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Daunorubicin (DNR) is an anthracycline antibiotic originating from soil-dwelling actinobacteria extensively used to treat malignant tumors. Over the decades, extensive attempts were made to enhance the production of anthracyclines by introducing genetic modifications and mutations in combination with media optimization, but the target production levels remain comparatively low. Developing an appropriate culture medium to maximize the yield of DNR and preventing autotoxicity for the producing organism remains a challenge. Our prospective review sheds light on a method involving perturbation that enhances the precursors to regulate the type II PKS pathway, enhancing cells’ capacity to increase secondary metabolite production. The suggested method also entails the preparation of culture media for the cultivation of Streptomyces sp. and enhanced yield of DNR, as well as making it inactive with iron or its reduced forms following efflux from the producer. The iron or iron–DNR complex is encapsulated by oleic acid or lipid micelle layers in the culture media, finally resulting in the generated inactive DNR and the DNR–iron–oil complex. This idea has the potential to protect the producer organism from autotoxicity and prevent the inhibition of metabolite production. The approach of substituting sugar with oil in culture media has a dual role wherein it promotes Streptomyces growth by utilizing lipids as an energy source and encapsulating the generated DNR–iron complex in the medium. In this review, we discussed aspects like anthracycline producers, biosynthesis pathways, and gene regulation; side effects of DNR; mechanisms for autotoxicity evasion; and culture media components for the enhancement of DNR production in Streptomyces sp. We anticipate that our work will help researchers working with secondary metabolites production and decipher a methodology that would enhance DNR yield and facilitate the extraction of the resulting DNR by lowering costs in large-scale fermentation.

Low-temperature plasma (LTP) is currently one of the non-invasive and environmentally friendly methods of seed treatment and is massively tested on various types of crops. For the needs of gene banks, the use of LTP technology represents the treatment of seeds before sowing to improve the germination and emergence of long-term stored seed samples. Seeds of four genotypes of wheat, oats, flax, and rapeseed stored in the gene bank for 1, 10, and 20 years were plasma treated for 20, 25, and 30 min. Standard germination parameters (SG3, SG7, GR, MGT, and GI), as well as predictive models, were used to evaluate the effect of plasma treatment on seeds, and the effect on seed metabolism was assessed by superoxide dismutase (SOD) activity. The plasma treatment had different effects on germination and on the enzymatic activity of the tested species, and the result was influenced by both the duration of the treatment and the crop species/genotype. The plasma treatment has a positive effect on germination parameters in flax and rapeseed; in some variants, as in wheat, oats generally reacted negatively. SOD activity was variable in wheat, while higher activity with increasing treatment time was found in other crops. The results of this first study focused on long-term stored seeds and showed the potential of plasma treatment of seeds of plant genetic resources, the possibility of stimulating the germination of stored PGRs, and the need to optimize treatment conditions for individual genotypes.

The gliding arc cold plasma treatment of wheat seeds is an alternative to chemical seed dressing, but this treatment is not very effective. This study was focused on testing the addition of salt, water and nitrogenous compounds during plasma treatment to increase the efficiency of the plasma seed treatment. The additives were not supplied in the gaseous state, as usual, but in an aqueous solution by spraying under a plasma nozzle during the treatment. The phytosanitary effect of the plasma treatment was evaluated based on the germination of Tilletia caries spores from artificially infected seeds on water agar. The viability of the seeds, growth and yield characteristics of the plasma-treated seeds were evaluated in the laboratory, greenhouse and even in field experiments with the primary goal of reducing spore vitality. The germination of T. caries spores was mostly limited to the variant with the addition of water during plasma treatment, where spore germination reached only 32.7% of the control variant while maintaining sufficient growth properties. The experiment demonstrated the suitability of using the addition of various substances during plasma treatment in the form of aqueous solutions and increasing the effectiveness of this treatment.

Members of the genus Armillaria are widespread forest pathogens against which effective protection has not yet been developed. Due to their longevity and the creation of large-scale cloning of Armillaria individuals, the use of mycoviruses as biocontrol agents (BCAs) against these pathogens could be an effective alternative. This work describes the detection and characterization of viruses in Armillaria spp. collected in the Czech Republic through the application of stranded total RNA sequencing. A total of five single-stranded RNA viruses were detected in Armillaria ostoyae and A. cepistipes, including viruses of the family Tymoviridae and four viruses belonging to the recently described “ambivirus” group with a circular ambisense genome arrangement. Both hammerhead (HHRz) and hairpin (HpRz) ribozymes were detected in all the ambiviricot sequences. Armillaria viruses were compared through phylogenetic analysis and confirmed their specific host by direct RT-PCR. One virus appears to infect both Armillaria species, suggesting the occurrence of interspecies transmission in nature.

In the past few years, huge economic and environmental losses have resulted by fungal pathogens in agriculture and food production worldwide. Fungal toxins also pose a threat for food production and animal health. In the last decade, the site-specific fungicides has been used in plant protection, providing highly efficient protection against phytopathogens. However, the use of pesticides boosted problems with resistance in the targeted organisms and increased negative environmental impact of insecticides, herbicides and fungicides. Therefore, new methods are needed to control resistant populations of plant pathogens, including fungi. Mycoviruses are fungal viruses, i.e. viruses infecting fungi. Some mycoviruses affect the growth and virulence of the host fungus and may prevent successful infection of plant host. They have potential to reduce phytopathogens. In this review, the successful examples how mycoviruses alter hypovirulence in well-known phytopathogens are discussed. Mycoviruses could be useful as a one of new next generation pest-control products.

We introduced here the first loop mediated isothermal amplification (LAMP) assay for the identification of honey fungus, Armillaria ostoyae, a basidiomycote playing an important role in spruce declines in the Palaearctic region. In total, 101 isolates, representing three Armillaria species, were used to develop a new LAMP assay to determine species specific identification. We have here described LAMP primers enhanced with fluorescent dye that are able to amplify A. ostoyae DNA and detect fungi in a fast single step reaction. The detection limit of LAMP was 1 pg of genomic DNA per reaction. We optimized a new LAMP assay for the rapid detection of A. ostoyae using the translation elongation factor 1-α (tef1) marker and fluorescence labelled oligonucleotide assimilating probe. The LAMP assay does not require any specialized equipment, hence it can be used in the field for the rapid detection of A. ostoyae even using the portable and mobile device. The specificity of the assay was confirmed by the use of A. ostoyae strains and Armillaria cepistipes and Armillaria gallica strains, respectively. In conclusion, the assay could be a rapid, specific, sensitive and low-cost tool for identification of A. ostoyae as well as the first step for expansion of this method in practical applications.

Microsatellite markers were used for the assessment of genetic diversity and genetic structure in a germplasm collection of yellow mustard, Sinapis alba L. The comprehensive collection of genetic resources represented 187 registered varieties, landraces, and breeding materials. Microsatellites generated 44 polymorphic alleles in 15 loci. Eleven of them were medium to highly polymorphic, and the high levels of observed heterozygosity (0.12–0.83) and Nei’s gene diversity index (0.11–0.68) indicated a high level of polymorphism. Based on PCoA and neighbor joining analyses, the genetic resources were divided into two groups. The range of genetic dissimilarity in the analysed collection was in the range of 0.00–1.00. The high level of dissimilarity between the accessions was documented by the high WAM value (33.82%). Bayesian clustering algorithms were performed in the STRUCTURE 2.3.4 software. The number of clusters was estimated at K = 2. The accessions were classified according to Q1/Q2 values. The low average values of the parameters Fst_1 (0.3482), Fst_2 (0.1916), and parameter alpha (0.0602) indicated substantial mating barriers between varieties and reproductive isolation due to the limited exchange of genetic resources between breeders. These results demonstrated the importance of extensive collections of genetic resources for the maintenance of genetic diversity and indicated considerable genetic differentiation among accessions.

The opium poppy’s ability to produce various alkaloids is both useful and problematic. Breeding of new varieties with varying alkaloid content is therefore an important task. In this paper, the breeding technology of new low morphine poppy genotypes, based on a combination of a TILLING approach and single-molecule real-time NGS sequencing, is presented. Verification of the mutants in the TILLING population was obtained using RT-PCR and HPLC methods. Only three of the single-copy genes of the morphine pathway among the eleven genes were used for the identification of mutant genotypes. Point mutations were obtained only in one gene (CNMT) while an insertion was obtained in the other (SalAT). Only a few expected transition SNPs from G:C to A:T were obtained. In the low morphine mutant genotype, the production of morphine was decreased to 0.1% from 1.4% in the original variety. A comprehensive description of the breeding process, a basic characterization of the main alkaloid content, and a gene expression profile for the main alkaloid-producing genes is provided. Difficulties with the TILLING approach are also described and discussed.

ABSTRACT: In recent decades, interest in non-traditionally colored eggs has increased. For breeders, this market interest means breeding lines of laying hens that lay eggs of varied colors, such as the blue-green eggshells (Dominant Greenshell) in this study. This study presents the results of genotyping the polymorphism of the O locus responsible for shell coloration and photometric measurement of eggshell color based on the CIELAb system, which was carried out on the unique Czech breeding population Dominant Greenshell. The aim was to use a combination of phenotyping using the CIELab System method and genotyping of the O locus using the end-point PCR approach with the main focus on the accuracy of distinguishing shell color genotypes, streamlining the selection of dominant homozygotes in the O locus, optimizing this technology for the most efficient and cost-effective selection procedure in practical hen breeding. The optometric method was able to reliably distinguish only dominant and recessive phenotypes and eliminate from the population only undesirable recessive homozygotes with a white colored shell. The parameter a* (redness/greenness) from the CIELab color space turned out to be absolutely key for distinguishing dominant and recessive phenotypes. Using the CART methodology, a classification tree built on discriminating optometric characteristics a-blunt was obtained, however, for the group of desirable O/O homozygotes, the selection approach would result in incorrect genotyping of 31% of individuals. Therefore, a combined approach based on rapid and simple elimination of recessive homozygotes using phenotyping (CIELab photometric measurement) and molecular identification of the EAV-HP insertion in the SLCO1B3 gene in dominant phenotypes, regardless of color intensity affected by laying time/order, and allowing reliable elimination, has proven to be the most effective method to distinguish heterozygotes from the breeding population. The combination of optometric and molecular selection methods then leads to more efficient selection, reduction of overall selection costs. This process led to the stabilization of the breeding population within one generation and the achievement of a pure homozygous line with regard to eggshell color.