Occurrence and Distribution Patterns of Plum Tree Viruses and Genetic Diversity of Sharka Isolates in Bosnia and Herzegovina

In order to fill in a decade-long information gap regarding the biological, serological and molecular data for plum tree viruses in Bosnia and Herzegovina, a three-phase study combining symptom evaluation, and serological and molecular assays with high-throughput sequencing (HTS) technology was conducted. The most frequently observed symptoms were discolorations in the form of ring patterns, bands and irregular shapes, as well as vein banding. Sharka-associated symptoms in the form of ring patterns and semicircles were prevalent. A total of 468 plum tree samples were tested by ELISA for the presence of PPV, ApMV, PDV, PNRSV, PBNSPaV, ACLSV and MLRSV. An overall infection incidence of 51.9% was detected, with PPV being the most prevalent (48.7%), followed by PDV (2.99%), PNRSV (0.21%) and mixed infections of PPV+PDV (1.71%). RT-PCR-assisted strain typing in 45 samples revealed PPV-D as the most common strain (22.22%), followed by PPV-REC (6.66%). Mixed infections of PPV-D+PPV-REC were detected (6.66%). HTS enabled the recovery of a 9743 nts long sequence of PPV-D (PPV_O7/80, MW412433), which shared the highest nucleotide and amino acid identities with isolates S13 (LC375131) from Serbia, SVN1 (LC375132) from Slovenia and N9 (LC375129) from Bulgaria. The phylogenetic analysis of the whole genome placed the isolate of the D strain in a distinctive group with the Slovenian isolate SVN1 (LC375132). In addition, the (Cter)NIb/(Nter)CP fragment of a PPV-REC isolate (MW412434) obtained in this survey formed a separate group with previously known isolates from Bosnia and Herzegovina (BOS64Pl and BOS257Pl).European Cooperation in Science and TechnologyPeer Reviewe

The root transcriptome dynamics reveals new valuable insights in the salt-resilience mechanism of wild grapevine (Vitis vinifera subsp. sylvestris)

Introduction: Most of elite cultivated grapevine varieties (Vitis vinifera L.), conventionally grafted on rootstocks, are becoming more and more affected by climate changes, such as increase of salinity. Therefore, we revisited the valuable genetic resources of wild grapevines (V. sylvestris) to elaborate strategies for a sustainable viticulture. Methods: Here, we compared physiological and biochemical responses of two salt-tolerant species: a wild grapevine genotype “Tebaba” from our previous studies and the conventional rootstock “1103 Paulsen”. Interestingly, our physio-biochemical results showed that under 150mM NaCl, “Tebaba” maintains higher leaf osmotic potential, lower Na+/K+ ratio and a significant peaked increase of polyphenol content at the first 8h of salinity stress. This behavior allowed to hypothesis a drastic repatterning of metabolism in “Tebaba’s” roots following a biphasic response. In order to deepen our understanding on the “Tebaba” salt tolerance mechanism, we investigated a time-dependent transcriptomic analysis covering three sampling times, 8h, 24h and 48h. Results: The dynamic analysis indicated that “Tebaba” root cells detect and respond on a large scale within 8h to an accumulation of ROS by enhancing a translational reprogramming process and inducing the transcripts of glycolytic metabolism and flavonoids biosynthesis as a predominate non-enzymatic scavenging process. Afterwards, there is a transition to a largely gluconeogenic stage followed by a combined response mechanism based on cell wall remodeling and lignin biosynthesis with an efficient osmoregulation between 24 and 48 h. Discussion: This investigation explored for the first time in depth the established cross-talk between the physiological, biochemical and transcriptional regulators contributing to propose a hypothetical model of the dynamic salt mechanism tolerance of wild grapevines. In summary, these findings allowed further understanding of the genetic regulation mechanism of salt-tolerance in V. sylvestris and identified specific candidate genes valuable for appropriate breeding strategies

A Framework for the Evaluation of Biosecurity, Commercial, Regulatory, and Scientific Impacts of Plant Viruses and Viroids Identified by NGS Technologies

Recent advances in high-throughput sequencing technologies and bioinformatics have generated huge new opportunities for discovering and diagnosing plant viruses and viroids. Plant virology has undoubtedly benefited from these new methodologies, but at the same time, faces now substantial bottlenecks, namely the biological characterization of the newly discovered viruses and the analysis of their impact at the biosecurity, commercial, regulatory, and scientific levels. This paper proposes a scaled and progressive scientific framework for efficient biological characterization and risk assessment when a previously known or a new plant virus is detected by next generation sequencing (NGS) technologies. Four case studies are also presented to illustrate the need for such a framework, and to discuss the scenarios.Peer reviewe

A Novel and Highly Inclusive Quantitative Real-Time RT-PCR Method for the Broad and Efficient Detection of Grapevine Leafroll Associated Virus 1

Grapevine (Vitis vinifera L.) is one of the most important crops in the world due to its economic and social impact. Like many other crops, grapevine is susceptible to different types of diseases caused by pathogenic microorganisms. Grapevine leafroll-associated virus 1 (GLRaV-1) is a virus associated with grapevine leafroll disease and it is considered at the national and European level as a pathogen that must be absent in propagative plant material. For this reason, the availability of specific, sensitive and reliable detection techniques to ascertain the sanitary status of the plants is of great importance. The objective of this research was the development of a new GLRaV-1 detection method based on a TaqMan quantitative real-time RT-PCR targeted to the coat protein genomic region and including a host internal control in a duplex reaction. To this end, three new GLRaV-1 full genomes were recovered by HTS and aligned with all sequences available in the databases. The method has been validated following EPPO standards and applied for the diagnosis of field plant material and transmission vectors. The new protocol designed has turned out to be highly sensitive as well as much more specific than the current available methods for the detection and absolute quantitation of GLRaV-1 viral titer

N2O Temporal Variability from the Middle Troposphere to the Middle Stratosphere Based on Airborne and Balloon-Borne Observations during the Period 1987–2018

Nitrous oxide (N2O) is the fourth most important greenhouse gas in the atmosphere and is considered the most important current source gas emission for global stratospheric ozone depletion (O3 ). It has natural and anthropogenic sources, mainly as an unintended by-product of food production activities. This work examines the identification and quantification of trends in the N2O concentration from the middle troposphere to the middle stratosphere (MTMS) by in situ and remote sensing observations. The temporal variability of N2O is addressed using a comprehensive dataset of in situ and remote sensing N2O concentrations based on aircraft and balloon measurements in the MTMS from 1987 to 2018. We determine N2O trends in the MTMS, based on observations. This consistent dataset was also used to study the N2O seasonal cycle to investigate the relationship between abundances and its emission sources through zonal means. The results show a longterm increase in global N2O concentration in the MTMS with an average of 0.89 ± 0.07 ppb/yr in the troposphere and 0.96 ± 0.15 ppb/yr in the stratosphere, consistent with 0.80 ppb/yr derived from ground-based measurements and 0.799 ± 0.024 ppb/yr ACE-FTS (Atmospheric Chemistry Experiment Fourier Transform Spectrometer) satellite measurements

N2_2O Temporal Variability from the Middle Troposphere to the Middle Stratosphere Based on Airborne and Balloon-Borne Observations during the Period 1987–2018

Nitrous oxide (N$_2$O) is the fourth most important greenhouse gas in the atmosphere and is considered the most important current source gas emission for global stratospheric ozone depletion (O$_3$). It has natural and anthropogenic sources, mainly as an unintended by-product of food production activities. This work examines the identification and quantification of trends in the N$_2$O concentration from the middle troposphere to the middle stratosphere (MTMS) by in situ and remote sensing observations. The temporal variability of N$_2$O is addressed using a comprehensive dataset of in situ and remote sensing N$_2$O concentrations based on aircraft and balloon measurements in the MTMS from 1987 to 2018. We determine N$_2$O trends in the MTMS, based on observations. This consistent dataset was also used to study the N$_2$O seasonal cycle to investigate the relationship between abundances and its emission sources through zonal means. The results show a long-term increase in global N$_2$O concentration in the MTMS with an average of 0.89 ± 0.07 ppb/yr in the troposphere and 0.96 ± 0.15 ppb/yr in the stratosphere, consistent with 0.80 ppb/yr derived from ground-based measurements and 0.799 ± 0.024 ppb/yr ACE-FTS (Atmospheric Chemistry Experiment Fourier Transform Spectrometer) satellite measurements