Bayesian QTL analyses using pedigreed families of an outcrossing species, with application to fruit firmness in apple.

KEY MESSAGE: Proof of concept of Bayesian integrated QTL analyses across pedigree-related families from breeding programs of an outbreeding species. Results include QTL confidence intervals, individuals' genotype probabilities and genomic breeding values. Bayesian QTL linkage mapping approaches offer the flexibility to study multiple full sib families with known pedigrees simultaneously. Such a joint analysis increases the probability of detecting these quantitative trait loci (QTL) and provide insight of the magnitude of QTL across different genetic backgrounds. Here, we present an improved Bayesian multi-QTL pedigree-based approach on an outcrossing species using progenies with different (complex) genetic relationships. Different modeling assumptions were studied in the QTL analyses, i.e., the a priori expected number of QTL varied and polygenic effects were considered. The inferences include number of QTL, additive QTL effect sizes and supporting credible intervals, posterior probabilities of QTL genotypes for all individuals in the dataset, and QTL-based as well as genome-wide breeding values. All these features have been implemented in the FlexQTL(™) software. We analyzed fruit firmness in a large apple dataset that comprised 1,347 individuals forming 27 full sib families and their known ancestral pedigrees, with genotypes for 87 SSR markers on 17 chromosomes. We report strong or positive evidence for 14 QTL for fruit firmness on eight chromosomes, validating our approach as several of these QTL were reported previously, though dispersed over a series of studies based on single mapping populations. Interpretation of linked QTL was possible via individuals' QTL genotypes. The correlation between the genomic breeding values and phenotypes was on average 90 %, but varied with the number of detected QTL in a family. The detailed posterior knowledge on QTL of potential parents is critical for the efficiency of marker-assisted breeding.

Monitoring and occurrence of new races of Venturia inaequalis on apple in Belgium.

Since 1999, inside the framework of the European research project--FAIR5 CT97--3898 'Durable Apple Resistance in Europe'--a monitoring of scab infection on Leaves on 25 apple cuttivars (cvs)--including 10 differential hosts for scab races and European old cvs with partial scab resistance--was carried on trees in an unsprayed scab trap orchard planted at Gembloux, Belgium. The occurrence of scab races was determined by the use of a global assessment scale from 1 (no symptom) to 9 (maximum susceptibility) taking into account of both incidence and severity of the disease. Since 2004, scab was also scored according to a second evaluation scale based on qualitative symptoms. On the basis of the occurrence of symptoms on the various differential cvs, seven described physiological races of V. inaequalis were progressively identified in the orchard. During this period, important differences in incidence and severity of scab were observed on the different cvs. The susceptible cvs 'Golden Delicious', 'Gala' and 'Fiesta' were the most infected. On 13 polygenic varieties, sporulation was more or less important according to the disease pressure and their degree of susceptibility. The presence of race 7 on trees of Malus floribunda 821 has already been reported since 2000 year. Races 5, 6, 7 and 13 sporulated rather abundantly on their specific hosts according to the season's conditions although races 2 and 4 incited limited infection on their differential hosts and only under high disease pressure. First occurrence of race 3 in Belgium was observed in 2008.

A new versatile database created for geneticists and breeders to link molecular and phenotypic data in perennial crops: the AppleBreed DataBase.

OBJECTIVE: AppleBreed DataBase (DB) aims to store genotypic and phenotypic data from multiple pedigree verified plant populations (crosses, breeding selections and commercial cultivars) so that they are easily accessible for geneticists and breeders. It will help in elucidating the genetics of economically important traits, in identifying molecular markers associated with agronomic traits, in allele mining and in choosing the best parental cultivars for breeding. It also provides high traceability of data over generations, years and localities. AppleBreed DB could serve as a generic database design for other perennial crops with long economic lifespans, long juvenile periods and clonal propagation. RESULTS: AppleBreed DB is organized as a relational database. The core element is the GENOTYPE entity, which has two sub-classes at the physical level: TREE and DNA-SAMPLE. This approach facilitates all links between plant material, phenotypic and molecular data. The entities TREE, DNA-SAMPLE, PHENOTYPE and MOLECULAR DATA allow multi-annual observations to be stored as individual samples of individual trees, even if the nature of these observations differs greatly (e.g. molecular data on parts of the apple genome, physico-chemical measurements of fruit quality traits, and evaluation of disease resistance). AppleBreed DB also includes synonyms for cultivars and pedigrees. Finally, it can be loaded and explored through the web, and comes with tools to present basic statistical overviews and with validation procedures for phenotypic and marker data to certify data quality. AppleBreed DB was developed initially as a tool for scientists involved in apple genetics within the framework of the European project, 'High-quality Disease Resistance in Apples for Sustainable Agriculture' (HiDRAS), but it is also applicable to many other perennial crops.

Comparison of different techniques for inoculation of "Candidatus Phytoplasma mali" on apple and periwinkle in biological indexing procedure.

As phytoplasmas are non cultivable micro-organisms, the research on phytoplasmal diseases can only be achieved with infected hosts. Biological indexing (by grafting) is the simplest detection method for phytoplasmal diseases. We tested four different grafting techniques for inoculation of apple trees or periwinkles in greenhouse, including whip graft, bark graft, budding and chip-budding. All techniques were tested on apple trees (six trees per phytoplasma isolates) in insect-proof greenhouse. The whip and bark grafting were not feasible for periwinkle plants, because of fineness and fragility of their tissues: only the chip-budding was performed (four plants per isolate). In apple trees, the best and soonest positive results were obtained by chip and bark grafting. Except for seven transplants not-grown after grafting, 100% efficiency of inoculation was obtained by both methods. Nevertheless, the transmission of phytoplasma from transplant not-grown to rootstock was sometimes recorded (28.6%). The earliest phytoplasma symptoms after whip or bark grafting appeared after 3 months. Symptoms were obtained much later with budding and chip-budding. In case of periwinkles, infected apple and periwinkle materials were used as inoculum sources. Transmission of phytoplasma from periwinkle to periwinkle was successfully carried out by chip-budding grafting. The symptoms were observed during the second month after inoculation. The transmission of phytoplasma from infected apple material to periwinkle (by chip-budding) was achieved for 60 % of the tested samples. Moreover, the latency period before symptom observation was longer. Finally, we perceived the apple trees are more convenient and rapid than periwinkle plants for biological indexing of apple materials.

Influence of supplementary lighting during artificial scab inoculation tests in an apple breeding programm focused on partial resistance.

In 1988, the Department of Biological Control and Plant Genetic Resources at the Walloon Agricultural Research Centre started an apple-breeding programme using local genetic resources and modern varieties. Our objective is to create high quality commercial cultivars with durable resistance to scab (Venturia inaequalis), powdery mildew (Podosphaera leucotricha) and canker (Nectria galligena). The breeding strategy is based on crossing old apple cultivars and landraces selected as parents for low disease susceptibility and possessing other desirable horticultural characteristics. The programme aims to develop an early and efficient selection methodology adapted to partial disease resistance. One of the objectives is to define the optimal screening limit for discarding individuals after artificial scab inoculation tests. Working with large populations of seedlings entails spacing the seedling scab tests throughout the year. In order to work during winter, seedlings were grown in controlled cabinet conditions and in a glasshouse with supplementary lighting. To assess the bias introduced by these conditions, two trials were conducted: the first one to compare the influence of both environments on the results of scab inoculation tests, and the second one to assess the influence of the duration of supplementary lighting. The results enabled us to evaluate the limits of artificial cultural systems.

Use of fungicides against powdery mildew during an artificial scab inoculation test in an apple breeding programme focused on partial resistance.

Our project aimed to develop an effective selection methodology for partial disease resistance. One of its objectives was to define the optimal selection threshold for discarding individual seedlings after artificial scab inoculation tests. Working with large populations of seedlings, it was necessary to conduct the scab tests at intervals throughout the season, with the risk of powdery mildew infection of the young plantlets during the summer. Because Podosphaera leucotricha symptoms prevent breeders from properly evaluating scab symptoms, a trial was conducted to assess the effect of specific anti-mildew fungicides on the results of the scab inoculation test. The results allowed the active ingredient and the optimum time for application in relation to the inoculation date to be determined.

First evidence of the breakdown in Belgium of the scab resistance conferred by the Vf gene.

Scab is the most important fungal disease of apple throughout the world. The main breeding programmes are focused on scab resistance conferred by monogenic sources of resistance, specifically the Vf gene originated from Malus floribunda 821. Since 1993, evidence of the breakdown of this resistance mechanism appeared in different countries like United Kingdom, Germany, The Netherlands and France. In 1997, a first evidence of the Vf gene breakdown in Belgium was observed on the cultivar 'Judeline' planted in an untreated orchard at Aubel where a cider making company, with our collaboration, search for cultivars adapted for juice and cider processing. Therefore, a glasshouse experiment was planned in order to characterize which scab race was present. In 1999, the artificial inoculation of some cultivars in controlled conditions showed that the scab race collected the year before at Aubel presented the same characteristics than race 7 described by other works. At the same time, an orchard was established by our Department as planned in an European FAIR project named 'Durable Apple Resistance in Europe' (D.A.R.E) with the aim to study the resistance durability of some partial scab resistant apple cultivars inside an European orchard network. Sixteen repetitions of 20 different cultivars are planted and are originating from 8 participating countries. Five apple clones with specific host resistance towards 4 specific scab races are also present. Incidence and severity assessments of the disease are made 3 times a year on the whole tree according to two international standardized scales. For the first time in 2000, a specific scab race was observed on Malus floribunda 821 which caused very serious injuries. Those results confirm that new breeding strategies must be developed for reducing the risk of scab resistance breakdown in the future.