Pinus sylvestris bark represents a rich source of active compounds with antifungal, antibacterial, and antioxidant properties. The current study aimed to evaluate the antifungal potential of P. sylvestris bark against Botryosphaeria dothidea, Dothiorella sarmentorum, and Neofusicoccum parvum (Botryosphaeriaceae) through its chemical (water extracts) and biological (Trichoderma spp. isolated from the bark) components. The water bark extracts were prepared at two temperatures (80 and 120 °C) and pH regimes (7 and 9). The presence of bark extracts (30%) caused inhibition of mycelial growth of B. dothidea and D. sarmentorum for 39 to 44% and 53 to 60%, respectively. Moreover, we studied the antagonistic effect of three Trichoderma isolates originating from the pine bark. Trichoderma spp. reduced growth of B. dothidea by 67%–85%, D. sarmentorum by 63%–75% and N. parvum by 55%–62%. Microscopic examination confirmed typical mycoparasitism manifestations (coiling, parallel growth, hook-like structures). The isolates produced cellulase, β-glucosidase and N-acetyl-β-glucosaminidase. The volatile blend detected the emission of several volatile compounds with antimicrobial activity, including nonanoic acid, cubenene, cis-α-bergamotene, hexanedioic acid, and verticillol. The present study confirmed in vitro potential of P. sylvestris bark extracts and Trichoderma spp. against the Botryosphaeriaceae. The study is an important step towards the use of environmentally friendly methods of Botryosphaeriaceae disease control.

Some Trichoderma strains can persist in ecosystems with high concentrations of heavy metals. The aim of this research was to examine the variability of Trichoderma strains isolated from different ecosystems, based on their morphological properties and restriction analysis of ITS fragments. The fungal growth was tested on potato dextrose agar, amended with Cu(II) concentrations ranging from 0.25 to 10 mmol/l, in order to identify copper-resistant strains. The results indicate that some isolated strains of Trichoderma sp. show tolerance to higher copper concentrations. Further research to examine the ability of copper bioaccumulation by tolerant Trichoderma strains is needed.

Adding compost to soil can result in plant disease suppression through the mechanisms of antagonistic action of compost microflora against plant pathogens. The aim of the study was to select effective antagonists of Pythium aphanidermatum from compost, to assess the effect of its extracellular metabolites on the plant pathogen, and to characterize antifungal metabolites. The fungal isolate selected by a confrontation test was identified as Aspergillus piperis A/5 on the basis of morphological features and the internal transcribed spacer (ITS) region, ?-tubulin and calmodulin partial sequences. Liquid chromatography-mass spectroscopy (LC-MS) analysis showed that gluconic and citric acid were the most abundant in the organic culture extract. However, the main antifungal activity was contained in the aqueous phase remaining after the organic solvent extraction. The presence of considerable amounts of proteins in both the crude culture extract as well as the aqueous phase remaining after solvent extraction was confirmed by SDS-PAGE. Isolated Aspergillus piperis A/5 exhibits strong antifungal activity against the phytopathogen Pythium aphanidermatum. It secretes a complex mixture of metabolites consisting of small molecules, including gluconic acid, citric acid and itaconic acid derivatives, but the most potent antifungal activity was associated with proteins resistant to heat and organic solvents. Our findings about the activity and characterization of antagonistic strain metabolites contribute to the understanding of the mechanism of interaction of antifungal metabolites as well as fungal-fungal interaction. The obtained results provide a basis for further application development in agriculture and food processing.

Botryosphaeriales represent a diverse order of fungal pathogens of various woody plant species. In Serbia, these fungi are important pathogens of forest, ornamental, and fruit trees causing die-back, cankers, leaf blights, fruit, and root rot. The aim of this study was to evaluate the antifungal activity of Pinus sylvestris bark extracts and Trichoderma spp. against Botryosphaeria dothidea, Dothiorella sarmentorum, and Neofusicoccum parvum (Ascomycota, Botryosphaeriales) isolated from Picea abies, Thuja occidentalis, and Prunus laurocerasus trees planted in urban areas in Serbia. Bark extracts were prepared in water solution at two temperatures (80 and 120 °C). The extracts were tested using two concentrations (20% and 30%). Moreover, two Trichoderma isolates obtained from P. sylvestris bark were tested against Botryosphaeriales and their antagonistic potential was estimated in vitro using a confrontation test. Mycelial growth of B. dothidea and D. sarmentorum was significantly inhibited in the presence of bark extracts, while N. parvum showed no growth inhibition. Botryosphaeria dothidea growth was inhibited by 35 to 39% in the case of 20% extracts and by 39 to 44% in the case of 30% extracts. The growth inhibition of D. sarmentorum was between 48% and 56% in the case of 20% extracts and between 53% and 60% in the case of 30% extracts. The two Trichoderma isolates showed antifungal activity against the selected pathogens. An isolate BKG 4 showed the highest inhibition level, and it inhibited the growth of B. dothidea, D. sarmentorum, and N. parvum by 85%, 75%, and 62%, respectively. Preliminary results suggest that both P. sylvestris bark extracts and Trichoderma spp. could be used as biocontrol agents against B. dothidea, D. sarmentorum, and N. parvum, and this should be a further studied.

Selenium is a trace mineral chemically related to sulfur and tellurium. In the body selenium combines with protein molecules to form selenoproteins and it is distributed in low concentrations and unequally in air, soil and water all over the world. Edible mushrooms are known to be selenium accumulators. Since mushrooms contain relatively high protein levels, and they can accumulate large amounts of selenium, it is reasonable to expect that selenium could be incorporated into proteins. The growth of mycelia and fruit body formation of different medicinal mushroom strains of Pleurotus ostreatus (Hk-35 and P70) over the wide range of concentrations of inorganic form of selenium were examined. Mushrooms were cultivated on agar base media and on substrates based on sawdust. Vegetative growths of mycelium were measured as colony diameter in pure cultures supplemented with inorganic form of Se supplements, prepared as Na2SeO4 and Na2SeO3 in concentrations of: 1, 10, 25, 50, 75, 100 and 150 mg/l. Inorganic form of Se supplements, showed stimulation effects (in concentration of 1-50 mg/l) and toxic effects in higher concentration. On the standard industrial sawdust based substrate, supplemented with 100 mg/kg Na2SeO4 and Na2SeO3, accumulation of Se in fruit bodies was determined by the method of flameless atomic absorption spectrophotometer. The readings were performed on Varian SpectrAA-10 spectrophotometer equipped with VGA-76. Se as Na2SeO4 and Na2SeO3 was effectively taken up from substrates and accumulated in fruit bodies. Mushrooms accumulated selenium between 120 and 250 mg/kg dry weight. In mushrooms cultivated without Se supplement, Se contents were only about 1 mg/kg and in substrate about 0.1 mg/kg.