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The present work focuses on the antifungal effect of volatile organic compounds (VOCs) produced by Bacillus amyloliquefaciens CPA-8 against Monilinia laxa, M. fructicola and Botrytis cinera, three postharvest fruit pathogens of sweet cherry fruit. VOCs were evaluated with a double petri dish assay against mycelial and colony growth of target pathogens. For this purpose, CPA-8 was grown on different media and cultured for 24 and 48 h at 30 °C before assays. Data showed that mycelial growth inhibition was higher when CPA-8 was grown on Tryptone Soya Agar (TSA) while no differences were generally observed when CPA-8 was cultured for either, 24 and 48 h. Moreover, no effects were observed on colony growth. The main volatile compounds emitted by CPA-8 were identified by solid-phase microextraction (SPME)-gas chromatography as 1,3 pentadiene, acetoin (3-hydroxy-2-butanone) and thiophene. Pure compounds were also tested in vitro on mycelial growth inhibition and their EC50 values against the three pathogens were estimated. Thiophene was the most effective VOC, showing more than 82% suppression of mycelial growth at the highest concentration (1.35 μL/mL headspace) and EC50 values ranging from 0.06 to 6.67 μL/mL headspace. Finally, the effectiveness of thiophene and CPA-8 VOCs was evaluated against artificially inoculated cherry fruits. Among the target pathogens, M. fructicola was clearly controlled by CPA-8 with less than 25% of rotten fruits compared to the control (65% disease incidence) and for all pathogens, less than 37.5% of CPA-8 treated decayed fruits produced spores (disease sporulation). Otherwise, pure thiophene showed no effect against any pathogen on disease incidence and disease sporulation. The results indicated that VOCs produced by B. amyloliquefaciens CPA-8 could develop an additive antifungal effect against postharvest fruit pathogens on stone fruit. This research was supported by the European project BIOCOMES FP7-612713 and by the Catalan government (Generalitat de Catalunya) for the PhD grant 2016-FI-B2 00143 (Amparo M. Gotor).

Metabolism and in vitro fungitoxicity of perinaphthenone against three economically important fungi of the citrus, Botryodiplodia spp., Botrytis spp. and Fusarium spp. were investigated. Perinaphthenone exhibited significant antifungal activity at 62.5 μM and above. Even, the inhibitory effect against Fusarium spp. was significantly enhanced by exposure to direct light. In addition, the metabolism of perinaphthenone by the three fungi was studied. Results show that perinaphthenone was transformed almost completely during the first 24 h; two major products, whose concentration increased progressively during the twelve days of the test, were isolated and identified as 2,3-dihydro-1H-phenalen-1-ol and 2,3-dihydro-phenalen-1-one. Although both metabolic products displayed a considerable fungistatic effect, their slightly lower activities in comparison to perinaphthenone indicate that the transformation was a detoxification process. Studies on the relationship between the effect of some substituents in the perinaphthenone core and the mycelial growth inhibition of Botryodiplodia spp. were also carried out. Results show that the α, β-unsaturated carbonyl system is an important structural requirement but not the only to be necessary for the strong antifungal activity of perinaphthenone. In general, the antifungal properties of perinaphthenone may be modulated through the incorporation of substituents in the naphthalene core or in the α, β-unsaturated carbonyl system. It is concluded that perinaphthenone could be used as an antifungal agent or as a structural template for the development of new fungicide compounds., Phenalenones; Botrytis spp.; Botryodiplodia spp.; Fusarium spp.; Antifungal activity; Metabolism; Reduction; Derivatives.

Phytopathogens cause undeniably serious damage in agriculture by harming fruit cultivations and lowering harvest yields, which as a consequence substantially reduces food production efficiency. Fungi of the Botrytis, Colletotrichum and Verticillium genera are a main concern in berry production. However, no rapid detection method for detecting all of these pathogens simultaneously has been developed to date. Therefore, in this study, a multiplex real-time PCR assay for this purpose was established. Universal fungal primers for the D2 region of the large subunit ribosomal DNA and three multiplexable fluorogenic probes specific for the chosen fungi were designed and deployed. The triplex approach for the molecular detection of these fungi, which was developed in this study, allows for the rapid and effective detection of crucial berry pathogens, which contributes to a more rapid implementation of protective measures in plantations and a significant reduction in losses caused by fungal diseases.