Shelf Life Extension of Fresh Fruit and Vegetables by Chitosan Treatment

This art icle was downloaded by: [ 189.226.100.40] On: 06 June 2015, At : 08: 02 Publisher: Taylor & Francis I nform a Lt d Regist ered in England and Wales Regist ered Num ber: 1072954 Regist ered office: Mort im er House, 37- 41 Mort im er St reet , London W1T 3JH, UK Critical Reviews in Food Science and Nutrition Publicat ion det ails, including inst ruct ions f or aut hors and subscript ion inf ormat ion: ht t p: / / www. t andf online. com/ loi/ bf sn20 Shelf Life Extension of Fresh Fruit and Vegetables by Chitosan Treatment a a b Gianf ranco Romanazzi , Erica Feliziani , Silvia Baut ist a Baños & Dharini Sivakumar c a Depart ment of Agricult ural, Food and Environment al Sciences, Marche Polyt echnic Universit y, Via Brecce Bianche, 60131 Ancona, It aly b Cent ro de Desarrollo de Product os Biót icos, Inst it ut o Polit écnico Nacional Carr, Yaut epecJoj ut la km 6, San Isidro Yaut epec Morelos 62731, Mexico c Depart ment of Crop Sciences, Tshwane Universit y of Technology, Pret oria West , Pret oria 0001, Sout h Af rica Accept ed aut hor version post ed online: 05 Jun 2015. Click for updates To cite this article: Gianf ranco Romanazzi, Erica Feliziani, Silvia Baut ist a Baños & Dharini Sivakumar (2015): Shelf Lif e Ext ension of Fresh Fruit and Veget ables by Chit osan Treat ment , Crit ical Reviews in Food Science and Nut rit ion, DOI: 10. 1080/ 10408398. 2014. 900474 To link to this article: ht t p: / / dx. doi. org/ 10. 1080/ 10408398. 2014. 900474 D iscla im e r : This is a version of an unedit ed m anuscript t hat has been accept ed for publicat ion. As a service t o aut hors and researchers we are providing t his version of t he accept ed m anuscript ( AM) . Copyedit ing, t ypeset t ing, and review of t he result ing proof will be undert aken on t his m anuscript before final publicat ion of t he Version of Record ( VoR) . During product ion and pre- press, errors m ay be discovered which could affect t he cont ent , and all legal disclaim ers t hat apply t o t he j ournal relat e t o t his version also. PLEASE SCROLL DOWN FOR ARTI CLE Taylor & Francis m akes every effort t o ensure t he accuracy of all t he inform at ion ( t he “ Cont ent ” ) cont ained in t he publicat ions on our plat form . However, Taylor & Francis, our agent s, and our licensors m ake no represent at ions or warrant ies what soever as t o t he accuracy, com plet eness, or suit abilit y for any purpose of t he Cont ent . Any opinions and views expressed in t his publicat ion are t he opinions and views of t he aut hors, and are not t he views of or endorsed by Taylor & Francis. The accuracy of t he Cont ent should not be relied upon and should be independent ly verified wit h prim ary sources of inform at ion. Taylor and Francis shall not be liable for any losses, act ions, claim s, proceedings, dem ands, cost s, expenses, dam ages, and ot her liabilit ies what soever or howsoever caused arising direct ly or indirect ly in connect ion wit h, in relat ion t o or arising out of t he use of t he Cont ent . This art icle m ay be used for research, t eaching, and privat e st udy purposes. Any subst ant ial or syst em at ic reproduct ion, redist ribut ion, reselling, loan, sub- licensing, syst em at ic supply, or dist ribut ion in any form t o anyone is expressly forbidden. Term s & Condit ions of access and use can be found at ht t p: / / www.t andfonline.com / page/ t erm s- and- condit ions ACCEPTED MANUSCRIPT Critical Reviews in Food Science and Nutrition G. ROMANAZZI ET AL. Shelf Life Extension of Fresh Fruit and Vegetables by Chitosan Treatment GIANFRANCO ROMANAZZI1,*, ERICA FELIZIANI1, SILVIA BAUTISTA BAÑOS2, and DHARINI SIVAKUMAR3 1 Department of Agricultural, Food and Environmental Sciences, Marche Polytechnic University, Downloaded by [189.226.100.40] at 08:02 06 June 2015 Via Brecce Bianche, 60131 Ancona, Italy 2 Centro de Desarrollo de Productos Bióticos, Instituto Politécnico Nacional Carr, Yautepec- Jojutla km 6, San Isidro Yautepec Morelos 62731, Mexico 3 Department of Crop Sciences, Tshwane University of Technology, Pretoria West, Pretoria 0001, South Africa * Address correspondence to: G. Romanazzi, Department of Agricultural, Food and Environmental Sciences, Marche Polytechnic University, Via Brecce Bianche, 60131 Ancona, Italy. Phone: +39-071-2204336, Fax: +39-071-2204856. E-mail: g.romanazzi@univpm.it Among alternatives that are currently under investigation to replace the use of synthetic fungicides to control postharvest diseases in fresh produce and to extend their shelf life, chitosan application has shown promising disease control, at both preharvest and postharvest stages. Chitosan shows a dual mode of action, on the pathogen and on the plant, as it reduces the growth of decay-causing fungi and foodborne pathogens and induces resistance responses in the host tissues. Chitosan coating forms a semipermeable film on the surface of fruit and vegetables, thereby delaying the rate of respiration, decreasing weight loss, maintaining the overall quality, 1 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT and prolonging the shelf life. Moreover, the coating can provide a substrate for incorporation of other functional food additives, such as minerals, vitamins or other drugs or nutraceutical compounds that can be used to enhance the beneficial properties of fresh commodities, or in some cases the antimicrobial activity of chitosan. Chitosan coating has been approved as GRAS substance by USFDA, and its application is safe for the consumer and the environment. This review summarizes the most relevant and recent knowledge in the application of chitosan in Downloaded by [189.226.100.40] at 08:02 06 June 2015 postharvest disease control and maintenance of overall fruit and vegetable quality during postharvest storage. Keywords Edible coating, edible film, induced resistance, foodborne pathogens, antimicrobial activity, postharvest storage 2 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT INTRODUCTION Somewhere between 15% and 50% of fruit and vegetables produced on the global scale is lost after harvest (FAO, 2011), mainly due to microbiological spoilage (Kader, 2005). This percentage is greatly increased in developing countries, where the correct technologies for storage of fruit and vegetable are lacking (FAO, 2011). The susceptibility of fresh produce to postharvest diseases and deterioration of quality attributes increases after harvest and during Downloaded by [189.226.100.40] at 08:02 06 June 2015 prolonged storage, as a result of physiological and biochemical changes in the commodities. These changes can favor the development of postharvest pathogens and the incidence of postharvest diseases, which are the major cause of losses through the supply chain. Therefore, the development of decay-control measures that aim to maintain the quality of fruit and vegetables and to provide protection against postharvest diseases after removal from cold storage at the retailer‟s market shelf will be beneficial to reduce these postharvest losses. On the other hand, postharvest disease control for fresh horticultural produce should begin at the farm, and this involves the cultural practices and fungicide applications used. The adverse effects of synthetic fungicide residues on human health and the environment, and the possibility of the development of fungicide-resistant pathogens, have led to intensified worldwide research efforts to develop alternative control strategies. In addition, the current consumer trend is more towards „green‟ consumerism, with the desire for fewer synthetic additives in food, together with increased safety, excellent nutritional and overall quality, and improved shelf-life. Furthermore, there is the potential for foodborne outbreaks due to contamination of fruit in the field through dirty irrigation water or treatments, or at postharvest through human handling or improper sanitation (Beuchat, 2002). 3 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Application of chitosan treatment at the preharvest or postharvest stages has been considered as a suitable alternative treatment to replace the use of synthetic fungicides. This can help to prevent postharvest fruit diseases and to extend storage life, while maintaining the overall quality of the different fresh commodities (Bautista-Baños et al., 2006). Chitosan (poly b-(1-4)Nacetyl-d-glucosamine) has been identified as providing an ideal coating, with antimicrobial properties that can induce plant defense responses when applied to vegetal tissues (Devlieghere Downloaded by [189.226.100.40] at 08:02 06 June 2015 et al., 2004). On the other hand, chitosan coating also provides a substrate for incorporation of other functional natural food additives, which might improve its antimicrobial properties and prevent deterioration of fruit quality (Vargas et al., 2008). Chitosan treatment in the fresh produce industry is safe for the consumer and the environment, and chitosan has been approved by the United State Food and Drug Administration (USFDA) as a „Generally Recognized As Safe‟ (GRAS) food additive (USFDA, 2013). Nowadays, commercial chitosan formulations are available on the market. Some commercial formulations have been tested for the control of postharvest diseases in different fresh produce commodities, as shown in Table 1. The commercial formulations used in plant disease management, not only for the control of postharvest decay of fruit, include: Chitogel (Ecobulle, France) (Ait Barka et al., 2004; Elmer and Reglinski, 2006); Biochikol 020 PC (Gumitex, Lowics, Poland) (Nawrocki, 2006); Armour-Zen (Botry-Zen Limited, Dunedin, New Zealand) (Reglinski et al., 2010); Elexa 4 Plant Defense Booster (Plant Defense Booster Inc., USA) (Elmer and Reglinski, 2006); and Kendal Cops (Iriti et al., 2011). The main differences between practical grade chitosan solutions and commercial chitosan formulations arise from the techniques used for their preparation and application, which is more immediate for the 4 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT commercial formulations. Indeed, while practical grade chitosan needs to be dissolved in an acid medium some hours before use, the commercial formulations can be quickly dissolved in water (Romanazzi et al., 2013). However, nowadays, chitosan-based formulations used either at preharvest or postharvest are not registered as plant protectant products, but as growth adjuvants. The aim of this review is to summarize the most recent published and relevant advances in the application of chitosan for fresh horticultural produce, in terms of postharvest disease Downloaded by [189.226.100.40] at 08:02 06 June 2015 control, maintenance of overall product quality, use as a health promoting compound, and food safety issues. For better clarity, the data obtained for the in vivo applications of chitosan are divided into sections that consider temperate fruit, tropical fruit, and vegetables, as the environment and the way of cultivation differ across these categories. INFLUENCE OF PREHARVEST CHITOSAN APPLICATION OF ON POSTHARVEST DISEASE CONTROL Although many studies have reported on the effectiveness of chitosan treatments at the postharvest stage, the research findings on the evaluation of the preharvest application of chitosan on the control of postharvest decay in fresh produce is limited (Tables 2-4). However, chitosan applications prior to harvest might be suitable for fruit, such as table grapes and strawberries, because these fruit have a bloom on the surface and/or can suffer postharvest wetting or handling. Moreover, preharvest treatment can provide a preventive effect against pathogens, as the development of postharvest disease often arises from an inoculum that survives and accumulates on the fruit surface in the field or in the packaging line after the harvest. Table grape bunches sprayed in the field with solutions of practical grade chitosan at three different concentrations (1%, 0.5%, 0.1%), as once (21 days) or twice (21, 5 days) before 5 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT harvest, showed significantly reduced gray mold infections caused by Botrytis cinerea after 30 days storage at 0 °C, followed by 4 days of market shelf life (Romanazzi et al., 2002). Chitosan treatment showed postharvest disease control that is as effective as procymidone field treatment and SO2 fumigation of grapes after low temperature storage (Romanazzi et al., 2002). Berries sprayed with chitosan preharvest have shown decreased incidence and severity of gray mold in artificially inoculated fruits, with the best control of gray mold obtained 1-2 days after the Downloaded by [189.226.100.40] at 08:02 06 June 2015 application (Romanazzi et al., 2006). Postharvest disease has also been reduced by preharvest chitosan treatment and by postharvest UV-C irradiation (0.36 J/cm2 for 5 min), with the combination of these treatments providing a synergistic interaction (Romanazzi et al., 2006). Application of the antagonistic fungus Cryptococcus laurentii combined with 1% chitosan on the day before harvest significantly reduced natural decay in table grapes stored at 0 °C for 42 days, and thereafter held at 20 °C for 3 days under market-simulation conditions (Meng et al., 2010b). In another study, three different commercial formulations containing chitosan (Armour-Zen, OIIYS, Chito Plant) were compared in a field trial in which they were applied four times during the development of „Thompson Seedless‟ grapes (berry set, pre-bunch closure, veraison, and 2 weeks before harvest). The natural incidence of postharvest gray mold after storage at 2 °C for 5 weeks was reduced by the chitosan, regardless of the commercial formulation used among the three that were tested. Other rot diseases that were mainly caused by Alternaria spp. and Penicillium spp. were mainly reduced by the OII-YS chitosan formulation, which was even more effective than the fungicide program (Feliziani et al., 2013a). Strawberries sprayed with chitosan at full bloom or at the green-fruit or whitening fruit stages have shown decreased incidence of gray mold and Rhizopus rot infections using natural 6 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT inocula of B. cinerea and Rhizopus stolonifer, as seen after 10 days of storage at 0 °C followed by 4 days under market-simulation conditions. The disease control with 1% chitosan was more effective than the currently used chemical fungicides: procymidone (40 g hl-1 a.i.) used at the full bloom and green fruit stages; and pyrimethanil used at the whitening fruit stage (Romanazzi et al., 2000). Preharvest treatments with 1% and 2% chitosan decreased the incidence of postharvest gray mold from a natural inoculum, and after preharvest and postharvest inoculation, Downloaded by [189.226.100.40] at 08:02 06 June 2015 these applications performed significantly better than a fungicide. This treatment with 1% chitosan also performed better than that with 2% chitosan, which was occasionally phytotoxic (Mazaro et al., 2008). Preharvest spraying with 0.2%, 0.4% and 0.6% chitosan decreased postharvest gray mold and maintained the kept quality of strawberries during storage at 3 °C and 13 °C. Here, the incidence of disease decreased with increased chitosan concentration (Reddy et al., 2000a). Sweet cherries treated 7 days before harvest date with 0.1%, 0.5% and 1% chitosan showed decreased incidence of gray mold and brown rot after 2 weeks of storage at 0 °C followed by 7 days of shelf life, as compared to the untreated controls (Romanazzi et al., 1999). At the highest chitosan concentration (1%), the disease reduction was not different with respect to that seen after application of tebuconazole. Similar results were obtained when 1% chitosan was applied 3 days before harvest, as it reduced the incidence of postharvest disease in sweet cherries to the same level as the commercially applied synthetic fungicide fenhexamid (Feliziani et al., 2013b). Chitosan (1%) application 7 days before harvest and postharvest hypobaric treatments at 0.25 atm or 0.50 atm for 4 h showed synergistic effects in the control of total rot 7 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT diseases in sweet cherries stored at 0 °C for 14 days, and thereafter held at 20 °C for a 7-day shelf life (Romanazzi et al., 2003). Fornes et al. (2005) reported that „Clemenules‟ mandarin fruit treated 86 days before harvest and at a postharvest stage with low concentrations of chitosan ( 0.0125% to 0.125%) showed reduced water-spot incidence associated with fruit senescence. All of these treatments reduced the number of injured fruit, and the best results were achieved with the highest chitosan Downloaded by [189.226.100.40] at 08:02 06 June 2015 concentration (0.125%), which reduced water spot incidence by 52%. EFFECT OF POSTHARVEST CHITOSAN APPLICATION ON DISEASE CONTROL Use of chitosan for postharvest disease control in temperate fruit was investigated in the 1990s in many studies. These studies concerned the application of chitosan in general or focused in a group of chitosans, such as oligochitosan that are characterized by low molecular weight. El Ghaouth et al. (1991a; 1992a) and Zhang and Quantick (1998) reported that the control of gray mold and Rhizopus rot in chitosan-coated strawberries was similar to synthetic fungicide application. Cladosporium spp. and Rhizopus spp. infections were also reported to decrease in artificially inoculated strawberry fruit coated with chitosan and stored at 4 °C to 6 °C for 20 days (Park et al., 2005). Similar results were obtained for table grapes, as small bunches dipped in 0.5% and 1% chitosan solutions, and thereafter artificially inoculated with a B. cinerea conidial suspension (by spraying), and stored at low (0 °C) or room (20 °C) temperatures. The chitosan treatment decreased the spread of gray mold infection from one berry to the other berries (nesting) (Romanazzi et al., 2002). Li and Yu (2001) reported that 0.5% and 0.1% chitosan significantly reduced the incidence of brown rot caused by Monilinia fructicola in peach stored at 23 °C, compared to the untreated fruit. Similarly, application of 1% chitosan reduced 8 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT postharvest diseases of sweet cherry (Feliziani et al., 2013b). Treatments with chitosan and oligochitosan reduced disease incidence caused by Alternaria kikuchiana and Physalospora piricola and inhibited lesion expansion of the pear fruit stored at 25 °C. These disease-control effects of chitosan and oligochitosan were concentration dependent and weakened over the incubation time. Indeed, at the lowest chitosan concentration, its effectiveness was the lowest for disease control especially after 5 days of storage at ambient temperatures, compared to the Downloaded by [189.226.100.40] at 08:02 06 June 2015 beginning of storage (Meng et al., 2010a). For vegetables such as tomatoes, the infection diameter caused by R. stolonifer was 15% less than for the control when treated with chitosan at 1.0%, 1.5% and 2.0%, regardless of the molecular weight (Bautista-Baños and Bravo-Luna, 2004). The recent advances concerning chitosan application on postharvest temperate fruit have aimed to combine the biopolymer with other alternatives to fungicides, such as decontaminating agents, plant extracts, essential oils, biocontrol agents, or physical treatments, to provide improved synergistic interactions for the control of postharvest diseases, compared to chitosan alone. Chitosan has been applied in combination with various biocontrol agents, such as Candida satoiana or Cryptococcus laurentii, which are microorganisms that have antagonistic actions against postharvest pathogens (El-Ghaouth et al., 2000; De Capdeville et al., 2002; Yu et al., 2007; 2012; Meng et al., 2010b). Spraying of the antagonistic yeast, C. laurentii, followed by postharvest chitosan coating significantly reduced the natural decay of table grapes stored at 0 °C. The chitosan coating enhanced the effectiveness of the preharvest spray (Meng et al., 2010b). C. laurentii associated with 0.5% chitosan and calcium chloride was effective for the reduction 9 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT of postharvest blue mold caused by Penicillium expansum in pear as well. This combination resulted in more effective mold control than chitosan or C. laurentii alone, although chitosan at 0.5% inhibited the growth of the biocontrol yeast in vitro and in vivo. Moreover, after 6 days of incubation, the combined treatment with C. laurentii, chitosan and calcium chloride inhibited mold decay by nearly 89%, which was significantly higher than the treatments with C. laurentii, chitosan or calcium chloride alone, and with the combinations of C. laurentii and chitosan, and Downloaded by [189.226.100.40] at 08:02 06 June 2015 C. laurentii and calcium chloride (Yu et al., 2012). The combination of chitosan and C. laurentii on apple resulted in synergistic inhibition of blue mold rot, which was the most effective treatment at the optimal concentration of 0.1% chitosan (Yu et al., 2007). In tropical fruit, the application of the bacterium Lactobacillus plantarum alone or in combination with 2% chitosan preserved the quality characteristics of rambutan fruit (Martínez-Castellanos et al., 2009). Similarly, the combination of Candida saitoana with 0.2% glycolchitosan was more effective in controlling gray and blue mold of apple and green mold caused by Penicillium digitatum of oranges and lemons than the yeast or glycolchitosan alone (El-Ghaouth et al., 2000). On the contrary, the combination of chitosan with C. saitoana or with UV-C had no synergistic effects on the progress of blue mold of apple, although a single treatment provided significant reductions (De Capdeville et al., 2002). Extracts obtained from many plants have recently gained popularity and scientific interest for their antimicrobial properties, and thus their activities against decay-causing fungi on fruit and vegetables have been investigated (Gatto et al., 2011). Chitosan coating can be used as a carrier to incorporate plant essential oils or extracts that have antifungal activities or neutraceutical properties. Chitosan incorporated with limonene, a major component of lemon 10 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT essential oils, which has also been given the GRAS status by the USFDA, promoted the preservation of strawberry fruit during their shelf life (Vu et al., 2011). The addition of lemon essential oils enhanced chitosan antifungal activities both in in in-vitro tests and during cold storage of strawberries inoculated with a spore suspension of B. cinerea (Perdones et al., 2012). On table grapes, the combination of 1% chitosan and a grapefruit seed extract improved decay control with respect to single applications of chitosan and maintained the quality of table grapes Downloaded by [189.226.100.40] at 08:02 06 June 2015 (Xu et al., 2007b). Similarly, chitosan coatings that contained bergamot oil or cinnamon oil improved the quality of stored table grapes (Sánchez-González et al., 2011) and of sweet peppers (Xing et al., 2011a), respectively. Chitosan coating without or with essential oils (bergamot, thyme and tea-tree oil) was applied to oranges as preventive or curative treatments against blue mold. In all cases, the addition of the essential oils improved the antimicrobial activities of chitosan; however, the preventive and curative antimicrobial treatments with coatings containing tea-tree oil and thyme, respectively, were the most effective in the reduction of the microbial growth, as compared to the uncoated samples (Cháfer et al., 2012). On the other hand, in another study, combinations of cinnamon extract and chitosan were not compatible, as the cinnamon extract reduced the effectiveness of chitosan in the control of banana crown rot caused by a fungal complex, Colletotrichum musae, Fusarium spp. and Lasiodiplodia theobromae and in delaying fruit senescence during storage (Win et al., 2007). Treatments of papaya with 0.5% or 1.5% chitosan, or with the combination of 1.5% chitosan with an aqueous extract of papaya seed, controlled the development of anthracnose diseases of fruit inoculated with Colletotrichum gloeosporioides. However, no synergistic effects were obtained with the combination of chitosan at 1.5% and the aqueous extract of papaya for the control of the fungal growth (Bautista-Baños et 11 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT al., 2003). Similarly, limited control of R. stolonifer was observed for chitosan-coated tomatoes in combination with beeswax and lime essential oils (Ramos-García et al., 2012). In some trials chitosan was combined with oleic acid. Coatings based on chitosan either without or with oleic acid at different percentages delayed the appearance of natural fungal infections in comparison to uncoated strawberries. When oleic acid was added to the chitosan coating, there were fewer signs of fungal infection during strawberry storage, especially when Downloaded by [189.226.100.40] at 08:02 06 June 2015 the coatings contained the higher levels of oleic acid, which enhanced the antimicrobial properties of chitosan (Vargas et al., 2006). The postharvest application of chitosan has been combined with physical means for the control of postharvest decay of fruit and vegetables, such as UV-C irradiation, hypobaric treatment, and heat curing. Shao et al. (2012) studied the effects of heat-treatment at 38 °C for 4 days before and after coating apples with 1% chitosan. As well as complete control of blue mold and gray mold on these artificially inoculated apples during storage, chitosan coating followed by heat treatment improved the quality of the stored fruit. Moreover, the presence of chitosan coating prevented the occurrence of heat damage on the fruit surface (Shao et al., 2012). In another investigation, the development of postharvest brown rot on peaches and nectarines was controlled through the heating of fruit to 50 °C for 2 h under 85% relative humidity, which eradicated pre-existing Monilinia spp. infections that came from the field, with the application of 1% chitosan at 20 °C then protecting the fruit during handling in the packaging houses and until consumer use (Casals et al., 2012). The combination of immersion in hot water (46.1 °C for 90 min) and in 2% chitosan was beneficial to the storage qualities of mango, compared to untreated mangoes or to fruit treated only with hot water or chitosan (Salvador-Figueroa et al., 2011). 12 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Sweet cherries dipped in 1% chitosan and exposed soon after to hypobaric treatment (0.50 atm for 4 h) showed significant reductions in postharvest natural brown rot, gray mold, and total rot diseases, in comparison with the control and with each treatment applied alone. This combination produced a synergistic effect in its reduction of brown rot and total rots (Romanazzi et al., 2003). Chitosan was also applied as a technology to improve benefits obtained with modified atmosphere packaging. The combination of chitosan coating and modified atmosphere Downloaded by [189.226.100.40] at 08:02 06 June 2015 packaging was effective in preventing decay and browning, and in retaining the pericarp color in litchi fruit (De Reuck et al., 2009). To improve its efficacy in controlling postharvest decay of fruit and vegetables, chitosan has been combined with decontaminating agents. The combination of 0.5% chitosan with 10% or 20% ethanol, which is commonly used in the food industry for its antifungal properties, improved decay control with respect to the single treatments in B. cinerea-inoculated table grapes, as single berries or as clusters (Romanazzi et al., 2007). Application of natamycin, which is a common food additive that is used against mold and yeast growth, in combination with a bilayer coating that contained chitosan and polyethylene wax microemulsion, extended the shelf life of Hami melon, with decreases in weight loss and decay (Cong et al., 2007). Chitosan alone or in combination with sodium bicarbonate or ammonium carbonate significantly reduced the severity of anthracnose for both inoculated and naturally infected papaya fruit. The effects of chitosan combined with ammonium carbonate on the incidence and severity of anthracnose was greater than chitosan alone, and than chitosan with sodium bicarbonate (Sivakumar et al., 2005b). Similarly, the combination of chitosan with potassium metabisulfite was tested in litchi 13 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT fruit. Both chitosan and the combination of chitosan and potassium metabisulfite decreased postharvest decay of these litchi fruit (Sivakumar et al., 2005a). It is also worth mentioning the combination of chitosan with arabic gum, which is a common polysaccharide that is frequently used as an additive in the food industry; this combination controlled banana anthracnose caused by C. musae both in vitro and in vivo, and it enhanced the shelf-life of banana fruit (Maqbool et al., 2010a; 2010b). Downloaded by [189.226.100.40] at 08:02 06 June 2015 In some other studies the most suitable acids were tested for the dissolving of chitosan powder, and it was shown that practical grade chitosan should be dissolved in an acid solution to activate its antimicrobial and eliciting properties. Chitosan dissolved in 10 different acids (as 1% solutions of acetic, L-ascorbic, formic, L-glutamic, hydrochloric, lactic, maleic, malic, phosphoric, and succinic acids) was effective in reducing gray mold incidence on single table grape berries (Romanazzi et al., 2009). However, the greatest reduction of gray mold (about 70%, compared with the control) was observed after immersion of the berries in chitosan dissolved in acetic acid or formic acid, whereas there was intermediate effectiveness with chitosan dissolved in hydrochloric, lactic, L-glutamic, phosphoric, succinic, and L-ascorbic acids. The least effective treatments were chitosan dissolved in maleic or malic acids (Romanazzi et al., 2009). MODE OF ACTION OF CHITOSAN AGAINST THE POSTHARVEST PATHOGENS Due to the wide range of antifungal activities against postharvest pathogens (Table 5), chitosan coating can be applied as a biocoating to prolong the postharvest life of fresh produce (BautistaBaños et al., 2006). 14 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT The antimicrobial activities of chitosan appear to rely on electrostatic interactions between positive chitosan charges and the negatively charged phospholipids in the fungal plasma membrane. Chitosan first binds to the target membrane surface and covers it, and in a second step, after a threshold concentration has been reached, chitosan causes membrane permeabilization and the release of the cell contents (Palma-Guerrero et al., 2010). There are usually low levels of Ca2+ in fungi cytosol, due to the barrier formed by the plasma membrane, Downloaded by [189.226.100.40] at 08:02 06 June 2015 which has hermetic seals that regulate the passage of Ca2+ gradients. This process also involves the homeostatic mechanism, where the Ca2+ concentration regulates itself within the cytosol, and it sends the excess Ca2+ out of the cell or stores it in the intracellular organelles. Thus, as chitosan is applied, the homeostatic mechanism becomes drastically transformed, because as it forms channels in the membrane, it allows the free passage of Ca2+ down its gradients, which cause instabilities in the cells that can lead to death of the cell itself (Palma-Guerrero et al., 2009). In addition, inhibitory effects of chitosan on the H+-ATPase in the plasma membrane of R. stolonifer has been reported. García-Rincón et al. (2010) suggested that the decrease in H+ATPase activity can induce the accumulation of protons inside the cell, which would result in inhibition of the chemiosmotic driven transport that allows H +/K+ exchange. Moreover, a rapid efflux of potassium from cells of R. stolonifer has been reported as an effect of chitosan treatment; this was combined with an increase in pH of the culture medium, which was chitosanconcentration dependent. Both of these phenomena were related to the leaking of internal cellular metabolites (García-Rincón et al., 2010). Similarly, when R. stolonifer was grown in media containing chitosan, the release of proteins by the fungal cells increased significantly. It was 15 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT proposed that this release of proteins from the cell to the supernatant is because there are sites where the cell membrane is damaged by chitosan (Guerra-Sánchez et al., 2009). Besides its capacity for membrane permeabilization, chitosan can also penetrate into fungal cells. Fluorescent labeled chitosan was detected in fungal conidia and it was hypothesized that chitosan itself permeabilizes the plasma membrane to allow its entry into the cytoplasm (Palma-Guerrero et al., 2008; 2009). Another study used fluorescence visualization to Downloaded by [189.226.100.40] at 08:02 06 June 2015 demonstrate that oligochitosan can penetrate the cell membrane of Phytophthora capsici, and that, as it is positively changed, chitosan can bind to intracellular targets, such as DNA and RNA, which are negatively charged (Xu et al., 2007a). Similarly, observations made on Aspergillus niger have revealed the presence of labeled chitosan both inside and outside the cells, and the permeated chitosan was suggested to block DNA transcription, and therefore to inhibit the growth of the fungus (Li et al., 2008). Several studies have described the morphological changes on fungal hyphae and reproductive structures that can be induced by chitosan. Scanning electron microscopy observations of Fusarium sulphureum treated with chitosan have revealed effects on hypha morphology. The growth of hyphae treated with chitosan was strongly inhibited, and they were tightly twisted and formed rope-like structures. Spherical or club-shaped abnormally inflated ends were observed on the twisted hyphae, which were swollen and showed excessive branching. Further transmission electron microscopy observations have indicated ultrastructural alterations of the hyphae by chitosan. These changes included cell membrane disorganization, cell-wall disruption, abnormal distribution of the cytoplasm, non-membranous inclusion bodies in the cytoplasm, considerable thickening of the hyphal cell walls, and very frequent septation with 16 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT malformed septa (Li et al., 2009). Examination of ultrasections of the hyphae and conidia of chitosan-treated Alternaria alternata revealed marked alterations to the cell wall. The chitosantreated mycelia showed predominantly loosened cell walls, and in some areas, there was also lysis. The conidia exposed to chitosan were intensely damaged, and usually eroded, with broken cell walls seen that contained in some cases no cytoplasm (Sánchez-Domínguez et al., 2011). R. stolonifer subjected to the formulation of chitosan with beeswax and lime essential oils showed Downloaded by [189.226.100.40] at 08:02 06 June 2015 no development of the typical reproductive structures, and its mycelia were distorted and swollen (Ramos-García et al., 2012). In another investigation, chitosan-treated spores of R. stolonifer showed numerous and deeper ridge formations that were not observed on non-treated spores (Hernández-Lauzardo et al., 2008). Chitosan induced morphological changes of the mycelia of B. cinerea and R. stolonifer that were characterized by excessive hyphal branching, as compared to the control (El Ghaouth et al., 1992a). This was confirmed in another study, in which there was the induction of marked morphological changes and severe structural alterations in chitosantreated cells of B. cinerea. Microscopic observations showed coagulation in the fungus cytoplasm that was characterized by the appearance of small vesicles in the mycelia treated with chitosan. In other cases, the mycelia contained larger vesicles, or even empty cells, which were devoid of cytoplasm (Ait Barka et al., 2004). The area and the elliptical form of the spores was significantly different when C. gloeosporioides was grown on potato dextrose agar with added chitosan, compared to potato dextrose agar alone (Bautista-Baños et al., 2003). Similarly, the hyphal and germ-tube morphology of C. gloeosporioides growing on chitosan showed malformed hyphal tips with thickened walls. Many swellings occurred in the hyphae or at their tips, whereas in the controls cells the walls and germ tubes were smooth with no swellings or 17 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT vacuolation (Ali and Mahmud, 2008; Ali et al., 2010). The scanning electron micrographs showed normal growth of hyphae in the untreated controls for C. gloeosporioides, whereas there was hyphal agglomeration and formation of large vesicles in the mycelia in samples treated with chitosan-loaded nanoemulsions (Zahid et al., 2012). The fungal mycelia of Sclerotinia sclerotiorum exposed to chitosan were deformed, twisted and branched, or indeed, dead, with no visible cytoplasm in the fungal cells, whereas the untreated mycelia were normal in appearance Downloaded by [189.226.100.40] at 08:02 06 June 2015 (Cheah et al., 1997). Not all fungi show the same sensitivity to chitosan, which might be due their intrinsic characteristics. New findings relating to the permeabilization of the plasma membrane of different cell types of the fungi Neurospora crassa and the membrane composition among various resistant and non-resistant chitosan fungi appear to provide important factors (PalmaGuerrero et al., 2008; 2009; 2010). By imaging fluorescently labeled chitosan using confocal microscopy, it was seen that chitosan binds to the conidial surfaces of all of the species tested, although it only consistently permeabilized the plasma membranes of some of the fungi. Some of the other fungi formed a barrier to the chitosan. Analysis of the main plasma membrane components revealed important differences in the fatty acid compositions between the chitosansensitive and chitosan-resistant fungi. The cell membranes of chitosan-sensitive fungi showed higher content of the polyunsaturated fatty acid linolenic acid, higher unsaturation index, and lower plasma membrane fluidity. Chitosan binding should induce an increase in membrane rigidity in the regions to which it attaches. This interaction will enhance the differences in fluidity between the different membrane regions, which can cause membrane permeabilization. In a saturated, more rigid membrane, the changes in rigidity induced by chitosan binding would 18 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT be much lower, with little permeabilization, even in the presence of negatively charged phospholipid headgroups (Palma-Guerrero et al., 2010). The antifungal activities of chitosan have been reported to vary according to its molecular weight and concentration. It has also been noted that, in general, fungal growth inhibition increases as the concentration of chitosan increases in the cases of B. cinerea (El Ghaouth et al., 1992a; 2000; Ben-Shalom et al., 2003; Chien and Chou, 2006; Liu et al., 2007), R. stolonifer (El Downloaded by [189.226.100.40] at 08:02 06 June 2015 Ghaouth et al., 1992a), Penicillium citrinum (Xing et al., 2011b); P. digitatum (Chien and Chou, 2006), Penicillium italicum (Chien and Chou, 2006), P. expansum (El Ghaouth et al., 2000; Liu et al., 2007; Yu et al., 2007), M. fructicola (Yang et al., 2010; 2012), Botrydiplodia lecanidion (Chien and Chou, 2006), C. gloeosporioides (Jitareerat et al., 2007; Muñoz et al., 2009; Ali and Mahmud, 2008; Abd-Alla and Haggar, 2010; Ali et al., 2010), Fusarium solani (Eweis et al., 2006), A. kikuchiana (Meng et al., 2010a) and P. piricola (Meng et al., 2010a), although it decreases in the case of A. niger (Li et al., 2008). In some studies, the antifungal activity of chitosan decreased with an increase in molecular weight, within the range of 50 kDa to 1000 kDa (Li et al., 2008). The highest inhibitory effect against the growth of R. stolonifer was observed with low molecular weight chitosan, while the high molecular weight chitosan showed a greater effect on the development of the spores (Hernández-Lauzardo et al., 2008). High molecularweight chitosan had the lowest inhibitory effects on B. cinerea growth, compared to the low molecular weight chitosan (Badawy and Rabea, 2009). In the case of S. sclerotiorum, there was a negative correlation between mycelial growth inhibition and chitosan molecular weight (Ojaghian et al., 2013). Spore germination and germ-tube elongation of A. kikuchiana and P. piricola were significantly inhibited by chitosan and oligochitosan, although when compared to 19 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT chitosan, oligochitosan was more effective for the inhibition of spore germination (Meng et al., 2010a). However, other investigations have shown fungal growth inhibition by chitosan, regardless of the type of chitosan (Chien and Chou, 2006), without any fungicidal or fungistatic patterns among low, medium, and high molecular weight chitosans tested with different isolates of C. gloeosporioides (Bautista-Baños et al., 2005) and R. stolonifer (Guerra-Sánchez et al., 2009). Downloaded by [189.226.100.40] at 08:02 06 June 2015 INDUCTION OF RESISTANCE BY CHITOSAN IN FRUIT TISSUES Plant resistance towards pathogens occurs through hypersensitive responses that result in cell death at the penetration site, structural alterations, accumulation of reactive oxygen species (ROS), synthesis of secondary metabolites and defense molecules, and activation of pathogenesis-related (PR) proteins (Van-Loon and Van-Strien, 1999). The application of external elicitors to vegetative tissue can trigger plant resistance, by simulating the presence of a pathogen. Several studies have reported that chitosan can induce a series of enzyme activities and the production of various compounds that are correlated with plant defense reactions to pathogen attack (Bautista-Baños et al., 2006) (Tables 6-8). Chitosan can increase PR gene function through multiple modes, which includes activation of cell surface or membrane receptors, and internal effects on the plant DNA conformation, which can, in turn, influence gene transcription (Hadwiger, 1999). Histochemical staining of chitosan polymers indicates that chitosan accumulates in the plant cell wall, cytoplasm, and nucleus. The accumulation of positively charged chitosan along with its high affinity for negatively charged DNA suggests that it has a direct effect on the regulation of plant 20 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT defense responses, with influences on mRNA and protein synthesis (Hadwiger and Loschke, 1981). Phenylalanine ammonia lyase (PAL) is the key enzyme in the phenol synthesis pathway (Cheng and Breen, 1991), and the accumulation of phenols that act as phytoalexins is considered the primary inducible response in plants against a number of biotic and abiotic stresses (Bhattacharya et al., 2010). Chitosan application has been reported to increase PAL activity in Downloaded by [189.226.100.40] at 08:02 06 June 2015 treated fruit tissue. Table grape bunches with preharvest spraying with chitosan showed a threefold increase in PAL activity in the berry skin 24 h and 48 h after chitosan application (Romanazzi et al., 2002). PAL elicitation by chitosan was confirmed with table grapes sprayed in the vineyard without or with C. laurentii and coated with chitosan postharvest, and then stored at 0 °C (Meng et al., 2008; 2010b; Meng and Tian, 2009). Chitosan treatments induced the activity of PAL in sweet cherry (Dang et al., 2010) and strawberry (Romanazzi et al., 2000; Landi et al., 2014), thus enhancing the fruit defense responses. Chitinase and β-1,3-glucanase are two PR proteins that participate in defense against pathogens, as these can partially degrade the fungal cell wall (Van-Loon and Van-Strien, 1999). Increases in the activities of chitinase and β-1,3-glucanase were demonstrated as a result of chitosan application in „Valencia‟ oranges, 24 h after the chitosan treatment. It was proposed that these changes in the enzyme activities might have contributed to the reduction of black spot in the orange fruit (Canale Rappussi et al., 2009). Similarly, chitosan coating significantly reduced the decay of strawberry and raspberry, and induced a significant increase in chitinase and β-1,3glucanase activities of the berries, as compared to the controls (Zhang and Quantick, 1998; Landi et al., 2014). Compared to the untreated fruit, the high chitinase and β-1,3-glucanase activities in 21 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT chitosan-treated strawberries reinforced the microbial defense mechanism of the fruit and accentuated the resistance against fungal invasion (Zhang and Quantick, 1998; Wang and Gao, 2012). The chitinase and β-1,3-glucanase activities of papaya and mango subjected to chitosan treatment were much higher than in the untreated fruit (Jitareerat et al., 2007; Hewajulige et al., 2009), and oligochitosan treatment significantly enhanced the activities of chitinase and β-1,3glucanase in pear fruit (Meng et al., 2010a). In table grapes, preharvest chitosan treatments from Downloaded by [189.226.100.40] at 08:02 06 June 2015 three different commercial formulations induced the activity of endochitinase, while two of the chitosan formulations induced exochitinase activity (Feliziani et al., 2013a). In fruit tissue, the high activity of pectic enzymes, such as polygalacturonase, cellulase and pectate lyase, was shown to be closely associated with the weakening of the plant cell wall, thus resulted in softening of the fruit and greater susceptibility to storage rots (Stevens et al., 2004). Down-regulation of polygalacturonase resulted in firmer fruit (Atkinson et al., 2012). In peach fruit, the chitosan treatments somewhat inhibited polygalacturonase activity throughout the storage period. In particular, the combination consisted of a coating of chitosan and calcium chloride, the polyethylene packaging, and intermittent warming, with markedly inhibited polygalacturonase activity at the end of the refrigerated storage (Ruoyi et al., 2005). The macerating enzyme activities in tomato tissue, such as polygalacturonase, pectate lyase, and cellulose, in the vicinity of lesions caused by the pathogen A. alternata were less than half in chitosan-treated fruit, compared with untreated fruit. Chitosan inhibited the development of black mold rot of tomatoes and reduced the production of pathogenic factors by the fungus (Reddy et al., 2000b). 22 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Chitosan treatment might induce fruit disease resistance through regulation of ROS levels, antioxidant enzymes, and the ascorbate–glutathione cycle. ROS, such as H2O2 and O2-, are the earliest events that correlate plant resistance to pathogens (Baker and Orlandi, 1995); these are involved in the development of disease resistance in fruit (Torres et al., 2003). Although ROS might contribute to an enhancement of the plant defense, high level of ROS can cause lipid peroxidation and lead to the loss of membrane integrity of plant organs. To prevent Downloaded by [189.226.100.40] at 08:02 06 June 2015 harmful effects of excess ROS on plant tissues, the ROS can be detoxified by an antioxidant system. This consists of non-enzymatic antioxidants, such as ascorbic acid, glutathione, and phenolic compounds, and antioxidant enzymes, such as superoxide dismutase, peroxidases and catalases. Chitosan application was reported to reduce ROS in tissues of treated fruit, such as pear (Li et al., 2010a) and guava (Hong et al., 2012), and to lower the hydrogen peroxide content in litchi (Sun et al., 2010), pear (Li et al., 2010a), table grapes (Feliziani et al., 2013a) and strawberry (Romanazzi et al., 2013). This might be due to direct effects, as chitosan itself has antioxidant activity and scavenges hydroxyl radicals (Yen et al., 2008), or to indirect effects, as chitosan induces the plant antioxidant system. Higher levels of glutathione were reported after chitosan treatment in litchi (Sun et al., 2010), strawberry (Wang and Gao, 2012) and orange (Zeng et al., 2010). Higher quantities of ascorbic acid have also been reported after chitosan treatments in fruit tissues of strawberry (Wang and Gao, 2012), peach (Li and Yu, 2001; Ruoyi et al., 2005), sweet cherry (Dang et al., 2010; Kerch et al., 2011), jujube (Qiuping and Wenshui, 2007; Xing et al., 2011b), orange (Zeng et al., 2010), citrus (Chien and Chou, 2006), longan (Jiang and Li, 2001), guava (Hong et al., 2012), mango (Jitareerat et al., 2007; Zhu et al., 2008) and litchi (Sun et al., 2010). The reduction 23 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT of ascorbic acid loss in chitosan-coated sweet cherries was proposed to be due to the low oxygen permeability of the chitosan coating around the fruit surface, which lowers the oxygen level and reduces the activity of the ascorbic acid oxidase enzymes, which prevents the oxidation of ascorbic acid (Dang et al., 2010). The presence of antioxidants, such as the phenols, can substantially reduce the ROS content of plant tissues, as their hydroxyl groups and unsaturated double bonds make them very Downloaded by [189.226.100.40] at 08:02 06 June 2015 susceptible to oxidation (Rice-Evans et al., 1997). Moreover, phenolic compounds are involved in plant responses against biotic and abiotic stresses (Lattanzio et al., 2006; Bhattacharya et al., 2010). Chitosan coating was effective in the intensification of total antioxidant capacity of treated apricot, with increases in the phenolic compounds in the fruit tissue (Ghasemnezhad et al., 2010). In tomato, the content of phenolic compounds increased in chitosan-treated fruit compared to the untreated fruit (Liu et al., 2007), and this increase was directly proportional to the chitosan concentration used (Badawy and Rabea, 2009). Table grapes treated with chitosan had higher phenolic compound contents (Shiri et al., 2012; Feliziani et al., 2013a). Anthocyanin, flavonoid and total phenolics contents of chitosan treated litchi decreased more slowly than in untreated fruit (Zhang and Quantick, 1997; Jiang et al., 2005; De Reuck et al., 2009 ). Kerch et al. (2011) reported that total phenols and anthocyanin content increased in chitosan-treated sweet cherry after 1 week of cold storage, while their contents decreased in chitosan-treated strawberry stored under the same conditions. Similarly, in strawberry, chitosan-coated fruit had lower anthocyanin content, as the anthocyanins were synthesized at a slower rate than for the nontreated berries (El Ghaouth et al., 1991a), and the rate of pigment development was lower with an increase in chitosan concentration (Reddy et al., 2000a). The anthocyanin contents 24 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT significantly decreased throughout storage in strawberries coated with chitosan combined with oleic acid, whereas no significant changes were seen in the control samples at the end of the storage (Vargas et al., 2006). On the contrary, Wang and Gao (2012) reported that strawberries treated with chitosan maintained better fruit quality, with higher levels of phenolics, anthocyanins and flavonoids. In another study, the application of chitosan to strawberry increased the expression of genes involved in the biosynthesis of flavonoid compounds, such as Downloaded by [189.226.100.40] at 08:02 06 June 2015 chalcone isomerase, flavonol synthase, anthocyanidin synthase (Landi et al., 2014). Several factors, such as the cultivar of the studied commodity, the stage of maturation, the storage conditions, could account to explain the different responses to chitosan application concerning phenolic compounds accumulation in fruit tissues. Chitosan treatment has been reported to have an influence on antioxidant enzyme activities in the tissues of both temperate and tropical fruit and vegetables (Tables 6-8). Compared to untreated strawberries, those treated with chitosan maintained higher levels of antioxidant enzyme activities, such as catalase, glutathione-peroxidase, guaiacol peroxidase, dehydroascorbate reductase, and monodehydroascorbate reductase (Wang and Gao, 2012). Ascorbate peroxidase and glutathione reductase activities increased in pear treated with chitosan (Lin et al., 2008; Li et al., 2010a). Compared to the tissue of uncoated fruit, higher activities of superoxide dismutase, catalase, and peroxidase were reported after chitosan application to pear (Lin et al., 2008; Li et al., 2010a), sweet pepper (Xing et al., 2011a), and tropical fruit, such as guava (Hong et al., 2012). In addition, increased peroxidase activity after chitosan application has been reported for several other commodities, such as table grapes (Meng et al., 2008), pear (Meng et al., 2010a), sweet cherry (Dang et al., 2010), orange (Canale Rappussi et al., 2009), 25 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT tomato (Liu et al., 2007), and potato (Xiao-Juan et al., 2008). Conversely, in other studies, decreased peroxidase activity was reported in litchi fruit after chitosan application, whether or not it was combined with other treatments (Zhang and Quantick, 1997; De Reuck et al., 2009; Sun et al., 2010). Meanwhile, treatment of litchi fruit with a combination of chitosan and ascorbic acid increased the activities of superoxide dismutase and catalase, and the contents of ascorbic acid and glutathione (Sun et al., 2010). Treatments with chitosan alone or in Downloaded by [189.226.100.40] at 08:02 06 June 2015 combination with C. laurentii decreased the superoxide dismutase activity in table grape tissues (Meng et al., 2008; 2010b; Meng and Tian, 2009). Treatments of navel oranges with 2% chitosan effectively enhanced the activities of peroxidase, superoxide dismutase and ascorbate peroxidase, but decreased the activities of catalase and the content of ascorbic acid (Zeng et al., 2010). Physiological changes concerning polyphenol oxidase (PPO) activity have been observed after application of chitosan to fruit and vegetables (Tables 6-8). This has great impact on fruit quality; indeed, PPO is a phenol-related metabolic enzyme that catalyzes the oxidation of phenolic compounds that are involved in plant defense against biotic and abiotic stresses and in pigmentation/ browning of fruit and vegetable tissues (Lattanzio et al., 2006; Bhattacharya et al., 2010). In some investigations, chitosan decreased PPO activity, and its inhibitory effects are probably a consequence of the adsorption of suspended PPO, its substrates, or its products by the positive charges of chitosan (Badawy and Rabea, 2009). The other possibility is that the selective permeability to gases due to the chitosan coating generates low levels of oxygen around the fruit surface, which can delay the deteriorative oxidation reactions, and partially inhibit the activities of oxidases such as PPO (Ayranci and Tunc, 2003). The chitosan coating markedly reduces PPO activity and delays skin browning during fruit shelf life. The maintenance of the skin color of the 26 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT litchi fruit after chitosan treatment can be accounted for by the higher level of anthocyanin content in the skin that results from inhibition of PPO activity (Zhang and Quantick, 1997; Jiang et al., 2005; De Reuck et al., 2009). Similarly, the activities of PPO and peroxidase, and the related browning in the pericarp, were markedly lowered by treatment of harvested litchi fruits with ascorbic acid and 1% chitosan (Sun et al., 2010). In chitosan-treated tomato (Badawy and Rabea, 2009) and jujube (Wu et al., 2010; Xing et al., 2011b), the decreases in the PPO activities Downloaded by [189.226.100.40] at 08:02 06 June 2015 were concomitant with the enhanced phenolic content, and in sweet cherry (Dang et al., 2010), with the reduction in tissue browning. The combination of chitosan, calcium chloride and intermittent warming decreased the PPO activity in the tissues of peach that had been cold stored for 50 days (Ruoyi et al., 2005). However, in other investigations, PPO activities of fruit tissue increased after chitosan treatment. Chitosan treatment enhanced the activities of PPO in the flesh around the wound of a pear (Meng et al., 2010a). An increase in the activity of PPO was demonstrated as a result of chitosan application in „Valencia‟ oranges, which was seen 24 h after chitosan treatment (Canale Rappussi et al., 2009). Chitosan application in tomato fruit stored at 25 °C and 2 °C increased the content of the phenolic compounds and induced the activities of PPO, the levels of which were almost 1.5-fold those in the wounded control fruit at the same time (Liu et al., 2007). In this study, there was no direct relationship between the PPO activities and the content of phenolic compounds, although the phenolic compounds can be oxidized by the actions of PPO and peroxidase, to produce quinones (Campos-Vargas and Saltveit, 2002). It is likely that regulation of phenolic metabolism by the action of other enzymes, such as PAL, which participates in the biosynthesis of phenolic compounds, also has an important role (Liu et al., 2007). This could even explain the reason why in some investigations the PPO levels of fruit 27 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT tissue after chitosan application are variable. Preharvest spraying with C. laurentii combined with postharvest chitosan coating increased the activities of PPO in table grapes during storage, but after 3 days of shelf life, the PPO activities in the treated fruit were lower than in the untreated fruit (Meng et al., 2010b). During cold storage, the PPO activity of litchi fruit coated with chitosan increased slowly, reached a peak, and then decreased (Zhang and Quantick, 1997). EFFECT OF CHITOSAN TREATMENT ON MAINTENANCE OF FRUIT QUALITY AND Downloaded by [189.226.100.40] at 08:02 06 June 2015 RETENTION OF HEALTH-PROMOTING COMPOUNDS Chitosan coating provides a semipermeable film around the fruit surface, which modifies the internal atmosphere by reducing oxygen and/or elevating carbon dioxide levels, which decreases the fruit respiration level and metabolic activity, and hence delays the fruit ripening and senescence processes (Özden and Bayindirli, 2002; Olivas and Barbosa-Cánovas, 2005; Romanazzi et al., 2007; 2009; Vargas et al., 2008). A suppressed respiration rate slows down the synthesis and the use of metabolites, which results in lower soluble solids due to the slower hydrolysis of carbohydrates to sugars (Ali et al., 2011; Das et al., 2013). However, there are numerous confounding factors that can contribute to the soluble solids concentrations in fruit tissues; e.g., the fruit studied, its stage of ripeness, the storage conditions, and the thickness of the chitosan coating (Ali et al., 2011). On the other hand, as organic acids, such as malic and citric acid, are substrates for the enzymatic reactions of plant respiration, increased acidity and reduced pH would be expected in low-respiring fruit (Yaman and Bayindirli, 2001). Above all, the chitosan coating with its filmogenic properties has been used as a water barrier, to minimize water and weight loss of fruit during storage (Vargas et al., 2008; Bourlieu et al., 2009). All of 28 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT these physiological changes have been reported in fruit and vegetables treated with chitosan (Tables 6-8). For temperate fruit (Table 6), the chitosan coating minimized weight loss of stored apples, and its combination with heat treatment showed the lowest respiration rate, and significantly reduced pH and increased titratable acidity (Shao et al., 2012). Chitosan treatments of pears during storage reduced their vital activities, and in particular their respiration rate, which Downloaded by [189.226.100.40] at 08:02 06 June 2015 maintained the fruit quality and prolonged the shelf life. Compared with the control samples, chitosan-coated pears showed reduced weight loss (Zhou et al., 2008). Again in pear, chitosan coating alone and in combination with ascorbic acid resulted in decreased respiration rate, delayed weight loss, and retention of greater total soluble solids and titratable acidity (Lin et al., 2008). Chitosan-treated peaches showed lower respiration rates and higher titratable acidity than control peaches (Li and Yu, 2001). Chitosan forms a coating film on the outside surface of sweet cherries that effectively delayed the loss of water and promoted changes in titratable acidity and total soluble solids of the sweet cherries (Dang et al., 2010). Strawberries treated with chitosan alone or in combined with calcium gluconate showed reduced weight loss and respiration, which delayed the ripening and the progression of fruit decay due to senescence. Regardless of the addition of calcium gluconate to the chitosan, the coated strawberries had higher titratable acidity, and lower pH and soluble solids (Hernández-Muñoz et al., 2008). A chitosan coating without or with added calcium or vitamin E decreased weight loss and delayed the changes in pH and titratable acidity of strawberries and red raspberries during cold storage (Han et al., 2004; 2005). Chitosan application combined with bergamot oil provided a water vapor barrier for cold-stored table 29 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT grapes, which reduced the fruit weight losses. Due to its hydrophobic nature, the addition of bergamot oil lowered this phenomenon further (Sánchez-González et al., 2011). Similarly, weight loss reductions in chitosan-coated table grapes were observed when this was combined with putrescine (Shiri et al., 2012) and grape seed extract (Xu et al., 2007b). The complex of zinc(II) and cerium(IV) with chitosan film-forming material that was applied to preserve the quality of Chinese jujube fruit reduced the fruit respiration rate and weight loss, while it Downloaded by [189.226.100.40] at 08:02 06 June 2015 increased the fruit total soluble solids, as compared to the uncoated fruit (Wu et al., 2010). In another study, after 42 days of storage at 13 °C, chitosan-coated citrus fruit showed less weight loss and higher titratable acidity and total soluble solids, compared to the control fruit. The weight loss of these citrus fruit decreased as the concentration of chitosan was increased (Chien and Chou, 2006). Coating tomato fruit with chitosan solutions reduced the respiration rate and ethylene production, with greater effects with 2% chitosan than 1% chitosan. The chitosan coating increased the internal CO2 and decreased the internal O2 levels of the tomatoes. These chitosan-coated tomatoes were also higher in titratable acidity (El Ghaouth et al., 1992b). Similar changes in respiration, weight loss, pH, titratable acidity, and soluble solids content have been reported after chitosan treatment of tropical fruit (Table 7). Polysaccharidebased coatings, including chitosan, applied to banana fruit reduced the carbon dioxide evolution, loss of weight, and titratable acidity. Moreover, the reducing sugar content and the total soluble solids of the coated fruit were lower than with the untreated fruit, which suggests that the coated fruit synthesized reducing sugars at a slower rate, through the slowed metabolism (Kittur et al., 2001). Similarly in bananas, chitosan alone or in combination with 1-methylcyclopropene reduced the rate of respiration (by 32%) compared to untreated banana, and decreased titratable 30 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT acidity and increased total soluble solids (Baez-Sañudo et al., 2009). The composite coating of Arabic gum and chitosan provided an excellent semipermeable barrier around the banana fruit, which reduced weight loss, modified the internal atmosphere, and suppressed ethylene evolution, thus reducing respiration and delaying the ripening process. After 33 days of storage, the soluble solids concentrations of the treated banana fruit were lowered, whereas the titratable acidity was increased by the chitosan and Arabic gum coating (Maqbool et al., 2010a; 2010b; 2011). The Downloaded by [189.226.100.40] at 08:02 06 June 2015 application of chitosan delayed changes in eating quality, reduced respiration rate and weight loss, and increased total soluble solid and titratable acidity of stored longan (Jiang and Li, 2001) and guava (Hong et al., 2012) fruit. In mango fruit, the decline in respiration rate, fruit weight, and titratable acidity were all effectively inhibited by chitosan (Jitareerat et al., 2007), while the increase in total soluble solids was delayed during storage (Zhu et al., 2008). Mango fruit coated with chitosan and subjected to hydrothermal treatment had less weight loss, lower pH and soluble solids, but higher acidity, regardless of the hydrothermal process (Salvador-Figueroa et al., 2011). The CO2 concentration in the internal cavity of chitosan-treated papaya was significantly higher than that of the untreated fruit. The formation of a chitosan film on the fruit acted as a barrier for O2 uptake, and slowed the rate of respiration and the metabolic activity, and consequently the ripening process (Hewajulige et al., 2009). Again in papaya, chitosan provided effective control of weight loss, and delayed the changes in soluble solids concentrations over 5 weeks of storage. The titratable acidity of the papaya fruit declined throughout the storage period, although at a slower rate in the chitosan-coated fruit, as compared to the untreated fruit (Bautista-Baños et al., 2003; Ali et al., 2010; 2011). Chitosan coating without or with calcium infiltration markedly slowed the ripening of papaya, as shown by their lack of weight loss, delay 31 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT in titratable acidity decrease, and increase in soluble solids and pH (Al Eryani et al., 2008). In litchi fruit during storage, chitosan treatment produced an effective coating that reduced the respiration and transpiration of the fruit during storage (Lin et al., 2011), and reduced the decreases in the concentrations of total soluble solids and in the titratable acidity (Jiang et al., 2005). Similar results were obtained with the combination of chitosan with ascorbic acid, which significantly increased the titratable acidity and total soluble solids of stored litchi fruit (Sun et Downloaded by [189.226.100.40] at 08:02 06 June 2015 al., 2010). Firmness is a major attribute that dictates the postharvest quality of fruit (Barrett et al., 2010). Fruit softening is a biochemical process that is normally attributed to the deterioration of the cell-wall composition, which involves the hydrolysis of pectin by enzymes; e.g., polygalacturonase (Atkinson et al., 2012). Low levels of oxygen and higher levels of carbon dioxide restricts the activities of these enzymes and promotes the retention of fruit firmness during storage (Maqbool et al., 2011). Moreover, due to reduced transpiration, the water retention provides turgor to the fruit cells. Banana fruit treated with composite edible coatings of chitosan and Arabic gum showed significantly higher firmness than untreated bananas at the end of the storage period, and this firmness decreased as the concentration of the coating decreased (Maqbool et al., 2011). Chitosan coatings had beneficial effects on strawberry firmness, such that by the end of the storage period, the treated fruit had higher flesh firmness values than the untreated fruit (Hernández-Muñoz et al., 2008). In several other studies, chitosan coating maintained the firmness during storage of table grapes (Xu et al., 2007b; Sánchez-González et al., 2011), apple (Shao et al., 2012), pear (Lin et al., 2008), peach (Li and Yu, 2001), jujube (Qiuping and Wenshui, 2007), orange (Chien and Chou, 2006; Cháfer et al., 2012), banana 32 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT (Kittur et al., 2001; Win et al., 2007; Baez-Sañudo et al., 2009), mango (Zhu et al., 2008; Salvador-Figueroa et al., 2011), papaya (Bautista-Baños et al., 2003; Sivakumar et al., 2005b; Ali et al., 2010; 2011), rambutan (Martínez-Castellanos et al., 2009), guava (Hong et al., 2012) and tomato (El Ghaouth et al., 1992b) (Tables 6-8). In several studies, panelists were asked to observe and then rate the overall appearance, or just the flavor, of fruit treated or not with chitosan, using hedonic scales (Tables 6-8). These Downloaded by [189.226.100.40] at 08:02 06 June 2015 studies showed that chitosan can preserve the taste of pear fruit, which after cold storage was similar to the taste of the fresh fruit (Zhou et al., 2008). Similar results were obtained with the combination of chitosan and cinnamon oil coating, which retained sweet pepper quality, without the development of off-flavors (Xing et al., 2011a). Consumer acceptance based on color, flavor, texture, sweetness and acidity was improved by chitosan coating and/or heat treatment of apple fruit (Shao et al., 2012). For table grapes, chitosan alone and in combination with putrescine prolonged the maintenance of the original sensory quality, in comparison with the decline in the untreated grapes (Shiri et al., 2012). The combination of chitosan with grape seed extract delayed rachis browning and dehydration, and maintained the visual aspect of the berry without detrimental effects on taste or flavor (Xu et al., 2007b). In sweet cherries, chitosan coating had a strong effect on the maintenance of quality attributes, such as visual appearance, color, taste and flavor, as it had protective effects in preventing surface browning, cracking, and the leaking of juice (Dang et al., 2010). On strawberry, results from consumer sensory evaluations indicated that chitosan increased the appearance and acceptance of the strawberries (Devlieghere et al., 2004), whereas coatings containing chitosan and vitamin E developed a waxy-and-white surface on the coated fruits (Han et al., 2005). In strawberries, the aroma and flavor of chitosan-coated 33 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT fruit was considered less intense than those of the uncoated fruit, which were preferred by the panelists (Vargas et al., 2006). Likewise, panelists detected an untypical oily aroma in samples coated with the combination of chitosan and oleic acid (Vargas et al., 2006). On bananas, BaezSañudo et al. (2009) reported that chitosan coating did not affect the sensory quality of the fruit. In another case, banana fruit treated with 10% Arabic gum and 1% chitosan improved fruit quality during storage and received the highest sensory scores for taste, pulp color, texture, Downloaded by [189.226.100.40] at 08:02 06 June 2015 flavor, and overall acceptability (Maqbool et al., 2011). However, the fruit coated with high concentrations of Arabic gum, as 15% or 20%, combined with 10% chitosan did not ripen fully after about 1 month of storage, developed poor pulp color and inferior texture, and were offflavored (Maqbool et al., 2011). Similarly, the sensory evaluation of papaya for taste, peel color, pulp color, texture, and flavor revealed that the fruit treated with 1.5% chitosan attained maximum scores from the panelists in all of the tested parameters. The untreated fruit and those treated with 0.5% chitosan ripened after 3 weeks of storage, and then began to decompose, while the fruit treated with 2% chitosan did not ripen fully after more than 1 month of cold storage. This was because of the thickness of the chitosan coating, which blocked the lenticels and caused fermentation inside, and in both cases the fruit were discarded from the evaluation due to the unacceptable quality. The flavor of the fruit with 1.5% chitosan coating was rated as excellent, because the pulp was not only sweet and pleasant, but also had a characteristic aroma (Ali et al., 2010; 2011). Litchi fruit subjected to chitosan treatment either alone or combined with carbonate salts showed good eating quality (Sivakumar et al., 2005a). Several other investigations have reported changes after chitosan application to the color of the fruit peel, which were revealed either by technical instrumentation or by visual appearance 34 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT (Tables 6-8). The application of chitosan coating in longan fruit delayed the fruit peel discoloration, which was related to the concomitant inhibition of PPO activity, the enzyme responsible for polyphenol oxidation (Jiang and Li, 2001). Papaya fruit treated with chitosan underwent light changes in peel color, as indicated by the slower increase in lightness and chroma values, as compared to uncoated fruit. The delay of color development for the papaya fruit treated with 1.0% 1.5% and 2.0% chitosan might be attributable to the slow rate of Downloaded by [189.226.100.40] at 08:02 06 June 2015 respiration and reduced ethylene production, which leads to delayed fruit ripening and senescence (Ali et al., 2011). Similarly, the combination of calcium and chitosan delayed surface color changes of papaya fruit, as noted from the lower values of lightness and chroma and the higher value of hue angle in treated papaya, compared to untreated papaya (Al Eryani et al., 2008). During storage, chitosan coating delayed color changes in banana (Kittur et al., 2001; Win et al., 2007; Baez-Sañudo et al., 2009; Maqbool et al., 2011), litchi fruit (Zhang and Quantick, 1997; Caro and Joas, 2005; Joas et al., 2005; Ducamp-Collin et al., 2008; De Reuck et al., 2009; Sun et al., 2010), mango (Zhu et al., 2008; Salvador-Figueroa et al., 2011), citrus (Canale Rappussi et al., 2011), strawberry (Han et al., 2004; 2005; Hernández-Muñoz et al., 2008), and tomato (El Ghaouth et al., 1992b). Sensory analyses also revealed beneficial effects of chitosan coating in terms of delaying rachis browning and maintenance of the visual aspects of table grape berries (Xu et al., 2007b; Sánchez-González et al., 2011). Fruit and vegetables treated with chitosan have a higher nutritional value, because chitosan can retain the contents of the ascorbic and phenolic compounds (Tables 6-8), which are positively correlated with antioxidant capacity (Rapisarda et al., 1999). Moreover, chitosan can be used as a vehicle for the incorporation of functional ingredients, such as other antimicrobials, 35 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT minerals, antioxidants and vitamins. Some of these combinations can enhance the effects of chitosan or reinforce the nutritional value of the commodities (Vargas et al., 2008). Chitosanbased coatings can also carry high concentrations of calcium or vitamin E, thus significantly increasing the content of these nutrients in fresh and frozen strawberry and raspberry. Incorporation of calcium or vitamin E into chitosan-based coatings did not alter its antifungal properties, while it enhanced the nutritional value of these fresh and frozen strawberry and Downloaded by [189.226.100.40] at 08:02 06 June 2015 raspberry (Han et al., 2004). In addition, incorporation of calcium chloride in chitosan coating increased the stability of the cell wall and middle lamella of the strawberry tissue, and improved its resistance to the pectic enzymes produced by fungal pathogens (Hernández-Muñoz et al., 2006; 2008). Calcium chloride has been added to chitosan coating for papaya (Al Eryani et al., 2008), pear (Yu et al., 2012) and peach (Ruoyi et al., 2005). Core browning is a major problem during storage in pear, and Lin et al. (2008) reported that the combination of chitosan with ascorbic acids not only controlled the core browning of pear, but also increased the ascorbic acid content and the antioxidant capacity of the pear. The combination of chitosan with ascorbic acid showed similar results as for pear (Lin et al., 2008) when applied to litchi fruit (Sun et al., 2010). In the food industry, chitosan shows potential for application to food packaging, as a surrogate for petrochemical based films and as an innovative environmentally friendly material. This arises from its physico-chemical properties, its biodegradability, and its antifungal and antibacterial properties, with nontoxic and nonresidual effects (Porta et al., 2011; Schreiber et al., 2013). Considering the health conscious consumers and the carbon footprint on the environment, modern food packaging needs to address the application of bio-based active films or biopolymers, and chitosan shows potential as a bioagent or additive for the preparation of active 36 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT films. Cervera et al. (2004) reported that chitosan films show higher oxygen barrier properties but lower water vapor barrier properties, mainly due to their hydrophilic nature. The water vapor permeability of chitosan films was shown to increase as a result of water interacting with the hydrophilic chitosan polymer. Incorporation of essential oils reduced the water vapor permeability and the films showed resistance to breaking and were less glossy and deformable; at the same time, the essential oils increased the antimicrobial properties of the coating (Zivanovic Downloaded by [189.226.100.40] at 08:02 06 June 2015 et al., 2005; Hosseini et al., 2008; Sánchez-González et al., 2011). Incorporating nanoparticles into the chitosan film (Qi et al., 2004), such as ZnO (Li et al., 2010b) or Ag (Pinto et al., 2012) nanoparticles, improved the mechanical and barrier properties (Pereira de Abreu et al., 2007) and the thermal stability of the films (de Moura et al., 2009). EFFECTS OF CHITOSAN ON FOODBORNE PATHOGENS Foodborne illnesses are diseases that are caused by agents that enter the human body through the ingestion of food. In 2011, the Center for Disease Control and Prevention (CDC) estimated that in the United States each year there are 48 million foodborne illnesses that are responsible for 128,000 hospitalizations and 3,000 deaths (CDC, 2011). The World Health Organization (WHO) estimates that in 2005, 1.5 million people died worldwide from diarrheal diseases, with a great proportion of these cases being foodborne (WHO, 2006). Furthermore, in the future, with the growth of populations and movement of goods and people at the global scale, this might make the control of foodborne infections more difficult. Recent investigations have identified fruit and vegetables, and in particular leafy greens, as important vehicles for the transmission of many foodborne disease outbreaks (Berger et al., 2010). Nowadays, there is increasing demand for fresh, minimally processed vegetables, such as 37 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT „ready-to-eat‟ salads, which retain much of their indigenous microflora following their minimal processing. All types of produce have the potential to harbor pathogens, and Salmonella spp., Shigella spp., Escherichia coli, Campylobacter spp., Listeria monocytogenes, Yersinia enterocolitica, Bacillus cereus, Clostridium spp., Aeromonas hydrophila, some viruses, and other parasites are of the greatest public health interest (Beuchat, 2002). Fruit and vegetables can be contaminated by these microorganisms during the preharvest stage, mainly by contaminated Downloaded by [189.226.100.40] at 08:02 06 June 2015 water or sewage and faeces, or during the postharvest stage, in the handling and storage of the horticultural products. The growth of microorganisms on fresh-cut produce can also occur during the cutting and slicing operations (Beuchat, 2002). As well as its potentiality as a mechanical barrier, an edible chitosan coating can be used for its antimicrobial properties, to preserve fresh fruit and vegetables after harvest (Vargas et al., 2008). Some studies have reported on the antibacterial activities of chitosan films against foodborne pathogens of fresh fruit and vegetables (Table 9). Inatsu et al. (2010) evaluated different sanitizers to prevent growth of four strains of E. coli on the surface of tomato fruit, and they found that 0.1% chitosan was effective when applied after a sodium chloride washing treatment. However, in this case, other combinations of sanitizers were more effective (e.g., 0.1% lactic acid with 0.05% sodium chloride). Chitosan coating reduced the native microflora on the surface of litchi fruit (Sivakumar et al., 2005a) and strawberry (Ribeiro et al., 2007), but not for table grapes (Romanazzi et al., 2002). However, several additives can be incorporated into the chitosan coating, which can provide more specific functions, such as antimicrobial activity that is aimed at either preventing or reducing the growth of foodborne microorganisms (Vargas et al., 2008). Coatings of chitosan and allyl isothiocyanate 38 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT on cantaloupe reduced the Salmonella presence down to the limit of detection after 2 weeks of storage (Chen et al., 2012). Also when recontamination of cantaloupe with Salmonella was simulated, the results indicated that the chitosan-allyl isothiocyanate coating not only reduced the Salmonella more than the current practice based on acid washing, but it also maintained its antibacterial activity for longer periods of time. Furthermore, the native microflora monitored by the microbial counts for total aerobic bacteria, yeast and mold on the cantaloupe surface during Downloaded by [189.226.100.40] at 08:02 06 June 2015 storage were reduced by the chitosan and allyl isothiocyanate coating (Chen et al., 2012). Essential oils are among the antimicrobial agents that can be incorporated into chitosan coatings (Vargas et al., 2008; Antunes and Cavaco, 2010). A coating of chitosan and bergamot oil reduced the counts of molds, yeast, and mesophiles of table grape berries, as compared to the untreated fruit. The addition of bergamot oil enhanced the antimicrobial activities of the pure chitosan (Sánchez-González et al., 2011). In another study, growth of E. coli DH5α did not take place when the bacterium was incubated on substrates with added chitosan and beeswax, without or with added thyme or lime essential oils (Ramos-García et al., 2012). The antimicrobial activity of chitosan appears to be due to its polycationic characteristics, which allow chitosan to interact with the electronegative charges on the cell surface of fungi and bacteria. This can result in increased microbial cell permeability, internal osmotic disequilibrium, and cell leakage (Helander et al., 2001; Rabea et al., 2003; Liu et al., 2004; Raafat et al., 2008; Mellegård et al., 2011). A 12-h exposure period to chitosan resulted in higher levels of glucose and protein in the supernatant of cell suspensions of Staphylococcus aureus than observed for the medium without chitosan. The reactive amino groups in chitosan might conceivably interact with a multitude of anionic groups on the cell surface, to alter cell permeability and cause leakage of 39 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT intracellular components, such as glucose and protein, which will lead to cell death (Chung et al., 2011). Furthermore, the possibility of a direct interaction of chitosan with negatively charged nucleic acids of microorganisms, and consequently of chitosan interference in RNA and protein synthesis, has been proposed (Rabea et al., 2003). In contrast, Raafat et al. (2008) considered the probabilities of the penetration of chitosan into the nuclei of bacteria to be relatively low, as the size of a molecule of hydrated chitosan is bigger than the cell wall pores. Thus Raafat et al. Downloaded by [189.226.100.40] at 08:02 06 June 2015 (2008) examined cell damage of Staphylococcus simulans after exposure to chitosan, and they found irregular structures that protruded from the cell wall and a „vacuole-like‟ structure that possibly resulted from disruption of the equilibrium of the cell-wall dynamics, such as the ion and water efflux, and decreased the internal pressure; however, on the other hand, the cell membrane remained intact. These results show how chitosan appears not to interact directly with internal structures of the bacteria, but to just interact with external cell-wall polymers. Other mechanisms proposed for the chitosan antimicrobial activity are based on the strong affinity of chitosan for nutritionally essential metal ions. Rabea et al. (2003) reported that the binding of bacterial trace metals by chitosan inhibited both microbial growth and the production of bacterial toxins. The susceptibility of foodborne microorganisms to chitosan also depends on the characteristics of the microorganisms themselves. As the antimicrobial activity of chitosan relies on electrostatic interactions, the nature of the bacterial cell wall can influence the inhibition of microorganism growth by chitosan. The main important foodborne microorganisms are Gramnegative and Gram-positive bacteria. E. coli, Salmonella spp., Shigella spp., A. hydrophila, C. jejuni and Y. enterocolitica, are Gram-negative, and they are characterized by an outer cell wall 40 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT that consists essentially of lipopolysaccharides that contain phosphate and pyrophosphate groups that cover their surface with negative charges. The gram-positive bacteria, such as L. monocytogenes, B. cereus, and C. botulinum, have a cell wall that is composed essentially of peptidoglycan associated to polysaccharides and teichoic acids, which are also negatively charged. According to several studies, Gram-positive bacteria are more susceptible to chitosan than Gram-negative bacteria (No et al., 2002; Takahashi et al., 2008; Jung et al., 2010; Tayel et Downloaded by [189.226.100.40] at 08:02 06 June 2015 al., 2010), while according to others, the opposite is the case (Devlieghere et al., 2004). A recent study reported the effectiveness of chitosan and its derivatives against well-established biofilms formed by foodborne bacteria, which are assumed to be resistant to cleaning and disinfection practices. The results showed that a 1 h exposure to chitosan resulted in reductions in viable cells on mature L. monocytogenes biofilms, and in the attached populations of the other organisms tested, as B. cereus, Salmonella enterica and Pseudomonas fluorescens, except for S. aureus (Orgaz et al., 2011). In the food industry, chitosan is frequently used as an antioxidant, a clarifying agent, and an inhibitor of enzymatic browning. When applied to food, the antimicrobial activities of chitosan can be affected by the pH or the matrix. Indeed, the pKa of chitosan, where half of its amino group are protonated and half are not, is around 6.5; therefore, this means that at pH <6.5, the protonated form of chitosan predominates, which results in a greater positive charge density, and leads to stronger and more frequent electrostatic interactions, and thus to greater antimicrobial effectiveness (Helander et al., 2001; Devlieghere et al., 2004; Jung et al., 2010; Kong et al., 2010). This was illustrated by the growth of Candida lambica, which was completely inhibited at pH 4.0, while at pH 6.0, the same chitosan concentration led to a 41 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT relatively small decrease in growth rate (Devlieghere et al., 2004). Furthermore, this also explains why chitosan is less soluble in water alone than in solutions with acids, where chitosan shows more positive charges, and therefore a greater number of interactions. Chitosan with a higher degree of deacetylation, which has greater numbers of positive charges, would also be expected to have stronger antibacterial activities (Jung et al., 2010; Kong et al., 2010; Tayel et al., 2010). On the other hand, numerous studies have generated different results relating to Downloaded by [189.226.100.40] at 08:02 06 June 2015 correlations between the chitosan bactericidal activities and its molecular weight. In some studies, lower molecular weight chitosans (ranging from 2.7 ×104 to 5.5 ×104 Da) was more effective against bacteria than higher molecular weight chitosans (Liu et al., 2006; Tayel et al., 2010; Kim et al., 2011). In other studies, this trend was observed against Gram-negative bacteria, but not against Gram-positive bacteria (No et al., 2002; Zheng and Zhu, 2003). According to Benhabiles et al. (2012), when the molecular weight of chitosan is low, its polymer chains have greater flexibility to create more bonds, and they can thus better interact with the microbial cells. In other studies, no trends were reported for the antibacterial actions related to increased or decreased molecular weights of chitosan (Jung et al., 2010; Mellegård et al., 2011). CONCLUSIONS AND FUTURE TRENDS This review reports on the recent and most relevant studies concerning preharvest spraying and postharvest application of chitosan for fruit and vegetables. These studies have shown that this biopolymer can effectively maintain the fruit and vegetable quality, and can control their postharvest decay during storage and shelf life. Studies dealing with the mechanisms of action of chitosan as an antimicrobial against postharvest fungi and foodborne bacteria are also summarized here. The film-forming properties, antimicrobial activities, and induction of plant 42 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT resistance of chitosan appear to be the main factors in its success. With its intrinsic properties, and because of its double activity on the host and the pathogen, chitosan can be considered as the first of a new class of plant-protection products (Bautista-Baños et al., 2006). Moreover, chitosan has been under considerable investigation for applications in biomedicine, pharmacology, biotechnology, and in the food industry, due to its biocompatibility, biodegradability, and bioactivity (Synowiecki and Al-Khateeb, 2003; Tharanathan and Kittur, 2003; Wu et al., 2005). Downloaded by [189.226.100.40] at 08:02 06 June 2015 Chitosan is not toxic to humans and its safe use as a pharmaceutical carrier has been reported (Baldrick, 2010; USFDA, 2013). Chitosan has been reported to be a potentially viable alternative for fruit and vegetable preservation. Multicomponent edible coatings can be produced with suitable ingredients for the product to provide the desired barrier protection, while also serving as a vehicle for the incorporation of specific additives that can enhance the functionality, such as antioxidants and antimicrobials, thus avoiding pathogen or foodborne microorganism growth on the surface of fruit and vegetable products (Valencia-Chamorro et al., 2011). The combination of chitosan with minerals, vitamins or other nutraceutical compounds can reinforce the nutritional value of the commodities, without reducing the taste acceptability. This new generation of edible coatings is being especially designed to allow incorporation and/or controlled release of antioxidants, vitamins, nutraceuticals, and natural antimicrobial agents (Vargas et al., 2008; McClements et al., 2009). The availability of commercial chitosan products that are easily dissolvable in water now provides an alternative to synthetic fungicides for growers, for the control of diseases of fruit and vegetables. However, at present, none of the formulations of chitosan are registered as plant 43 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT protectant products. The present review summarizes the application of chitosan either preharvest or postharvest. Here, postharvest treatment is not advisable for fruit that are characterized by a bloom on the surface, such as table grapes, or that have a thin waxy pericarp and succulent flesh, such as strawberries, which can be easily damaged during harvest and postharvest handling. On these commodities, preharvest treatment (even 1-2 days before harvest) can be considered as a promising approach to control the postharvest decay of these fruit under storage. Although a lot Downloaded by [189.226.100.40] at 08:02 06 June 2015 of information regarding the effectiveness of chitosan in the control of postharvest decay of fruit and vegetables is available, its application to large-scale tests and its integration into commercial agricultural practices are key points that need to be investigated further. In addition, more studies concerning the exact mechanisms of action of chitosan are needed. Also, several mechanisms relating to its antifungal and antibacterial activities remain unclear. New knowledge about these aspects will provide the necessary information to support decisions relating to the preparation of the chitosan, which molecular weight chitosan to use, and the kind of commercial formulation. ACKNOWLEDGEMENT This work was supported by EUBerry Project (EU FP7 KBBE 2010-4, Grant Agreement No. 265942). 44 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT REFERENCES Abbasi, N.A., Iqbal, Z., Maqbool, M., and Hafiz, I.A. (2009). Postharvest quality of mango (Mangifera indica L.) fruit as affected by chitosan coating. Pak. J. Bot. 41 : 343-357. Abd-Alla, M.A., and Haggag, W.M. (2010). New safe methods for controlling anthracnose disease of mango (Mangifera indica L.) fruits caused by Colletotrichum gloeosporioides (Penz.). J. Am. Sci. 8 : 361-367. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Ait Barka, E., Eullaffroy, P., Clement, C., and Vernet, G. (2004). Chitosan improves development, and protects Vitis vinifera L. against Botrytis cinerea. Plant Cell Rep. 22 : 608-614. Al Eryani, A.R., Mahmud, T.M.M., Omar, S.R.S., and Zaki, A.R.M. (2008). Effects of calcium infiltration and chitosan coating on storage life and quality characteristics during storage of papaya (Carica papaya L.). Int. J. Agric. Resour. 3 : 296-306. Ali, A., and Mahmud, T.M.M. (2008). The potential use of locally prepared chitosan to control in vitro growth of Colletotrichum gloeosporioides isolated from papaya fruits. Acta Hortic. 804 : 177-182. Ali, A., Muhammad, M.T.M., Sijam, K., and Siddiqui, Y. (2010). Potential of chitosan coating in delaying the postharvest anthracnose (Colletotrichum gloeosporioides Penz.) of Eksotika II papaya. Int. J. Food Sci. Tech. 45 : 2134-2140. Ali, A., Muhammad, M.T.M., Sijam, K., and Siddiqui, Y. (2011). Effect of chitosan coatings on the physicochemical characteristics of Eksotika II papaya (Carica papaya L.) fruit during cold storage. Food Chem. 124 : 620-626. 45 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Antunes, M.D.C., and Cavaco, A.M. (2010). The use of essential oils for postharvest decay control. A review. Flavour Fragr. J. 25 : 351-366. Atkinson R.G., Sutherland P.W., Johnston S.L., Gunaseelan K., Hallett I.C., Mitra D., Brummell D.A., Schroder R., Johnston J.W., and Schaffer R.J. (2012). Down-regulation of polygalacturonase 1 alters firmness, tensile strength and water loss in apple (Malus x domestica) fruit. BMC Plant Biol. 12 : 129. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Ayranci, E., and Tunc, S. (2003). A method for the measurement of the oxygen permeability and the development of edible films to reduce the rate of oxidative reactions in fresh foods. Food Chem. 80 : 423-431. Badawy, M.E.I., and Rabea, E.I. (2009). Potential of the biopolymer chitosan with different molecular weights to control postharvest gray mold of tomato fruit. Postharvest Biol. Technol. 51 : 110-117. Badawy, M.E.I., Rabea, E., Rogge, T.M., Stevens, C.V., Smagghe, G., Steurbaut, W., and Höfte, M. (2004). Synthesis and fungicidal activity of new N,O-Acyl Chitosan derivatives. Biomacromolecules 5 : 589-595. Baez-Sañudo, M., Siller-Cepeda, J., Muy-Rangel, D., and Heredia, J.B. (2009). Extending the shelf-life of bananas with 1-methylcyclopropene and a chitosan-based edible coating. J. Sci. Food Agric. 89 : 2343-2349. Baker, C.J., and Orlandi, E.W. (1995). Active oxygen in plant pathogenesis. Annu. Rev. Phytopathol. 33 : 299-321. Baldrick, P. (2010). The safety of chitosan as a pharmaceutical excipient. Regul. Toxicol. Pharm. 56 : 290-299. 46 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Barrett, D.M., Beaulieu, J.C., and Shewfelt, R. (2010). Color, flavor, texture, and nutritional quality of fresh-cut fruits and vegetables: desirable levels, instrumental and sensory measurement, and the effects of processing. Crit. Rev. Food Sci. Nutr. 50 : 369-389. Bautista-Baños, S., and Bravo-Luna, L. (2004). Evaluación del quitosano en el desarrollo de la pudrición blanda del tomate durante el almacenamiento. Rev. Iberoamericana de Tecnología Postcosecha 6 : 63-67. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Bautista-Baños, S., Hernández-López, M., and Bosquez-Molina, E. (2004). Growth inhibition of selected fungi by chitosan and plant extracts. Mex. J. Phytopathol. 22 : 178-186. Bautista-Baños, S., Hernandez-Lopez, M., Bosquez-Molina, E., and Wilson, C.L. (2003). Effects of chitosan and plant extracts on growth of Colletotrichum gloeosporioides, anthracnose levels and quality of papaya fruit. Crop Prot. 22 : 1087-1092. Bautista-Baños, S., Hernandez-Lauzardo, A.N., Velazquez-del Valle, M.G., Hernandez-Lopez, M., Ait Barka, E., Bosquez-Molina, E., and Wilson, C.L. (2006). Chitosan as a potential natural compound to control pre and postharvest diseases of horticultural commodities. Crop Prot. 25 : 108-118. Bautista-Baños, S., Hernández-López, M., Trejo-Tapia, J.L., Hernández-Lauzardo, A.N., Bautista-Cerón, M.K., and Melo-Giorgana, G.E. (2005). Effect of chitosan on in vitro development and morphology of two isolates of Colletotrichum gloeosporioides Penz. Mex. J. Phytopathol. 23 : 62-67. Benhabiles, M.S., Salah, R., Lounici, H., Drouiche, N., Goosen, M.F.A., and Mameri, N. (2012). Antibacterial activity of chitin, chitosan and its oligomers prepared from shrimp shell waste. Food Hydrocoll. 29 :48-56. 47 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Ben-Shalom, N., Ardi, R., Pinto, R., Aki, C., and Fallik, E. (2003). Controlling gray mould caused by Botrytis cinerea in cucumber plants by means of chitosan. Crop Prot. 22 : 285290. Berger, C.D., Sodha, S. V., Shaw, R.K., Griffin, P.M., Plnk, D., Hand, P., and Frankel, G. (2010). Fresh fruits and vegetables as vehicle for the transmission of human pathogens. Environ. Microbiol. 12 : 2385-2397. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Beuchat, L.R. (2002). Ecological factors influencing survival and growth of human pathogens on raw fruits and vegetables. Microbes Infect. 4 : 413-423. Bhattacharya, A., Sood, P., and Citovsky, V. (2010). The roles of plant phenolics in defence and communication during Agrobacterium and Rhizobium infection. Mol. Plant Pathol. 11 :705-719. Bourlieu C., Guillard, V., Vallès-Pamiès, B., Guilbert, S., and Gontard, N. (2009). Edible moisture barrier: how to assess of their potential and limits in food products shelf-life extension? Crit. Rev. Food Sci. Nutr. 49 : 474-499. Calvo-Garrido, C., Viñas, I., Elmer, P.A.G., Usall, J., and Teixidò, N. (2013). Suppression of Botrytis cinerea on necrotic grapevine tissues by early season applications of natural products and biological control agents. Pest Manag. Sci. in press. Campos-Vargas, R., and Saltveit, M.E. (2002). Involvement of putative chemical wound signals in the induction of phenolic metabolism in wounded lettuce. Physiol. Plant. 114 : 73-84. Canale Rappussi, M.C., Pascholati, S.F., Aparecida Benato, E., and Cia, P. (2009). Chitosan reduces infection by Guignardia citricarpa in postharvest “Valencia” oranges. Braz. Arch. Biol. Technol. 52 : 513-521. 48 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Canale Rappussi, M.C., Benato, E.A., Cia, P., and Pascholati, S.F. (2011). Chitosan and fungicides on postharvest control of Guignardia citricarpa and on quality of “Pêra Rio” oranges. Summa Phytopathol. 37 : 142-144. Caro, Y., and Joas, J. (2005). Postharvest control of litchi pericarp browning (cv. Kwaï Mi) by combined treatment of chitosan and organic acids II. Effect of the initial water content of pericarp. Postharvest Biol. Technol. 38 : 137-144. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Casals, C., Elmer, P.A.G., Viñas, I., Teixidó, N., Sisquella, M., and Usall, J. (2012). The combination of curing with either chitosan or Bacillus subtilis CPA-8- to control brown rot infections caused by Monilinia fructicola. Postharvest Biol. Technol. 64 : 126-132. Cè, N., Noreña, C.P.Z., and Brandelli, A. (2012). Antimicrobial activity of chitosan films containing nisin, peptide P34, and natamycin. J. Food 10 : 21-26. Centers for Diseases Control and Prevention (CDC). Estimates of foodborne illness in the United States. Available at: http://www.cdc.gov/foodborneburden/PDFs/FACTSHEET_A_FINDINGS.pdf. Accessed January 14, 2011. Cervera, M.F., Heinämäki, J., Räsänen, M., Maunu, S.L., Karjalainen, M., Acosta, O.M.N., Colarte, A.I., and Yliruusi, J. (2004). Solid-state characterization of chitosans derived from lobster chitin. Carbohydr. Polym. 58 : 401-408. Cháfer, M., Sánchez-González, L., González-Martínez, C., and Chiralt, A. (2012). Fungal decay and shelf life of oranges coated with chitosan and bergamot, thyme, and tea tree essential oils. J. Food Sci. 77 : 182-187. 49 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Cheah, L.H., Page, B.B.C., and Shepherd, R. (1997).Chitosan coating for inhibition of sclerotinia rots of carrots. New Zeal. J. Crop Hort. 25 : 89-92. Chen, W., Jin, T.Z., Gurtler, J.B., Geveke, D.J. and Fan, X. (2012). Inactivation of Salmonella on whole cantaloupe by application of an antimicrobial coating containing chitosan and allyl isothiocyanate. Int. J. Food Microbiol. 155 : 165-170. Cheng, G.W., and Breen, P.J. (1991). Activity of phenylalanine ammonia-lyase (PAL) and Downloaded by [189.226.100.40] at 08:02 06 June 2015 concentrations of anthocyanins and phenolics in developing strawberry fruit. J. Amer. Soc. Hort. Sci. 116 : 865-869. Chien, P.J., and Chou, C.C. (2006). Antifungal activity of chitosan and its application to control post-harvest quality and fungal rotting of Tankan citrus fruit (Citrus tankan Hayata). J. Sci. Food Agric. 86 : 1964-1969. Chung, Y.C., Yeh, J.Y., and Tsai, C.F. (2011). Antibacterial characteristics and activity of watersoluble chitosan derivatives prepared by the Maillard reaction. Molecules 16 : 8504-8514. Cong, F., Zhang, Y., and Dong, W. (2007). Use of surface coatings with natamycin to improve the storability of Hami melon at ambient temperature. Postharvest Biol. Technol. 46 : 7175. Dang, Q.F., Yan, J.Q., Li, Y., Cheng, X.J., Liu, C.S., and Chen, X.G. (2010). Chitosan acetate as an active coating material and its effects on the storing of Prunus avium L. J. Food Sci. 75 : 125-131. Das, D.K., Dutta, H., and Mahanta, C.L. (2013). Development of a rice starch-based coating with antioxidant and microbe-barrier properties and study of its effect on tomatoes stored at room temperature. LWT – Food Sci. Technol. 50 : 272-278. 50 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT De Capdeville, G., Wilson, C.L., Beer, S.V., and Aist, J.R. (2002). Alternative disease control agents induce resistance to blue mold in harvested “Red Delicious” apple fruit. Phytopathology 92 : 900-908. De Moura, M.R., Aouada, F.A., Avena-Bustillos, R.J., McHugh, T.H., Krochta, J.M., and Mattoso, L.H.C. (2009). Improved barrier and mechanical properties of novel hydroxypropyl methylcellulose edible films with chitosan/tripolyphosphate nanoparticles. Downloaded by [189.226.100.40] at 08:02 06 June 2015 J. Food Eng. 92 : 448-453. De Reuck, K., Sivakumar, D., and Korsten, L. (2009). Effect of integrated application of chitosan coating and modified atmosphere packaging on overall quality retention in litchi cultivars. J. Sci. Food Agric. 89 : 915-920. Devlieghere, F., Vermeulen, A., and Debevere, J. (2004). Chitosan: antimicrobial activity, interactions with food components and applicability as a coating on fruit and vegetables. Food Microbiol. 21 : 703-714. Du, J., Gemma, H., and Iwahori, S. (1997). Effects of chitosan coating on the storage of peach, Japanese pear and kiwifruit. J. Japan. Soc. Hort. Sci. 66 : 15-22. Duan, J., Wu, R., Strik, B.C., and Zhao, Y. (2011). Effect of edible coating on the quality of fresh blueberry (Duke and Elliott) under commercial storage conditions. Postharvest Biol. Technol. 59 : 71-79. Ducamp-Collin, M.N., Ramarson, H., Lebrun, M., Self, G., and Reynes, M. (2008). Effect of citric acid and chitosan on maintaining red colouration of litchi fruit pericarp. Postharvest Biol. Technol. 49 : 241-246. 51 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Eikemo, H., Stensvand, A., and Tronsmo, A.M. (2003). Induced resistance as a possible means to control diseases of strawberry caused by Phytophthora spp. Plant Dis. 87 : 345-350. El Ghaouth, A., Arul, J., Ponnampalam, R., and Boulet, M. (1991a). Chitosan coating effect on storability and quality of fresh strawberries. J. Food Sci. 56 : 1618-1620. El Ghaouth, A., Arul, J., Ponnampalam, R., and Boulet, M. (1991b). Use of chitosan coating to reduce water loss and maintain quality of cucumber and bell pepper fruits. J. Food Downloaded by [189.226.100.40] at 08:02 06 June 2015 Process. Pres. 15 : 359-368. El Ghaouth, A., Arul, J., Grenier, J., and Asselin, A. (1992a). Antifungal activity of chitosan on two postharvest pathogens of strawberry fruits. Phytopatology 82 : 398-402. El Ghaouth, A., Ponnampalam, R., Castaigne, F., and Arul, J. (1992b). Chitosan coating to extend the storage life of tomatoes. Hortscience 27 : 1016-1018. El Ghaouth, A., Smilanick, J.L., and Wilson, C.L. (2000). Enhancement of the performance of Candida saitoana by the addition of glycolchitosan for the control of the postharvest decay of apple and citrus fruit. Postharvest Biol. Technol. 19 : 103-110. Elmer, P.A.G., and Reglinski, T. (2006). Biosuppression of Botrytis cinerea in grapes. Plant Pathol. 55 : 155-177. El-Mougy, N.S., Abdel-Kader, M.M., and Aly, M.H. (2012). Effect of a new chemical formula on postharvest decay incidence in citrus fruit. J. Plant Prot. Res. 52 : 156-164. Eweis, M., Elkholy, S.S., and Elsabee, M.Z. (2006). Antifungal efficacy of chitosan and its thioures derivatives upon the growth of some sugar-beet pathogens. Int. J. Biol. Macromol. 38 : 1-8. 52 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT FAO. 2011. Global food losses and food waste. In: International Congress “SAVE FOOD!”, 1617 May, Düsseldorf, Germany. Feliziani, E., Smilanick, J.L., Margosan, D.A., Mansour, M.F., Romanazzi, G., Gu, S., Gohil, H.L., and Rubio Ames, Z. (2013a). Preharvest fungicide, potassium sorbate, or chitosan use on quality and storage decay of table grapes. Plant Dis. 97 : 307-314. Feliziani, E., Santini, M., Landi, L., and Romanazzi, G. (2013b). Pre and postharvest treatment Downloaded by [189.226.100.40] at 08:02 06 June 2015 with alternatives to synthetic fungicides to control postharvest decay of sweet cherry. Postharvest Biol. Technol. 78 : 133-138. Fornes, F., Almela, v., Abad, M., and Agustí, M. (2005). Low concentrations of chitosan coating reduce water spot incidence and delay peel pigmentation of Clementine mandarin fruit. J. Sci. Food Agric. 85 : 1105-1112. Gao, P., Zhu, Z., and Zhang, P. (2013). Effects of chitosan-glucose complex coating on postharvest quality and shelf life of table grapes. Carbohyd. Polym. 95 : 371-378. García-Rincón, J., Vega-Pérez, J., Guerra-Sánchez, M.G., Hernández-Lauzardo, A.N., PeñaDíaz, A., and Velázquez-Del Valle, M.G. (2010). Effect of chitosan on growth and plasma membrane properties of Rhizopus stolonifer (Ehrenb.:Fr.) Vuill. Pestic. Biochem. Physiol. 97 : 275-278. Gerasimova, N.G., Pridvorova, S.M., and Ozeretskovskaya, O.L. (2005). Role of LPhenylalanine ammonia lyase in the induced resistance and susceptibility of potato plants. Appl. Biochem. Microbiol. 41 : 103-105. 53 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Gatto, M.A., Ippolito, A., Linsalata, V., Cascarano, N.A., Nigro, F., Vanadia, S., and Di Venere, D. (2011). Activity of extracts from wild edible herbs against postharvest fungal diseases of fruit and vegetables. Postharvest Biol. Technol. 61 : 72-82. Ghasemnezhad, M., Shiri, M.A., and Sanavi, M. (2010). Effect of chitosan coatings on some quality indices of apricot (Prunus armeniaca L.) during cold storage. Caspian J. Env. Sci. 8 : 25-33. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Guerra-Sánchez, M.G., Vega-Pérez, J., Velázquez-del Valle, M.G., and Hernández-Lauzardo, A.N. (2009). Antifungal activity and release of compounds on Rhizopus stolonifer (Ehrenb.:Fr.) Vuill. by effect of chitosan with different molecular weights. Pestic. Biochem. Physiol. 93 : 18-22. Hadwiger, L.A. (1999). Host-parasite interactions: elicitation of defense responses in plant with chitosan. Experientia Suppl. 87 : 185-200. Hadwiger, L.A., and Loschke D.C. (1981). Molecular communication in host-parasite interactions: hexosamine polymers (chitosan) as regulator compounds in race-specific and other interactions. Phytopathology 71 : 756-762. Han, C., Zhao, Y., Leonard, S.W., and Traber, M.G. (2004). Edible coating to improve storability and enhance nutritional value of fresh and frozen strawberry (Fragaria x ananassa) and raspberries (Rubus ideaus). Postharvest Biol. Technol. 33 : 67-78. Han, C., Lederer, C., McDaniel, M., and Zhao, Y. (2005). Sensory evaluation of fresh strawberry (Fragaria x ananassa) coated with chitosan-based edible coatings. J. Food Sci. 70 : 172178. 54 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Helander, I.M., Nurmiaho-Lassila, E.L., Ahvenainen, R., Rhoades, J., and Roller, S. (2001). Chitosan disrupts the barrier properties of the outer membrane of Gram-negative bacteria. Int. J. Food Microbiol. 71 : 235-244. Hernández-Lauzardo, A.N., Velázquez-Del Valle, M.G., Veranza-Castelán, L., Melo-Giorgana, G.E., and Guerra-Sánchez, M.G. (2008). Effect of chitosan and three isolates of Rhizopus stolonifer obtained from peach, papaya and tomato. Fruits 65 : 245-253. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Hernández-Muñoz, P., Almenar, E., Ocio, M.J., and Gavara, R. (2006). Effect of calcium dips and chitosan coatings on postharvest life of strawberries (Fragaria x ananassa). Postharvest Biol. Technol. 39 : 247-253. Hernández-Muñoz, P., Almenar, E., del Valle, V., Velez, D., and Gavara, R. (2008). Effect of chitosan coating combined with postharvest calcium treatment on strawberry (Fragaria x ananassa) quality during refrigerated storage. Food Chem. 110 : 428-435. Hewajulige, I.G.N., Sultanbawa, Y., and Wijesundara, R.L.C. (2009). Mode of action of chitosan coating on anthracnose disease control in papaya. Phytoparasitica 37 : 437-444. Hong, K., Xie, J., Zhang, L., Sun, D., and Gong, D. (2012). Effects of chitosan coating in postharvest life and quality of guava (Psidium guajava L.) fruit during cold storage. Sci. Hortic. 144 : 172-178. Hosseini, M.H., Razavi, S.H., Mousavi, S.M.A., Yasaghi, S.A.S., and Hasansaraei, A.G. (2008). Improving antibacterial activity of edible films based on chitosan by incorporating thyme and clove essential oils and EDTA. J. Appl. Sci. 8 : 2895-2900. 55 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Inatsu, Y., Kitagawa, T., Bari, M.L., Nei, D., Juneja, V., and Kawamoto, S. (2010). Effectiveness of acidified sodium chloride and other sanitizers to control Escherichia coli O157:H7 on tomato surfaces. Foodborne Pathog. Dis. 7 : 629-635. Iriti, M., Vitalini, S., Di Tommaso, G., D‟Amico, S., Borgo, M., and Faoro, F. (2011). New chitosan formulation prevents grapevine powdery mildew infection and improves polyphenol content and free radical scavenging activity of grape and wine. Aust. J. Grape Downloaded by [189.226.100.40] at 08:02 06 June 2015 Wine Res. 17 : 263-269. Jiang, Y., and Li, Y. (2001). Effects of chitosan coatings on postharvest life and quality of longan fruit. Food Chem. 73 : 139-143. Jiang, Y., Li, J., and Jiang, W. (2005). Effects of chitosan on shelf life of cold-stored litchi fruit at ambient temperature. LWT- Food Sci. Technol. 57 : 757-761. Jitareerat, P., Paumchai, S., Kanlayanarat, S., and Sangchote, S. (2007). Effect of chitosan on ripening, enzymatic activity, and disease development in mango (Mangifera indica) fruit. New Zeal. J. Crop Hort. 35 : 211-218. Joas, J., Caro, Y., Ducamp, M.N., and Reynes, M. (2005). Postharvest control of pericarp browning of litchi fruit (Litchi chinensis Sonn cv Kwaï Mi) by treatment with chitosan and organic acids I. Effect of pH and pericarp dehydratation. Postharvest Biol. Technol 38 : 128-136. Jung, E.J., Youn, D.K., Lee, S.H., No, H.K., Ha, J.G., and Prinyawiwatkul, W. (2010). Antibacterial activity of chitosans with different degrees of deacetylation and viscosities. Int. J. Food Sci. Technol. 45 : 676-682. 56 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Kader, A.A. (2005). Increasing food availability by reducing postharvest losses of fresh produce. Acta Hortic. 682 : 2169-2175. Kerch, G., Sabovics, M., Kruma, Z., Kampuse, S., and Straumite, E. (2011). Effect of chitosan and chitooligosaccharide on vitamin C and polyphenols contents in cherries and strawberries during refrigerated storage. Eur. Food Res. Technol. 233 : 351-358. Kim, K.W., Min, B.J., Kim, Y.T., Kimmel, R.M., Cooksey,K., and Park, S.I. (2011). Downloaded by [189.226.100.40] at 08:02 06 June 2015 Antimicrobial activity against foodborne pathogens of chitosan biopolymer films of different molecular weights. LWT – Food Sci. Technol. 44 : 565-569. Kittur, F.S., Saroja, N., Habibunnisa, and Tharanathan, R.N. (2001). Polysaccharide-based composite formulations for shelf-life extension of fresh banana and mango. Eur. Food Res. Technol. 213 : 306-311. Kong, M., Chen, X.G., Xing, K., and Park, H.J. (2010). Antimicrobial properties of chitosan and mode of action: a state of the art review. Int. J. Food Microbiol. 144 : 51-63. Kurzawińska, H., and Mazur, S. (2007). The effect of Pythium oligandrum and chitosan used in control of potato against late blight and the occurrence of fungal disease on tuber peel. Commun. Agric. Appl. Biol. Sci. 72 : 967-971. Landi, L., Feliziani, E. and Romanazzi, G. (2014). Expression of defense genes in strawberry fruit treated with different resistance inducers. J. Agric. Food Chem. (in press). Lattanzio, V., Lattanzio, V.M.T. and Cardinali, A. (2006). Role of phenolics in the resistance mechanisms of plants against fungal pathogens and insects. In: Phytochemistry Advances in Research (Imperato, F. ed.), pp. 23–67. India: Research Signpost. 57 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Li, H., and Yu, T. (2001). Effect of chitosan on incidence of brown rot, quality and physiological attributes of postharvest peach fruit. J. Sci. Food Agric. 81 : 269-274. Li, X.F., Feng, X.Q., Yang, S., and Wang, T.P. (2008). Effects of molecular weight and concentration of chitosan on antifungal activity against Aspergillus niger. Iran. Polym. J. 17 : 843-852. Li, Y.C., Sun, X.J., Bi, Y., Ge, Y.H., and Wang, Y. (2009). Antifungal activity of chitosan on Downloaded by [189.226.100.40] at 08:02 06 June 2015 Fusarium sulphureum in relation to dry rot of potato tuber. Agr. Sci. China 8 : 597-604. Li, J., Yan, J., Wang, J., Zhao, Y., Cao, J., and Jiang, W. (2010a). Effects of chitosan coating on oxidative stress in bruised Yali pears (Pyrus bretschneideri Rehd.). Int. J. Food Sci. Tech. 45 : 2149-2154. Li, L.H., Deng, J.C., Deng, H.R., Liu, Z.L., and Xin, L. (2010b). Synthesis and characterization of chitosan/ZnO nanoparticle composite membranes. Carbohydr. Res. 345 : 994-998. Lin, L., Wang, B., Wang, M., Cao, J., Zhang, J., Wu, Y., and Jiang, W. (2008). Effects of a chitosan-based coating with ascorbic acid on post-harvest quality and core browning of “Yali” pear (Pyrus bertschneideri Rehd.). J. Sci. Food Agric. 88 : 877-884. Lin, B., Du Y., Liang X., Wang, X., Wang, X., and Yang, J. (2011). Effect of chitosan coating on respiratory behavior and quality of stored litchi under ambient temperature. J. Food Eng. 102 : 94-99. Lira-Saldivar, R.H., Hernández-Suárez, M., and Hernández-Castillo, F.D. (2006). Activity of Larrea tridentata (D.C.) Coville L. extracts and chitosan against fungi that affect horticultural crops. Rev. Chapingo Ser. Hortic. 12 : 211-216. 58 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Liu, J., Tian, S., Meng, X., and Hu, Y. (2007). Effects of chitosan on control of postharvest diseases and physiological responses of tomato fruit. Postharvest Biol. Technol. 44 : 300306. Liu, N., Chen, X.G., Park, H.J., Liu, C.G., Liu, C.S., Meng, X.H., and Yu, L.J. (2006). Effect of MW and concentration of chitosan on antibacterial activity of Escherichia coli. Carbohyd. Polym. 64 : 60-65. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Liu, H., Du, Y., Wang, X., and Sun, L. (2004). Chitosan kills bacteria through cell membrane damage. Int. J. Food Microbiol. 95 : 147-155. Maqbool, M., Ali, A., Ramachandran, S., Smith, D.R., and Alderson, P.G. (2010a). Control of postharvest antrachnose of banana using a new edible composite film. Crop Prot. 29 : 1136-1141. Maqbool, M., Ali, A., and Alderson, P.G. (2010b). A combination of gum arabic and chitosan can control antrachnose caused by Colletotrichum musae and enhance the shelf-life of banana fruit. J. Hortic. Sci. Biotech. 85 : 432-436. Maqbool, M., Ali, A., Alderson, P.G., Zahid, N., and Siddiqui, Y. (2011). Effect of a novel edible composite coating based on gum Arabic and chitosan on biochemical and physiologic responses of banana fruit during cold storage. J. Agr. Food Chem. 59 : 54745482. Martínez-Castellanos, G., Shirai, K., Pelayo-Zaldívar, C., Pérez-Flores, L., and SepúlvedaSánchez, J.D. (2009). Effect of Lactobacillus plantarum and chitosan in the reduction of browning of pericarp Rambutan (Nephelium lappaceum). Food Microbiol. 26 : 444-449. 59 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Mazaro, S.M., Deschamps, C., May De Mio, L.L., Biasi, L.A., De Gouvea, A., and Kaehler Sautter, C. (2008). Postharvest behaviour of strawberry fruits after pre harvest treatment with chitosan and acibenzolar-s-methyl. Rev. Bras. Frutic. 30 : 185-190. McClements, D.J., Decker, E.A., Park, Y., and Weiss, J. (2009). Structural design principles for delivery of bioactive components in nutraceuticals and functional foods. Crit. Rev. Food Sci. 49 : 577-606. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Mellegård H., Strand, S.P., Christensen, B.E., Granum, P.E., and Hardy, S.P. (2011). Antibacterial activity of chemically defined chitosans: Influences of molecular weight, degree of acetylation and test organism. Int. J. Food Microbiol. 148 : 48-54. Meng, X., Li, B., Liu, J., and Tian, S. (2008). Physiological responses and quality attributes of table grape fruit to chitosan preharvest spray and postharvest coating during storage. Food Chem. 106 : 501-508. Meng, X., and Tian., S. (2009). Effects of preharvest application of antagonistic yeast combined with chitosan on decay and quality of harvested table grape fruit. J. Sci. Food Agric. 89 : 1838-1842. Meng, X., Yang, L., Kennedy, J.F., and Tian, S. (2010a). Effects of chitosan and oligochitosan on growth of two fungal pathogens and physiological properties in pear fruit. Carbohyd. Polym. 81 : 70-75. Meng, X.H., Qin, G.Z., and Tian, S.P. (2010b). Influences of preharvest spraying Cryptococcus laurentii combined with chitosan coating on postharvest disease and quality of table grapes in storage. LWT – Food Sci. Technol. 43 : 596-601. 60 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Molloy, C., Cheah, L.H., and Koolaard, J.P. (2004). Induced resistance against Sclerotinia sclerotiorum in carrots treated with enzymatically hydrolysed chitosan. Postharvest Biol. Technol. 33 : 61-65. Muñoz, Z., Moret, A., and Garcés, S. (2009). Assessment of chitosan for inhibition of Colletotrichum sp. on tomatoes and grapes. Crop Prot. 28 : 36-40. Nawrocki, J. (2006). The protection of parsley seedlings (Petroselinum sativum Hoffm. ssp. Downloaded by [189.226.100.40] at 08:02 06 June 2015 microcarpum) against damping-off. Commun. Agric. Appl. Biol. Sci. 71 : 993-997. No, H.K., Park, N.Y., Lee, S.H., and Meyers, S.P. (2002). Antibacterial activity of chitosan and chitosan oligomers with different molecular weights. Int. J. of Food Microbiol. 74 : 6572. Ojaghian, M.R., Amoneafy, A.A., Cui, Z.Q., Xie, G.L., Zhang, J., Shang, C., and Li., B. (2013). Application of acetyl salicylic acid and chemically different chitosans against storage carrot rot. Postharvest Biol. Technol. 84 : 51-60. Olivas, G.I., and Barbosa-Cánovas, G.V. (2005). Edible coatings for fresh-cut fruits. Crit. Rev. Food Sci. 45 : 657-670. Orgaz, B., Lobete, M.M., Puga, C.H., and San Jose, C. (2011). Effectiveness of chitosan against mature biofilms formed by food related bacteria. Int. J. Mol. Sci. 12 : 817-828. Özden, Ç., and Bayindirli, L. (2002). Effects of combinational use of controlled atmosphere, cold storage and edible coating applications on shelf life and quality attributes of green peppers. Eur. Food Res. Technol. 214 : 320-326. 61 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Palma-Guerrero, J., Jansson, H., Salinas, J., and Lopez-Llorca, L. (2008). Effect of chitosan on hyphal and spore germination of plant pathogenic and biocontrol fungi. J. Applied Microbiol. 104 : 541-553. Palma-Guerrero, J., Huang, I., Jansson, H., Salinas, J., Lopez-Llorca, L., and Read, N. (2009). Chitosan permeabilizes the plasma membrane and kills cells of Neurospora crassa in an energy dependent manner. Fungal Genet. Biol. 46 : 585-594. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Palma-Guerrero, J., Lopez-Jimenez, J., Pérez-Berna, A., Huang, I., Jansson, H., Salinas, J., Villalaín, J., Read, N., and Lopez-Llorca, L. (2010). Membrane fluidity determines sensitivity of filamentous fungi to chitosan. Mol. Microbiol. 75 : 1021-1032. Park, S.I., Stan, S.D., Daescheler, M.A., and Zhao, Y. (2005). Antifungal coatings on fresh strawberries (Fragaria x ananassa) to control mold growth during cold storage. J. Food Sci. 70 : 202-207. Perdones, A., Sánchez-González, L., Chiralt, A., and Vargas, M. (2012). Effect of chitosanlemon essential oil coatings on storage-keeping quality of strawberry. Postharvest Biol. Technol. 70 : 32-41. Pereira de Abreu, D.A., Paseiro Losada, P., Angulo, I., and Cruz, J.M. (2007). Development of new polyolefin films with nanoclays for application in food packaging. Eur. Polym. J. 43 : 2229-2243. Pinto, R.J.B., Fenandes, S.C.M., Freira, C.S.R., Sadocco, P., Causio, J., Neto, C.P., and Trindade, T. (2012). Antibacterial activity of optically transparent nanocomposite films based on chitosan or its derivatives and silver nanoparticles. Carbohydr. Res. 348 : 77-83. 62 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Plascencia-Jatomea, M., Viniegra, G., Olayo, R., Castillo-Ortega, M.M., and Shirai, K. (2003). Effect of chitosan and temperature on spore germination of Aspergillus niger. Macromol. Biosci. 3 : 582-586. Porta, R., Mariniello, L., di Pierro, P., Sorrentino, A., and Giosafatto, C.V.L. (2011). Transglutaminase crosslinked pectinand chitosan-based edible films: a review. Crit. Rev. Food Sci. 51 : 223-238. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Qiuping, Z., and Wenshui, X. (2007). Effect of 1-methylcycloprpene and/or chitosan coating treatments on storage life and quality maintenance of Indian jujube fruit. LWT – Food Sci. Technol. 40 : 404-411. Qi, L., Xu, Z., Jiang, X., Hu, C., and Zou, X. (2004). Preparation and antibacterial activity of chitosan nanoparticles. Carbohydr. Polym. 339 : 2693-2700. Raafat, D., von Bargen, K., Hass, A., and Sahl, H.G. (2008). Insights into the mode of action of chitosan as an antimicrobial compound. Appl. Environ. Microbiol. 74 : 3764-3773. Rabea, E.I., Badawy, M.E.T., Stevens ,C.V., Smagghe, G., and Steurbaut, W. (2003). Chitosan as antimicrobial agent: application and mode of action. Biomacromolecules 4 : 14571465. Rabea, E.I., and Badawy, M.E.I. (2012). Inhibitory effects on microbial growth of Botrytis cinerea and Erwinia carotovora on potato using of a biopolymer chitosan at differnt molecular weights. Arc. Phytopathol. Plant Prot. 1-11. Rapisarda, P., Tomaino, A., Lo Cascio, R., Bonina, F., De Pasquale, A., and Saijia, A. (1999). Antioxidant effectiveness as influenced by phenolic content of fresh orange juices. J. Agric. Food Chem. 47 : 4718-4723. 63 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Ramos-García, M., Bosquez-Molina, E., Hernández-Romano, J., Zavala-Padilla, G., TerrésRojas, E., Alia-Tejacal, I., Barrera-Necha, L., Hernández-López, M., and Bautista-Baños, S. (2012). Use of chitosan-based edible coatings in combination with other natural compound, to control Rhizopus stolonifer and Escherichia coli DH5α in fresh tomatoes. Crop Prot. 38 : 1-6. Reddy, B.M.V., Belkacemi, K., Corcuff, R., Castaigne, F., and Arul, J. (2000a). Effect of pre- Downloaded by [189.226.100.40] at 08:02 06 June 2015 harvest chitosan sprays on post-harvest infection by Botrytis cinerea and quality of strawberry fruit. Postharvest Biol. Technol. 20 : 39-51. Reddy, B.M.V., Angers, P., Castaigne, F., and Arul, J. (2000b). Chitosan effect on blackmold rot and pathogenic factors produced by Alternaria alternata in postharvest tomatoes. J. Am. Soc. Hortic. Sci. 125 : 742-747. Reglinski, T., Elmer, P.A.G., Taylor, J.T., Wood, P.N., and Hoyte, S.M. (2010). Inhibition of Botrytis cinerea growth and suppression of botrytis bunch rot in grape using chitosan. Plant Pathol. 59 : 882-890. Ribeiro, C., Vicente, A.A., Teixeira, J.A., and Miranda, C. (2007). Optimization of edible coating composition to retard strawberry fruit senescence. Postharvest Biol. Technol. 44 : 63-70. Rice-Evans, C. A., Miller, N. J., and Paganga, G. (1997). Antioxidant properties of phenols. Trends Plant Sci. 2 : 152-159. Romanazzi, G., Schena, L., Nigro, F., and Ippolito, A. (1999). Preharvest chitosan treatments for the control of postharvest decay of sweet cherries and table grapes. J. Plant Pathol. 81 : 237. 64 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Romanazzi, G., Nigro, F., and Ippolito, A. (2000). Effetto di trattamenti pre e postraccolta con chitosano sui marciumi della fragola in conservazione. Frutticoltura 62 : 71-75. Romanazzi, G., Nigro, F., Ippolito, A., Di Venere, D., and Salerno, M. (2002). Effects of pre and postharvest chitosan treatments to control storage grey mold of table grapes. J. Food Sci. 67 : 1862-1867. Romanazzi, G., Nigro, F., and Ippolito, A. (2003). Short hypobaric treatments potentiate the Downloaded by [189.226.100.40] at 08:02 06 June 2015 effect of chitosan in reduction storage decay of sweet cherries. Postharvest Biol. Technol. 29 : 73-80. Romanazzi, G., Mlikota Gabler, F., and Smilanick, J.L. (2006). Preharvest chitosan and postharvest UV irradiation treatments suppress gray mold of table grapes. Plant Dis. 90 : 445-450. Romanazzi, G., Karabulut, O.A., and Smilanick, J.L. (2007). Combination of chitosan and ethanol to control postharvest gray mold of table grapes. Postharvest Biol. Technol. 45 : 134-140. Romanazzi, G., Mlikota Gabler, F., Margosan, D.A., Mackey, B.E., and Smilanick, J.L. (2009). Effect of acid used to dissolve chitosan on its film forming properties and its ability to control postharvest gray mold of table grapes. Phytopathology 99 : 1028-1036. Romanazzi, G., Feliziani, E., Santini, M., and Landi, L. (2013). Effectiveness of postharvest treatment with chitosan and other resistance inducers in the control of storage decay of strawberry. Postharvest Biol. Technol. 75 : 24-27. 65 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Ruoyi K., Zhifang Y., and Zhaoxin L. (2005). Effect of coating and intermittent warming on enzymes, soluble pectin substances and ascorbic acid of Prunus persica (Cv. Zhonghuashoutao) during refrigerated storage. Food Res. Int. 38 : 331-336. Sánchez-Domínguez, D., Ríos M.Y., Castillo-Ocampo, P., Zavala-Padilla, G., Ramos-García, M., and Buatista-Baños, S. (2011). Cytological and biochemical changes induces by chitosan in the pathosystem Alternaria alternata-tomato. Pesticide Biochem. Physiol. 99 : Downloaded by [189.226.100.40] at 08:02 06 June 2015 250-255. Sánchez-González, L., Pastor, C., Varga, M., Chiralt, A., González-Martínez, C., and Cháfer, M. (2011). Effect of hydroxypropylmethylcellulose and chitosan coatings with and without bergamot essential oil on quality and safety of cold-stored grapes. Postharvest Biol. Technol. 60 : 57-63. Salvador-Figueroa, M., Aragón-Gómez, W.I., Hernández-Ortiz, E., Vázquez-Ovando, J.A., and Adriano-Anaya M. (2011). Effect of chitosan coating on some characteristics of mango (Mangifera indica L.) “Ataulfo” subjected to hydrothermal process. Afr. J. Agric. Resour. 6 : 5800-5807. Schreiber, S.B., Bozell, J.J., Hayes, D.G., and Zivanovic, S. (2013). Introduction of primary antioxidant activity to chitosan for application as a multifunctional food packaging material. Food Hydrocolloid. 33 : 207-214. Shao, X.F., Tu, K., Tu, S., and Tu, J. (2012). A combination of heat treatment and chitosan coating delays ripening and reduces decay in “Gala” apple fruit. J. Food Qual. 35 : 83-92. Shiri, M.A., Ghasemnezhad, M., Bakhshi, D., and Sarikhani, H. (2012). Effect of postharvest putrescine application and chitosan coating on maintaining quality of table grape cv. 66 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT “Shahroudi” during long-term storage. J. Food Process. Pres. 37 : 999-1007.Sivakumar, D., Regnier, T., Demoz, B., and Korsten, L. (2005a). Effect of different post-harvest treatments on overall quality retention in litchi fruit during low temperature storage. J. Hortic. Sci. Biotech. 80 : 32-38. Sivakumar, D., Sultanbawa, J., Ranasingh, N., Kumara, P., and Wijesundera, R.L.C. (2005b). Effect of the combined application of chitosan and carbonate salts on the incidence of Downloaded by [189.226.100.40] at 08:02 06 June 2015 anthracnose and on the quality of papaya during storage. J. Hortic. Sci. Biotech. 80 : 447452. Stevens, C., Liu, J., Khan, V.A., Lu, J.Y., Kabwe, M.K., Wilson, C.L., Igwegbe, E.C.K., Chalutz, E., and Droby, S. (2004). The effects of low-dose ultraviolet light-C treatment on polygalacturonase activity, delay ripening and Rhizopus soft rot development of tomatoes. Crop Prot. 23 : 551-554. Sun, D., Liang, G., Xie, J., Lei, X., and Mo, Y. (2010). Improved preservation effects of litchi fruit by combining chitosan coating with ascorbic acid treatment during postharvest storage. Afr. J. Biotechnol. 9 : 3272-3279. Synowiecki, J., and Al-Khateeb, N. (2003). Production, properties, and some new applications of chitin and its derivatives. Crit. Rev. Food Sci. Nutr. 43 : 145-171. Takahashi, T., Imai, M., Suzuki, I., and Sawai, J. (2008). Growth inhibition effect on bacteria of chitosan membranes regulated with deacetylation degree. Biochem. Eng. J. 40 : 485-491. Tayel, A.A., Moussa, S., Opwis, K., Knittel, D., Schollmeyer, E., and Nickisch-Hartfiel, A. (2010). Inhibition of microbial pathogens by fungal chitosan. Int. J. Biol. Macromol. 47 : 10-14. 67 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Tharanathan, R.N., and Kittur, F.S. (2003). Chitin – The undisputed biomolecule of great potential. Crit. Rev. Food Sci. 43 : 61-87. Torres, R., Valentines, M.C., Usall, J., Vinas, I., and Larrigaudiere, C. (2003). Possible involvement of hydrogen peroxide in the development of resistance mechanisms in „Golden Delicious‟ apple fruit. Postharv. Biol. Technol. 27 : 235–242. USFDA, 2013. GRAS notice inventory. GRN No. 397. www.fda.gov Downloaded by [189.226.100.40] at 08:02 06 June 2015 Valencia-Chamorro, S., Palou, L., del Rio, M.A., and Pérez-Gago, M.B. (2011). Antimicrobial edible coating for fresh and minimally processed fruits and vegetables: a review. Crit. Rev. Food Sci. 51 : 872-900. Van-Loon, L.C., and Van-Strien, E.A. (1999). The families of pathogenesis-related proteins, their activities, and comparative analysis of PR-1 type proteins. Physiological and Molecular Plant Pathology 55 : 85-97. Vargas, M., Albors, A., Chiralt, A., and Gonzalez-Martinez, C. (2006). Quality of cold-stored strawberries as affected by chitosan-oleic acid edible coatings. Postharvest Biol. Technol. 41 : 164-171. Vargas, M., Pastor, C., Chiralt, A., McClements, J., and González-Martínez, C. (2008). Recent advances in edible coatings for fresh and minimally processed fruits. Recent advances in edible coatings for fresh and minimally fruits. Crit. Rev. Food Sci. 48 : 496.511. Vu, D.K., Hollingsworth R.G., Leroux E., Salmieri S., and Lacroix M. (2011). Development of edible bioactive coating based on modified chitosan for increasing the shelf life of strawberries. Food Res. Int. 44 : 198-203. 68 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Wang, S.Y., and Gao, H. (2012). Effect of chitosan-based edible coating on antioxidants, antioxidant enzyme system, and postharvest fruit quality of strawberries (Fragaria x ananassa Duch.). LWT – Food Sci. Technol. 52 : 71-79. Win, N.K.K., Jitareerat, P., Kanlayanarat, S., and Sangchote, S. (2007). Effects of cinnamon extract, chitosan coating, hot water treatment and their combinations on crown rot disease and quality of banana fruit. Postharvest Biol. Technol. 45 : 333-340. Downloaded by [189.226.100.40] at 08:02 06 June 2015 World Health Organization (WHO). (2006). WHO consultation to develop a strategy to estimate the global burden of foodborne diseases: taking stock and charting the way forward. Geneva, Switzerland: WHO, September 25–27, 2006. Wu, H., Wang, D., Shi, J., Xue, S., and Gao, M. (2010). Effect of complex of Zinc (II) and Cerium (IV) with chitosan on the preservation quality and degradation of organophosphorus pesticides in Chinese jujube (Zizyphus jujube Mill. Cv. Dongzao). J. Agric. Food Chem. 58 : 5757-5762. Wu, T., Zivanovic, S., Draughon, F.A., Conway, W.S., and Sams, C.E. (2005). Physiochemical properties and bioactivity of fungal chitin and chitosan. J. Agr. Food Chem. 53 : 38883894. Yaman, Ö., and Bayindirli, L. (2001). Effects of an edible coating and cold storage on shelf-life and quality of cherries. LWT – Food Sci. Technol. 35 : 146-150. Yang, L., Zhao, P., Wang, L., Filippus, I., and Meng, X. (2010). Synergistic effect of oligochitosan and silicon on inhibition of Monilia fructicola infections. J. Sci. Food Agric. 90 : 630-634. 69 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Yang, L., Zhang, J.L., Bassett, C.L., and Meng, X.H. (2012). Difference between chitosan and oligochitosan in growth of Monilinia fructicola and control of brown rot in peach fruit. LWT – Food Sci. Technol. 46 : 254-259. Yen, M.-T., Yang, J.-H., and Mau, J.-L. 2008. Antioxidant properties of chitosan from crab shells. Carbohyd. Polym. 74 : 840-844 Yong-Cai, L., Xiao-Juan, S., Yang, B., Yong-Hong G., and Yi, W. (2009). Antifungal activity of Downloaded by [189.226.100.40] at 08:02 06 June 2015 chitosan on Fusarium sulphureum in relation to dry rot of potato tuber. Agr. Sci. China 8 : 597-604. Yu, T., Li, H.Y., and Zheng, X.D. (2007). Synergistic effect of chitosan and Cryptococcus laurentii on inhibition of Penicillium expansum infections. Int. J. Food Microbiol. 114 : 261-266. Yu, T., Yu, C., Chen, F., Sheng, K., Zhou, T., Zunun, M., Abudu, O., Yang, S., and Zheng, X. (2012). Integrated control of blue mold in pear fruit by combined application of chitosan, a biocontrol yeast and calcium chloride. Postharvest Biol. Technol. 69 : 49-53. Xiao-Juan, S., Yang, B., Yong-Cai, L., Rui-Feng, H., and Yong-Hong, G. (2008). Postharvest chitosan treatment induces resistance in potato against Fusarium sulphureum. Agr. Sci. China 7 : 615-621. Xing, Y., Li, X., Xu, Q., Yun, J., Lu, Y., and Tang, Y. (2011a). Effect of chitosan coating enriched with cinnamon oil on qualitative properties of sweet pepper (Capsicum annuum L.). Food Chem. 124 : 1443-1450. Xing, Y., Xu, Q., Che, Z., Li, X., and Li, W. (2011b). Effects of chitosan-oil coating on blue mold disease and quality attributes of jujube fruits. Food Funct. 2 : 466-474. 70 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Xu, J., Zhao, X., Wang, X., Zhao, Z., and Du, Y. (2007a). Oligochitosan inhibits Phytophtora capsici by penetrating the cell membrane and putative binding to intracellular targets. Pesticide Biochem. Physiol. 88 : 167-175. Xu, W.T., Huang, K.L., Guo, F., Qu, W., Yang, J.J., Liang, Z.H., and Luo, Y.B. (2007b). Postharvest grapefruit seed extract and chitosan treatments of table grapes to control Botrytis cinerea. Postharvest Biol. Technol. 46 : 86-94. Downloaded by [189.226.100.40] at 08:02 06 June 2015 Zahid, N., Ali, A., Manickam, S., Siddiqui, Y., and Maqbool, M. (2012). Potential of chtiosanloaded nanoemulsions to control different Colletotrichum spp. and maintain quality of tropical fruits during cold storage. J. Appl. Microbiol. 113 : 925-939. Zeng, K., Deng, Y., Ming, J., and Deng, L. (2010). Induction of disease resistance and ROS metabolism in navel oranges by chitosan. Sci. Hortic. 126 : 223-228. Zivanovic, S., Chi, S., and Draughon, A.F. (2005). Antimicrobial activity of chitosan films enriched with essential oils. J. Food Sci. 70 : 45-51. Zhang, D., and Quantick, P.C. (1997). Effect of chitosan coating on enzymatic browning and decay during postharvest storage of litchi (Litchi chinensis Sonn.) fruit. Postharvest Biol. Technol. 12 : 195-202. Zhang, D., and Quantick, P.C. (1998). Antifungal effects of chitosan coating on fresh strawberries and raspberries during storage. J. Hortic. Sci. Biotechnol. 73 : 763-767. Zheng, L.Y., and Zhu, J.F. (2003). Study on antimicrobial activity of chitosan with different molecular weights. Carbohyd. Polym. 54 : 527-530. 71 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Zhou, R., Mo, Y., Li, Y., Zhao, Y., Zhang, G., and Hu, Y. (2008). Quality and internal characteristics of Huanghua pear (Pyrus pyrifolia Nakai, cv. Huanghua) treated with different kinds of coatings during storage. Postharvest Biol. Technol. 49 : 171-179. Zhu, X., Wang, Q., Cao, J., and Jiang, W. (2008). Effects of chitosan coating on postharvest quality of mango (Mangifera indica L. cv. Tainong) fruits. J. Food Process. Pres. 32 : Downloaded by [189.226.100.40] at 08:02 06 June 2015 770-784. 72 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 1. Chitosan-based commercial products that are available for the control of postharvest diseases. Product trade name Chito Plant Downloaded by [189.226.100.40] at 08:02 06 June 2015 OIIYS ArmourZen Biorend FreshSeal Company (Country) Formulation a.i. (%) Fruit/vegetable Reference ChiPro GmbH (Bremen, Germany) Venture Innovations (Lafayette, LA, USA) Botry-Zen Limited (Dunedin, New Zealand) Bioagro S.A. (Chile) Powder 99.9 Liquid 5.8 Table grapes, sweet cherry, strawberry Table grapes Feliziani et al., 2013a; 2013b; Romanazzi et al., 2013 Feliziani et al., 2013a Liquid 14.4 Peach, table grapes Liquid 1.25 Liquid n.d. Clementine, mandarin fruit Banana Casals et al., 2012; Calvo-Garrido et al., 2013; Feliziani et al., 2013a Fornes et al., 2005 Powder 100 Rambutan fruit Martínez-Castellanos et al., 2009 Powder 100 Mango Jitarrerat et al., 2007 Liquid 2 Potato Kurzawińska and Mazur, 2007 BASF Corporation (Mount Olive, NJ, USA) ChitoClear Primex ehf (Siglufjordur, Iceland) Bioshield Seafresh (Bangkok, Thailand) Biochikol Gumitex 020 PC (Lowics, Poland) a.i., active ingredient 73 Baez-Sañudo et al., 2009 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Downloaded by [189.226.100.40] at 08:02 06 June 2015 Table 2. Chitosan treatments with other applications for storage decay of temperate fruit. Fruit Decay Table grape Gray mold Strawberry Raspberry Integration to chitosan - References (application time) Romanazzi et al., 2002 (pre- and postharvest) Acid solutions Romanazzi et al., 2009 (postharvest) Ethanol Romanazzi et al., 2007 (postharvest) Grape seed extract Xu et al., 2007b (postharvest) Gray mold and blue UV Romanazzi et al., 2006 (preharvest) mold Decay in general Cryptococcus Meng and Tian, 2009 (preharvest); laurentii 2010a (postharvest) Gray mold El Ghaouth et al., 1991a; 1992a (postharvest); Zhang and Quantick, 1998 (postharvest); Romanazzi et al., 2000 (pre and postharvest); Reddy et al., 2000a (preharvest); Mazaro et al., 2008 (preharvest) Lemon essential Perdones et al., 2012 (postharvest) oil Red thyme, Vu et al., 2011(postharvest) oregano extract, limonene, peppermint Rhizopus rot El Ghaouth et al., 1992a (postharvest); Zhang and Quantick, 1998 (postharvest); Romanazzi et al., 2000 (pre and postharvest); Park et al., 2005 (postharvest) Cladosporium rot Park et al., 2005 (postharvest) Decay in general Calcium lactate + Han et al., 2004 (postharvest) calcium gluconate, vitamin E Calcium gluconate Hernández-Muñoz et al., 2006 (postharvest); 2008 (postharvest) Oleic acid Vargas et al., 2006 (postharvest) Decay in general Calcium lactate, Han et al., 2004 (postharvest) calcium gluconate, vitamin E 74 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Fruit Decay Gray mold and Rhizopus rot Blueberry Apple Decay in general Blue mold Downloaded by [189.226.100.40] at 08:02 06 June 2015 Gray mold Pear Blue mold Peach Brown rot Sweet cherry Decay in general Orange Blue mold Tankan citrus fruit Clementine mandarin fruit Integration to chitosan - References (application time) UV-C, Candida satoiana, harpin Cryptococcus laurentii Candida satoiana Heat treatment Candida satoiana Heat treatment Calcium chloride + Cryptococcus laurentii Heat treatment Hypobaric treatment - Duan et al., 2011 (postharvest) De Capdeville et al., 2002 (postharvest) Yu et al., 2007 (postharvest) Zhang and Quantick, 1998 (postharvest) El Ghaout et al., 2000 (postharvest) Shao et al., 2012 (postharvest) El Ghaout et al., 2000 (postharvest) Shao et al., 2012 (postharvest) Yu et al., 2012 (postharvest) Li and Yu, 2001 (postharvest) Casals et al., 2012 (postharvest) Romanazzi et al., 2003 (pre- and postharvest) Romanazzi et al., 1999 (preharvest); Feliziani et al., 2013b (pre- and postharvest) Cháfer et al., 2012 (postharvest) Black spot disease Bergamot, thyme, tea tree essential oil - Decay in general - Canale Rappussi et al., 2009; 2011 (postharvest) Chien and Chou, 2006 (postharvest) Decay in general - Fornes et al., 2005 (pre- or postharvest) 75 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 3. Chitosan treatments with other applications for storage decay of tropical fruit. Fruit Decay Banana Anthracnose Mango Downloaded by [189.226.100.40] at 08:02 06 June 2015 Papaya Dragon fruit Litchi fruit Longan fruit Integration to chitosan Arabic gum References (application time) Zahid et al., 2012 (postharvest) Maqbool et al., 2010a; 2010b (postharvest) Crown rot Cinnamon extract Win et al., 2007 (postharvest) Anthracnose Zhu et al., 2008 (postharvest); Abd-Alla and Haggag, 2010 (postharvest) Irradiation Abbasi et al., 2009 (postharvest) Anthracnose Hewajulige et al., 2009 (postharvest); Ali et al., 2010 (postharvest); Zahid et al., 2012 (postharvest) Aqueous extract of Bautista-Baños et al., 2003 papaya seeds (postharvest) Ammonium carbonate, Sivakumar et al., 2005b sodium bicarbonate (postharvest) Anthracnose Zahid et al., 2012 (postharvest) Blue mold and Potassium Sivakumar et al., 2005a Cladosporium rot metabisulfite (postharvest) Decay in general Jiang and Li, 2001 (postharvest) 76 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 4. Chitosan treatments with other applications for storage decay of vegetables. Vegetable Decay Tomato Gray mold Gray mold and blue mold Blackmold Rot Decay in general Cinnamon oil Fusarium rot and alternaria rot Natamycin References (application time) El Ghaouth et al., 1992b (postharvest); Badawy and Rabea, 2009 (postharvest) Liu et al., 2007 (postharvest) Reddy et al., 2000b (postharvest) Xing et al., 2011a (postharvest) Cong et al., 2007 (postharvest) Downloaded by [189.226.100.40] at 08:02 06 June 2015 Sweet pepper Melon Integration to chitosan 77 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 5. Growth inhibition of chitosan on decay-causing fungi that affect the produce during Downloaded by [189.226.100.40] at 08:02 06 June 2015 storage. Fungus Alternaria alternata Alternaria kikuchiana Aspergillus phoenicus Aspergillus niger Botrydiplodia lecanidion Botrytis cinerea Infected species Tomato Reference Sánchez-Domínguez et al., 2011 Pear Meng et al., 2010a Pear Cè et al., 2012 Tankan citrus fruit Plascencia-Jatomea et al., 2003 Chien and Chou, 2006 Cladosporium spp. Colletotrichum gloeosporioides Litchi fruit, strawberry Mango, papaya Colletotrichum musae Colletotrichum spp. Fusarium solani Fusarium sulphureum Fusarium spp. Geotricum candidum Guignardia citricarpa Lasiodiplodia theobromae Monilinia fructicola Banana Tomato, potato, bell pepper fruit, cucumber, peach, strawberries, table grapes, pear, apple, Tankan citrus fruit Table grapes and tomato El Ghaouth et al., 1992a; 2000; Du et al., 1997; Romanazzi et al., 2002; Ben-Shalom et al., 2003; Ait Barka et al., 2004; Badawy et al., 2004; Chien and Chou, 2006; Lira-Saldivar et al., 2006; Elmer and Reglinski, 2006; Liu et al., 2007; Xu et al., 2007b; Badawy and Rabea, 2009; Rabea and Badawy, 2012 Park et al., 2005; Sivakumar et al., 2005a Bautista Baños et al., 2003; Sivakumar et al., 2005b; Jitareerat et al., 2007; Ali and Mahmud, 2008; Hewajulige et al., 2009; AbdAlla and Haggar, 2010; Ali et al, 2010; Zahid et al., 2012 Win et al., 2007; Maqbool et al., 2010a; 2010b; Zahid et al., 2012 Muñoz et al., 2009 Potato Eweis et al., 2006 Yong-Cai, et al., 2009 Banana Win et al., 2007 El-Mougy et al., 2012 Orange Canale Rappussi et al., 2009; 2011 Banana Win et al., 2007 Apple, peach Yang et al., 2010; 2012 78 ACCEPTED MANUSCRIPT Downloaded by [189.226.100.40] at 08:02 06 June 2015 ACCEPTED MANUSCRIPT Fungus Monilinia laxa Penicillium citrinum Penicillium digitatum Infected species Sweet cherry Jujube Reference Feliziani et al., 2013b Xing et al., 2011b Orange, lemon, Tankan citrus fruit Penicillium expansum Litchi fruit, strawberries, apple, pear, tomato Tankan citrus fruit El Ghaouth et al., 2000; Bautista Baños et al., 2004; Chien and Chou, 2006; El-Mougy et al., 2012 El Ghaouth et al., 2000; Sivakumar et al., 2005a; Liu et al., 2007; Yu et al., 2007 Penicillium italicum Penicillium stolonifer Phytophthora cactorum Physalospora piricola Rhizopus stolonifer Sclerotinia sclerotiorum Chien and Chou, 2006; El-Mougy et al., 2012 Pear Cè et al., 2012 Strawberries Eikemo et al., 2003 Pear Meng et al., 2010a Peach, strawberries, papaya, tomato El Ghaouth et al., 1992a; Bautista Baños et al., 2004; Park et al., 2005; Guerra-Sánchez et al., 2009; García-Rincón et al., 2010; HernándezLauzardo et al., 2010; Ramos-García et al., 2012 Cheah et al., 1997; Molloy et al., 2004; Ojaghian et al., 2013 Carrot 79 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 6. Physiological changes that can occur in temperate fruit after chitosan treatment. Fruit Physiological change Table grapes Phenylalanine ammonia-lyase Integration to chitosan - Peroxidase Cryptococcus laurentii - Downloaded by [189.226.100.40] at 08:02 06 June 2015 Polyphenol oxidase, superoxide dismutase Cryptococcus laurentii Putrescine Bergamot oil Chitinase, myricetin Quercetin Respiration Trans-resveratrol UV Bergamot oil Soluble solids content Cryptococcus laurentii Glucose Cryptococcus laurentii Putrescine Bergamot oil Titratrable acidity Total phenolic content Weight loss, color, texture Putrescine Grape seed extract Glucose Putrescine Grape seed extract - Shattering and cracking Strawberries Titratable acidity 80 References Romanazzi et al., 2002; Meng et al., 2008 Meng and Tian, 2009; Meng et al., 2010b Meng et al., 2008; Gao et al., 2013 Meng et al., 2008; Gao et al., 2013 Meng and Tian, 2009; Meng et al., 2010b Feliziani et al., 2013a Feliziani et al., 2013a Shiri et al., 2012 Sánchez-González et al., 2011 Romanazzi et al., 2006 Feliziani et al., 2013a Meng et al., 2008 Sánchez-González et al., 2011 Meng et al., 2010b Gao et al., 2013 Meng et al., 2008 Meng et al., 2008 Meng et al., 2010b Shiri et al., 2011 Sánchez-González et al., 2011 Shiri et al., 2012 Xu et al., 2007b Gao et al., 2013 Shiri et al., 2012 Xu et al., 2007b El Ghaouth et al., 1991a; Zhang and Quantick, 1998; Reddy et al., 2000a ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Downloaded by [189.226.100.40] at 08:02 06 June 2015 Fruit Physiological change Integration to chitosan Vitamin E Calcium gluconate pH Calcium gluconate Antocyanin content Vitamin E - Total polyphenol Soluble solids content Colour Oleic acid Vitamin E Calcium gluconate Firmness Vitamin E Calcium gluconate - Vitamin C content - Glutathion Chitinase - β-1,3 glucanase - Phenilalanine ammonia-lyase - Weight loss Respiration Vitamin E - Chalcone isomerase, flavonol synthase, anthocyanidin synthase, calcium-dependent protein kinase, potassium - 81 References Han et al., 2004; 2005 Hernández- Muñoz et al., 2008 Hernández-Muñoz et al., 2008 Han et al., 2004 El Ghaouth et al., 1991a; Zhang and Quantick, 1998; Reddy et al., 2000a Vargas et al., 2006 Kerch et al., 2011 Han et al., 2005 Hernández-Muñoz et al., 2008 Han et al., 2004; 2005 Hernández-Muñoz et al., 2008 El Ghaouth et al., 1991a Zhang and Quantick, 1998; Kerch et al., 2011; Wang and Gao, 2012 Wang and Gao, 2012 Zhang and Quantick, 1998; Landi et al., 2014 Zhang and Quantick, 1998; Landi et al., 2014 Romanazzi et al., 2000; Landi et al., 2014 Han et al., 2004 El Ghaouth et al., 1991a; Vargas et al., 2006 Landi et al., 2014 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Fruit Downloaded by [189.226.100.40] at 08:02 06 June 2015 Raspberry Apple Pear Apricot Peach Sweet cherry Physiological change channel, PR-1, polygalacturonase, polygalacturonase inhibiting protein Catalase, glutathione-peroxidase, guaiacol peroxidase, dehydroascorbate reductase, monodehydroascorbate reductase Weigth loss, color, pH, titratable acidity Ascorbic acid, titratable acidity, firmness, antocyanin content Respiration, firmness, weicht loss, titratable acidity Polyphenol oxidase, chitinase, β1,3 glucanase, ROS, catalase, superoxide dismutase, ascorbate peroxidase, glutathione reductase Peroxidase Respiration, permeability of cell membrane, weight loss Soluble solid contents, titratable acidity, firmness Total phenolic content, antioxidant activity, weight loss Titratable acidity, ascorbic acid, respiration, firmness, ethylene and malondialdehyde production, superoxide dismutase Polyphenol oxidase, peroxidase, ascorbic acid oxidase, polygalacturonase, vitamic C Integration to chitosan References - Wang and Gao, 2012 Vitamin E Han et al., 2004 Heat Zhang and Quantick, 1998 Shao et al., 2012 - Meng et al., 2010b Li et al., 2010a Ascorbic acid Ascorbic acid Titratable acidity, soluble solid, catalase, peroxidase, 82 Meng et al., 2010b; Li et al., 2010a Zhou et al., 2008 Lin et al., 2008 Lin et al., 2008 - Ghasemnezhad et al., 2010 - Li and Yu, 2001 CaCl2 coating + PEpackage + intermittent warming - Ruoyi et al., 2005 Dang et al., 2010 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Fruit Physiological change polyphenol oxidase, phenilalanine ammonia-lyase, respiration Ascorbic acid Orange Phenols content, antocyanin content Water loss, firmness Downloaded by [189.226.100.40] at 08:02 06 June 2015 References - Dang et al., 2010; Kerch et al., 2011 Kerch et al., 2011 Bergamot, thyme, tea tree essential oil - Color Tankan citrus fruit Jujube Integration to chitosan Cháfer et al., 2012 Chitinase, b-1,3-glucanase, polyphenol oxidase Peroxidase - Superoxide dismutase, catalase, ascorbate peroxidase, glutathione reductase, hydrogen peroxide content, ascorbate content Firmness, weight loss, titratable acidity, ascorbic acid, soluble solids Polyphenol oxidase, phenolic compounds Ascorbic acid - Canale Rappussi et al., 2011 Canale Rappussi et al., 2009 Canale Rappussi et al., 2009; Zeng et al., 2010 Zeng et al., 2010 - Chien and Chou, 2006 Zinc, cerium 1methylcycloprope ne 1methylcycloprope ne 1methylcycloprope ne Zinc, cerium Zinc, cerium Xing et al., 2011b Wu et al., 2010 Xing et al., 2011b Qiuping and Wenshui, 2007 - Firmness Weight loss Respiration, soluble solids 83 Qiuping and Wenshui, 2007 Qiuping and Wenshui, 2007 Wu et al., 2010 Wu et al., 2010 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 7. Physiological changes that can occur in tropical fruit after chitosan treatment. Fruit Physiological changes Banana Titratable acidity Respiration Downloaded by [189.226.100.40] at 08:02 06 June 2015 Firmness, soluble solids content Integration to chitosan 1-methylcyclopropene Arabic gum 1-methylcyclopropene Arabic gum 1-methylcyclopropene Arabic gum Longan fruit Mango Papaya Color change - Weight loss 1-methylcyclopropene Arabic gum Arabic gum Respiration, weight loss, color change, polyphenol oxidase, titratable acidity, total soluble solids, ascorbic acid Titratable acidity, weight loss Total soluble solids, firmness, color change pH Chitinase, b-1,3glucanase Respiration, ascorbic acid Titratable acidity, total soluble solids Ascorbic acid Weight loss, color - - References Kittur et al., 2001 Baez-Sañudo et al., 2009 Maqbool et al., 2010a, 2010b Kittur et al., 2001 Baez-Sañudo et al., 2009 Maqbool et al., 2011 Kittur et al., 2001; Win et al., 2007 Baez-Sañudo et al., 2009 Maqbool et al., 2010a; 2010b; 2011 Kittur et al., 2001; Win et al., 2007 Baez-Sañudo et al., 2009 Maqbool et al., 2011 Maqbool et al., 2010a; 2010b; 2011 Jiang and Li, 2001 Hydrothermal process Hydrothermal process Jitareerat et al., 2007; Zhu et al., 2008 Salvador-Figueroa et al., 2011 Zhu et al., 2008 Salvador-Figueroa et al., 2011 Hydrothermal process - Salvador-Figueroa et al., 2011 Jitareerat et al., 2007 - Jitareerat et al., 2007; Zhu et al., 2008 Ali et al., 2010; 2011 Al Eryani et al., 2008 Ali et al., 2011 Al Eryani et al., 2008 Ali et al., 2011 Calcium infiltration Calcium infiltration - 84 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Fruit Physiological changes change Firmness Downloaded by [189.226.100.40] at 08:02 06 June 2015 Chitinase, b-1,3glucanase Respiration Litchi fruit Weight loss Integration to chitosan Ammonium carbonate, sodium bicarbonate Calcium infiltration Ammonium carbonate, sodium bicarbonate Aqueous extract of papaya seeds - - Organic acids Titratable acidity Organic acids Total phenolic content, flavonoid content Anthocyanin content Respiration rate Color change - Modified atmosphere packaging Organic acids Total soluble solid Peroxidase Ascorbic acid Modified atmosphere packaging Ascorbic acid - 85 References Sivakumar et al., 2005b Al Eryani et al., 2008 Ali et al., 2010; 2011 Sivakumar et al., 2005b Bautista-Baños et al., 2003 Hewajulige et al., 2009 Hewajulige et al., 2009, Ali et al., 2011 Zhang and Quantick, 1997; Jiang and Li, 2001; Sivakumar et al., 2005a; Sun et al., 2010; Lin et al., 2011 Joas et al., 2005; Caro and Joas, 2005 Jiang et al., 2005; Sivakumar et al., 2005a; Sun et al., 2010 Joas et al., 2005; Caro and Joas, 2005 Zhang and Quantick, 1997; Sivakumar et al., 2005a Zhang and Quantick, 1997; Jiang et al., 2005; Sivakumar et al., 2005a; De Reuck et al., 2009 Lin et al., 2011 Zhang and Quantick, 1997; Ducamp-Collin et al., 2008 Caro and Joas, 2005; Joas et al., 2005 Sun et al., 2010 De Reuck et al., 2009 Jiang et al., 2005 Sun et al., 2010 Zhang and Quantick, 1997; Dong et al., 2004 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Fruit Physiological changes Polyphenol oxidase Integration to chitosan Ascorbic acid Modified atmosphere packaging - Downloaded by [189.226.100.40] at 08:02 06 June 2015 Ascorbic acid Modified atmosphere packaging Ascorbic acid Super oxide dismutase, catalase, hydrogen peroxide, malondialdeyde; ascorbic acid content Rambutan Firmness, soluble solid, Lactobacillus titratable acidity plantatum Guava Firmness, peroxidase superoxide dismutase, catalase, inhibition of superoxide free radical production, titratable acidity, ascorbic acid, weight loss, soluble solids, chlorophyll and malondialdehyde content 86 References Sun et al., 2010 De Reuck et al., 2009 Zhang and Quantick, 1997; Jiang et al., 2005; Lin et al., 2011 Sun et al., 2010 De Reuck et al., 2009 Sun et al., 2010 Martínez-Castellanos et al., 2009 Hong et al., 2012 ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Downloaded by [189.226.100.40] at 08:02 06 June 2015 Table 8. Physiological changes that can occur in vegetables after chitosan treatment. Vegetables Physiological changes Tomato Respiration rate, color change, ethylene production, firmness, titratable acidity Polyphenol oxidase, phenolic content Peroxidase Protein content Polygalacturonase, pectate lyase, cellulose, phytoalexin production, pH Peroxidase, polyphenol oxidase, flavonoid content, lignin content Phenylalanine ammonialyase Superoxide dismutase, peroxidase, catalase Respiration, weight loss, color Respiration, weight loss, color Weigth loss, ascorbic acid, pH Polyphenol oxidase, peroxidase, phenylalanine ammonia-layse Potato Sweet pepper Cucumber Melon Carrot Integration to chitosan - References - Liu et al., 2007; Badawy and Rabea, 2009 Liu et al., 2007 Badawy and Rabea, 2009 Reddy et al., 2000b - Xiao-Juan et al., 2008 - Gerasimova et al., 2005 Cinnamon oil Xing et al., 2011a - El Ghaouth et al., 1991b - El Ghaouth et al., 1991b Natamycin Cong et al., 2007 - Ojaghian et al., 2013 87 El Ghaouth et al., 1992b ACCEPTED MANUSCRIPT ACCEPTED MANUSCRIPT Table 9. Application of chitosan on fruit and vegetable to control foodborne microorganisms. Microorganism Escherichia coli Whole cantaloupe Integration to chitosan References Beeswax + lime essential oil Allyl isothiocyanate, nisin Inatsu et al., 2010 Ramos-García et al., 2012 Chen et al., 2012 Downloaded by [189.226.100.40] at 08:02 06 June 2015 Salmonella spp. Substrate of growth Tomato Tomato 88 ACCEPTED MANUSCRIPT