Carotenoid cleavage oxygenases (CCOs) enzymes play an important role in plant growth and development by producing a wide array of apocarotenoids and their derivatives. These compounds are vital for colouring flowers and fruits and synthesizing plant hormones such as abscisic acid and strigolactones. Despite their importance, the gene family responsible for CCO enzymes in sunflowers has not been identified. In this study, we identify the CCO genes of the sunflower plant to fill this knowledge gap. Phylogenetic and synteny analysis indicated that the Helianthus annuus CCO (HaCCO) genes were conserved in different plant species and they could be divided into three subgroups based on their conserved domains. Analysis using MEME tool and multiple sequence alignment identified conserved motifs in the HaCCO gene sequence. Cis-regulatory elements (CREs) analysis of the HaCCO genes indicated the presence of various responsive elements related to plant hormones, development, and responses to both biotic and abiotic stresses. This implies that these genes may respond to plant hormones, developmental cues, and drought stress, offering potential applications in the development of more resistant crops. Genes belonging to the 9-cis-epoxy carotenoid dioxygenases (NCED) subgroups predominantly exhibited chloroplast localization, whereas the genes found in other groups are primarily localized in the cytoplasm. These 21 identified HaCCOs were regulated by 60 miRNAs, indicating the crucial role of microRNAs in gene regulation in sunflowers. Gene expression analysis under drought stress revealed significant up-regulation of HaNCED16 and HaNCED19, genes that are pivotal in ABA hormone biosynthesis. During organ-specific gene expression analysis, HaCCD12 and HaCCD20 genes exhibit higher activity in leaves, indicating a potential role in leaf pigmentation. This study provides a foundation for future research on the regulation and functions of the CCO gene family in sunflower and beyond. There is potential for developing molecular markers that could be employed in breeding programs to create new sunflower lines resistant to biotic and abiotic stresses.

Introduction: Mediterranean forests are currently facing a surge in abiotic stressors such as droughts and massive fires as a result of climate crisis and human pressure. Susceptibility to biotic stressors has also increased, including a variety of pests and pathogens capable of weakening and potentially killing forest flora. Biodiversity of microbiome protects forests against declines as it increases trees’ resilience and adaptability.Objectives: With the objective of analyzing the relationship between health status and fungal diversity, in the present work the mycobiota of declined and healthy specimens of keystone Mediterranean tree species is described and compared.Methods: To this end, bark and wood from declining Spanish forests of Castanea sativa (chestnut), Quercus ilex (holm oak), Quercus suber (cork oak), and Quercus pyrenaica (pyrenean oak) were sampled and the Internal Transcribed Spacer (ITS1) genomic region was sequenced.Results: Results showed a predominance of Ascomycota, Basidiomycota, and Mucoromycota in all samples. Alpha diversity at genus level was not affected by health status and was characterized by uneven, poorly distributed fungal communities dominated by a few genera. Differentially abundant (DA) genera between healthy and declined samples were found in chestnut (15), holm oak (6), and pyrenean oak (4) trees, but not in cork oak. Some genera known for their plant protection potential, such as Trichoderma, were found exclusively in healthy trees. Functional profiles revealed a predominance of phytopathogens and saprobes in all tree species, irrespective of their health status.Discussion: This study emphasizes the importance of Mediterranean forests as biodiversity refuges and highlights the value of above-ground tissues as a valid approach to assess shifts in forests’ microbiome diversity in response to biotic and abiotic stressors.

Carotenoid cleavage oxygenases (CCOs) enzymes play an important role in plant growth and development by producing a wide array of apocarotenoids and their derivatives. These compounds are vital for colouring flowers and fruits and synthesizing plant hormones such as abscisic acid and strigolactones. Despite their importance, the gene family responsible for CCOs in sunflowers has not been studied. In this study, we identify the CCO genes of the sunflower plant to fill this knowledge gap. Phylogenetic and synteny analysis indicated that the Helianthus annnus CCO (HaCCO) proteins are conserved in different plant species and they can be divided into three subgroups based on their conserved domains. MEME analysis and multiple sequence alignment showed that conserved motifs exist in the sequence of HaCCO genes. Cis-regulatory elements (CREs) analysis of the HaCCO genes revealed the presence of multiple plant hormones and abiotic and biotic responsive elements in their promoters, suggesting that these genes might be responsive to plant hormones, developmental and drought stress, which could be utilized to develop more resilient crops. Genes belonging to the 9-cis-epoxy carotenoid dioxygenases (NCED) subgroups predominantly exhibit chloroplast localization, whereas the genes found in other groups are primarily localized in the cytoplasm. These 21 identified HaCCOs were regulated by 60 miRNAs indicating the crucial role of microRNAs in gene regulation in sunflowers. Gene expression analysis under drought stress revealed significant up-regulation of HaNCED16 and HaNCED19 genes that provide pivotal role in ABA hormone biosynthesis. While during organ-specific gene expression analysis HaCCD12 and HaCCD20 genes exhibit higher activity in leaves, indicating a potential role in leaf pigmentation. These studies provide a foundation for future research on the regulation and functions of this gene family. It could potentially be utilized to develop molecular markers that could be used in breeding programs to develop new sunflower lines.

European chestnut (Castanea sativa) is threatened by the invasive fungus Cryphonectria parasitica, which causes chestnut blight. The virulence of the fungus can be reduced by a group of mycoviruses that can spread among vegetatively compatible strains through hyphal anastomosis. Hypovirulent isolates are used as biocontrol agents, but their efficiency can be diminished by restricted hyphal anastomosis if the variability of vegetative compatibility (vc) types in a population is high. Sexual reproduction could increase the vc type diversity and further complicate biocontrol in a region. Therefore, knowledge of genetic diversity of C. parasitica is important to assess the effectiveness of a biological control program. The present study was performed in the Autonomous Province of Bozen-Bolzano (South Tyrol) in northern Italy, where chestnut cultivation provides an additional income to farmers. The genetic characterization of C. parasitica isolates from 35 chestnut stands and one forest population in different districts of South Tyrol was performed based on the analysis of vegetative incompatibility loci, the mating type locus, and the internal transcribed spacer (ITS) region. In this study, a total of 23 different vc types were found all over South Tyrol with a Shannon diversity index of 1.86. EU-2, EU-1, and EU-13 were the most widespread vc types comprising 51%, 13%, and 9% of the fungal isolates, respectively. Both mating types were present in the region with a ratio close to 1:1. Three different haplotypes were identified based on ITS sequence analysis, which pointed to two introduction events of the fungus to the region and allowed placing C. parasitica from South Tyrol into a larger phylogeographic context.

Cryphonectria parasitica, which causes chestnut blight, is one of the most important pathogens of forest trees. In Europe, mycovirus-mediated biocontrol is the most efficient method to control the disease but can be impeded by the lack of information about the population structure of the fungus within a region. In particular, sexual reproduction and the new introduction of the pathogen can complicate biocontrol strategies. For this reason, this study aimed to determine the population structure of C. parasitica, which causes chestnut blight, in the northern Italian region of South Tyrol, using eleven multilocus microsatellite markers. Fifty-one haplotypes were found across South Tyrol, belonging to three divergent clusters. Recombinant genotypes demonstrated that sexual reproduction occurs across the different clusters. The most dominant genotypes in the region were also the most dominant in neighboring areas, such as Switzerland, northern Italy and France. All of the clusters from South Tyrol were related to the Italian genotype pool and are thought to have been introduced from northern Italian and other European populations due to naturally occurring gene flow or human-mediated introduction. At least three separate introduction events of C. parasitica might have happened in South Tyrol that could be separated by time. This study demonstrated a high genetic diversity of C. parasitica in South Tyrol and helped to shed light on the sexual reproduction and introduction events in the local populations.