Sean Myles’s research while affiliated with Dalhousie University and other places

Apples (Malus X. domestica Borkh.) are an economically important fruit species and are primarily diploid, although other ploidy levels exist. The impact of ploidy on agricultural traits in apple is not well understood but is an important factor to consider for breeding and production of apples. We used heterozygosity to infer ploidy for 970 apple accessions from a diverse collection. Next, we contrasted inferred diploid and inferred triploid apples across 10 agriculturally important traits. After correction for multiple testing, we determined that triploids have significantly higher phenolic content, but do not significantly differ from diploids for any other trait. We also determined that triploid varieties have significantly earlier release dates than diploids, suggesting that contemporary breeding programs primarily release diploid varieties. Ultimately, our results provide evidence that phenotypic differences between trees of differing ploidy are subtle and often insignificant and that there are limited measurable benefits to the use of triploids for apple production.

Premise Fruit quality is crucial in breeding new apple accessions. Before tasting, consumers assess freshness and flavour based on the physical appearance of fruit. Understanding how fruit quality traits such as shape and size vary across diverse apples provides a foundation for future breeding efforts. Methods We analyzed images of 5724 apples representing 743 different trees and 534 unique accessions from Canada’s Apple Biodiversity Collection to quantify variation in fruit shape and size. To achieve this, we used a pseudo-landmarking approach paired with traditional linear measurements including length, width, area, solidity, and aspect ratio. We also incorporated previously collected fruit weight measurements from the same trees. Results Using a comprehensive measure of shape, we determined that the primary source of variation in apple fruit shape was the width to length (aspect) ratio of the fruit. This variation was not significantly correlated with differences in fruit size from its area and harvest weight. Conclusions Our findings indicate that two critical aspects of morphological variation in apple—fruit shape and size—are independent, suggesting it is possible to select for a diverse range of fruit shapes while maintaining a consistent and marketable size.

Apples (Malus X. domestica Borkh.) are an economically important fruit species and the focus of continuing breeding efforts around the world. While most apple varieties are diploid, ploidy levels vary across the species, and triploids may be used in breeding despite poor fertility. The impact of ploidy on agricultural traits in apple is not well understood but is an important factor to consider when breeding new apple varieties. Here, we use mean heterozygosity values to categorize 970 apple accessions as diploid or triploid and then contrast apples of varying ploidy levels across 10 agriculturally important traits with sample sizes ranging from 427 to 928 accessions. After correction for multiple testing, we determine that triploids have significantly higher phenolic content. By examining historical release dates for apple varieties, our findings suggest that contemporary breeding programs are primarily releasing diploid varieties, and triploids tend to be older varieties. Ultimately, our results suggest that phenotypic differences between diploids and triploids are subtle and often insignificant indicating that triploids may not provide substantial benefit above diploids to apple breeding programs.

Apple quality traits such as fruit texture, sugar content, and firmness retention during storage are key targets for breeders. Understanding the genetic control of fruit quality traits can enable the development of genetic markers, useful for marker-assisted breeding of new apple cultivars. We made use of over 260,000 single nucleotide polymorphisms (SNPs) genotyped across 1,054 apple accessions from Canada's Apple Biodiversity Collection to perform genome-wide association for 21 fruit quality and phenology traits. We identified two loci on chromosome 15 and 16 associated with phenolic content and a locus on chromosome 10 associated with softening. In addition, we determined that allelic variation at the NAC18.1 transcription factor was associated with numerous traits including harvest date, firmness at harvest, and firmness after storage. Our analyses suggest that NAC18.1 independently acts as a high level regulator of multiple ripening related traits and we propose a model for the allelic effects at NAC18.1 on apple ripening and softening.

An apple's aroma is a major determinant of its desirability by consumers. To better understand the aroma of apples, 2-dimensional gas-chromatography mass-spectrometry (2D-GCMS) was used to quantify 106 volatile organic compounds (VOCs) from 515 apple varieties. We identified esters and aldehydes as the most abundant classes of VOCs, with butyl acetate and hexyl acetate being present in nearly every variety. Principal component analysis (PCA) revealed that the primary axis of variation in the apple volatilome is correlated with harvest date, with early-harvested apples expressing a greater number and higher concentration of VOCs compared to late-harvested apples. Genome-wide association studies (GWAS) using 250,579 single nucleotide polymorphisms (SNPs) identified a significant association between SNPs near the alcohol acyltransferase (AAT1) locus and the abundance of several esters. Additionally, strong associations were observed between SNPs at the NAC18.1 transcription factor locus and the abundances of 1-hexanol and 1-butanol, which serve as precursors for hexyl acetate and butyl acetate, respectively. These findings provide a foundation for understanding the genetic basis of apple aroma production and pave the way for the genomics-assisted enhancement of the aroma profiles of apple varieties to meet consumer preferences.

Ripening time, softening, and phenolic content are phenotypes of considerable commercial importance in apples. Identifying causal genetic variants controlling these traits not only advances marker-assisted breeding, but it is also an essential step for the application of gene editing technologies in apples. To advance the discovery of genetic variants associated with these phenotypes, we examined allele frequency differences between groups of phenotypically extreme individuals from Canada’s Apple Biodiversity Collection using pooled whole genome sequencing (pool-seq). We sequenced pooled DNA samples to an average read depth of 150x and scanned the genome for allelic differentiation between pools. For each phenotype, we identified >20 million genetic variants and identified numerous candidate genes. We identified loci on chromosomes 3 and 4 associated with ripening time, the former suggesting that regulatory variants upstream of a previously identified transcription factor NAC18.1 may be causal. Our analysis identified candidate regions on chromosomes 4, 8, and 16 associated with phenolic content, and suggested a cluster of UDP-Glycosyltransferase family genes as candidates for polyphenol production. Further, we identified regions on chromosomes 17 and 10 associated with softening and suggest a Long-chain fatty alcohol dehydrogenase family gene as putatively causal.

Societal Impact Statement Apples are among the most widely consumed fruits in the world, with a third of all apples being pressed into apple juice or fermented into cider. Cider has grown in popularity in Canada and the United States, and North American cider makers are increasingly interested in using traditional European ‘cider apples’ rather than using commonly grown ‘dessert apples’ that are grown primarily for fresh consumption. While we find that commonly grown dessert apples do differ from European cider apples for a small number of cider making characteristics, our results show that dessert apples are most often indistinguishable from cider apples across 10 traits. Our work provides a first step towards quantifying the differences between these two apple types.

An understanding of the relationship between the cultivated apple (Malus domestica) and its primary wild progenitor species (M. sieversii) not only provides an understanding of how apples have been improved in the past, but may be useful for apple improvement in the future. We measured 10 phenotypes in over 1000 unique apple accessions belonging to M. domestica and M. sieversii from Canada’s Apple Biodiversity Collection. Using principal components analysis (PCA), we determined that M. domestica and M. sieversii differ significantly in phenotypic space and are nearly completely distinguishable as two separate groups. We found that M. domestica had a shorter juvenile phase than M. sieversii and that cultivated trees produced flowers and ripe fruit later than their wild progenitors. Cultivated apples were also 3.6 times heavier, 43% less acidic, and had 68% less phenolic content than wild apples. Using historical records, we found that apple breeding over the past 200 years has resulted in a trend towards apples that have higher soluble solids, are less bitter, and soften less during storage. Our results quantify the significant changes in phenotype that have taken place since apple domestication, and provide evidence that apple breeding has led to continued phenotypic divergence of the cultivated apple from its wild progenitor species.

The apple ( Malus domestica ) is one of the world’s most commercially important perennial crops and its improvement has been the focus of human effort for thousands of years. Here, we genetically characterise over 1000 apple accessions from the United States Department of Agriculture (USDA) germplasm collection using over 30,000 single-nucleotide polymorphisms (SNPs). We confirm the close genetic relationship between modern apple cultivars and their primary progenitor species, Malus sieversii from Central Asia, and find that cider apples derive more of their ancestry from the European crabapple, Malus sylvestris , than do dessert apples. We determine that most of the USDA collection is a large complex pedigree: over half of the collection is interconnected by a series of first-degree relationships. In addition, 15% of the accessions have a first-degree relationship with one of the top 8 cultivars produced in the USA. With the exception of ‘Honeycrisp’, the top 8 cultivars are interconnected to each other via pedigree relationships. The cultivars ‘Golden Delicious’ and ‘Red Delicious’ were found to have over 60 first-degree relatives, consistent with their repeated use by apple breeders. We detected a signature of intense selection for red skin and provide evidence that breeders also selected for increased firmness. Our results suggest that Americans are eating apples largely from a single family tree and that the apple’s future improvement will benefit from increased exploitation of its tremendous natural genetic diversity.