Genome editing for healthy crops: traits, tools and impacts.

Crop cultivars in commercial use have often been selected because they show high levels of resistance to pathogens. However, widespread cultivation of these crops for many years in the environments favorable to a pathogen requires durable forms of resistance to maintain "healthy crops". Breeding of new varieties tolerant/resistant to biotic stresses by incorporating genetic components related to durable resistance, developing new breeding methods and new active molecules, and improving the Integrated Pest Management strategies have been of great value, but their effectiveness is being challenged by the newly emerging diseases and the rapid change of pathogens due to climatic changes. Genome editing has provided new tools and methods to characterize defense-related genes in crops and improve crop resilience to disease pathogens providing improved food security and future sustainable agricultural systems. In this review, we discuss the principal traits, tools and impacts of utilizing genome editing techniques for achieving of durable resilience and a "healthy plants" concept.

CRISPR/Cas-mediated plant genome editing: outstanding challenges a decade after implementation.

The discovery of the CRISPR/Cas genome-editing system has revolutionized our understanding of the plant genome. CRISPR/Cas has been used for over a decade to modify plant genomes for the study of specific genes and biosynthetic pathways as well as to speed up breeding in many plant species, including both model and non-model crops. Although the CRISPR/Cas system is very efficient for genome editing, many bottlenecks and challenges slow down further improvement and applications. In this review we discuss the challenges that can occur during tissue culture, transformation, regeneration, and mutant detection. We also review the opportunities provided by new CRISPR platforms and specific applications related to gene regulation, abiotic and biotic stress response improvement, and de novo domestication of plants.

The Genus Heterogynis Rambur, 1866 (Heterogynidae, Lepidoptera): Congruence of Molecular, Morphological and Morphometric Evidence Reveal New Species in Serbia.

The Heterogynidae are a small family of moths consisting of a single genus Heterogynis and sixteen described species distributed in the Mediterranean region. A species new to science, Heterogynis serbica sp. nov., is described from the locality of Srebrenac, Mt. Kopaonik, Republic of Serbia, Balkan Peninsula, by applying an integrative taxonomic approach using morpho-anatomical characteristics, wing morphometics and DNA barcoding. Male genitalia, scanning electron micrographs of adult male head anatomy, abdominal tergites/sternites, cocoons and habitats of the closely related species H. serbica sp. nov. and H. zikici are discussed and illustrated. Photographs of adult males and females, cocoons, plants in which the cocoons were found and habitats are shown. Importantly, marked differences in genital structure and other morphological characters were noted. These differences were confirmed with forewing morphometrics and COI-based DNA barcoding results. Additionally, DNA barcodes for H. serbica sp. nov. and H. zikici were compared against previously available data for the genus to evaluate the phylogenetic relationships. We conclude that deep, previously unknown and unexpected intrageneric morphological diversity exists in the genus Heterogynis.

Ectomycorrhizal Fungi Modulate Pedunculate Oak's Heat Stress Responses through the Alternation of Polyamines, Phenolics, and Osmotica Content.

The physiological and biochemical responses of pedunculate oaks (Quercus robur L.) to heat stress (HS) and mycorrhization (individually as well in combination) were estimated. One-year-old Q. robur seedlings were grown under controlled conditions in a pot experiment, inoculated with a commercial inoculum of ectomycorrhizal (ECM) fungi, and subjected to 72 h of heat stress (40 °C/30 °C day/night temperature, relative humidity 80%, photoperiod 16/8 h) in a climate chamber, and they were compared with seedlings that were grown at room temperature (RT). An in-depth analysis of certain well-known stress-related metrics such as proline, total phenolics, FRAP, ABTS, non-protein thiols, and lipid peroxidation revealed that mycorrhized oak seedlings were more resistant to heat stress (HS) than non-mycorrhized oaks. Additionally, levels of specific polyamines, total phenolics, flavonoids, and condensed tannins as well as osmotica (proline and glycine betaine) content were measured and compared between four treatments: plants inoculated with ectomycorrhizal fungi exposed to heat stress (ECM-HS) and those grown only at RT (ECM-RT) versus non-mycorrhized controls exposed to heat stress (NM-HS) and those grown only at room temperature (NM-RT). In ectomycorrhiza inoculated oak seedlings, heat stress led to not only a rise in proline, total phenols, FRAP, ABTS, non-protein thiols, and lipid peroxidation but a notable decrease in glycine betaine and flavonoids. Amounts of three main polyamines (putrescine, spermine, and spermidine) were quantified by using high-performance liquid chromatography coupled with fluorescent detection (HPLC/FLD) after derivatization with dansyl-chloride. Heat stress significantly increased putrescine levels in non-mycorrhized oak seedlings but had no effect on spermidine or spermine levels, whereas heat stress significantly increased all inspected polyamine levels in oak seedlings inoculated with ectomycorrhizal inoculum. Spermidine (SPD) and spermine (SPM) contents were significantly higher in ECM-inoculated plants during heat stress (approximately 940 and 630 nmol g-1 DW, respectively), whereas these compounds were present in smaller amounts in non-mycorrhized oak seedlings (between 510 and 550 nmol g-1 DW for Spd and between 350 and 450 nmol g-1 DW for Spm). These findings supported the priming and biofertilizer roles of ectomycorrhizal fungi in the mitigation of heat stress in pedunculate oaks by modification of polyamines, phenolics, and osmotica content.

Diversity and Distribution of Macrofungi in Protected Mountain Forest Habitats in Serbia and Its Relation to Abiotic Factors.

Fungal diversity is one of the most important indicators of overall forest biodiversity and its health. However, scarce information exists on the state of macrofungal communities of mountain forests in Serbia, making it one of the countries with the least-published mycological data in the Mediterranean and Balkan region of Europe. This paper presents the results of the first comprehensive, long-term study of macrofungal communities in some of the most important mountain forest ecosystems in Serbia (Tara, Kopaonik and Vidlic). In the course of three consecutive years, the sampling of five permanent experimental plots resulted in 245 species of macrofungi, classified into three functional groups (terricolous saprothrophs, lignicolous, and mycorrhizal fungi). Special attention was given to protected and indicator species, which point out the great value of studied forest habitats and the importance of their conservation. It was found that precipitation, habitat humidity, and temperature significantly influence the occurrence and distribution, primarily of mycorrhizal and lignicolous group of fungi. Thus, the continuation of long-term monitoring is crucial in order to more precisely determine which groups/species of macrofungi would, and to what extent they would, adapt to a rapidly changing climate.