Adriaan W. van Heusden

Exploring genetic variation in the tomato (Solanum section Lycopersicon) clade by whole-genome sequencing.

We explored genetic variation by sequencing a selection of 84 tomato accessions and related wild species representative of the Lycopersicon, Arcanum, Eriopersicon and Neolycopersicon groups, which has yielded a huge amount of precious data on sequence diversity in the tomato clade. Three new reference genomes were reconstructed to support our comparative genome analyses. Comparative sequence alignment revealed group-, species- and accession-specific polymorphisms, explaining characteristic fruit traits and growth habits in the various cultivars. Using gene models from the annotated Heinz 1706 reference genome, we observed differences in the ratio between non-synonymous and synonymous SNPs (dN/dS) in fruit diversification and plant growth genes compared to a random set of genes, indicating positive selection and differences in selection pressure between crop accessions and wild species. In wild species, the number of single-nucleotide polymorphisms (SNPs) exceeds 10 million, i.e. 20-fold higher than found in most of the crop accessions, indicating dramatic genetic erosion of crop and heirloom tomatoes. In addition, the highest levels of heterozygosity were found for allogamous self-incompatible wild species, while facultative and autogamous self-compatible species display a lower heterozygosity level. Using whole-genome SNP information for maximum-likelihood analysis, we achieved complete tree resolution, whereas maximum-likelihood trees based on SNPs from ten fruit and growth genes show incomplete resolution for the crop accessions, partly due to the effect of heterozygous SNPs. Finally, results suggest that phylogenetic relationships are correlated with habitat, indicating the occurrence of geographical races within these groups, which is of practical importance for Solanum genome evolution studies.

Perspectives on deciphering mechanisms underlying plant heat stress response and thermotolerance

Global warming is a major threat for agriculture and food safety and in many cases the negative effects are already apparent. The current challenge of basic and applied plant science is to decipher the molecular mechanisms of heat stress response (HSR) and thermotolerance in detail and use this information to identify genotypes that will withstand unfavorable environmental conditions. Nowadays X-omics approaches complement the findings of previous targeted studies and highlight the complexity of HSR mechanisms giving information for so far unrecognized genes, proteins and metabolites as potential key players of thermotolerance. Even more, roles of epigenetic mechanisms and the involvement of small RNAs in thermotolerance are currently emerging and thus open new directions of yet unexplored areas of plant HSR. In parallel it is emerging that although the whole plant is vulnerable to heat, specific organs are particularly sensitive to elevated temperatures. This has redirected research from the vegetative to generative tissues. The sexual reproduction phase is considered as the most sensitive to heat and specifically pollen exhibits the highest sensitivity and frequently an elevation of the temperature just a few degrees above the optimum during pollen development can have detrimental effects for crop production. Compared to our knowledge on HSR of vegetative tissues, the information on pollen is still scarce. Nowadays, several techniques for high-throughput X-omics approaches provide major tools to explore the principles of pollen HSR and thermotolerance mechanisms in specific genotypes. The collection of such information will provide an excellent support for improvement of breeding programs to facilitate the development of tolerant cultivars. The review aims at describing the current knowledge of thermotolerance mechanisms and the technical advances which will foster new insights into this process.

The Bemisia tabaci species complex: Additions from different parts of the world

Bemisia tabaci is one of the most threatening pests in many crops. We sequenced part of the mitochondrial cytochrome oxidase I gene from fifty whitefly populations collected in Indonesia, Thailand, India and China. Nineteen unique sequences (haplotypes) of the cytochrome oxidase I were identified in these populations. They were combined with sequences available in databases, resulting in a total of 407 haplotypes and analyzed together with nine outgroup accessions. A phylogenetic tree was calculated using the maximum likelihood method. The tree showed that all groups that were found in previous studies were also present in our study. Additionally, seven new groups were identified based on the new haplotypes. Most B. tabaci haplotypes grouped based on their geographical origin. Two groups were found to have a worldwide distribution. Our results indicate that our knowledge on the species complex around B. tabaci is still far from complete.

Mapping and characterization of novel parthenocarpy QTLs in tomato

Parthenocarpy is the development of the fruit in absence of pollination and/or fertilization. In tomato, parthenocarpy is considered as an attractive trait to solve the problems of fruit setting under unfavorable conditions. We studied the genetics of parthenocarpy in two different lines, IL5-1 and IVT-line 1, both carrying Solanum habrochaites chromosome segments. Parthenocarpy in IL5-1 is under the control of two QTLs, one on chromosome 4 (pat4.1) and one on chromosome 5 (pat5.1). IVT-line 1 also contains two parthenocarpy QTLs, one on chromosome 4 (pat4.2) and one on chromosome 9 (pat9.1). In addition, we identified one stigma exsertion locus in IL5-1, located on the long arm of chromosome 5 (se5.1). It is likely that pat4.1, from IL5-1 and pat4.2, from IVT-line 1, both located near the centromere of chromosome 4 are allelic. By making use of the microsynteny between tomato and Arabidopsis in this genetic region, we identified ARF8 as a potential candidate gene for these two QTLs. ARF8 is known to act as an inhibitor for further carpel development in Arabidopsis, in absence of pollination/fertilization. Expression of an aberrant form of the ArabidopsisARF8 gene, in tomato, has been found to cause parthenocarpy. This candidate gene approach may lead to the first isolation of a parthenocarpy gene in tomato and will allow further use in several crop species.

Three QTLs for Botrytis cinerea resistance in tomato

Tomato (Solanum lycopersicum) is susceptible to grey mold (Botrytis cinerea). Partial resistance to this fungus was identified in accessions of wild relatives of tomato such as S. habrochaites LYC4. In order to identify loci involved in quantitative resistance (QTLs) to B. cinerea, a population of 174 F2 plants was made originating from a cross between S. lycopersicum cv. Moneymaker and S. habrochaites LYC4. The population was genotyped and tested for susceptibility to grey mold using a stem bioassay. Rbcq1, a QTL reducing lesion growth (LG) and Rbcq2, a QTL reducing disease incidence (DI) were identified. Rbcq1 is located on Chromosome 1 and explained 12% of the total phenotypic variation while Rbcq2 is located on Chromosome 2 and explained 15% of the total phenotypic variation. Both QTL effects were confirmed by assessing disease resistance in two BC2S1 progenies segregating for either of the two QTLs. One additional QTL, Rbcq4 on Chromosome 4 reducing DI, was identified in one of the BC2S1 progenies. F2 individuals, homozygous for the Rbcq2 and Rbcq4 alleles of S. habrochaites showed a reduction of DI by 48%. QTLs from S. habrochaites LYC4 offer good perspectives for breeding B. cinerea resistant tomato cultivars.

Resistance to Bemisia tabaci in tomato wild relatives

Bemisia tabaci is one of the most threatening pests in agriculture, particularly in Solanaceous crops such as tomato and pepper that are cultivated in the open field. Pesticide application is often not effective and hazardous to humans and environment. The exploitation of plant natural defenses that are present in wild relatives of tomato, may offer a solution. To evaluate resistance parameters and to identify plant material with high levels of resistance, we screened a number of accessions of tomato wild relatives using three methods; a free-choice test in a screenhouse in Indonesia, a no-choice test with clip-on cages in a greenhouse and a leaf disc test in a climate-room in the Netherlands. Antibiosis resulting in low adult survival was the major component for resistance in tomato. However, other resistance component(s) may play a role as well. In some accessions there was a change in the resistance level over time. Several resistance parameters used in the different tests were well correlated. The best resistance source was an accession of Solanum galapagense, which had not been identified as being resistant in the past. This is of particular interest as this species is closely related to the cultivated tomato, which may facilitate introgression of the resistance component(s). Whitefly non-preference and resistance were associated with the presence of type IV trichomes. Other mechanisms might be involved since some accessions without type IV trichomes showed low nymphal density. The leaf disc test is a good in vitro alternative for the clip-on cage whitefly resistance screening, as shown by the high correlation between the results obtained with this test and the clip-on cage test. This offers breeders the possibility to carry out tests more efficiently.

Tomato breeding in the genomics era: insights from a SNP array

BackgroundThe major bottle neck in genetic and linkage studies in tomato has been the lack of a sufficient number of molecular markers. This has radically changed with the application of next generation sequencing and high throughput genotyping. A set of 6000 SNPs was identified and 5528 of them were used to evaluate tomato germplasm at the level of species, varieties and segregating populations.ResultsFrom the 5528 SNPs, 1980 originated from 454-sequencing, 3495 from Illumina Solexa sequencing and 53 were additional known markers. Genotyping different tomato samples allowed the evaluation of the level of heterozygosity and introgressions among commercial varieties. Cherry tomatoes were especially different from round/beefs in chromosomes 4, 5 and 12. We were able to identify a set of 750 unique markers distinguishing S. lycopersicum ‘Moneymaker’ from all its distantly related wild relatives. Clustering and neighbour joining analysis among varieties and species showed expected grouping patterns, with S. pimpinellifolium as the most closely related to commercial tomatoesearlier results.ConclusionsOur results show that a SNP search in only a few breeding lines already provides generally applicable markers in tomato and its wild relatives. It also shows that the Illumina bead array generated data are highly reproducible. Our SNPs can roughly be divided in two categories: SNPs of which both forms are present in the wild relatives and in domesticated tomatoes (originating from common ancestors) and SNPs unique for the domesticated tomato (originating from after the domestication event). The SNPs can be used for genotyping, identification of varieties, comparison of genetic and physical linkage maps and to confirm (phylogenetic) relations. In the SNPs used for the array there is hardly any overlap with the SolCAP array and it is strongly recommended to combine both SNP sets and to select a core collection of robust SNPs completely covering the entire tomato genome.

Differences in insect resistance between tomato species endemic to the Galapagos Islands

BackgroundThe Galapagos Islands constitute a highly diverse ecosystem and a unique source of variation in the form of endemic species. There are two endemic tomato species, Solanum galapagense and S. cheesmaniae and two introduced tomato species, S. pimpinellifolium and S. lycopersicum. Morphologically the two endemic tomato species of the Galapagos Islands are clearly distinct, but molecular marker analysis showed no clear separation. Tomatoes on the Galapagos are affected by both native and exotic herbivores. Bemisia tabaci is an important introduced insect species that feeds on a wide range of plants. In this article, we address the question whether the differentiation between S. galapagense and S. cheesmaniae may be related to differences in susceptibility towards phloem-feeders and used B. tabaci as a model to evaluate this.ResultsWe have characterized 12 accessions of S. galapagense, 22 of S. cheesmaniae, and one of S. lycopersicum as reference for whitefly resistance using no-choice experiments. Whitefly resistance was found in S. galapagense only and was associated with the presence of relatively high levels of acyl sugars and the presence of glandular trichomes of type I and IV. Genetic fingerprinting using 3316 SNP markers did not show a clear differentiation between the two endemic species. Acyl sugar accumulation as well as the climatic and geographical conditions at the collection sites of the accessions did not follow the morphological species boundaries.ConclusionOur results suggest that S. galapagense and S. cheesmaniae might be morphotypes rather than two species and that their co-existence is likely the result of selective pressure.

Exploring the Natural Variation for Seedling Traits and Their Link with Seed Dimensions in Tomato

The success of germination, growth and final yield of every crop depends to a large extent on the quality of the seeds used to grow the crop. Seed quality is defined as the viability and vigor attribute of a seed that enables the emergence and establishment of normal seedlings under a wide range of environments. We attempt to dissect the mechanisms involved in the acquisition of seed quality, through a combined approach of physiology and genetics. To achieve this goal we explored the genetic variation found in a RIL population of Solanum lycopersicum (cv. Moneymaker) x Solanum pimpinellifolium through extensive phenotyping of seed and seedling traits under both normal and nutrient stress conditions and root system architecture (RSA) traits under optimal conditions. We have identified 62 major QTLs on 21 different positions for seed, seedling and RSA traits in this population. We identified QTLs that were common across both conditions, as well as specific to stress conditions. Most of the QTLs identified for seedling traits co-located with seed size and seed weight QTLs and the positive alleles were mostly contributed by the S. lycopersicum parent. Co-location of QTLs for different traits might suggest that the same locus has pleiotropic effects on multiple traits due to a common mechanistic basis. We show that seed weight has a strong effect on seedling vigor and these results are of great importance for the isolation of the corresponding genes and elucidation of the underlying mechanisms.

Identification and mapping of quantitative resistance to late blight (Phytophthora infestans) in Solanum habrochaites LA1777

Late blight (Phytophthora infestans) can have devastating effects on tomato production over the whole world. Most of the commercial cultivars of tomato, Solanum lycopersicum, are susceptible. Qualitative and quantitative resistance has been described in wild relatives of tomato. In general qualitative resistance can more easily be overcome by newly evolved isolates. Screening of three S. habrochaites accessions (LA1033, LA2099 and LA1777) through a whole plant assay showed that accession LA1777 had a good level of resistance to several isolates of P. infestans. To explore the potential in this wild species, an introgression line (IL) population of S. habrochaites LA1777 was used to screen individual chromosome regions of the wild species by a detached leaf assay. Two major isolates (T1,2 and T1,2,4) were used and two parameters were measured: lesion size (LS), and disease incidence (DI). Substantial variation was observed between the individual lines. QTLs were identified for LS but not for DI. The presence of five QTLs derived from LA1777 (Rlbq4a, Rlbq4b, Rlbq7, Rlbq8 and Rlbq12) results in unambiguous higher levels of resistance. All QTLs co-localized with previously described QTLs from S. habrochaites LA2099 except QTL Rlbq4b, which is therefore a novel QTL.