitative trait locus, fs10.1, for fruit-shape index (ratio of fruit length to fruit width), that also segregated in the F(2) populations. This finding verified the observation of Peterson in 1959 of linkage between fruit color and fruit-shape genes in a cross between round and elongated-fruited parents. The linkage relationship in pepper resembles similar linkage in potato, in which anthocyanin and tuber-shape genes were found linked to each other in a cross of round and elongated-tuber parents. It is therefore possible that the shape pattern of distinct organs such as fruit and tuber in pepper and potato is controlled by a similar gene(s).     

The R1 resistance gene cluster contains three groups of independently evolving, type I R1 homologues and shows substantial structural variation among haplotypes of Solanum demissum

Cultivated and wild potatoes contain a major disease-resistance cluster on the short arm of chromosome V, including the R1 resistance (R) gene against potato late blight. To explore the functional and evolutionary significance of clustering in the generation of novel disease-resistance genes, we constructed three approximately 1 Mb physical maps in the R1 gene region, one for each of the three genomes (haplotypes) of allohexaploid Solanum demissum, the wild potato progenitor of the R1 locus. Totals of 691, 919 and 559 kb were sequenced for each haplotype, and three distinct resistance-gene families were identified, one homologous to the potato R1 gene and two others homologous to either the Prf or the Bs4 R-gene of tomato. The regions with R1 homologues are highly divergent among the three haplotypes, in contrast to the conserved flanking non-resistance gene regions. The R1 locus shows dramatic variation in overall length and R1 homologue number among the three haplotypes. Sequence comparisons of the R1 homologues show that they form three distinct clades in a distance tree. Frequent sequence exchanges were detected among R1 homologues within each clade, but not among those in different clades. These frequent sequence exchanges homogenized the intron sequences of homologues within each clade, but did not homogenize the coding sequences. Our results suggest that the R1 homologues represent three independent groups of fast-evolving type I resistance genes, characterized by chimeric structures resulting from frequent sequence exchanges among group members. Such genes were first identified among clustered RGC2 genes in lettuce, where they were distinguished from slow-evolving type II R-genes. Our findings at the R1 locus in S. demissum may indicate that a common or similar mechanism underlies the previously reported differentiation of type I and type II R-genes and the differentiation of type I R-genes into distinct groups, identified here.     

CR1 — a dispersed repeated element associated with the Cab-1 locus in tomato

Cab-1 is a complex genetic locus in tomato consisting of four clustered genes encoding chlorophyll a/b-binding polypeptide. Southern blot analysis of total tomato DNA with genomic clones corresponding to the Cab-1 locus has revealed the presence of a repetitive element in the 3 kb spacer regions between two of these genes. This repetitive element, named CR1, has been characterized via sequencing, genetic mapping and hybridization to related solanaceous species. Results indicate that there are as many as 30 copies of this element in the tomato genome and that most, if not all, are found at independent loci. Sites corresponding to 12 of the repeats have been located on different regions of chromosomes 2, 4, 5, 7, 10 and 11. A 1.6 kb PstI-EcoRI fragment from the Cab-1 locus containing the element was sequenced and found to be 75% AT-rich. No open reading frames larger than 150 bp were detected. Several imperfect inverted repeats flanked by direct repeats could be found at the ends of the element. This arrangement is reminiscent of known transposons. Southern hybridization analysis indicates that multiple copies of CR1 exist in all species of the genus Lycopersicon as well as in Solanum lycopersicoides and S. tuberosum (potato), but not in eggplant, pepper, petunia, Datura or tobacco. Melt-off experiments indicate that members of the CR1 family in the tomato genome are more closely related to one another than to homologous members in the genomes of S. lycopersicoides or S. tuberosum, suggesting some type of concerted evolution.     

Euchromatin and pericentromeric heterochromatin: comparative composition in the tomato genome

Eleven sequenced BACs were annotated and localized via FISH to tomato pachytene chromosomes providing the first global insights into the compositional differences of euchromatin and pericentromeric heterochromatin in this model dicot species. The results indicate that tomato euchromatin has a gene density (6.7 kb/gene) similar to that of Arabidopsis and rice. Thus, while the euchromatin comprises only 25% of the tomato nuclear DNA, it is sufficient to account for approximately 90% of the estimated 38,000 nontransposon genes that compose the tomato genome. Moreover, euchromatic BACs were largely devoid of transposons or other repetitive elements. In contrast, BACs assigned to the pericentromeric heterochromatin had a gene density 10-100 times lower than that of the euchromatin and are heavily populated by retrotransposons preferential to the heterochromatin-the most abundant transposons belonging to the Jinling Ty3/gypsy-like retrotransposon family. Jinling elements are highly methylated and rarely transcribed. Nonetheless, they have spread throughout the pericentromeric heterochromatin in tomato and wild tomato species fairly recently-well after tomato diverged from potato and other related solanaceous species. The implications of these findings on evolution and on sequencing the genomes of tomato and other solanaceous species are discussed.     

Autonomous cell death, temperature sensitivity and the genetic control associated with resistance to cucumber mosaic virus (CMV) in diploid potatoes (Solanum spp.)

Cucumber mosaic virus (CMV) is a commonly occurring plant virus that causes severe damage in many crops, including the diploid crop species tomato and pepper (Lycopersicon spp. and Capsicum spp., respectively) of the family Solanaceae, but it is neither common nor economically important in cultivated potatoes (Solanum tuberosum; Solanaceae). Resistance to CMV was examined in two diploid (2n=2x=24), highly heterozygous potato populations (Solanum spp.; Solanaceae) consisting of 76 and 126 progeny. Resistance to long-distance transport of CMV controlled by one locus with a major effect and functional at a low temperature (18°C) but overcome at a high temperature (28°C) was identified in one population. In the other population, resistance was controlled by two loci with major effects. In both populations, additional genes with minor effects were probably also involved. Induced resistance to CMV, associated with autonomously developing cell death lesions (Anl) previously not known in potato, was expressed in one parental line. The mechanisms of resistance to CMV may be associated with an inherent or developmental lack of host factors required for compatible CMV-host interactions in viral long distance transport and/or inability of CMV to efficiently suppress the host gene silencing mechanism in potatoes. Polyploidy (gene dose) and high heterozygosity (multiple homologous genes) of potato cultivars may be significant in conferring the durable resistance to CMV. These data provide explanations why CMV is not common and economically important in cultivated potatoes, even though CMV commonly occurs in other crops, weeds and wild plants in potato production areas. Received: 11 February 1999 / Accepted: 25 March 1999