Ana María Ramos-Cabrer

Variation in grafted European chestnut and hybrids by microsatellites reveals two main origins in the Iberian Peninsula

This is the first known large-scale molecular study of simple sequence repeats (SSR) loci based on samples from grafted trees found in the Iberian Peninsula and Canary Islands and hybrids. Interspecific hybrids resistant to ‘ink disease’ (Phytophthora spp.) were obtained in France, Portugal and Spain, although difficult to distinguish by morphology. This study focuses on genetic variation using 10 SSRs (11 unlinked loci) from clonally propagated cultivars (574 accessions) of European chestnut (Castanea sativa) and hybrids (71 accessions). They were compared with a representative sample of exotic chestnut species present in the Atlantic area, 47 accessions of Castanea crenata, 37 of Castanea mollissima and 33 of Castanea henryi. Accessions were analysed using a model-based Bayesian procedure (Structure), factorial correspondence analysis and analysis of molecular variance. The main chestnut species, hybrids and alien introgressions were differentiated. Two main origins of variability in European cultivated chestnut were found in the Iberian Peninsula, one in the North and a second in the Centre. Andalusian and Canary Island accessions could be assigned to both of these zones, which indicate that they could have been colonised with cultivars originating from either zone, in the case of the Canary Islands from the sixteenth century on.

Genetic variability and diversification process in local pear cultivars from northwestern Spain using microsatellites

This is the first known large-scale molecular study of simple sequence repeats loci based on pear cultivars from the northwestern Iberian Peninsula. Most of the Spanish pear crop (one of the largest in Europe) is based on the Spanish cultivar Blanquilla and various other foreign cultivars. However, local cultivars can still be found in old orchards in northwestern Spain. Between 1978 and 1981, the Centro de Investigaciones Agrarias de Mabegondo (Xunta de Galicia) established a Germplasm Bank of local pear cultivars containing 221 accessions. In the current study, these were analysed and compared with 20 commercial cultivars of Pyrus spp. using 19 microsatellites. We identified 127 genotypes out of 221 accessions with an average of 43% clonality in this collection. Genotypes were analysed using a model-based Bayesian procedure (Structure), factorial correspondence analysis and molecular variance analysis, and Jaccard coefficients were estimated. Four reconstructed populations were identified by Structure, one related to Asian cultivars, two to French and English cultivars and one to Galician cultivars. The four identified groups of pears had evolved independently. This study explains the diversification process in pear cultivars from northwestern cultivars based on hybridization (16%), selection of triploids (38%) and the introgression of commercial cultivars in the collection (4%).

Breeding Apple (Malus x Domestica Borkh)

The apple tree is a hybrid originating from a combination of wild species (Malus sieversii is supposed to be the main contributor). Growers at first selected the best specimens by seedlings, but when grafting was discovered as a mean of vegetative propagation, improvement in fruit quality became faster. Apple is cultivated in most of the temperate regions due to the fruit’s quality, its easiness to propagate, and its natural aptitude to bear. Apples are considered a healthy fruit, as the saying goes ‘an apple a day keeps the doctor away’. An apple tree can reach up to 10 m height above its own roots, having a globose canopy and the longevity between 60 and 100 years. Depending on the rootstock and the age of the tree, the roots can occupy between 2 and 104 m, although most frequently they range between 10 and 30 m (Atkinson 1980).

Morphology and microsatellites in Spanish apple collections

Summary The population of apple, Malus pumila Mill., indigenous to northwest Spain shows a wide range of ripening periods and fruit quality and is an unexploited resource for breeding programmes. The main purpose of this study was to fingerprint these accessions and to construct a molecular database including the cultivars commonly grown in Spain. A total of 77 indigenous and 26 international accessions were analysed using ten microsatellite [simple sequence repeat (SSR)] markers. Some of the main morphological and agronomic characteristics, such as fruit weight, colour, shape, sweetness in ºBrix, flavour type according to content of malic and tannic acids, and harvest time were recorded. The 77 Spanish cultivars showed a unique fingerprint. We propose to preserve these 77 genetically different cultivars in our Germplasm Bank, and recommend that 31 of them should be considered for a core collection to maintain all the diversity recorded. Some 29% of the cultivars showed three alleles at several loci and were confirmed, cytometrically, as triploids. These tended to produce larger fruit (15% by weight) than diploids. The use of ten polymorphic microsatellite markers provided a useful technique for fingerprinting apple cultivars in the Spanish germplasm resource and for indicating triploids. The fingerprints indicated that the famous triploid Dutch cultivar ‘Belle de Boskoop’ may be derived from an unreduced gamete of ‘Reinette de Caux’.

Genetic Relationship between Castanea sativa Mill. Trees from North-western to South Spain Based on Morphological Traits and Isoenzymes

This study is focused in the main chestnut growing areas from north- to south-western Spain. We studied 50 accessions corresponding to 22 cultivars named by local growers; 11 from Andalucía, 9 from Castilla-León, and 2 from Extremadura. These were compared with 22 types classified previously in Galicia (82 accessions). Ten morphological traits and five isoenzyme systems were used to classify them and establish genetic relationships. A lower genetic variability has been found in Extremadura (middle Spain) and Andalucía (south) than in Galicia (north western) which may be because of the lower importance of the crop in those areas and the geographic barriers between them. Galicia could be the origin of the types found in Castilla-León, Extremadura, and south-western Andalucía, and the types found in Central Andalucía were closer to those found in Castilla-León and Extremadura. Most accessions studied (117/132) could be classified positively, making 37 out of 44 cultivars found from northern to southern Spain. Three groups of synonymies were found in Castilla-León: ‘Brava’ and ‘Negral’, ‘Gallego’ and ‘Injerto’, and ‘Rapega’ and ‘Verdello’. Most of the Spanish types have been propagated as a principal clone. Variability within the types is important and must be considered to select the best clones as cultivars.

Analysis of the genetic diversity and structure of the Spanish apple genetic resources suggests the existence of an Iberian genepool

87 Pags.- 3 Tabls.- 5 Figs.- 1 Suppl. Fig.- 5 Suppl. Tabls. The definitive version is available at: http://onlinelibrary.wiley.com/journal/10.1111/(ISSN)1744-7348

New Synonyms and Homonyms for Cultivars from Northwestern Spain

The main grapevine germplasm bank of Vitis spp. at the Estación de Viticultura y Enología de Galicia in northwestern Spain includes 272 accessions. Eight SSRs differentiated 66 different genotypes, with 27 groups of synonyms and six homonyms. Brancellao Blanco (white berries) could not be differentiated from Brancellao (black berries). Eleven genotypes had not been previously described. The eight SSRs used in this study confirmed the existence of two groups of cultivars from Galicia: the Caíño group in the west and the Merenzao group in the east.

Database of European chestnut cultivars and definition of a core collection using simple sequence repeats

We have evaluated 271 accessions corresponding to 118 European cultivars, 96 from Spain, 16 from Italy, four from France and two from Portugal with the following objectives: (1) to provide a European database based on reference simple sequence repeats (SSRs) and (2) to define a core collection. A set of 24 highly polymorphic SSRs were used for the genetic analysis. Two main clusters were identified using a model-based Bayesian procedure, which correspond to Spanish and Italian cultivar clusters, with the latter showing a higher genetic diversity. An additional genetic substructure was observed among five different groups of cultivars. A core collection with a minimum of 37 cultivars was selected. We provided a database including 132 European accessions with unique genotypes evaluated with 24 SSRs as a reference for distinction, registering and traceability. Finally, we found that a core collection based on 14% of the total accessions conserves all allelic diversity.

Apple (Malus spp.) Breeding: Present and Future

Apple breeding has been extremely successful in providing a highly diverse fruit crop. Recent (>50 millions years ago) genome-wide duplication (GWD) resulted in the 17 chromosomes in the Pyreae and confirmed the origin of cultivated apple on Malus sieversii being the same species as M. × domestica. Malus, as many other species in the family Rosaceae, shows gametophytic self-incompatibility (GSI), which forces outcrossing. GSI at the pistil is regulated by extacellular ribonuclease, S-RNase, which is encoded by S locus. Growers and agronomists have provided multiple cultivars with different colors, shapes, resistances, climatic adaptation or industrial aptitudes. The aim in apple breeding was the combination of different kinds of resistance and good fruit quality to produce dessert cultivars and cultivars for processing. Some of the best of these cultivars display resistance to scab (Venturia inaequalis), mildew (Podosphaera leucotricha), fire blight (Erwinia amylovora), bacterial canker (Pseudomonas syringae), red spider mite (Panonychus ulmi), winterfrost and good fruit quality. Different scab resistance sources of wild species (Vf, Vr, VA) were combined in the new series of cultivars. Multiple efforts worldwide have conserved most of that variation, the pillar for the traditional and new techniques profiting from the analysis of the apple genome, the genome-wide association studies (GWAS), identifying SNPs and genes, the analysis of genes differentially expressed (GDE) identified by qRT-PCR and microarray analysis, and the recent molecular genetic tool CRISPR/Cas9 to edit and correct the genome.

Development of a standardized methodology for phenotypical characterizations in apple

4 Pags.- 2 Tabls. Articles derived from XIV EUCARPIA Symposium on Fruit Breeding and Genetics (Bologna,Italy. June 14-18 2015) . The definitive version is available at: http://www.actahort.org/index.htm