Portuguese propolis: A potential source of environmentally friendly biofungicides

[Excerpt] The need to increase food production to feed an increasingly growing world population has led to an intensive use of pesticides. Globally, 4.6 million tons of chemical pesticides are sprayed into the environment every year1 . Out of the total consumption of pesticides, 17.5% are in the form of fungicides. However, pesticides have undesirable characteristics, such as the development of pathogen resistance and long degradation periods, and negative impacts on the environment, like soil and water pollution, bioaccumulation in the food chain, toxicity to non-target organisms and public health problems2 . Therefore, new environmentally friendly natural products, which are effective, safe, biodegradable and leave no harmful residues are needed. [...]The need to increase food production to feed an increasingly growing world population has led to an intensive use of pesticides. Globally, 4.6 million tons of chemical pesticides are sprayed into the environment every year1 . Out of the total consumption of pesticides, 17.5% are in the form of fungicides. However, pesticides have undesirable characteristics, such as the development of pathogen resistance and long degradation periods, and negative impacts on the environment, like soil and water pollution, bioaccumulation in the food chain, toxicity to non-target organisms and public health problems2 . Therefore, new environmentally friendly natural products, which are effective, safe, biodegradable and leave no harmful residues are needed. Propolis is a natural resinous mixture made by honeybees (particularly Apis mellifera L.) from various plant sources and processed with salivary secretions. More than 300 different compounds have been identified so far in propolis, including several bioactive compounds such as flavonoids and phenolic acids and derivatives, which are characteristic of plant secondary metabolism and are related with diverse biological properties like antimicrobial and antioxidant activities3 . In this work we evaluated the antifungal activity of ethanolic extracts (EE) from Portuguese propolis samples against phytopathogenic fungi species responsible for considerable damage in several economically important crops. Results showed that different propolis samples have different fungitoxic efficiencies but also that different species exhibit different susceptibilities. Globally, these findings suggest that propolis EE can be explored as source of biofungicides for application in a more sustainable agriculture management. In order to evaluate this propolis application, its phytotoxicity will be studied using in vitro flax plants (Linum usitatissimum L.)

Potential of Portuguese propolis as source of biofungicides

Agriculture provides livelihoods for 40% of worldwide population1 . Due to the rapid human population growth, it is estimated that global food production will need to increase by 60% to feed over 9.8 billion people by 20502 . The use of pesticides contributes to high crop yields and effective farm management3 . Globally, 4.6 million tons of chemical pesticides are sprayed into the environment every year4,5. Out of the total consumption of pesticides, 17.5% are in the form of fungicides5 . Although efficient, these chemical controls have undesirable characteristics, such as the development of pathogen resistance, environmental pollution, long degradation periods, bioaccumulation in the food chain and toxicity to non-target organisms6 . Therefore, alternative paths to the intensive use of crop protection chemicals are needed, such as the use of natural products which are effective, biodegradable and leave no harmful residues. Propolis is a natural resinous mixture made by honeybees (particularly Apis mellifera L.) from various plant sources and bee wax. Hundreds of different compounds have been identified so far in propolis, including several bioactive compounds such as flavonoids and phenolic acids and terpene derivatives, generally associated to its antibacterial, antifungal, antiviral and antioxidant activities, among others7 . Previous work showed that propolis ethanol extracts (EEs) can be explored as source of biofungicides for application in a sustainable agriculture management 8. In this study we evaluated the antifungal activity of EEs from Portuguese propolis samples against a range of phytopathogenic fungal species that affect economically important crops in Portugal. Interestingly, results showed that different propolis samples have distinct fungitoxic activities but also that different fungi species exhibit distinct susceptibilitiesFEDER/COMPETE/POCI– Operational Competitiveness and Internationalization Programme, under Project POCI-01-0145-FEDER-006958 and National Funds by FCT - Portuguese Foundation for Science and Technology, under the project UID/AGR/04033/201

Mechanistic studies of cytotoxicity induced by a Portuguese propolis extract, using Saccharomyces cerevisiae as eukaryotic cell model

Propolis is a natural complex mixture produced by honey bees (particularly Apis mellifera L.) by collecting exudates from various plant sources. Characterized by a plethora of chemicals, propolis is generally rich in flavonoids, phenolic acids and terpene derivatives, bioactive compounds associated to it’s antimicrobial, anti-inflammatory, antimutagenic and antioxidant activities1 . Previous work had shown that the ethanol extract of a sample from the Portuguese region of Beira Alta exhibited unique dual genotoxic and antigenotoxic effects using the yeast S. cerevisiae eukaryotic model2 . In this work we prepared two ethanol extracts (EE) of propolis samples from Pereiro (P) - Beira Alta - collected in 2010 (P10.EE) and 2017 (P17.EE) to investigate the mechanisms of cytotoxicity and genotoxicity using specific S. cerevisiae mutants. While P17.EE didn’t show any toxic effect, yeast cells exposed to P10.EE showed a considerable decreased viability along time, assessed by colony-forming units. Interestingly, the oxidative stress response-defective mutant yap1 was more resistant than the wild type, suggesting that this cytotoxic effect was not mediated by oxidative stress. P.EE’s genotoxicity was also analysed by the nucleus-cytosolic translocation of NHP6A protein, considered a marker of necrosis. P10.EE induced NHP6A protein translocation to the cytoplasm, observed by fluorescence microscopy, suggesting that cytotoxicity of this extract was indeed mediated by necrosis. Although P17.EE didn’t seem to induce necrotic cell death, both extracts induced plasma membrane integrity loss, assessed by flow cytometry, using propidium iodide as marker. As recently observed for erythroleukemic cells with Brazilian propolis3 , here we present the first evidence that also Portuguese propolis have necrotic-mediated cytotoxicity in yeast cellsEDER/COMPETE/POCI– Operational Competitiveness and Internationalization Programme, under Project POCI-01-0145- FEDER-006958 and National Funds by FCT - Portuguese Foundation for Science and Technology, under the project UID/AGR/04033/201

Portuguese propolis: a source of valuable bioactivities

To FEDER/COMPETE/POCI– Operational Competitiveness and Internationalization Programme, under Project POCI-01-0145-FEDER-006958 and FCT - Portuguese Foundation for Science and Technology, under the project UID/AGR/04033/2013