Dunja Bandelj

Validation of candidate reference genes in RT-qPCR studies of developing olive fruit and expression analysis of four genes involved in fatty acids metabolism

Olive (Olea europaea L.) is an evergreen Mediterranean oil fruit tree with great economic, cultural and historical importance. For accurate gene expression studies of specific genes, reverse transcriptase–quantitative polymerase chain reaction (RT-qPCR) is often the method of choice, using suitable reference genes (RGs). This study identified RGs for RT-qPCR studies of developing olive fruit from 29 RG candidates. We used 12 sampling points to cover the five stages of olive fruit development. According to the results of the geNorm algorithm, the two best RGs were TIP41-like family protein (TIP41) and TATA binding protein (TBP), while several classical RGs proved not to be suitable. Using the two new RGs, four genes involved in the metabolism of fatty acids were studied and showed distinct expression patterns associated with mesocarp development and ripening stages. In addition to identifying two RGs for future analysis of gene expression in olive fruit, our results also provide a list of potential RGs that can be easily tested in other studies of olive gene expression in different developmental stages or in biologically challenged olive tissues. The results are also valuable for future research on genes that influence the synthesis and accumulation of olive fruit metabolites.

Characterization and defining of a core set of novel microsatellite markers for use in genotyping and diversity study of Adriatic fig (Ficus carica L.) germplasm

To support studies of the genetic diversity and population structure of Ficus carica L., 16 additional microsatellite markers were developed from genomic libraries enriched for GA/TC and GT/AC motifs. A total of 87 alleles were amplified in 83 analyzed fig samples. The number of alleles per locus ranged from three to eight, with a mean value of 5.44 alleles per locus. Mean values of observed and expected heterozygosity were 0.575 and 0.626, respectively. Calculated PIC values varied among loci in a range from 0.369 to 0.761. All markers were successfully cross-amplified in five closely related species belonging to the Moraceae family: Ficus (F. barteri Sprague, F. rubiginosa Desf. ex Vent., F. benjamina L.) and Morus (M. alba L. and M. nigra L.). These novel microsatellite markers represent a new valuable tool for further genome investigation, population studies, and management of genetic resources of wild, feral, and domesticated Ficus carica L. samples and other related species of Moraceae family, distributed mostly in tropical and subtropical regions throughout the world.

Paternity Analysis of the Olive Variety “Istrska Belica” and Identification of Pollen Donors by Microsatellite Markers

The leading olive variety in Slovenia is “Istrska belica” (Olea europaea L.), which currently represents 70% of all olive trees in productive orchards. Paternity analysis based on microsatellite markers was used for genotyping and identification of the potential pollen donors of “Istrska belica” and for assessing the proportion of self-fertilization in monovarietal olive orchards in the Slovene Istria. Seven microsatellite loci were used for genotyping thirty-one olive embryos from “Istrska belica” trees and for all potential pollen donor varieties, which are grown in the region and could participate as pollinators. Genotyping results and allele identification were performed using the FaMoz software. The most probable pollen donor was assigned to 39% of all analyzed embryos. Among all analyzed embryos no single case of self-fertilization was confirmed. According to the present results, the variety “Istrska belica” was in all cases fertilized by foreign pollen. The results will contribute to defining the new guidelines for farmers regarding the proper management and growing practice in monovarietal olive groves.

Population Fluctuation of Adult Males of the Olive Fruit Fly Bactrocera oleae (Rossi) Analysis in Olive Orchards in Relation to Abiotic Factors

ABSTRACT Very few publications have documented the effects of climatic conditions on population fluctuation of Bactrocera oleae in the northeast part of the Mediterranean Basin. A field study (2005-2009) was carried out to analyze the population dynamics of B. oleae in the northern Balkans and to determine the potential correlation of population fluctuations of S. oleae and fruit infestation with abiotic parameters. The results of this study showed that despite the fact that climatic conditions had little direct influence on the population dynamics of adult males of B. oleae, minimum temperatures and precipitation have a significant effect on fruit infestation by B. oleae in the northern Balkans. The results further indicate that the annual population dynamic of the B. oleae in the northern Balkans is triple-peaked and is different from those observed in the southern Balkans. Better knowledge of seasonal population fluctuations of B. oleae in the northern Balkans could optimize the timing of insecticide applications in integrated control and thus reduce control costs and the risk of non-target effects and environmental contamination.

DNA profiling of figs ( Ficus carica L.) from Slovenia and Californian USDA collection revealed the uniqueness of some North Adriatic varieties

A set of 23 local varieties from Slovenia and 218 fig accessions from Californian fig germplasm collection were compared to determine the identity of genotypes and their possible genetic relationships. Figs were genotyped using twelve microsatellite loci. One hundred alleles were identified over all microsatellite loci with an average of 8.33 alleles per locus and a polymorphic information content of 0.557 per locus. DNA genotyping demonstrated a relatively high level of genetic diversity between analysed figs. Comparison of fig genotypes from Slovenia and California demonstrated that only six Slovenian varieties shared identical DNA profiles with figs from the Californian collection, while the other 17 Slovenian varieties were unique and characteristic to the North Adriatic region. The information obtained will contribute to a better management of fig genetic resources.

Impact of weather conditions and drought stress on primary and secondary metabolites of olives from Slovenian Istra

Sugars, the primary metabolites of plants, play an important role in the process of olive ripening, as they provide energy for metabolic changes and serve as a source for the biosynthesis of fatty acids. Phenolic compounds, which are secondary metabolites, can be synthesized in plants as a reflection of stressful situations such as tissue damage and UV irradiation. The objective of this study was to assess the impact of different irrigation treatments (0, 15, 33, 40, and 100% crop evapotranspiration (ETc)) on primary metabolites (sugars, sugar alcohols) and secondary metabolites (phenolic compounds). In different phenological periods, the levels of oleuropein, sugars and sugar alcohols were determined in olives and leaf samples in the crop years 2014, 2015, and 2016. The levels of oleuropein in olive fruits and leaves were highest under the irrigation treatment of 40% ETc and decreased noticeably under the irrigation treatment of 100% ETc. The median levels of glucose were higher compared with the mannitol and fructose levels in olive fruits. Total determined sugars in olive fruits during maturation decreased. While significant correlations were found in olive fruits between determined sugars and sugar alcohols and different irrigation treatments, we did not observe any correlation between determined sugars and sugar alcohols in olive leaves and irrigation treatments. Our research confirmed the complexity of the content of oleuropein and sugars or sugar alcohols in the leaves and fruits of olive trees from Slovenian Istra, which depend on various factors, not just drought stress. Weaker and less obvious correlations between the studied compounds and drought stress might be a consequence of unusual rainfall in the studied crop years.

Different Quantities of Applied Water on Olea europaea L. Cultivated under Humid Conditions

AbstractDue to increased occurrence and intensity of agricultural droughts in humid Mediterranean regions, monitored irrigation is becoming an increasingly inevitable element of agricultural practi...

Genetic, Biochemical, Nutritional and Antimicrobial Characteristics of Pomegranate (Punica granatum L.) Grown in Istria

This study characterises the genetic variability of local pomegranate (Punica granatum L.) germplasm from the Slovenian and Croatian areas of Istria. The bioactive components and antioxidant and antimicrobial properties of ethanol and water extracts of different parts of pomegranate fruit were also determined, along with their preliminary nutritional characterisation. Twenty-six different genotypes identified with microsatellite analysis indicate the great diversity of pomegranate in Istria. The pomegranate fruit ethanol extracts represent rich sources of phenolic compounds (mean value of the mass fraction in exocarp and mesocarp expressed as gallic acid is 23 and 16 mg/g, respectively). The ethanol extracts of pomegranate exocarp and mesocarp showed the greatest antimicrobial activity against Candida albicans, Candida parapsilosis, Rhodotorula mucilaginosa, Exophiala dermatitidis and Staphylococcus aureus, and the same water extracts against S. aureus and Escherichia coli. To the best of our knowledge, this study represents the first report of the characterisation of pomegranate genetic resources from Istria at different levels, including the molecular, chemical, antimicrobial and nutritional properties.