Perennial crops, such as fruit trees, are infected by many viruses, which are transmitted through vegetative propagation and grafting of infected plant material. Some of these pathogens cause severe crop losses and often reduce the productive life of the orchards. Detection and characterization of these agents in fruit trees is challenging, however, during the last years, the wide application of high-throughput sequencing (HTS) technologies has significantly facilitated this task. In this review, we present recent advances in the discovery, detection, and characterization of fruit tree viruses and virus-like agents accomplished by HTS approaches. A high number of new viruses have been described in the last 5 years, some of them exhibiting novel genomic features that have led to the proposal of the creation of new genera, and the revision of the current virus taxonomy status. Interestingly, several of the newly identified viruses belong to virus genera previously unknown to infect fruit tree species (e.g., Fabavirus, Luteovirus) a fact that challenges our perspective of plant viruses in general. Finally, applied methodologies, including the use of different molecules as templates, as well as advantages and disadvantages and future directions of HTS in fruit tree virology are discussed.

Naturally contaminated basil seeds were treated by a surface dielectric barrier discharge driven in the humid air by an amplitude modulated AC high voltage to avoid heat shock. In order to avoid direct contact of seeds with microdischarge filaments, the seeds to be treated were placed at sufficient distance from the surface discharge. After treatment, the seeds were analyzed in comparison with control samples for their microbial contamination as well as for the capability of germination and seedling growth. Moreover, chemical modification of seed surface was observed through the elemental energy dispersive x-ray analysis and wettability tests. We found that treatment applied at 20% duty cycle (effective discharge duration up to 20 s) significantly decreases microbial load without reducing the viability of the seeds. On the other side, seedling growth was considerably accelerated after the treatment, and biometric growth parameters of seedlings (total length, weight, leaf extension) considerably increased compared to the controls. Interestingly, scanning electron microscopy images taken for the different duration of treatment revealed that seed radicle micropylar regions underwent significant morphological changes while the coat was substantially undamaged. Inside the seed, the embryo seemed to be well preserved while the endosperm body was detached from the epithelial tegument. A total of 9 different genera of fungi were recovered from the analyzed seeds. Scanning electron microscopy images revealed that conidia were localized especially in the micropylar region, and after plasma treatment, most of them showed substantial damages. Therefore, the overall effect of the treatment of naturally contaminated seeds by reactive oxygen and nitrogen species produced by plasma and the consequent changes in surface chemistry and microbial load can significantly improve seed vigor.

Basil ( Ocimum basilicum ) seeds were treated for different exposure times with a non-equilibrium plasma produced by a volume dielectric barrier discharge in humid air at atmospheric pressure. Plasma treatment did not change the seed structure and morphology, as visualized by high-resolution computed x-ray microtomography. A faster and higher germination rate was observed with plasma treatment of 1 and 3 min. Plantlets grown in sand, after both 2 and 3 weeks, showed a more developed root apparatus and better biometric parameters, compared to plants developing from non-treated seeds. After the plasma treatment, internal redistribution of macro and micronutrients was observed by using micro x-ray fluorescence spectroscopy. In particular P, K and Mg concentrated in the radicle, moving from the endosperm and cotyledons, while Zn, initially concentrated in specific tissues of the cotyledon, appeared more homogeneously distributed inside the whole seed after the plasma treatment. Significant variations in electrical impedance spectra were also observed after plasma treatment. This element redistribution in the seed was caused by the intense electrical field generated by the volume dielectric barrier discharge plasma, causing a movement of important micro and macronutrients from the storage regions of the seed towards the radicle tissues. This ion movement could explain the observed faster germination of the plasma-treated seeds. Indeed, such movement of ions is similar to what is generally observed in seed tissues during germination. The plasma treatment therefore somehow anticipates and implements the mobilization of key nutrients towards the radicle, resulting in faster and higher germination of the seeds as well as improved characteristics of the basil plantlet, especially at the root level.

Plum pox virus (PPV), agent of Sharka disease, is the most important quarantine pathogen of peach (P. persica L. Batsch). Extensive evaluation of peach germplasm has highlighted the lack of resistant sources, while suggesting the presence of a quantitative disease resistance, expressed as reduction in the intensity of symptoms. Unravelling the genetic architecture of peach response to PPV infection is essential for pyramiding resistant genes and for developing more tolerant varieties. For this purpose, a genome-wide association (GWA) approach was applied in a panel of accessions phenotyped for virus susceptibility and genotyped with the IPSC peach 9 K SNP Array, and coupled with an high-coverage resequencing of the tolerant accession ‘Kamarat’. Genome-wide association identified three highly significant associated loci on chromosome 2 and 3, accounting for most of the reduction in PPV-M susceptibility within the analysed peach population. The exploration of associated intervals through whole-genome comparison of the tolerant accession ‘Kamarat’ and other susceptible accessions, including the PPV-resistant wild-related species P. davidiana, allow the identification of allelic variants in promising candidate genes, including an RTM2-like gene already characterized in A. thaliana. The present study is the first effort to identify genetic factors involved in Sharka disease in peach germplasm through a GWA approach. We provide evidence of the presence of quantitative resistant loci in a collection of peach accessions, identifying major loci and highly informative SNPs that could be useful for marker assisted selection. These results could serve as reference bases for future research aimed at the comprehension of genetic mechanism regulating the complex peach-PPV interaction.