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The release of new olive cultivars with an increased squalene content in their virgin olive oil is considered an important target in olive breeding programs. In this work, the variability of the squalene content in a core collection of 36 olive cultivars was first studied, revealing two olive cultivars, 'Dokkar' and 'Klon-14', with extremely low and high squalene contents in their oils, respectively. Next, four cDNA sequences encoding squalene synthases (SQS) were cloned from olive. Sequence analysis and functional expression in bacteria confirmed that they encode squalene synthases. Transcriptional analysis in distinct olive tissues and cultivars indicated that expression levels of these four SQS genes are spatially and temporally regulated in a cultivar-dependent manner and pointed to OeSQS2 as the gene mainly involved in squalene biosynthesis in olive mesocarp and, therefore, in the olive oil. In addition, the biosynthesis of squalene appears to be transcriptionally regulated in water-stressed olive mesocarp.

Olive is one of the most cultivated species in the Mediterranean Basin and beyond. Despite being extensively studied for its commercial relevance, the origin of cultivated olive and the history of its domestication remain open questions. Here, we present a genealogical and kinship relationships analysis by mean of chloroplast and nuclear markers of different genera, subgenus, species, subspecies, ecotypes, cultivated, ancient and wild types, which constitutes one of the most inclusive research to date on the diversity within Olea europaea species. A complete survey of the variability across the nuclear and plastid genomes of different genotypes was studied through single nucleotide polymorphisms, indels (insertions and deletions), and length variation.

The fruit size of a cultivated olive tree is consistently larger than its corresponding wild relatives because fruit size is one of the main traits associated with olive tree domestication. Additionally, large fruit size is one of the main objectives of modern olive breeding programs. However, as the long juvenile period is one main hindrance in classic breeding approaches, obtaining genetic markers associated with this trait is a highly desirable tool. For this reason, GWAS analysis of both genetic markers and the genes associated with fruit size determination, measured as fruit weight, was herein carried out in 50 genotypes, of which 40 corresponded to cultivated and 10 to wild olive trees. As a result, 113 genetic markers were identified, which showed a very high statistically significant correlation with fruit weight variability, p < 10−10. These genetic markers corresponded to 39 clusters of genes in linkage disequilibrium. The analysis of a segregating progeny of the cross of “Frantoio” and “Picual” cultivars allowed us to confirm 10 of the 18 analyzed clusters. The annotation of the genes in each cluster and the expression pattern of the samples taken throughout fruit development by RNAseq enabled us to suggest that some studied genes are involved in olive fruit weight determination.

In Spain, several local studies have highlighted the likely presence of unknown olive cultivars distinct from the approximately 260 ones previously described in the literature. Furthermore, recent advancements in identification techniques have significantly enhanced in terms of efficacy and precision. This scenario motivated a new nationwide prospecting effort aimed at recovering and characterizing new cultivated germplasm using high-throughput molecular markers. In the present study, the use of 96 EST-SNP markers allowed the identification of a considerable amount of new material (173 new genotypes) coming from areas with low intensification of production in different regions of Spain. As a result, the number of distinct national genotypes documented in the World Olive Germplasm Bank of IFAPA, Córdoba (WOGBC-ESP046) increased to 427. Likewise, 65 and 24 new synonymy and homonymy cases were identified, respectively. This rise in the number of different national cultivars allowed to deepen the knowledge about the underlying genetic structure. The great genetic variability of Spanish germplasm was confirmed, and a new hot spot of diversity was identified in the northern regions of La Rioja and Aragon. Analysis of the genetic structure showed a clear separation between the germplasm of southern and northern-northeastern Spain and indicated a significantly higher level of admixture in the latter. Given the expansion of modern olive cultivation with only a few cultivars, this cryptic germplasm is in great danger of disappearing. This underlines the fact that maintaining as many cultivars as possible will increase the genetic variability of the olive gene pool to meet the future challenges of olive cultivation.

Olive, the emblematic Mediterranean fruit crop, owns a great varietal diversity, which is maintained in ex situ field collections, such as the World Olive Germplasm Bank of Córdoba (WOGBC), Spain. Accurate identification of WOGBC, one of the world's largest collections, is essential for efficient management and use of olive germplasm. The present study is the first report of the use of a core set of 96 EST-SNP markers for the fingerprinting of 1273 accessions from 29 countries, including both field and new acquired accessions. The EST-SNP fingerprinting made possible the accurate identification of 668 different genotypes, including 148 detected among the new acquired accessions. Despite the overall high genetic diversity found at WOGBC, the EST-SNPs also revealed the presence of remarkable redundant germplasm mostly represented by synonymy cases within and between countries. This finding, together with the presence of homonymy cases, may reflect a continuous interchange of olive cultivars, as well as a common and general approach for their naming. The structure analysis revealed a certain geographic clustering of the analysed germplasm. The EST-SNP panel under study provides a powerful and accurate genotyping tool, allowing for the foundation of a common strategy for efficient safeguarding and management of olive genetic resources.

Los microorganismos endófitos representan una estrategia innovadora para el manejo de las enfermedades vasculares en el olivo actuando como una defensa natural innata para hacer frente a la infección por patógenos que colonizan el xilema como son Verticillium dahliae y Xylella fastidiosa. Sin embargo, pocos estudios han caracterizado el efecto de la variedad y la edad de la planta en la estructura y diversidad microbiana del xilema del olivo. Por lo tanto, este trabajo se ha centrado en la caracterización mediante un enfoque metagenómico de las comunidades bacterianas y fúngicas presentes en el fluido xilemático de plantones jóvenes y olivos adultos de las variedades de olivo ‘Picual’, ‘Arbequina’, ‘Arbosana’, ‘Frantoio’ y ‘Hojiblanca’. Se han identificado un total de 440 ASVs bacterianos y 96 ASVs fúngicos que variaron en función de la edad de la planta alcanzando una diversidad (alfa) microbiana mayor en plantas adultas que en los plantones jóvenes y presentando una estructura microbiana (beta-diversidad) diferente en función de la edad, pero no de la variedad. Por otra parte, las variedades ‘Arbosana’ y ‘Hojiblanca’ mostraron una mayor riqueza bacteriana y fúngica, respectivamente. Globalmente, Meiothermus (24.61%), Anoxybacillus (18.21%) y Paenibacillus (13.15%) fueron los géneros bacterianos más representativos mientras que Aureobasidium fue el hongo más abundante (50.51%), seguido de Plectosphaerella (6.94%) y Malassezia (6.38%). Estos resultados contribuyen a mejorar la caracterización y el conocimiento de la variabilidad del microbioma xilemático de olivo, generando una información muy útil para optimizar el manejo de las enfermedades vasculares del olivo mediante el uso de sus propios microrganismos endofíticos como agentes control biológico.

[Background] Olive orchards are threatened by a wide range of pathogens. Of these, Verticillium dahliae has been in the spotlight for its high incidence, the difficulty to control it and the few cultivars that has increased tolerance to the pathogen. Disease resistance not only depends on detection of pathogen invasion and induction of responses by the plant, but also on barriers to avoid the invasion and active resistance mechanisms constitutively expressed in the absence of the pathogen. In a previous work we found that two healthy non-infected plants from cultivars that differ in V. dahliae resistance such as ‘Frantoio’ (resistant) and ‘Picual’ (susceptible) had a different root morphology and gene expression pattern. In this work, we have addressed the issue of basal differences in the roots between Resistant and Susceptible cultivars., [Results] The gene expression pattern of roots from 29 olive cultivars with different degree of resistance/susceptibility to V. dahliae was analyzed by RNA-Seq. However, only the Highly Resistant and Extremely Susceptible cultivars showed significant differences in gene expression among various groups of cultivars. A set of 421 genes showing an inverse differential expression level between the Highly Resistant to Extremely Susceptible cultivars was found and analyzed. The main differences involved higher expression of a series of transcription factors and genes involved in processes of molecules importation to nucleus, plant defense genes and lower expression of root growth and development genes in Highly Resistant cultivars, while a reverse pattern in Moderately Susceptible and more pronounced in Extremely Susceptible cultivars were observed., [Conclusion] According to the different gene expression patterns, it seems that the roots of the Extremely Susceptible cultivars focus more on growth and development, while some other functions, such as defense against pathogens, have a higher expression level in roots of Highly Resistant cultivars. Therefore, it seems that there are constitutive differences in the roots between Resistant and Susceptible cultivars, and that susceptible roots seem to provide a more suitable environment for the pathogen than the resistant ones.

Fatty acid composition of olive oil has an important effect on the oil quality to such an extent that oils with a high oleic and low linoleic acid contents are preferable from a nutritional and technological point of view. In the present work, we have first studied the diversity of the fatty acid composition in a set of eighty-nine olive cultivars from the Worldwide Olive Germplasm Bank of IFAPA Cordoba (WOGBC-IFAPA), and in a core collection (Core-36), which includes 28 olive cultivars from the previously mentioned set. Our results indicate that oleic and linoleic acid contents displayed the highest degree of variability of the different fatty acids present in the olive oil of the 89 cultivars under study. In addition, the independent study of the Core-36 revealed two olive cultivars, Klon-14 and Abou Kanani, with extremely low and high linoleic acid contents, respectively. Subsequently, these two cultivars were used to investigate the specific contribution of different fatty acid desaturases to the linoleic acid content of mesocarp tissue during olive fruit development and ripening. Fatty acid desaturase gene expression levels, together with lipid analysis, suggest that not only OeFAD2-2 and OeFAD2-5 but also the different specificities of extraplastidial acyltransferase enzymes are responsible for the variability of the oleic/linoleic acid ratio in olive cultivars. All this information allows for an advancement in the knowledge of the linoleic acid biosynthesis in different olive cultivars, which can impact olive breeding programs to improve olive oil quality.