Are biological antagonists an alternative to synthetic fungicides for preventing postharvest diseases of fruits and vegetables?

The objectives of this research were to develop control measures of postharvest diseases of citrus and deciduous fruits by using naturally-occurring, non-antibiotic-producing antagonists; study the mode of action of effective antagonists and optimize their application methods. Several antagonists were found against a variety of diseases of fruits and vegetables. One particularly effective yeast antagonist (US-7) was chosen for more in-depth studies. This antagonist outcompetes rot pathogens at the wound site for nutrients and space; it is better adapted than the pathogen to extreme environmental conditions such as temperature, humidity and osmotic changes, and is relatively resistant to common postharvest fungicides. Our data suggests that other modes of action may also be involved. These are induction of host resistance by the antagonists or its products, and direct interaction between the antagonists and the pathogen with the possible involvement of an extracellular material and/or cell wall degrading enzymes produced by the antagonist. However, these interactions were not fully elucidated. The antagonistic activity of US-7 and other biocontrol agents isolated, was enhanced by calcium salts. While the mode of action is not known, the addition of these salts had a significant effect both in laboratory experiments and in large-scale tests. Compatibility of the yeast antagonist with present packinghouse treatments and procedures was determined. An integrated control procedure was developed, utilizing the antagonists together with ultra-low dosages of fungicides and activity-enhancing additives. This cooperative research resulted in numerous publications describing the antagonistic agents. their mode of action and possible commercial application. Patents were developed from this research and a commercial company is pursuing the licensing of these patents and the testing of the procedure on a commercial scale. Our research findings have expanded the potential for using non-antibiotic-producing antagonistic microorganisms in the control of postharvest diseases of fruits and vegetables; thus meeting a critical need to find alternatives to the use of synthetic fungicides on food products.

Postharvest diseases of pome fruit are typically caused by a wide diversity of fungal pathogens, and the list of confirmed causal agents is still growing. There is considerable knowledge on the epidemiology of wound pathogens, such as Botrytis cinerea and Penicillium expansum. In contrast, knowledge on the occurrence of the different postharvest diseases caused after latent (quiescent) infections during long-term storage and their epidemiology is limited. Well-known pathogens causing postharvest losses after latent infections are Neofabraea spp. and Colletotrichum spp., but in many cases the causal agents that occur in a specific region remain unknown and their control relies on the routine use of fungicide applications. However, due to the growing concern over the use of synthetic fungicides, alternative control measures are highly desired. Over the past years the use of physical treatments, natural compounds, and biocontrol agents have been investigated as alternatives. However, no single method has emerged that can robustly and reliably control postharvest diseases of pome fruit in practice. In this review it is argued to approach latent postharvest diseases as complex problems that require multiple interventions at different stages of the disease process in a systems intervention approach for their control. Such approach requires a deep understanding of the epidemiology of the causal agents in the orchard, fruit defence mechanisms against pathogens, and the molecular biology of host-pathogen interactions in order to develop novel disease control methods in which the deployment of resistant cultivars can be a cornerstone.

Postharvest diseases of fruits and vegetables cause serious losses of fresh produce-world-wide. Most rot pathogens are controlled by various methods such as refrigeration, controlled atmosphere and fungicides. Biological control strategies are emerging as promising alternatives to the use of synthetic fungicides. Several antagonistic microorganisms have been found that can effectively inhibit postharvest diseases of fruits and vegetables.

Disease management may require multiple decisions by growers on whether to apply a pesticide, the frequency of applications, and also the type of pesticide to use. We developed models for estimating the probability of switching behavior by hop growers related to use of nonsynthetic or synthetic fungicides, or mixtures thereof, for hop powdery mildew (Podosphaera macularis). Growers used nonsynthetic fungicides alone in 61.5% of their applications, later switching to synthetic fungicides or mixtures of the two in 21.5 or 17.0% of their total applications, respectively. Binary logistic regression predicted that the likelihood of switching fungicide type was associated with use of a nonsynthetic fungicide in the first application of the year and how early in the season growers made that application. The predicted probability of switching increased proportionate to the incidence of plants with powdery mildew but with a weaker effect. Multinomial logistic regression modeling of switching between nonsynthetic, synthetic, or mixtures of fungicides indicated a complex pattern of switching behavior. A random forest classification algorithm identified the most important variables in the multinomial logistic regression model as time of year, individual grower effects, the date of the first application, seasonal mean disease incidence, thoroughness of spring pruning, and cultivar susceptibility to two races of the fungus. Overall, our analyses indicate that growers switch from nonsynthetic fungicides to more active (and more expensive) synthetic fungicides or mixtures thereof at critical periods of crop susceptibility and when expected or observed disease incidence increases. Reducing the use of synthetic fungicide inputs may be closely linked to disease risk mitigation, as risk mitigation is indissolubly linked to growers' decisions to switch to these more potent fungicides or fungicide mixtures.

Postharvest loss of sweet pepper and other fresh produce is a major challenge throughout the world. The control of the loss of these valuable farm produce is primarily based on the use of synthetic fungicides. Nevertheless, there are concerns based on the impact of these chemicals on the environment and human health. Hence, call for a much safer and environmentally friendly alternative. Among the various biological alternatives, the use of bacterial antagonistic strain is becoming popular throughout the globe. Bacterial antagonists are now controlling a number of postharvest pathogens. Several modes of action have been suggested by which microbial antagonists inhibit the growth of postharvest pathogens. However, very little is known about the overall diversity of microbial communities on harvested produce; and how these communities can serve as a foundation for research on postharvest biocontrol. Competition for nutrients and space is the most widely accepted mechanism of action for bacterial antagonists. In addition, antibiosis through the production of antibiotics, mycoparasitism through the production of cell wall lytic enzymes, production of volatile organic compounds and induced resistance are other modes of bacterial antagonist actions by which they suppress the activity of postharvest pathogens.

As an alternative to the use of synthetic chemical fungicides to control plant disease, aluminium‐containing salts were evaluated for their effects on the mycelial growth of various fungal or fungus‐like pathogens and their ability to control carrot cavity spot (Pythium sulcatum) and potato dry rot (Fusarium sambucinum). Results showed that various aluminium‐containing salts provided strong inhibition of all the tested pathogens (Alternaria solani, Botrytis cinerea, F. sambucinum, P. sulcatum and Rhizopus stolonifer) with minimal inhibitory concentration of 1–10 mM. Aluminium chloride and aluminium sulphate were generally the most effective, inhibiting mycelial growth of pathogens by as much as 47% and 100%, respectively, at a salt concentration of 1 mM. Applied at 5 mM, aluminium sulphate also provided 28% and 100% inhibition of dry rot and cavity spot, respectively. Aluminium chloride (5 mM) reduced dry rot by 25% whereas aluminium lactate (5 mM) decreased cavity spot lesions by 86%. These results indicate that various aluminium‐containing salts may provide an alternative to the use of synthetic fungicides to control these pathogens.

17 - Natural antimicrobials for preserving fresh fruit and Vegetables

Endophytes: Saviour of apples from post-harvest fungal pathogens

Postharvest diseases in fruits cause serious losses of fresh produce worldwide. The application of synthetic fungicides for the control of postharvest diseases such as anthracnose in mangoes can cause adverse effects on the environment and on human and animal health and has generated phytopathogen resistance. Biological control with the application of marine yeasts and chlorine dioxide (ClO2) to reduce the use of synthetic fungicides can be an alternative to prevent anthracnose in Ataulfo mango fruits. The results showed that different doses of ClO2 inhibited the mycelium growth and spore germination of Colletotrichum gloeosporioides in vitro . When ClO2 and the marine yeasts Debaryomyces hansenii and Rhodotorula minuta were applied to mango fruits, no signs of anthracnose disease incidence and lesion diameter were observed (LSD, p<0.05). Therefore, the application of ClO2 plus antagonist yeasts provides excellent control of anthracnose disease in Ataulfo mango fruits.

In “Rocha” pear, postharvest diseases, blue and grey moulds caused by Penicillium expansum and Botrytis cinerea, respectively, are particularly important. The control of postharvest decay has been based on the use of synthetic fungicides. However, there are risks to consumer health and the environment. In previous work, the antagonistic activity of the yeast-like fungus Aureobasidium pullulans (isolated from the surfaces of fruit, leaves and branches of “Rocha” pear trees), when applied as a wound treatment, was able to significantly reduce postharvest decay caused by P. expansum under storage conditions, and during simulated shelf-life. For more effective prevention and reduction of postharvest diseases in fruits it is necessary to understand the complex interactions in biological control between host, pathogen and antagonist, for which information is currently lacking. Thus, the aims of this study were to evaluate the biocontrol activity of the yeast-like fungus A. pullulans against P. expansum and to elucidate the resistance mechanism involved in the antagonistic activity of A. pullulans to oxidative stress. Fruits of Pyrus communis L. cv. Rocha were stored at −0.5°C, 95% relative humidity for 5.5 months. Fruit maturity was measured in terms of pericarp colour, sugar analysis, Streif index and flesh firmness. Assays of carbon dioxide and ethylene, electrolyte leakage, ascorbate, glutathione and antioxidant enzymes were used to evaluate oxidative stress. Furthermore, malondialdehyde, considered as a final product of lipid peroxidation, and the levels of hydrogen peroxide and methanesulfonic acid were determined and linked to oxidative stress impact on the lipid phase of cell membranes. During a 5-month period, fruit firmness, value of hue angle and colour, and Streif index changed significantly. Sucrose content decreased whereas fructose, glucose and sorbitol levels increased. After inoculation with A. pullulans and infection with P. expansum, a fungistatic, rather than fungicidal, effect was detected from 2.5 months onwards. Three months after storage, the higher superoxide dismutase activity and hydrogen peroxide levels caused by A. pullulans plus P. expansum were possibly associated with additional unsaturated fatty acids, as reflected by increased acyl lipid peroxidation. After five months of storage, in tissues inoculated with A. pullulans followed by infection with P. expansum, oxidative stress was linked to a general increase in the antioxidant peroxidase system activity. Glutathione and ascorbate content also increased along with the loss of cellular membrane selectivity. The results revealed a fungistatic effect of A pullulans on P. expansum. The prevalence of oxidative stress on “Rocha” pear wounds during the storage period was characterized, taking into account the interactions between A. pullulans and P. expansum. The information gained forms the basis for a biocontrol strategy, and the development of successful formulations for application in the orchard and after harvest. This strategy could eradicate and protect fruit against postharvest diseases, and is thus a promising alternative to the use of synthetic pesticides, being safe for the environment and for human health.

BackgroundFungi are one of the microorganisms that cause most damage to fruits worldwide, affecting their quality and consumption. Chemical controls with pesticides are used to diminish postharvest losses of fruits. However, biological control with microorganisms or natural compounds is an increasing alternative to protect fruits and vegetables. In this study, the antifungal effect of Streptomyces sp. CACIS-1.5CA on phytopathogenic fungi that cause postharvest tropical fruit rot was investigated.Main bodyAntagonistic activity was evaluated in vitro by the dual confrontation over fungal isolates obtained from grape, mango, tomato, habanero pepper, papaya, sweet orange, and banana. The results showed that antagonistic activity of the isolate CACIS-1.5CA was similar to the commercial strain Streptomyces lydicus WYEC 108 against the pathogenic fungi Colletotrichum sp., Alternaria sp., Aspergillus sp., Botrytis sp., Rhizoctonia sp., and Rhizopus sp. with percentages ranging from 30 to 63%. The bioactive extract obtained from CACIS-1.5 showed a strong inhibition of fungal spore germination, with percentages ranging from 92 to 100%. Morphological effects as irregular membrane border, deformation, shrinkage, and collapsed conidia were observed on the conidia. Molecularly, the biosynthetic clusters of genes for the polyketide synthase (PKS) type I, PKS type II, and NRPS were detected in the genome of Streptomyces sp. CACIS-1.5CA.ConclusionsThis study presented a novel Streptomyces strain as a natural alternative to the use of synthetic fungicides or other commercial products having antagonistic microorganisms that were used in the postharvest control of phytopathogenic fungi affecting fruits.

For plant protection, reliance on agrochemicals is not a sustainable measure, because of the inadequate bioavailability of their active ingredients. Use of synthetic fungicides in controlling plant fungal diseases is routine. The permeability and solubility of active compounds in fungicides determine their bioavailability. Extensive use of fungicides for disease management results in pollution and hazardous impacts on the ecosystem. In this era of modern technologies, nanotechnology provides a smart solution to this problem at the nanoscale level. Synthesis of nanoparticles through physical, chemical, and biological methods is an emergent field of nanotechnology. Nanoparticles are used in making different nanoformulations, nanoemulsions, and nanocapsules. Among all nanoparticles, use of metallic nanoparticles in formulating nanofungicides is common. These nanofungicides offer targeted delivery, enhanced bioavailability due to greater solubility and permeability, small doses, less dose-dependent toxicity, and controlled release. The kingdom Fungi is a diverse group of organisms, and synthetic fungicides are not always effective against all phytopathogenic fungi. Thus, in this scenario, nanoparticles can be used as alternative control measures because of their ability to permeate through biomembranes and their large surface areas, which are attributed to their extremely small size. Among metallic nanoparticles, silver nanoparticles have strong antifungal potential. Silver ions inactivate thiol groups in the fungal cell wall, resulting in cell lysis and DNA mutation, thereby disrupting membrane processes (such as the membrane electron transport chain and transmembrane energy metabolism) and dissociating enzyme complexes, and ultimately blocking the respiratory chain in the fungus. The relationship between the size of nanoparticles and their efficiency and antifungal potential is an inverse one. This chapter provides comprehensive knowledge of nanoparticles, metallic nanoparticles, nanoformulations, nanofungicides, and their roles in disease management in plants. It also provides insight into the miraculous properties of these particles at the nanoscale.

Antifungal activity and chemical composition of twenty essential oils against significant indoor and outdoor toxigenic and aeroallergenic fungi

Post-harvest development of anthracnose in pepper (Capsicum spp): Etiology and management strategies

Fresh fruits and vegetables are susceptible to a large variety of spoilage agents before and after harvest. Among these, fungi are mostly responsible for the microbiological deteriorations that lead to economically significant losses of fresh produce. Today, synthetic fungicides represent the first approach for controlling postharvest spoilage in fruits and vegetables worldwide. However, the emergence of fungicide-resistant pathogen biotypes and the increasing awareness of consumers towardthe health implications of hazardous chemicals imposed an urgent need to reduce the use of synthetic fungicides in the food supply; this phenomenon strengthened the search for alternative biocontrol strategies that are more effective, safer, nontoxic, low-residue, environment friendly, and cost-effective. In the last decade, biocontrol with antagonistic yeasts became a promising strategy to reduce chemical compounds during fruit and vegetable postharvest, and several yeast-based biocontrol products have been commercialized. Biocontrol is a multipartite system that includes different microbial groups (spoilage mold, yeast, bacteria, and nonspoilage resident microorganisms), host fruit, vegetables, or plants, and the environment. The majority of biocontrol studies focused on yeast-mold mechanisms, with little consideration for yeast-bacteria and yeast-yeast interactions. The current review focused mainly on the unexplored yeast-based interactions and the mechanisms of actions in biocontrol systems as well as on the importance and advantages of using yeasts as biocontrol agents, improving antagonist efficiency, the commercialization process and associated challenges, and future perspectives.