Exploitation of Yeasts as an Alternative Strategy to Control Post Harvest Diseases of Fruits-A Review

Transcription

1 World Applied Sciences Journal 31 (5): , 2014 ISSN IDOSI Publications, 2014 DOI: /idosi.wasj Exploitation of Yeasts as an Alternative Strategy to Control Post Harvest Diseases of Fruits-A Review V. Yeka Zhimo, Dawa Dolma Bhutia, Jayanta Saha and Birendranath Panja Department of Plant Pathology, Bidhan Chandra Krishi Viswavidyalaya, Mohanpur, Nadia , West Bengal, India Abstract: There is an increasing concern about the environmental effects and safety of chemical pesticides and fungicides all over the world. Microbial biocontrol agents possess a number of important advantages over traditional chemical pesticides which make their commercial outlook particularly promising and can be commercially developed with relative ease to control post harvest diseases of fruits. Biological control of different plant diseases was focused primarily using bacteria or filamentous fungi and so, application of yeasts as biocontrol agents acts as a new trend against different pathogens. Various observations suggest that competition for space and nutrients between yeasts and pathogens, antibiosis, parasitism, induction of host resistance and also resistance to oxidative stress are likely to be the main mechanisms of action. In order to enhance biocontrol activity of antagonists against fungal pathogens, certain strategies, such as adding calcium salts, carbohydrates, amino acids and other nitrogen compounds to biocontrol treatments are proposed and can prove more beneficial than using yeasts alone. A more thorough understanding of the mechanisms of action, interactions of fruit tissue-pathogen-biocontrol agent and effective formulation techniques are still needed to develop these yeasts into potential candidates in the management of post harvest diseases of fruits. Key words: Biological control Fruits Post-harvest diseases Yeasts INTRODUCTION environment (temperature, relative humidity, atmosphere composition, etc.), produce handling methods, post Post harvest diseases of fruits caused by pathogens harvest hygiene, produce maturity and ripeness stage, pose a major challenge throughout the fruit growing cultivar susceptibility to postharvest diseases and areas of the world accounting to about 20-25% of the treatments used for disease control. Fungicides are harvested during postharvest handling [1-4] in developed commonly used to control these post harvest decays. countries and even more exasperating in the developing However, though harvested fruits treated with fungicides countries, where it often exceeds over 35%, due to retard post harvest diseases, there is a greater likelihood inadequate storage, processing and transportation of direct human exposure to them. Synthetic chemicals facilities [5]. A side from direct economic considerations, are also discouraged due to their carcinogenicity, diseased produce poses a potential health risk. In most teratogenicity, high and acute residual toxicity, cases, fresh produce which is obviously diseased would environmental pollution and their effects on food and not be consumed. A number of fungal genera such as side effects on humans [6-7]. Besides these, phytoxicity Aspergillus, Penicillium, Alternaria and Fusarium are and off odour effects of some fungicides have limited their known to produce mycotoxins under certain conditions. use. Development of resistance to commonly used Generally speaking, the greatest risk of mycotoxin fungicides within populations of post harvest pathogens contamination occurs when diseased fruit produce is used has also become a significant problem. For example, in the production of processed food. Losses due to acquired resistance by Penicillium italicum and postharvest disease are affected by a great number of Penicillium digitatum to fungicides used for treating factors including: commodity type, the postharvest citrus fruits has been reported [8]. This calls for a new Corresponding Author: V. Yeka Zhimo, Department of Plant Pathology, Bidhan Chandra Krishi Viswavidyalaya, Mohanpur, Nadia , West Bengal, India. 785

2 paradigm shift from the use of synthetic fungicides to a sustain many of the pesticides used in the post-harvest safer and environmentally friendly alternative for environment and can grow rapidly on cheap substrates in reducing the postharvest decay in fruits and vegetables fermenters and are therefore easy to produce in large [9-10]. quantities. The suggested modes of action of biocontrol In the last few years, biological control of yeasts are not likely to constitute any hazard for the postharvest diseases of fruits has been developed as a consumer. Furthermore, yeast cells contain high amounts promising alternative to chemical control [11-12]. Among of vitamins, minerals and essential amino acids and the replete of bio-control approaches, the use of the several reports on the beneficial effect of yeast addition microbial antagonists like yeasts, fungi and bacteria is in both food and feed can be found in the literature quite promising and gaining popularity [2, 13-19]. Among [21-23]. the antagonistic microorganisms, natural yeasts have been widely used as biological control agents [20]. Mechanisms of Yeasts in Biological Control: Yeast based technologies and products (e.g Aspire The mechanism by which the yeasts inhibits the target containing Candida oleophila, Yield Plus containing pathogen is not clear and still incomplete due to the C. albidus) for post harvest disease management of fruits difficulties encountered during the study of the complex are already practiced and developed in foreign countries. interactions between host, pathogen, antagonist and Unfortunately, no such technology or product (s) have other microorganisms present making it extremely been developed in India although it is the major fruit difficult to construct experiments that can exclude all producing country in the world and very limited research other possible mechanisms in the complex biocontrol work on this aspect has been carried out. environment. However, several modes of action have The objective of this review is to critically assess the been suggested and so far no one single mechanism has use of yeasts as microbial antagonists for controlling been shown to be responsible for the whole biocontrol postharvest diseases of fruits, in order to reduce the use effect [15]. of chemical agents that may be harmful to humans and the environment. Competition for Space and Nutrition: Competition for space and nutrients is the primary mechanism of action of What are yeasts?: Yeasts are a heterogenous group of most of the biocontrol agents. Rapid colonization of fruits fungi that superficially appear to be homogeneous. by biocontrol agents is critical for decay control and They grow in a conspicuous unicellular form that manipulations leading to improved colonization enhance reproduces by fission, budding, or a combination of both. biocontrol [24]. They should be able to grow more rapidly True yeasts reproduce sexually, developing than the pathogen and should have the ability to survive ascospores or basidiospores under favourable conditions. even under conditions that are unfavorable to the The majority of ascomycetous and basidiomycetous pathogen. The biocontrol activity of some yeasts increase yeasts isolated in laboratories go unrecognized because with their increasing concentrations and decreasing most of them are heterothallic. In most instances, only one concentrations of pathogens. For example, Candida of the mating types is isolated and therefore no asci or 7 saitona was effective at a concentration of 10 CFU/ml for basidia are produced. Yeast-like fungi (imperfect yeasts) controlling Penicillium expansum on apples fruits and reproduce only by asexual means. The identification of less effective when concentrations were decreases [25]. these fungi is based upon a combination of morphological In another study, it was reported that for Candida and biochemical criteria. Morphology is primarily used to 8 saitona, a concentration of 10 CFU/ml was better in establish the genera, whereas biochemical assimilations controlling Penicillium expansum on apples [26]. are used to differentiate the various species. Several This qualitative relationship, however, is highly important properties of yeasts make them stand out dependent on the ability of the antagonists to multiply among others for biological control purposes: and grow at the wound site. This was demonstrated by (i)yeasts do not produce allergenic spores or using a mutant of Pichia guilliermondii, which lost its mycotoxins as many mycelial fungi do or antibiotic biocontrol activity against Penicillium digitatum on metabolites as possibly produced by bacterial antagonists grapefruit and against Botrytis cinerea on apples, even (ii). They generally have simple nutritional requirements when applied to the wounds at concentrations as high as and are able to colonize dry surfaces for long periods of CFU/ml [14]. The cell population of this mutant time.. (iii) They rapidly utilize available nutrients and can remained constant at the wound sites during incubation 786

3 period, while that of the wild type increased 10- to 20-fold, registered for use on food or feed. Moreover, similar within 24 h hours. Competition for nutrients has been problems with resistant pathogen strains, as experienced suggested as the mode of action of several yeasts like today with fungicides, would probably rapidly occur if a Pichia guilliermondii against Penicillium digitatum [27], one-substance effect was the only mechanism involved. Candida guilliermondii, Cryptococcus laurentii and Metschnikowia pulcherima against Botrytis cinerea and Parasitism: Parasitism by yeasts has been suggested Penicillium expansum [13, 28-31]. It was demonstrated as a fungus-inhibiting mechanism. The yeast Pichia that addition of exogenous nutrients reduced the guilliermondii inhibits B. cinerea and adheres strongly efficacy of P. guilliermondii against P. digitatum [27]. to the fungal mycelium [37]. It has also been shown that The antagonistic yeasts Candida lauretii and P. anomala strain K has a strong production of Sporobolomyces roseus had stronger sugar consumption ß-1,3-glucanase enzyme that degrades the fungal cell wall than the pathogen B. Cinerea by blocking fungus [38]. Aureobasidium pullulans in apple wounds conidial germination due to the deprivation of nutrients produces extracellular exochitinase and -1,3-glucanase [32]. However, no differences in sugar consumption were which could play a role in the biocontrol activity [39]. observed between yeasts with and without biocontrol activity, suggesting that additional factors are part of the Induction of Host Resistance: The characteristic defense inhibiting mechanisms. In fruit wounds, competition for response in fruit includes production of inhibitors of cell nutrients is probably extended to other nutrients, such as wall-degrading enzymes of the pathogen, activity of nitrogen compounds present in low concentration. Both antifungal compounds (such as phenolic compounds organic and inorganic sources of nitrogen can be utilized and phytolaexins), active oxygen species and by yeast for growth. Amino acids, amines and urea are all reinforcement of the cell wall of the host [40]. The suitable sources of nitrogen for all yeasts, as inorganic resistance induction is due to the antagonist ability ammonium salts while nitrate utilization is confined to a to elicit host plant defense responses. It involves certain species or genera of yeasts and this is a valuable several chemical or biochemical reactions in the host diagnostic character used for identification purposes tissue, including changes of tissue structure and too. Although very few species can hydrolyse production of pathogenesis-related proteins, which proteins extracellularly, short peptides can be transported can be expressed locally or systemically [41-42]. into the cell and utilized intracellularly [33]. In a tissue P. guillermondii has been shown to stimulate the culture plate system with membrane inserts separating production of ethylene in grapefruit [37]. Candida famata the organism, Janisiewicz et al., [34] were able to show (F35) stimulates the production of the phytoalexins, that the yeast Aureobasidium pullulans consumes the scoparone and scopoletin in the wound site of oranges amino acids and inhibits germination of P. expansum in [43]. Candida oleophila was found to induce resistance apple juice. A non-destructive method using tissue to P. digitatum when applied in the surface of both culture plates: a defusing membrane at the lower end of wounded and unwounded grapefruit [44]. Some Candida cylindrical inserts to o study the competition for nutrients strains are able to cause chemical and osmotic changes in separated from the competition for space has been apple tissues and thus favouring antagonist settlement developed [34]. [25]. P. guilliermondii strain has been shown to stimulate the production of ethylene, a hormone in grapefruit able Antibiosis: Production of antibiotic substances is a to activate the phenylalaninammoniumlyase, an enzyme commonly suggested mode of action for bacterial involved in the synthesis of phenols, phytoalexins and biocontrol agents [35]. It has been shown that the yeast lignins [37]. Accumulation of phytoalexins, i.e., scoparon Pseudozyma flocculosa produces extracellular fatty acids and scopoletin has been observed in citrus fruits treated that are detrimental to powdery mildew [36]. Biocontrol with yeast cells [45]. agents producing antibiotic metabolites could be effective against a wide range of target organisms, including Resistance to Oxidative Stress: Resistance to oxidative pathogens that have occurred prior to the application of stress is another suggested mode of action of some the biocontrol agent, as well as against latent infections. yeast. A model system consisting of two yeasts with However, given the current debate about the antibiotic higher (Cryptococcus laurentii LS-28) or lower resistance of human pathogens it is doubtful that an (Rhodotorula glutinis LS-11) antagonistic activity antibiotic-producing biocontrol agent would be against the postharvest pathogens Botrytis cinerea and 787

4 Penicillium expansum was analysed and found that Commercially Available Yeast Biocontrol Products: Cryptococcus laurentii LS-28 exhibited faster and greater There are currently three commercial yeast biocontrol colonization of wounds than Rhodotorula glutinis LS-11 products available on the market for combating [46]. In contrast to LS-28, the number of LS-11 cells post-harvest decays in fruit. Aspire (Ecogen, Inc., dropped 1 and 2 h after application and then increased Langhorne, Pa) is based on the yeast Candida oleophila only later. In vitro, LS-28 was more resistant to and is used as a spray or dip against post-harvest ROS-generated oxidative stress. The combined diseases on pome and citrus fruits [15, 65]. The product application of biocontrol yeasts and ROS-deactivating was introduced commercially in the United States in enzymes in apple wounds prevented the decrease in The commercial product Yield Plus with Cryptococcus number of LS-11 cells mentioned above and enhanced albidus as the active antagonist was introduced colonization and antagonistic activity of both biocontrol commercially on the South African market in 1997 by yeasts against B. cinerea and P. expansum. Anchor Yeast. Yield Plus is used as a biocontrol To summarize, the activity of a biocontrol agent product against Botrytis, Penicillium and Mucor on seems to be primarily dependent on its ability to rapidly apples and pears and is also under evaluation for other colonize the wound site and compete for nutrients, but crops (Lisa Picard, Anchor Yeast, Cape Town, South may also depend on its ability to attach firmly to hyphae Africa, pers. comm.). The recent product Shemer, of the pathogen, produce cell wall degrading enzymes registered in Israel, is based on a newly identified yeast or volatile compounds, or induce host resistance. Metschnikowia fructicola [66] and is effective against a An example: it has been demonstrated that direct wide range of pathogens of grape, strawberry and sweet attachment, competition for nitrogen and carbon sources, potato. secretion of hydrolytic enzymes and induction of host resistance play a role in the biocontrol mechanisms of Combination of Some Safe Compounds with Yeasts: P. guilliermondii M8 against B. cinerea [47]. Biological means cannot at the moment solve all the problems of postharvest rots during fruit storage and Yeasts as a Biological Control Candidate in the they must be considered instruments to be used in Management of Post Harvest Diseases: Yeasts are combination with other methods in an integrated vision particularly attractive as biological control candidates of disease management. For example, biocontrol agents and are encouraged by the fact that these microorganisms can be combined with waxes and fungicides applied not are frequently associated with foods and might be more only in post but also in pre-harvest [67]. In order to acceptable to consumers than exotic bacteria [48] and also enhance biocontrol activity of antagonists against fungal there is considerable information available with respect pathogens, certain strategies, such as adding calcium to techniques for genetic manipulation, production and salts, carbohydrates, amino acids and other nitrogen storage of yeast cells [49]. Moreover, yeasts as a major compounds to biocontrol treatments, are proposed component of the epiphytic microbial community on the [68-71]. Many researchers have shown that calcium surfaces of fruits and vegetables [50], may be a more plays an important role in the inhibition of postharvest effective biocontrol agent because they are decay of fruits [72-73]. Postharvest calcium treatment of phenotypically adapted to this niche and, thus, are able apples provided broad-spectrum protection against the to more effectively colonize and compete for nutrients postharvest pathogens of Penicillium expansum and and space on fruit surfaces. In the last two decades, Botrytis cinerea [74]. The addition of CaCl 2 (2% w/v) to several scientific studies have demonstrated the the formulation of the yeast biocontrol agent, Candida efficiency of yeasts as biocontrol agents against a wide oleophila, enhanced the ability of this yeast to protect variety of phytopathogens [51-60]. Among yeasts, apples against postharvest decay [75]. The efficacy of Candida spp. have been found to be highly effective controlling grey mould and blue mould rots in apples was against different fungal pathogens [25], Hanseniaspora enhanced when Trichosporon sp., even at a low uvarum against Rhizopus stolonifer and Botrytis concentration of 10 5 CFU ml 1, was applied in the cinerea [61], Cryptococcus spp. against grey mould, presence of CaCl 2 (2% w/v) in an aqueous suspension blue mould and Mucor rot [62]. Yeast strains from species, [76]. However, application of 68 mm CaCl 2 to grapefruit Pichia guilliermondii, Debaryomyces hansenii and reduced the incidence of green mold decay by 43% Kloeckera apiculata have been shown to have the and in combination with an antagonist P. guilliermondi ability to control post-harvest diseases of several fruits strain US-7 by 97% [77]. Other compounds like [25, 63-64]. sodium bicarbonate and ammonium molybdate exhibit 788

5 broad-spectrum antifungal activity [78-79]. Chitosan and field crop or soil-borne diseases, successful commercial its derivatives, including glycolchitosan, were also control of postharvest diseases of fruit and vegetables reported to inhibit fungal growth and to induce host- must be extremely efficient, in the range of 95-98%. As of defense responses in plants and harvested commodities today, such levels of control can be reliably reached by [80]. Combining 0.2% glycolchitosan with the antagonist bio- fungicides only when supplemented with low levels Candida saitoana was more effective in controlling green of synthetic chemical fungicides. A more thorough mould of oranges and lemons, caused by P. digitatum understanding of the microbial ecology of fruit surfaces and gray and blue molds of apples than either treatment will help us figure out which problems to work on, how to alone [81]. The addition of ammonium molybdate to the approach them, when and where to apply the biocontrol yeast Candida sake, significantly enhanced its biocontrol agent and predict situations in which biocontrol would efficacy in controlling postharvest decay in pear fruits not be expected to work. [82]. However, there was little reported use of sodium bicarbonate as an additive to improve the efficacy of REFERENCES yeasts against fungal diseases in fruits. 1. El-Ghaouth, A., C.L. Wilson and M.E. Wisniewski, Challenges for the Future and Concluding Remarks: Biologically based alternatives to synthetic Over the past 20 years, biocontrol research has evolved fungicides for the postharvest diseases of fruit toward being more integrated into a production systems and vegetables. In: Naqvi, S.A.M.H. (Ed.), Diseases approach with more awareness of industry concerns. of Fruit and Vegetables, vol. 2. Kluwer Academic As noted in this review, more research is needed in Publishers, The Netherlands, pp: many aspects of the science and technology of yeast 2. Droby, S., Improving quality and safety of postharvest biocontrol and in integrating the agents into fresh fruit and vegetables after harvest by the use combined pre- and postharvest production and handling of biocontrol agents and natural materials. Acta systems and combining with safe chemicals in preventing Horticulturae, 709: postharvest diseases and can be used to control mixed 3. Zhu, S.J., Non-chemical approaches to decay populations of fungicide-sensitive and fungicide-resistant control in postharvest fruit. In: Noureddine, B., Norio, pathogens [83]. Although several mechanisms of action S. (Eds.), Advances in Postharvest Technologies have been suggested for postharvest biocontrol agents, for Horticultural Crops. Research Signpost, a deeper understanding of the tritrophic interactions of Trivandrum, India, pp: fruit tissue-pathogen-biocontrol agent is still needed. 4. Singh, D. and R.R. Sharma, Postharvest Induced resistance has been postulated to be one of the diseases of fruit and vegetables and their mechanisms of action of yeasts as postharvest biocontrol management. In: Prasad, D. (Ed.), Sustainable Pest agents. However, information about elicitors/effectors of Management. Daya Publishing House, New Delhi, the antagonist involved and our ability to genetically and India. physiologically manipulate them is still lacking. Questions 5. Abano, E.E. and L.K. Sam-Amoah, Effects of related to the effect of host physiology on bio- control different pretreatments on drying characteristics of activity are also unresolved. More research effort is banana slices. Journal of Engineering and Applied needed in order to address the need to lower the effective Sciences, 6(3): biomass and the inherent production costs of 6. Lingk, W., Health risk evaluation of pesticide antagonistic microorganisms to be used in practical contaminations in drinking water. Gesunde Pflangen, applications and to enhance the efficacy of these 43: beneficial microbes. Suitable formulations of these 7. Unnikrishnan, V. and B.S. Nath, Hazardous agents could play a crucial role in their effectiveness by chemicals in foods. Indian J. Dairy Biosci, 11: increasing their dispersion and colonization on fruit skin, 8. Fogliata, G.M., L.G.J. Torres and L.D. Ploper, by prolonging their survival in practical conditions and Detection of imazalil resistant strains of Penicillium by enhancing the mechanisms of action underlying their digitatum Sacc. in citrus packing houses of Tacuman biological activity. The biological control of postharvest Province (Argentina) and their behaviour against diseases is viewed with caution and scepticism by many current employed and alternative fungicides. Revista in the agricultural community. Unlike the control of tree, Industrial Y Agrícola de Tucumán, 77:

6 9. Ragsdale, N.N. and H.D. Sisler, Social and 22. Bui, K. and P. Galzy, In: Spencer, J. F. T. and political implications of managing plant diseases Spencer, D. M. (Eds.), Yeast Technology. Springerwith decreased availability of fungicides in the Verlag, Berlin, Germany, pp: 407. United States. Annual Review of Phytopathology, 23. Hussein, H.S., R.I. Mackie, N.R. Merchen, D.H. Baker 32: and C.M. Parsons, Effects of oleaginous yeast 10. Mari, M., F. Neri and P. Bertolini, Novel on growth performance, fatty acid composition of Approaches to Prevent and Control Postharvest muscles and energy utilization by poultry. Diseases of Fruit. Stewart Postharvest Review, Bioresource Technology, 55: (6): 4. Stewart Postharvest Solutions Ltd., London, 24. Mercier, J.C.L. and Wilson, Colonization of UK. apple wounds by naturally occurring microflora and 11. Wilson, C.L. and M.E. Wisniewski, Biological introduced Candida oleophila and their effect on control of postharvest diseases of fruit and infection by Botrytis cinerea during storage. vegetables: an emerging technology Annual Biological Control, 4: Review of Phytopathology, 27: McLaughlin, R.J., C.L. Wilson, E. Chalutz, 12. Wisniewski, M.E. and C.L. Wilson, Biological W.F. Kurtzman and S.F. Osman, control of postharvest diseases of fruit and Characterization and reclassification of yeasts used vegetables: recent advances. HortScience, 27: for biological control of postharvest diseases of fruit 13. Roberts, R.G., Postharvest biological control of and vegetables. Applied Environmental gray mold of apple by Cryptococcus laurentii. Microbiology, 56: Phytopathology, 80: El-Ghaouth, A., C.L. Wilson and M.E. Wisniewski, 14. Droby, S., E. Chalutz and C.L. Wilson, Untrastructural and cytochemical aspects of Antagonistic microorganisms as biocontrol agents biocontrol activity of Candida saitona in apple fruit. of postharvest diseases of fruit and vegetables. Phytopathology, 88: Postharvest News Information, 2: Droby, S., E. Chalutz, C.L. Wilson and 15. Janisiewicz, W.J. and L. Korsten, Biological M.E. Wisniewski, Characterization of the control of postharvest diseases of fruit. Annual biocontrol activity of Debaryomyces hansenii in the Review of Phytopathology, 40: control of Penicillium digitatum on grapefruit. 16. Zhang, H., X. Zheng, C. Fu and Y. Xi, Canadian Journal of Microbiology, 35: Postharvest biological control of gray mold rot of 28. Elad, Y., J. Kohl and N.J. Fokkema, Control of pear with Cryptococcus laurentii. Postharvest infection and sporulation of Botrytis cinerea on bean Biology and Technology, 35(1): and tomato by saprophytic yeasts. Phytopathology, 17. Korsten, L., Advances in control of 84: postharvest diseases in tropical fresh produce. 29. Piano, S., V. Neyrotti, Q. Migheli and M.L. Gullino, International Journal of Postharvest Technology and Biocontrol capability of Metschnikowia Innovation, 1(1): pulcherrima against Botrytis postharvest rot of apple. 18. Sharma, R.R., D. Singh and R. Singh, Biological Postharvest Biology and Technology, 11: control of post harvest diseases of fruits and 30. Saligkarias, I.D., F.T. Gravanis and H.A.S. Epton, vegetables by microbial antagonists: A review Biological control of Botrytis cinerea on tomato Biological Control, 50: plants by the use of epiphytic yeasts Candida 19. Zhao, Y., X.F., Shao, K. Tu and J.K. Chen, guilliermondii strains 101 and US 7 and Candida Inhibitory effect of Bacillus subtilis B10 on the oleophila strain I-182: in vivo studies. Biological diseases of postharvest strawberry. Journal of Fruit Control, 25: Science, 24(3): Vero, S., P. Mondino, J.Burgueno, M. Soubes and 20. Irtwange, S., Application of Biological Control M.E. Wisniewski, Characterization of biocontrol Agents in Pre- and Post-harvest Operations. activity of two yeast strains from Uruguay against Agricultural Engineering International. 8, Invited blue mould of apple. Postharvest Biology and Overview 3, A and M University Press, Texas. Technology, 26: Stringer, D.A., Industrial development and 32. Filonow, A.B., Role of competition for sugars evaluation of new protein sources: microorganisms. by yeasts in the biocontrol of gray mold of apple. Proceedings of the Nutrition Society, 41: Biocontrol Science and Technology, 8:

7 33. Yamada, T. and D.M. Ogrydazaik, Extracellular 45. Rodov, V., S. Ben-Yehoshua, R. Albaglis and acid proteases produced by Saccharomycopsis D. Fang, Accumulation of phytoalexins lipolytic, J. Bacteriol., 154: scoparone and scopoletin in citrus fruit subjected to 34. Janisiewicz, W.J., T.J. Tworkoski and C. Sharer, various postharvest treatments. Acta Horticulturae, Characterizing the mechanism of biological control 381: of postharvest diseases on fruits with a simple 46. Castoria, R., L. Caputo, F. De Curtis and V. De Cicco, method to study competition for nutrients Biological Control: Resistance of Postharvest Phytopathology, 90: Biocontrol Yeasts to Oxidative Stress: A Possible 35. Raaijmakers, J.M., M. Vlami and J.T. de Souza, New Mechanism of Action. Phytopathology, Antibiotic production by bacterial biocontrol agents. 93(5): Antonie van Leeuwenhoek, 81: Zhang, D., D. Spadaro, A. Garibaldi and M.L. Gullino, 36. Avis, T.J. and R.R. Bélanger Mechanisms and Potential biocontrol activity of a strain of Pichia means of detection of biocontrol activity of guilliermondii against grey mold of apples and its Pseudozyma yeasts against plant-pathogenic fungi. possible modes of action. Biological control, FEMS Yeast Research, 2: : Wisniewski, M., C. Biles and S. Droby, The use 48. Eckert, J.W., Role of chemical fungicides and of yeast Pichia guilliermondii as a biocontrol agent: biological agents in post harvest disease control. characterization of attachment to Botrytis cinerea. In: Biological control of postharvest diseases of fruits Wilson, C.L., Chalutz, E. (Eds.), Biological Control of and vegetables. In: Workshop Proceedings, vol. 92, Postharvest Diseases of Fruit and Vegetables. Proc. Shepherdstown, VA, September US Department Workshop, US Department of Agriculture, ARS-92, of Agriculture, Agricultural Research Service pp: Publications, pp: Jijakli, M.H. and P. Lepoivre, Characterization of 49. Hofstein, R., B. Friedlender, E. Chalutz and S. Droby, an exo-1, 3-glucanase produced by Pichia anomala Large scale production and pilot testing of strain K, antagonist of Botrytis cinerea on apples. biocontrol agents of postharvest diseases. In: Phytopathology, 88: Wilson, C.L., Wisniewski, M. (Eds.), Biological 39. Castoria, R., F. de Curtis, G. Lima, L. Caputo, Control of Postharvest Diseases-Theory and S. Pacifico and V. de Cicco, Aureobasidium Practice. CRC Press Inc., Boca Raton, FL, USA, pullulans (LS-30), an antagonist of postharvest pp: pathogens of fruits: study on its mode of action. 50. Beech, F.W. and R.R. Davenport, The role of Postharvest Biology and Technology, 32: yeast in cidermaking. In: Rose, A.H., Harrison, 40. Jamalizadeh, M., H.R. Etebarian, H. Aminian and I.S. (Eds.), Yeast. Academic Press, London, A. Alizadeh, A review of mechanisms of action pp: of biological control organisms against postharvest 51. Wang, Y.F., Y.H. Bao, D.H. Shen, W. Feng, fruit spoilage. EPPO Bull., 41: T. Yu, J. Zhang and X.D. Zheng, Kloepper, J.W., S. Tuzun and J.A. Kuc, Biocontrol of Alternaria alternata on cherry Proposed definition related to induced disease tomato fruit by use of marine yeast resistance. Biocontrol Science Technology, Rhodosporidium paludigenum Fell and Tallman. 2: International Journal of Food Microbiology, 42. van Loon, L.C., Induced resistance in plants 123: and the role of pathogenesis-related proteins. 52. Hashem, M. and S. Alamri, The biocontrol of European Journal of Plant Pathology, 103: postharvest disease (Botryodiplodia theobromae) of 43. Arras, G., Mode of action of an isolate of guava (Psidium guajava L.) by the application of Candida famata in biological control of Penicillium yeast strains. Postharvest Biology and Technology, digitatum in orange fruit. Postharvest Biology and 53: Technology, 8: Rosa, M.M., S.M. Tauk-Tornisielo, P.E. Rampazzo 44. Droby, S., V. Vinokur, B. Weiss, L. Cohen, A. Daus and S.R. Ceccato-Antonini, Evaluation of the and E.E. Goldschmidt, Induction of resistance biological control by the yeast Torulaspora globosa to Penicillium digitatum in grapefruit by the yeast against Colletotrichum sublineolum in sorghum. biocontrol agent Candida oleophila. Phytopathology, World Journal of Microbiology and Biotechnology, 92: :

8 54. Zhang, D., D. Spadaro, A. Garibaldi and M.L. Gullino, 63. Chalutz, E., R. Ben-Arie, S. Droby, L. Cohen, B. Weiss Selection and evaluation of new antagonists for their efficacy against postharvest brown rot of peaches. Postharvest Biology and Technology, 55: Kong, Q., S. Shan, Q. Liu, X. Wang and F. Yu, Biocontrol of Aspergillus flavus on peanut kernels by use of a strain of marine Bacillus megaterium. International Journal of Food Microbiology, 139: Li, R., H. Zhang, W. Liu and X. Zheng, Biocontrol of postharvest gray and blue mold decay of apples with Rhodotorula mucilaginosa and possible mechanisms of action. Int. Journal of Food Microbiology, 146: Manso, T. and C. Nunes, Metschnikowia andauensis as a new biocontrol agent of fruit postharvest diseases. Postharvest Biology and Technology, 61: Robiglio, A., M.C. Sosa, M.C, Lutz, C.A. Lopes and M.P. Sangorrín, Yeast biocontrol of fungal spoilage of pears stored at low temperature. International Journal of Food Microbiology, 147: Rosa-Magri, M.M., S.M. Tauk-Tornisielo and S.R. Ceccato-Antonini, Bioprospection of yeasts as biocontrol agents against phytopathogenic molds. Brazilian Archives of Biology and Technology, 54: Zhang, D., D. Spadaro, A. Garibaldi and M. L. Gullino, Potential biocontrol activity of a strain of Pichia guilliermondii against grey mold of apples and its possible modes of action. Biological control, 57: Ben-Arie, R., S. Droby, J. Zutkhi, L. Cohen, B. Weiss, P. Sarig, M. Zeidman, A. Daus and E. Chalutz, Preharvest and postharvest biological control of Rhizopus and Botrytis bunch rots of table grapes with antagonistic yeasts. In: Wilson, C.L., Chalutz, E. (Eds.), Biological Control of Postharvest Diseases of Fruits and Vegetables. ARS Publication 92, USDA-ARS. Washington DC, USA, pp: Roberts, R.G., Characterization of postharvest biological control of deciduous fruit diseases by Cryptococcus spp. Biological Control of Postharvest and Diseases of Fruits and Vegetables Wrkshp. Proc., Shepherdstown, W.Va., Sept U.S. Dept. Agr.-Agr. Res. Serv. Publ., 92: and C.L. Wilson, Yeasts as biocontrol agents of postharvest diseases of fruit. Phytoparasitica, 16: Wilson, C.L. and E. Chalutz, Postharvest biocontrol of Penicillium rots of citrus with antagonistic yeasts and bacteria. Scientia Horticulturae, 40: Janisiewicz, W.J., B. Leverentz, W.S. Conway, R.A. Saftner, A.N. Reed and M.J. Camp, Control of bitter rot and blue mold of apples by integrating heat and antagonist treatments on 1-MCP treated fruit stored under controlled atmosphere conditions. Postharvest Biology and Technology, 29: Kurtzman, C.P. and S. Droby, Metschnikowia fructicola, a new ascosporic yeast with potential for biocontrol of postharvest fruit rots. Systematic and Applied Microbiology 24 (3), Conway, W.S Effect of postharvest calcium treatment on decay of delicious apples. Plant Disease, 66: Pusey, P.L., C.L. Wilson, M.W. Hotchkiss and J.D. Franklin, Compatibility of Bacillus subtilis for postharvest control of peach brown with commercial fruit waxes, dicloran and cold-storage conditions. Plant Disease, 70: Conway, W.S., Effect of postharvest calcium treatment on decay of delicious apples. Plant Disease, 66: Conway, W.S., G.M. Greene and K.D. Hickey, 1987a. Effects of preharvest and postharvest calcium treatments of peaches on decay caused by Monilinia fructicola. Plant Disease, 71: Conway, W.S., K.C. Gross and C.E. Sams, 1987b. Relationship of bound calcium and inoculum concentration to the effect of postharvest calcium treatment on decay of apples by Penicillium expansum. Plant Disease, 71: Janisiewicz, W.J., J. Usall and B. Boras, Nutritional enhancement of biocontrol of blue mold of apples. Phytopathology, 82: Conway, W.S. and C.E. Sams, Influence of fruit maturity on the effect of postharvest calcium treatment on decay of Golden Delicious apples. Plant Disease, 69: Conway, W.S., C.E. Sams, R.G. McGuire and A. Kelman, Calcium teatment of apples and potatoes to reduce postharvest decay. Plant Disease, 76:

9 74. Saftner, R.A., W.S. Conway and C.E. Sams, Nunes, C., J. Usall, N. Teixido, E. Fons and I. Vinas, Effects of some polyamine biosynthesis inhibitors 2002a. Postharvest biological control by Pantoea and calcium chloride on in vitro growth and decay agglomerans (CPA-2) on Golden Delicious apples. development in apples caused by Botrytis cinerea Journal of Applied Microbiology, 92(2): and Penicillium expansum. Journal of American 80. Allan, C.R. and L.A. Hadwinger, The fungicidal Society Horticultural Science, 122: effect of chitosan on fungi of varying cell 75. Wisniewski, M.E., S. Droby, E. Chalutz and Y. Eilam, wallcomposition. Experimental. Mycology, 3: Effect of Ca2+ and Mg2+ on Botrytis cinerea 81. El-Ghaouth, A., J.L. Smilanick and C.L. Wilson, and Penicillium expansum in vitro and on the Enhancement of the performance of Candida saitona biocontrol activity of Candida oleophilla. Plant by the addition of glycochitosan for control of Pathology, 44: postharvest decay of apple and citrus fruit. 76. Tian, S.P., Q. Fan, Y. Xu and Y. Wang, Effects Postharvest Biology and Technology, 19: of Trichosporon sp. in combination with calcium and 82. Nunes, C., J. Usall, N. Teixido, R. Torres and I. Vinas, fungicide on biocontrol of postharvest diseases in 2002b. Control of Penicillium expansum and Botrytis apple fruits. Acta Botanica Sinica, 43: cinerea on apples and pears with a combination of 77. Droby, S., M.E. Wisniewski, L. Cohen, B. Weisis, Candida sake (CPA-1) and Pantoea agglomerans. D. Toutou, Y. Eilam and E. Chalutz, Influence of Journal of Food Protection, 65: CaCl2 on Penicillium digitatum grapefruit peel tissue 83. Lima G., F.D. Curtis, D. Piedimonte, A.M. Spina and and biocontrol activity of Pichia guilliermondii. V.D. Cicco, Integration of biocontrol yeast and Phytopathology, 87: thiabendazole protects stored apples from fungiciude 78. Corral, L.G., L.S. Post and T.J. Montville, sensitive and resistant isolates of Botrytis cinerea. Antimicrobial activity of sodium bicarbonate. Postharvest Biology and Technology, 40: Journal of Food Science, 53: