Deficitni princip namakanja oljčnih nasadov v Slovenski Istri

The olive tree has anatomical-morphological and physiological adaptations which enable it to cope well with dry conditions and water deficits. However, if water shortage occurs during the development phases (shoot growth, flower bud development, bloom, fruit set, cell division and enlargement and oil accumulation), which are the most susceptible to stress, it can also have a negative effect on the growth and productivity of olive trees. The Slovenian Istria is facing with increased risk of drought. Due to increased occurrence and intensity of agricultural droughts controlled deficit irrigation will become an inevitable element of agricultural practice in Slovene Istria.</span

Određivanje DNA otiska različitih sorta masline pomoću mikrosatelitskih markera

Microsatellites combine several features of an ultimate molecular marker and they are used increasingly in various plant genetic studies and applications. In this work we report on the utilisation of fourteen previously developed olive microsatellite markers for the identification and differentiation of a set of nineteen olive varieties. All analysed microsatellite markers revealed a high level of polymorphism that allowed unique genotyping of the examined varieties. Ninety-six alleles were detected at all 14 loci, which multiplied into a large number of observed genotypes, giving high discrimination value for varietal identification. A minimum number of three microsatellite markers was chosen for the rapid and unambiguous varietal identification of nineteen olive varieties and only two markers were sufficient for differentiation of five local varieties. DNA fingerprints of olive cultivars by means of microsatellites provided meaningful data, which can be extended by additional olive varieties or new microsatellites and used for accurate inter-laboratory comparison. The data obtained can be used for the varietal survey and construction of a database of all olive varieties grown in Slovenia providing also additional genetic information on the agronomic and quality characteristics of the olive varieties.U različitim genetičkim istraživanjima biljaka sve se više koriste mikrosateliti kao najpogodniji molekularni marker. U četrnaest analiziranih lokusa otkriveno je ukupno 96 alela, što omogućava vrlo veliku razlikovnost pri identifikaciji pojedinih sorta. Tipiziranjem genoma različitih sorta masline pomoću mikrosatelita dobivene su vrlo korisne informacije koje se mogu dopuniti uporabom dodatnih markera. Tom se metodologijom mogu karakterizirati i nove sorte maslina, a i obavljati pouzdane međulaboratorijske usporedbe rezultata. Dobiveni podaci mogu se koristiti pri izradbi baze podataka za identifikaciju pojedinih sorta i njihovu rasprostranjenost na području Slovenije. Oni također omogućavaju da se agronomske značajke i kakvoća različitih vrsta maslina dopune dodatnim genetičkim informacijama

Identification and validation of novel EST-SSR markers in olives

The olive (Olea europaea L.) is a leading oil crop in the Mediterranean area. Limited information on the inheritance of agronomic significant traits hinders progress in olive breeding programs, which encourages the development of markers linked to the traits. In this study, we report on the development of 46 olive simple sequence repeat (SSR) markers, obtained from 577,025 expressed sequence tags (ESTs) in developing olive fruits generated in the framework of the Slovenian national olive transcriptome project. Sequences were de novo assembled into 98,924 unigenes, which were then used as a source for microsatellites searching. We identified 923 unigenes that contained 984 SSRs among which dinucleotide SSRs (36 %) were the most abundant, followed by tri- (33 %) and hexa- (21 %) nucleotides. Microsatellite repeat motif GA (37 %) was the most common among dinucleotides, while microsatellite repeat motif GAA was the most abundant trinucleotide SSR motif (16 %). Gene ontology annotations could be assigned to 27 % of the unigenes. A hundred and ten expressed sequence tag-derived-simple sequence repeats (EST-SSRs) with annotated genes were selected for primer designing and finally, 46 (42 %) polymorphic EST-SSRs were successfully amplified and used to validate genetic diversity among 24 olive varieties. The average number of alleles per locus, observed heterozygosity, expected heterozygosity, and polymorphic information content were 4.5, 0.649, 0.604 and 0.539, respectively. Twenty-seven EST-SSRs showed good diversity properties and were recommended for further olive genome investigation

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

INFLUENCE OF MICROBIAL AND ORGANIC FERTILIZERS ON BACTERIAL COMMUNITIES COMPOSITION DURING KEY GROWTH PHENOPHASES OF MAIZE

INTRODUCTION Maize is among the three world’s most important and widely grown cereals (Seyi-Amole & Onilude, 2021). The excessive and long-term application of agrochemicals for providing maize with essential nutrients, required for the development of all growth phenophases and for yield enhancement, leaves adverse consequences for human health and the environment (Khaliq et al., 2004). Although their use is inevitable to meet the increasing demand of the growing human population for a healthy food supply, organic fertilizers and biofertilizers (microbial fertilizers) are becoming recognized as effective, economically feasible, and environmentally sound alternatives for sustainable agriculture (Lawal & Babalola, 2014; Hui et al., 2017; Mahanty et al., 2017). The main objective of this study was to evaluate the influence of the microbial inoculant Phytobiotic (PHY), containing a consortium of Bacillus subtilis sp. subtilis and Microbacterium sp., on native maize microbiome during key growth phenophases (seedling, flowering, and harvesting) under field conditions, as well as to compare whether differences in efficacy between PHY, poultry manure (PM) and their combination (PHY_PM) exist, based on yield parameters. STATE OF THE ART Seeds, roots, and soil samples were taken for metabarcoding analysis during four growth phenophases (I-IV). Samples of uninoculated seeds and soil, poultry manure, and seeds inoculated with PHY were primarily taken before sowing (phenophase I). Further, during the growing season [phenophases II (seedling), III (flowering), and IV (harvesting)] the effect of PHY, PM, and PHY_PM on maize seeds, roots, and soil microbiome was evaluated in relation to concurrently sampled negative controls. A total DNA from the collected samples was isolated, amplified with primers 515F/ 806R targeting the V4 region of the 16S rRNA, and subjected to next-generation sequencing (NGS). The obtained sequence data were bioinformatically processed and used for the evaluation of alpha and beta diversity. Yield and associated parameters (number of grown and fallen/broken plants, rating fence, plant vigor, the occurrence of Ustilago sp., and grain moisture) were evaluated after harvest. RESULTS Seeds exibited lower bacterial diversity compared to the soil, root, and manure samples.The most abundant taxon in uninoculated seeds pre-harvest was Pantoea, while in seeds treated with PHY the most abundant was Acinetobacter, followed by Pantoea, Pseudomonas, and Bacillus. After harvest, Pantoea and Pseudomonas prevearled in seeds. Soil bacterial communities mostly remained unchanged, regardless of the treatment (PHY, PM, and PHY_PM) applied or the tested phenophase, with uncultured Gaiellales and Bacillus being the most abundant. Contrarily, root bacterial communities differed in distribution and relative abundance of different taxa between phenophases and between treatments. The most abundant taxa in roots during the inial phenophase (II) was Pseudomonas. In the flowering phenophase (III), Bacillus prevealed with two to three times higher relative abundance in treatments with PHY or PM compared to the negative control, while Lechevalieria dominated in harvesting phenophase (IV). A statistically significant increase in maize yield was obtained in the treatment with PHY, with an average value of 650 kg/ha compared to the negative control. The lowest yield was obtained in the treatment with PM. DISCUSSION The prevalence of Acinetobacter, Pantoea, Pseudomonas, and Bacillus in seeds treated with PHY pre-harvest, indicates that treatment with PHY is highly beneficial considering the known plant growth promoting potential of these genera, that were also previously confirmed as core maize inhabitants (Mehta et al., 2021). As core members, Pantoea and Pseudomonas remained present after harvest. The benefit of the application of Bacillus-based fertilizers to soil is the enhancement of the plant-available forms of nutrients and the inducement of pest and pathogens defense systems (Radhakrishnan, et al., 2017). It is of crucial importance that none of the three treatments applied in this study affected the composition of the indigenous soil bacterial communities during four tested phenophases, which is highly important when selecting suitable agricultural practices. Shifts in root microbiome over maize growth could be related to the production of different root metabolites over the growing season (Bourceret et al., 2022). Roots were especially rich with genera (Pseudomonas, Stenotrophomonas, Sphingobacterium, Achromobacter) known as phosphate solubilizers (Mehta et al., 2021). Furthermore, Bacillus was dominant in roots in flowering phenophase. This genus is known for its wide spectrum of beneficial effects on plants, like phosphate solubilization, biosynthesis of growth hormones, antimicrobial activity, induction of systemic resistance, etc. (Dimkic et al., 2022). CONCLUSIONS Considering the above-mentioned effect of PHY on maize yield incensement, its non-disruption effect on the core microbiome, and the positive effect on enhancing the presence of beneficial bacterial genera, this microbial inoculant could be proposed as a promising alternative to chemicals and organic fertilizers in maize cultivation

Bioinoculants in sustainable agriculture – the influence on maize characteristics and phytobiome

Maize (Zea mays L.) occupies a significant part of the world cultivated area. Beneficial microorganisms and organic fertilizers are powerful tools for sustainable agricultural management. Applying appropriate formulation either through seed coating or composting material together with seeds during the sowing was aimed to achieve optimal performance on plants and increase the crop yields. Also, a complete analysis of phytobiome including high-throughput amplicon and whole-genome sequencing, and computational biology of the treated and non-treated plants' roots and soil was applied. The statistically significant difference in bacterial communities between phenophases and treatments was much more prevalent in the roots than soil itself. In general, seedling phenophase (II) differed in terms of diversity from bacterial communities of flowering (III) and harvesting phenophases (IV), regardless of treatment, indicating good colonization with the treated substrate for up to one month. In the root samples, the dominance of the genus Pseudomonas was evident, in the initial stages (phase II), while this percentage decreased linearly over time. High relative abundance of the genera Achromobacter, Sphingobacterium, and Stenotrophomonas also characterized II phenophase. Contrary, a relative abundance of the genus Bacillus was characteristic for III phenophase, and especially twice or three times higher in treatments with phytobiotic or manure compared to the negative control. Furthermore, it was observed that with increasing inoculation of Bacillus isolate, the percentage of Pantoea genus decreases drastically in coated seeds. In conclusion, a statistically significant increase in maize yield was obtained in the treatment with phytobiotic with an average value of 650 kg/ha compared to the negative control

Utjecaj tehnologije prerade na razvoj spontane mikrobne populacije i kakvoću stolnih maslina slovenskoga dijela Istre

The objective of the research is to determine the leading microorganisms in spontaneous fermentations of table olives in Slovenian Istria. The influence of traditional regional and modified Spanish style technology on yeast and lactic acid bacteria population dynamics and on the quality of Istrska belica and Storta table olive varieties was studied during 180 days of fermentation. Apart from that, pH of the brine during fermentation and total biophenols in olive fruits before processing and after 60 and 180 days of fermentation were determined. The quality of the final product was determined with sensory analysis. Table olive fermentation was carried out by yeasts. Aureobasidium pullulans, Cryptococcus adeliensis, Metschnikowia pulcherrima, Rhodotorula mucilaginosa, Pichia anomala and Candida oleophila were isolated and identified using PCR-RFLP analysis of ITS regions and traditional phenotypic tests. High initial amount of total biophenols in olive fruits and their better preservation during traditional processing influenced microbial population dynamics and quality characteristics of table olives. The modified Spanish style technology was not confirmed as suitable for retaining positive characteristics of the product of traditional regional technology in Slovenian Istria.Svrha je istraživanja bila odrediti glavne mikroorganizme koji sudjeluju u spontanoj fermentaciji stolnih sorata maslina, uzgojenih u slovenskom dijelu Istre. Tijekom 180 dana fermentacije ispitan je utjecaj tradicionalnog i modificiranog španjolskog načina prerade na dinamiku populacije kvasaca i mliječno-kiselih bakterija, te na kakvoću stolnih maslina Istrska belica i Storta. Osim toga, mjerena je pH-vrijednost rasola i određeni su ukupni biofenoli u neprerađenim plodovima te u plodovima nakon 60 i 180 dana fermentacije. Senzorskom je analizom ispitana kakvoća stolnih maslina pa je utvrđeno da kvasci prevladavaju tijekom njihove fermentacije. Kombinacijom molekularno-biološke metode, PCR-RFLP analizom ITS područja, i tradicionalne metode identifikacije kvasaca identificirane su ove vrste: Aureobasidium pullulans, Cryptococcus adeliensis, Metschnikowia pulcherrima, Rhodotorula mucilaginosa, Pichia anomala i Candida oleophila. Veliki udio ukupnih biofenola u plodovima prije prerade i u stolnim maslinama, pripravljenim na tradicionalan način, utječe na dinamiku populacije mikroorganizama i kakvoću stolnih maslina. Modificirani španjolski način prerade nije prikladan za očuvanje pozitivnih značajka tradicionalne prerade stolnih maslina, uzgojenih u slovenskom dijelu Istre

Genetička, biokemijska, hranjiva i antimikrobna svojstva šipka (Punica granatum L.) uzgojenog u Istri

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.U ovom je radu opisana genetička raznolikost germplazme šipka (Punica granatum L.) s područja slovenskog i hrvatskog dijela Istre. U etanolnim i vodenim ekstraktima različitih dijelova ploda šipka pronađeni su bioaktivni spojevi, te su ispitana njihova antioksidacijska i antimikrobna svojstva i preliminarno određena hranjiva svojstva ekstrakata. Dvadeset i šest različitih genotipova utvrđenih analizom mikrosatelita upućuju na veliku raznolikost šipka u Istri. Etanolni ekstrakti ploda šipka predstavljaju bogat izvor fenolnih spojeva (srednja vrijednost masenog udjela, izraženog kao galna kiselina, bila je 23 mg/g u egzokarpu i 16 mg/L u mezokarpu). Etanolni ekstrakti egzokarpa i mezokarpa šipka imali su snažan antimikrobni učinak na sljedeće vrste: Candida albicans, Candida parapsilosis, Rhodotorula mucilaginosa, Exophiala dermatitidis i Staphylococcus aureus, dok su vodeni ekstrakti egzokarpa i mezokarpa uspješno suzbijali rast bakterija S. aureus i Escherichia coli. Prema našim spoznajama, ovo je prva karakterizacija genetičkih izvora šipka iz Istre, koja obuhvaća molekularna, kemijska, antimikrobna i hranjiva svojstva plodova