ReDivia - Repositorio Digital de l'Instit Valencià d'Investigacions Agràries
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Viroids as Companions of a Professional Career
Get PDFSince the early 1970s when "virus-like" agents were considered as the cause of two diseases (potato spindle tuber and citrus exocortis), their study and further characterization have been linked to the development and use of molecular biology tools. Sucrose density gradient centrifugation and polyacrylamide gel electrophoresis (PAGE) played a critical role in the pioneering studies of PSTVd and citrus exocortis viroid (CEVd). This was later modified by using other PAGEs (sequential PAGE, return PAGE, two-dimensional PAGE), and/or different staining methods (ethidium bromide, silver nitrate, etc.). Since then, disease-causing agents suspected to be viroids were usually subjected to a number of tests to define their: (i) Molecular nature (RNA or DNA; single stranded or double stranded; circular or linear RNA); (ii) molecular weight; (iii) secondary and tertiary structure. Further biological assays are also essential to establish the relationship of a viroid with plant disease and to fulfill Koch's postulates
PdMFS1 Transporter Contributes to Penicilliun digitatum Fungicide Resistance and Fungal Virulence during Citrus Fruit Infection
Get PDFA new Penicillium digitatum major facilitator superfamily (MFS) transporter (PdMFS1) was identified and functionally characterized in order to shed more light on the mechanisms underlying fungicide resistance. PdMFS1 can play an important role in the intensification of resistance to fungicides normally used in P. digitatum postharvest treatments. In the PdMFS1 disrupted mutants, a slight effect in response to chemical fungicides was observed, but fungicide sensitivity was highly affected in the overexpression mutants which became resistant to wide range of chemical fungicides. Moreover, P. digitatum knock-out mutants exhibited a lower rate of fungal virulence when infected oranges were stored at 20 degrees C. Disease symptoms were higher in the PdMFS1 overexpression mutants coming from the low-virulent P. digitatum parental strain. In addition, the gene expression analysis showed an induction of PdMFS1 transcription in all overexpression mutants regardless from which progenitor came from, and four-time intensification of the parental wild type strain during citrus infection reinforcing PdMFS1 role in fungal virulence. The P. digitatum MFS transporter PdMFS1 contributes not only to the acquisition of wide range of fungicide resistance but also in fungal virulence during citrus infection
Varietal Change Dominates Adoption of Technology in Spanish Citrus Production
Get PDFAfter describing the technology adopted and its influence on production and yield, the importance of varietal change, which is essential to meet demand, is underscored. The choice of a variety by growers depends on the expected price it will fetch along with the commercial and crop characteristics and the quality. Survival of a variety in the marketplace depends on the performance as expected and on the existence of other more competitive varieties. First, in Spanish citrus farming, the navel group predominate, with 41.10% of orange production, in which the main varieties are "Navelina" and "Lanelate." The group of late white oranges is also important. The mandarin orange group includes clementines and mandarin hybrids; lemons also feature prominently. In sweet oranges, the navel group and late white group has often been renewed with varieties from the same group. In very early clementines varieties, the survival of which is usually short, the supply of new varieties is excessive. Because of its commercial characteristics and harvesting time, the group with the highest expansion possibilities are the mandarin hybrids which are in demand by hypermarkets. In the introduction of varieties, in recent years we have seen varietal clubs being established, with area control of each variety and very strict rules on cultivation and marketing
Genome-wide analysis of AGO, DCL and RDR gene families reveals RNA-directed DNA methylation is involved in fruit abscission in Citrus sinensis
Get PDFBackground: Small RNAs regulate a wide variety of processes in plants, from organ development to both biotic and abiotic stress response. Being master regulators in genetic networks, their biogenesis and action is a fundamental aspect to characterize in order to understand plant growth and development. Three main gene families are critical components of RNA silencing: DICER-LIKE (DCL), ARGONAUTE (AGO) and RNA-DEPENDENT RNA POLYMERASE (RDR). Even though they have been characterized in other plant species, there is no information about these gene families in Citrus sinensis, one of the most important fruit species from both economical and nutritional reasons. While small RNAs have been implicated in the regulation of multiple aspects of plant growth and development, their role in the abscission process has not been characterized yet.Results: Using genome-wide analysis and a phylogenetic approach, we identified a total of 13 AGO, 5 DCL and 7 RDR genes. We characterized their expression patterns in root, leaf, flesh, peel and embryo samples using RNA-seq data. Moreover, we studied their role in fruit abscission through gene expression analysis in fruit rind compared to abscission zone from samples obtained by laser capture microdissection. Interestingly, we determined that the expression of several RNA silencing factors are down-regulated in fruit abscission zone, being particularly represented gene components of the RNA-dependent DNA Methylation pathway, indicating that repression of this process is necessary for fruit abscission to take place in Citrus sinensis.Conclusions: The members of these 3 families present characteristic conserved domains and distinct expression patterns. We provide a detailed analysis of the members of these families and improved the annotation of some of these genes based on RNA-seq data. Our data suggests that the RNA-dependent DNA Methylation pathway is involved in the important fruit abscission process in C. sinensis
The Altered Expression of microRNA408 Influences the Arabidopsis Response to Iron Deficiency
Get PDFMicroRNAs contribute to the adaptation of plants to varying environmental conditions by affecting systemic mineral nutrient homeostasis. Copper and iron deficiencies antagonistically control the expression of Arabidopsis thaliana microRNA408 (miR408), which post-transcriptionally regulates laccase-like multicopper oxidase family members LAC3, LAC12, and LAC13. In this work, we used miR408 T-DNA insertion mutants (408-KO1 and 408-KO2) and a previously characterized transgenic line overexpressing miR408 (35S:408-14) to explore how miR408 influences copper- and iron-dependent metabolism. We observed that the altered expression of miR408 diminished plant performance and the activation of the iron-regulated genes under iron-deficient conditions. Consistently with the low expression of the miR408-target laccases, we showed that the vascular bundle lignification of the 35S:408-14 plants diminished. The decrease in the phenoloxidase and ferroxidase activities exhibited by wild-type plants under iron deficiency did not occur in the 408-KO1 plants, probably due to the higher expression of laccases. Finally, we observed that the hydrogen peroxide levels under iron starvation were altered in both the 408-KO1 and 35S:408-14 lines. Taken together, these results suggest that Arabidopsis plants with modified miR408 levels undergo multiple deregulations under iron-deficient conditions
Effect of mass rearing on the genetic diversity of the predatory mite Amblyseius swirskii
Get PDFAmblyseius swirskii Athias-Henriot (Acari: Phytoseiidae) is a predatory mite used to control whiteflies and thrips in protected crops. This biocontrol agent, originating from the Eastern Mediterranean region, has been mass-reared for commercial use since 2005 and is widely used in augmentative biocontrol programs. As a polyphagous predator, it has to cope with different biotic and abiotic factors. However, possible adaptation to mass rearing for production might be hindering its resilience and capacity for optimum performance in the field. In this study, we investigated the effect of long-term mass rearing on the genetic diversity of A. swirskii. We identified six microsatellite loci from whole-genome nanopore sequencing of A. swirskii and used these in a comparative analysis of the genetic diversity and differentiation in eight wild populations collected from Israel in 2017 and a commercially available population. Our results indicate that the commercial population is 2.5x less heterozygous than the wild A. swirskii. Furthermore, the commercial population has the highest genetic differentiation from all the natural populations, as indicated by higher pairwise F-st values. Overall, we show that commercially reared A. swirskii have reduced genetic variation compared to their wild counterparts, which may reduce their performance when released to control pests in an integrated pest management (IPM) context
Assessing Ploidy Level Analysis and Single Pollen Genotyping of Diploid and Euploid Citrus Genotypes by Fluorescence-Activated Cell Sorting and Whole-Genome Amplification
Get PDFFlow cytometry is widely used to determine genome size and ploidy level in plants. This technique, when coupled with fluorescence-activated cell sorting (FACS), whole genome amplification and genotyping (WGA), opens up new opportunities for genetic studies of individualized nuclei. This strategy was used to analyze the genetic composition of single pollen nuclei of different citrus species. The flow cytometry and microscope observations allowed us to differentiate the populations of pollen nuclei present in the diploid and euploid genotypes analyzed, showing that citrus has binuclear pollen. We have identified in the "CSO" tangor an additional nuclei population composed by the vegetative plus generative nuclei. Genotyping of this nuclei population revealed that vegetative and generative nuclei show the same genetic configuration. In addition, we have demonstrated the presence of unreduced gametes in the diploid genotype "Mexican lime." Genomic amplification is a robust method for haploid nuclei genotyping with several molecular markers, whereas in diploid nuclei using heterozygous markers showed a bias towards one of the two alleles, limiting the use of this tool in this type of nuclei. We further discuss the importance and applications of single pollen genotyping in citrus genetic studies
Which came first: The disease or the pest? Is there a host mediated spread of Beauveria bassiana (Ascomycota: Hypocreales) by invasive palm pests?
Get PDFThe red palm weevil (RPW) Rhynchophorus ferrugineus (Olivier) (Coleoptera: Curculionidae) is threatening the palm family worldwide, causing important economic losses. Current tactics to manage the weevil are largely based on chemical control, although the use of pesticides is hampered by several environmental constraints. Since the first introduction of RPW in Spain in 1996 and during its progressive spread around the Mediterranean basin, the number of reports of natural infection of RPW populations by entomopathogenic fungi (EPF) has been rising for 15 years, and this rise could support a pest-mediated EPF spread. To challenge this hypothesis, we assessed the usefulness of the region of elongation factor 1-alpha (EF1-alpha), Bloc nuclear intergenic region (Bloc) and inter simple sequence repeat (ISSR) markers, alone or in combination, to infer the relationships among Mediterranean Beauveria and Metarhizium strains isolated from the RPW. Second, the effect of abiotic factors, such as temperature, humidity and UV-B radiation, on the germination and growth of these EPFs strains as a function of their genealogy and geographic origin were determined. Finally, the pathogenicity of strains from different genetic clades was evaluated against larvae and adults of R. ferrugineus. The phylogenetic analysis based on the EF-1 alpha gene identified eight different sequences among 24 fungal isolates of four fungal species. Similar clades were clustered when Bloc and ISSR analyses were performed. The results showed that strains of different origins were clustered in the same Glade, and this outcome could be explained by an RPW-mediated EPF spread that was also influenced by time, geographical and other RPW related factors. Neither the response to abiotic factors nor virulence to RPW larvae and adults were related to the sequence type, with all B. bassiana strains well adapted to Mediterraneam climatic conditions. Taken together, these findings may help to select the best strain for RPW management
Delaying berry ripening of Bobal and Tempranillo grapevines by late leaf removal in a semi-arid and temperate-warm climate under different water regimes
No full textBackground and Aims Climate change is advancing grape ripening and decoupling sugar and phenolic maturity, impacting wine typicity. The aim of this study was to test whether late leaf removal (LLR) under different watering regimes delayed harvest of two Spanish red cultivars in a semi-arid and temperate-warm climate. Methods and Results In two trials carried out in eastern Spain with the Bobal and Tempranillo cultivars, vines were partially defoliated above the bunch zone shortly before veraison under rainfed and deficit irrigation conditions during two seasons. The rate of grape ripening in both cultivars was significantly affected by LLR under either watering regime, consequently delaying harvest. Vine water status and leaf photosynthetic rate were improved by LLR. The reduction in leaf area-to-fruit ratio resulting from the LLR treatments was found to be more limiting for the accumulation of anthocyanin than for TSS. Consequently, LLR negatively affected wine colour intensity. In addition, yield was constrained by LLR in Tempranillo due to a reduction in bunch and berry mass. Conclusions The reduction in the rate of accumulation of grape TSS provoked by LLR did not necessarily result in a more balanced berry maturity. The effectiveness of the LLR technique appears to depend on its final impact on leaf area-to-fruit ratio and vine water status, the cultivar photosynthetic compensation capacity and the environmental conditions. Significance of the Study Late leaf removal might not be effective for coupling anthocyanin and TSS in berries under moderate water stress conditions, given the observed reductions in red wine colour
Influence of temperature on the progamic phase in Citrus
Get PDFTemperature in the progamic phase is critical for the success of plant sexual reproduction, and new knowledge is needed to optimise breeding programmes to obtain new varieties that adapt to a climate change scenario. Using three male donors and one female recipient in the genus Citrus, we evaluated the effect of four temperature regimes on each process in the progamic phase. An innovative method based on microscopic observations of cross sections from pollinated pistils collected daily allowed a comprehensive analysis of pollen tube growth (dynamics and kinetics) along the pistil. Pollen grain germination and stigmatic receptivity were evaluated directly on the stigma, which offers more accurate information than previously reported in vitro experiments. Our results showed that warm temperatures reduce the time needed by pollen tubes to reach ovules and accelerate pistil degeneration, while cold temperatures produce the opposite effects. Interestingly, we observed both pollen grain germination and pollen tube growth at 10 degrees C, which have not been observed in previous studies in citrus. At this temperature, the differences observed in both pollen grain germination and pollen tube growth for different genotypes reflect the adaptation of their sporophytic generation to low temperatures which would enable gametophytic screening to be used as a tool to select better adapted genotypes to different temperature conditions. The differences observed in the growth rates between pollen tubes in each genotype-temperature combination provide an opportunity to explore additional gametophytic selection in this reproductive phase. The capacity to respond to temperature changes in the progamic phase to ensure mating can be useful for breeding programs that focus on obtaining better adapted populations to different temperature conditions