Control of table grape storage rots by pre-harvest applications of salts

Postharvest Biology and Technology 42(2006)142–149Control of table grape storage rots by pre-harvest applications of saltsFranco Nigro ∗,Leonardo Schena,Angela Ligorio,Isabella Pentimone,Antonio Ippolito,Mario G.SalernoDipartimento di Protezione delle Piante e Microbiologia Applicata,Universit`a degli Studi di Bari,Via Amendola 165/A,70126Bari,ItalyReceived 30November 2005;accepted 11June 2006AbstractThe activity of 19inorganic and organic salts to control table grape storage rots was preliminarily assessed by in vitro and in vivo tests.Several salts reduced the growth of Botrytis cinerea Pers.on amended (0.1–2%,w/v)glucose–agar;however,only calcium chloride (CC),potassium carbonate (PC),sodium bicarbonate (SB)and sodium carbonate (SC)significantly reduced the incidence of grey mould on small table grape bunches (cv.Italia).PC,SB and SC showed a similar effect in the in vitro (inhibition of mycelial growth and conidia germination of B.cinerea )and in vivo tests (reduction of incidence of grey mould on table grape),whereas CC was effective only in vivo .Efficacy of salts applied before harvest was extensively evaluated in small-scale tests (CC,PC,SB,and SC),and in large-scale tests (CC,SB,SC).In both,small-and large-scale tests,field applications of salts resulted in a significant reduction of botrytis storage rots.Field rots (mainly sour rot)were significantly reduced when vines were sprayed at least 21days before harvest.In large-scale tests,simulating the practical commercial conditions in Southern Italy,two salt applications (30and 90days before harvest)of CC,SC,or SB significantly reduced postharvest grey mould from 63.8%among untreated controls to 22.5,31.2,and 29.5%,respectively.In the same conditions,field rots were 23.4%(untreated control),9.5%(CC),11.9%(SC)and 12.0%(SB).On the whole,salts showed an activity higher or similar to that of conventional chemical treatments.Finally,results from specific tests suggested that pH and inhibition of polygalacturonase activity of B.cinerea seem to play a role in the mode of action of SB,SC,PC and CC.©2006Elsevier B.V .All rights reserved.Keywords:Botrytis cinerea ;Table grape;Inorganic salts;Grey mould;Sour rot;Control1.IntroductionGrey mould,due to Botrytis cinerea Pers.,is one of the most important diseases of table grapes,because of the dam-age caused in the harvest season and during storage.Losses are severe,particularly in years when heavy rainfall dur-ing fruit ripening occurs.The pathogen can also develop at low temperature,shortening the duration of storage and marketing.Sour rot,a grape disease with an ambiguous and controversial aetiology (Guerzoni and Marchetti,1987),causes losses that exceed grey mould in Southern Italy.The disease,associated with Acetobacter spp.,and other bacte-ria and yeasts transmitted by the grape fruitfly (Drosophila melanogaster Mg.),is a serious problem in Apulia (South-∗Corresponding author.Tel.:+390805443055;fax:+390805442911.E-mail address:nigrof@agr.uniba.it (F.Nigro).ern Italy)where table grapes bunches are left on vines under plastic coverings to delay the harvest.The control of botry-tis storage rot and sour rot is very difficult since postharvest treatments with synthetic fungicides or food additives are not allowed by European legislation (European Parliament and Council Directive 95/2/EC,1995).Pre-harvest fungi-cide applications are not always effective to control botrytis storage rot and ineffective against sour rot.Furthermore,the use of conventional chemicals is becoming increasingly restricted because of concerns for the environment and human health,as well as the cost of developing new fungicides to overcome resistance developed by the pathogens.Therefore,there is a need for alternative disease-management practices that can reduce rots without risks for consumers and work-ers.Many alternative means have been proposed to manage postharvest disease of fruit and vegetables.These include biocontrol agents (Janisiewicz and Korsten,2002),natural0925-5214/$–see front matter ©2006Elsevier B.V .All rights reserved.doi:10.1016/j.postharvbio.2006.06.005F.Nigro et al./Postharvest Biology and Technology42(2006)142–149143substances(Ippolito and Nigro,2003),and physical treat-ments(Nigro et al.,1998).In the search for biocompatible products,which could be defined as chemicals exhibiting low mammalian and envi-ronmental toxicity,many salts have been recently tested as alternative control means,either alone(Smilanick et al., 1999)or in combination with physical(Palou et al.,2001)and biological treatments(Ippolito et al.,2005;Karabulut et al., 2005).Salts are inexpensive,easily accepted by consumers, non-toxic,with minor environmental impact at the effective concentrations,and usually used in the food industry.Several inorganic salts have been shown to be active antimicrobial agents against a range of phytopathogenic fungi.In particular,postharvest treatments with calcium chlo-ride(CC)and sodium bicarbonate(SB)have been proposed as safe and effective alternative means to control posthar-vest rots of fruit and vegetables.The activity of bicarbonate salts against fungal pathogens is well known(Hervieux et al.,2002)and recent postharvest applications reduced stor-age rots of melon(Aharoni et al.,1997),papaya(Sivakumar et al.,2002),citrus(Palou et al.,2002;Smilanick et al., 2005),banana(Alvindia et al.,2004),table grape(Mlikota Gabler and Smilanick,2001),and bell pepper(Fallik et al.,1997).Calcium,known for its ability to reduce or delay parasitic and/or physiological disorders in fruit and vegetables,also gave promising results in controlling stor-age rots when applied both as organic and inorganic salts (Punja and Grogan,1982;Biggs,1999;Conway et al., 1999).Most of the research regarding the use of carbonates and calcium salts to control postharvest rots has involved posthar-vest applications;however,they are inappropriate for fruit such as strawberries and table grapes since the cosmetic appeal of these fruit can be seriously reduced by posthar-vest handling(Ippolito and Nigro,2000).There are only few reports about pre-harvest applications of salts to control stor-age rots.Application of SB24h before harvest significantly reduced the incidence of botrytis storage rot on table grape (Karabulut et al.,2003).Field applications of CC increased the storage life of kiwifruit(Gerasopoulos et al.,1996)and reduced the incidence of russet on‘Golden Delicious’apples (Brown et al.,1996)and various rots on cactus pear(Schirra et al.,1999).Field applications of SB were effective for con-trolling Monilinia fructicola on organic apricots(McLaren and Fraser,2000).Similarly,pre-and postharvest applica-tion of CC on table grapes reduced the incidence of grey mould storage rot(Ippolito et al.,1997;Miceli et al.,1999).The present work was performed to evaluate the effective-ness of a range of salts,mostly common food preservatives, for the control of pre and postharvest rots of table grapes.In this regard,we conducted small-and large-scale trials over a4-year period with different pre-harvest application sched-ules of these salts to obtain the most effective postharvest decay control efficacy.Investigation of some possible mech-anisms by which the most effective salts reduce rot incidence was also carried out.2.Materials and methods2.1.In vitro and in vivo screening of saltsThe effect of19salts(Table1)on the mycelial growth of B. cinerea was evaluated using1%glucose agar(GA).An aque-ous solution of the salts was sterilised byfiltration(0.45␮m) and added to molten(45◦C)GA,to achieve0.1,0.25,0.5,1, and2%(w/v)final concentration;GA without salts served as a control.GA-salt solutions,poured into100mm Petri dishes, were inoculated in the centre with a5mm mycelial plug taken from the edge of actively growing colony of B.cinerea and incubated for5days at22◦C.Colony diameter was mea-sured as the average of the longest and the shortest diameter; the results were expressed as minimum inhibitory concentra-tion(MIC).Salts(Table2)were also tested for their activity against B.cinerea on table grape,cv.Italia,harvested at com-mercial maturity from groves located at Conversano,and Rutigliano(South-eastern Italy).Small bunches,of approxi-mately10berries,were sprayed with an aqueous salt solution (1%,w/v)or sterile distilled water(control)until dripping and left to dry.Before application the pH of all salt solu-tions was measured.The bunches were then sprayed with a conidial suspension of B.cinerea,prepared by removing the spores from a sporulating edge of the3-week-old culture with a sterile bacteriological loop and suspending them in sterile distilled water.The suspension wasfiltered through three sterile cheesecloth layers and spore concentration was adjusted to5×105mL−1using a haemocytometer.Treated bunches were arranged in plastic trays,covered with plas-tic sheet,and stored at0◦C(95–98%RH)for22days.The percentage of rotted berries was evaluated after7days shelf Table1Minimum inhibitory concentration(MIC)of salts for Botrytis cinerea in a colony growth assaySalt MIC(%,w/v) Sodium chloride(NaCl)>2.0 Sodium formate(HCOONa)>2.0 Calcium nitrate(Ca(NO3)2·4H2O)>2.0Sodium phosphate monobasic(NaH2PO4·H2O)>2.0 Sodium nitrate(NaNO3)>2.0 Calcium chloride(CaCl2)>2.0 Ammonium chloride(NH4Cl)>2.0 Sodium acetate(CH3COONa·3H2O)>2.0 Potassium chloride(KCl)>2.0 Potassium phosphate dibasic(K2HPO4)>2.0 Ammonium sulphate((NH4)2SO4)>2.0 Sodium phosphate dibasic(Na2HPO4·12H2O) 2.0 Sodium sulphate(Na2SO4)0.5 Ammonium phosphate dibasic((NH4)2HPO4)0.5 Sodium bicarbonate(NaHCO3)0.5 Sodium silicate(Na2Ox2SiO2·2H2O)0.5 Potassium carbonate(K2CO3)0.5 Sodium carbonate(Na2CO3)0.25 Ammonium bicarbonate(NH4HCO3)0.25 Diameter of B.cinerea colonies was determined after5days incubation at 22◦C on glucose agar amended with different salt concentrations(0.1,0.25, 0.5,1.0,and2%,w/v).144F.Nigro et al./Postharvest Biology and Technology42(2006)142–149Table2pH values and effect of19aqueous salt solutions(1%,w/v)on the devel-opment of rots on small table grape bunches artificially inoculated with B. cinereaSalts pH Rottedberries(%) Sodium phosphate dibasic(Na2HPO4·12H2O)8.497.0a Ammonium chloride(NH4Cl) 5.580.0b Control(distilled water) 5.778.0bc Sodium nitrate(NaNO3) 5.978.0bc Ammonium sulphate((NH4)2SO4) 5.677.0bc Sodium sulphate(Na2SO4) 5.977.0bc Sodium chloride(NaCl) 5.976.0bc Sodium acetate(CH3COONa·3H2O)7.673.0bc Sodium formate(HCOONa)7.173.0bc Ammonium phosphate dibasic((NH4)2HPO4)8.172.0bc Monobasic sodium phosphate(NaH2PO4·H2O) 4.371.0bc Ammonium bicarbonate(NH4HCO3)8.164.0bcd Potassium chloride(KCl) 5.963.0bcd Calcium nitrate(Ca(NO3)2·4H2O) 5.762.0bcd Potassium phosphate dibasic(K2HPO4)9.062.0bcd Sodium silicate(Na2Ox2SiO2·2H2O)11.660.0cd Calcium chloride(CaCl2) 6.048.0de Sodium bicarbonate(NaHCO3)8.545.0de Sodium carbonate(Na2CO3)11.341.0e Potassium carbonate(K2CO3)11.336.0e Values marked with different letters are statistically different according to Fisher’s protected LSD test(P≤0.05).life at20±1◦C and90–95%RH.In both screening tests a completely randomized experimental design,includingfive replicates(Petri dishes or small bunches),was utilised.An arcsine transformation was applied to data percentage prior to analysis of variance(ANOV A)and the mean values were compared by using Fisher’s protected LSD test.2.2.Small-and large-scale trialsField trials were conducted over a4-year period in com-mercial orchards of table grapes(cv.Italia)located in South-eastern Italy,involving pre-harvest applications of salts and conventional fungicides(chemical controls)as described in Table3.In particular,aqueous salt solutions(1%,w/v)were sprayed twice(21and5days before harvest)in thefirst3 years of the trials;a treatment receiving only one salts appli-cation(5days before harvest)was also included in the second year.For each year,the spraying time was in the last week of October or thefirst week of November,depending on the harvest date.According to the control schedule used in the commercial grove which hosted the trials,in the fourth year salts were sprayed90days(end of August)and30days(end of October)before harvest,in order to test their effective-ness under the practical control conditions commonly used in Southern Italy.A motor-driven back sprayer(delivering approximately10hL ha−1)and a hydro-pneumatic sprayer (delivering approximately7hL ha−1)were used for small-and large-scale tests,respectively.Harvested bunches were placed in plastic crates,covered with plastic sheet to maintain high relative humidity(95–98%)and stored.The incidence Table3Schematic representation of small-and large-scalefield testsYear Treatments Number ofapplicationsTiming a FirstUntreatedCC221and5SCSBPCProcymidone121 Second bUntreatedCC221and5SC15SBPCProcymidone121Large-scale testsThirdUntreatedCC221and5SCSBCiprodinil andfludioxonil121FourthUntreatedCC290and30SCSBCiprodinil andfludioxonilTrials provided pre-harvest sprays with1%(w/v)aqueous solution of cal-cium chloride(CC),sodium bicarbonate(SB),sodium carbonate(SC),and potassium carbonate(PC).Untreated table grapes bunches and bunches treated with procymidone(Sumisclex®,50%a.i.)at0.75g L−1or a mix-ture of ciprodinil andfludioxonil(Switch®,37.5and25%a.i.,respectively) at0.8g L−1,were used as controls.a Days before harvest.b For each salt,treatments with two applications(21and5days before harvest)and one application(5days before harvest)were performed.of naturally occurring decay(sour rot,grey mould,and sec-ondary rots)was evaluated on two different sets of bunches,in thefield(5days before harvest)and after storage at0±1◦C (1month)and shelf life(5–7days)at20±1◦C,90–95%RH. Decay assessment was done by using an empirical scale with the following values:(0)bunch without rots;(1)1–5%of rot-ted berries;(2)6–10%of rotted berries;(3)11–25%of rotted berries;(4)26–50%of rotted berries;(5)51–75%of rotted berries;(6)more than76%of rotted berries.This empiri-cal scale made it possible to calculate the McKinney index (McKinney,1923),expressing the weighted average of the disease severity as actual percentage in terms of the maximum disease severity.The McKinney index(MI)was calculated by means of the following formula:MI=[df/T n D]×100, where d is the degree of disease severity assessed on the bunch,f its frequency,T n the total number of the fruit exam-ined(healthy and diseased),and D the highest degree of disease intensity occurring on the empirical scale.All trials were arranged in a completely randomised block design with four replicates.In small-scale tests each repli-cation contained three grape vines,from which15bunchesF.Nigro et al./Postharvest Biology and Technology42(2006)142–149145were harvested;in large-scale tests each replication con-tained50grape vines,from which30bunches were randomly harvested.An arcsine transformation was applied to data per-centage prior to analysis of variance(ANOV A),and the mean values of treatments compared by using Fisher’s protected LSD test.2.3.Effect of salts and pH on the germination of B. cinerea conidiaThe ability of B.cinerea conidia to germinate was eval-uated in50mL of liquid medium containing2.4%potato dextrose broth(PDB)supplemented with1%of calcium chlo-ride(CC),potassium carbonate(PC),sodium bicarbonate (SB),or sodium carbonate(SC).PDB medium was utilised without modifying the pH determined by each salt or adjust-ing its value to pH6–10by means of NaOH,KOH or HCl. An equal set of PDB medium without salts was used as con-trol.Diluted conidial suspension of B.cinerea was added to the medium at afinal concentration of104cells mL−1. The percentage of germinated conidia(length/width ratio≥2) was evaluated after6and24h incubation on a rotary shaker (100rpm),at21◦C,in the dark.For each salt type,pH value and reading date,three separate replications of100conidia were evaluated.2.4.Effect of salts on polygalacturonase activity of B. cinereaTo evaluate the effect of salts on polygalacturonase activity of B.cinerea,the pathogen was grown in a minimal salt(MS) medium(KH2PO4,13.6g/L;NH4NO3,4g;MgSO4·7H2O, 2.6g/L;FeSO4·7H2O,20mg/L;CuSO4·5H2O,0.07mg/L;ZnSO4·7H2O,0.23mg/L;(NH4)2MoO4,0.18mg/L; MnSO4·4H2O,2mg/L;H3BO4, 3.1mg/L;pectin,5g/L; sodium polypectate,5g/L)containing CC,PC,SB,or SC at different concentrations(10,20,and40mM).MS medium without salts was used as a control.Two agar plugs (9mm in diameter)taken from the edge of actively growing mycelium of B.cinerea were used to inoculate50mL of medium.After7days incubation at25◦C on a rotary shaker at120rpm,the media were collected byfiltration and the concentration of polygalacturonase(PG)assessed measuring the concentration of reducing sugars using the 2-cyanoacetamide method described by Gross(1982).Three replicate samples were utilised for each salt and for each concentration.3.Results3.1.Preliminary screening of saltsThe in vitro tests showed a variable effect of salts on B. cinerea growth(Table1).After5days incubation at22◦C most salts reduced the colony diameter of the pathogen.A complete inhibition was achieved by ammonium bicarbon-ate and SC at0.25%and by ammonium phosphate,PC,SB, sodium silicate,and sodium sulphate at0.5%.CC did not affect the growth of B.cinerea,whereas it was enhanced on GA amended with calcium nitrate,sodium chloride,and sodium nitrate,as compared to the control(Table1).In the in vivo tests on small table grape bunches,salts showed a variable activity against B.cinerea(Table2).Aque-ous solutions(1%,w/v)of CC,SB,SC,and PC were the most effective,reducing the percentage of rotted berries by38.5, 42.3,47.4,and53.9,respectively,as compared to the water control.None of the salt solutions had a phytotoxic effect on the berries.The pH of salt solutions did not show any signif-icant correlation with the control activity(data not shown). High disease reductions were obtained with SC and PC for which the pH was11.3;however,a significant disease reduc-tion was obtained also by using CC,with a pH of6.00.3.2.Small-scale trialsIn small-scale trials conducted in thefirst year,CC,PC, SB,and SC reduced the incidence of total rots on table grape bunches treated21and5days before harvest(Fig.1).In thefield assessment(5days before harvest)all the salts sig-nificantly(P≤0.05)reduced rot incidence as compared to the untreated control,with reduction ranging from46.1to 64.6%(Fig.1).A significant(P≤0.05)rot reduction was also achieved after30days of storage followed by5days of shelf life at20◦C(Fig.1).CC,PC,SB,and SC reduced rot inci-dence from57.6to71.4%as compared to the untreatedcon-Fig.1.Small-scale tests conducted in thefirst year investigation.Rot inci-dence among table grape bunches treated in thefield21and5days before harvest with1%(w/v)aqueous solution of calcium chloride(CC),potas-sium carbonate(PC),sodium bicarbonate(SB),or sodium carbonate(SC). Untreated bunches and bunches treated with procymidone(Sumisclex®,50% a.i.)at0.75g L−1were used as controls.Rots were assessed in thefield5 days before harvest(A)and after30days storage at0◦C followed by5 days shelf life at20±1◦C(B).For each assessment,columns marked with the same letters are not statistically different according to Fisher’s protected LSD test(P≤0.05).146F.Nigro et al./Postharvest Biology and Technology42(2006)142–149Fig.2.Small-scale tests conducted in the second year investigation.Inci-dence of postharvest rots among table grapes bunches treated in thefield once(5days before harvest—A)or twice(21and5days before harvest—B) with calcium chloride(CC),potassium carbonate(PC),sodium bicarbonate (SB),or sodium carbonate(SC).Untreated bunches and bunches treated with procymidone(Sumisclex®,50%a.i.)at0.75g L−1were used as controls. Rots were assessed after30days storage at0◦C followed by5days shelf life at20±1◦C.Values followed by the same letters are not statistically different according to Fisher’s protected LSD test(P≤0.05).trol;moreover,SB was significantly(P≤0.05)more effective than the chemical control.In the second year of trials,all salts reduced storage rots as compared to the untreated control(Fig.2).Two salt appli-cations(5and21days before harvest)allowed higher levels of protection compared to a single salt spray(5days before harvest),although the values were not statistically different. In particular,on the bunches treated21and5days before har-vest,storage rots were reduced by54.8%(CC),50.1%(PC), 50.7%(SB),and50.6%(SC),as compared to the untreated control(Fig.2).In both years of trials,field rots were mainly ascribed to sour rot whereas storage rots were almost exclusively caused by B.cinerea(data not shown).rge-scale trialsIn large-scale trials conducted in the third year of trials, salts applications21and5days before harvest significantly reduced storage rots(Fig.3),but were ineffective in reducing the rot incidence in thefield(data not shown).After1-month storage at0◦C and5days at20±1◦C,storage rot incidence was reduced by49.3%(CC),53.6%(SC),and37.2%(SB)as compared to the untreated control.Chemical control was the most effective treatment in reducing storage rots,although not statistically different from SC(Fig.3).In the fourth year trials,the application of salts90and 30days before harvest resulted in a significant reduction of rots compared to untreated controls(Fig.4).In thefield assessment the three tested salts reduced sour rot incidence by 59.4%(CC),49.2%(SC),and48.6%(SB),whereas the chem-ical control was not statistically different from theuntreated rge-scale trials conducted in the third year investigation.Stor-age rot incidence on table grapes bunches treated in thefield21and5 days before harvest with calcium chloride(CC),potassium carbonate(PC), sodium bicarbonate(SB),or sodium carbonate(SC).Untreated bunches and bunches treated with a mixture of ciprodinil andfludioxonil(Switch®,37.5 and25%a.i.,respectively)at0.8g L−1were used as controls.Rot incidence was determined after30days storage at0◦C followed by7days shelf life at 20±1◦C.Columns marked with the same letter are not statistically different according to Fisher’s protected LSD test(P≤0.05).rge-scale trials conducted in the fourth year investigation.Rot inci-dence among table grape bunches treated in thefield30and90days before harvest with calcium chloride(CC),potassium carbonate(PC),sodium bicar-bonate(SB),or sodium carbonate(SC).Untreated bunches and bunches treated with a mixture of ciprodinil andfludioxonil(Switch®,37.5and25% a.i.,respectively)at0.8g L−1were used as controls.Rots were assessed in thefield(A)and after30days storage at0◦C followed by6days shelf life at20±1◦C(B).For each assessment,columns marked with the same let-ters are not statistically different according to Fisher’s protected LSD test (P≤0.05).F.Nigro et al./Postharvest Biology and Technology42(2006)142–149147Fig.5.Effect of different salt concentrations on the polygalacturonase activ-ity of :calcium chloride;PC:potassium carbonate;SB: sodium bicarbonate;SC:sodium carbonate.Bars represent the standard error (n=3).control.After1-month storage at0◦C and5days shelf life, storage rots were reduced by64.7%(CC),51.1%(SC),53.8% (SB),and40.4%(chemical control),as compared to the untreated control(Fig.4).In both years of trialsfield rots were mainly ascribed to sour rot,whereas storage rots were almost exclusively caused by B.cinerea,the incidence of secondary rots(due to Alternaria,Aspergillus,Cladosporium,Mucor,Penicillium, and Rhizopus spp.)being negligible(data not shown).3.4.Effect of salts and pH on conidial germination and polygalacturonase activity of B.cinereaThe pH had a variable influence on germination of B. cinerea conidia for the different salts.After6h incubation,a concentration of1%of PC,SB,and SC prevented germina-tion of B.cinerea spores in PDB from pH6to10and at the native pH of each salt solution.Germination of the spores in 1%CC occurred from pH6to10and was similar to that of the control.CC,PC,SB,and SC significantly inhibited polygalac-turonase activity of B.cinerea at the tested concentrations (Fig.5).In particular,CC was the most effective at all the tested concentrations,whereas SB resulted in an increasing reduction with increasing concentrations(Fig.5).4.DiscussionThe objective of the present study was selection of salts effective in controlling sour rot and botrytis storage rot of table grapes by means of pre-harvest applications.In prelim-inary screenings,several salts showed in vitro and/or in vivo inhibitory activity against B.cinerea,although no correla-tion was found between the two activities.Indeed,ammo-nium bicarbonate,ammonium phosphate,sodium acetate,and sodium sulphate strongly reduced B.cinerea mycelial growth but displayed an inconsistent activity against grey mould on small bunches.In particular,sodium phosphate reduced the growth of the pathogen in vitro,but increased rot incidence on small table grape bunches.As reported for other phosphates(Palou et al.,2002),presumably this salt provided additional nutrients and/or enhanced environmental conditions for the development of the pathogen.In addition, CC showed almost no in vitro inhibitory activity but strongly reduced grey mould incidence on table grapes.Other salts such as PC,SB,and SC showed the same behaviour,with both in vitro and in vivo assays.Therefore,the present study suggests that in vitro tests are inappropriate to predict the potential of a salt in controlling grey mould of table grapes as already reported for silver scurf on potato tubers(Hervieux et al.,2002).The different in vitro and in vivo behaviour of salts suggests that specific salt–host tissue interactions may involve biochemical reactions,such as defense mechanisms contributing to the control of grey mould(Hervieux et al., 2002).Furthermore,the interaction between salt and agar medium(Biggs et al.,1997)as well as the interaction between salt and environment(Punja and Grogan,1982)may play an important role.Results of the present work showed that pH of salt solu-tions has a minor role in the mode of action.High disease reductions were obtained with PC and SC(pH11.3),but also with SB(pH8.5),and CC(pH6.0).Furthermore,PC,SB,and SC completely inhibited the germination of B.cinerea inde-pendently of the pH,whereas in water and in CC solutions, conidial germination was reduced,but not completely inhib-ited by high pH.Although our results,in accordance with data reported by Palmer et al.(1997),demonstrate that pH alone cannot explain the inhibitory action of salts,it cannot be excluded that a change in pH value may affect the activity of salts(Punja and Grogan,1982;Mecteau et al.,2002).In the present work,results from small-and large-scale tests demonstrated that pre-harvest applications of salts are an effective strategy to reduce the incidence of sour rot and botrytis storage rot of table grapes.Although with some dif-ferences,CC,PC,SB,and SC in small-scale tests and CC,SB, and SC in large-scale tests,provided similar or higher level of protection compared to conventional fungicides.Higher control levels were achieved with two applications(21and 5days before harvest)as compared to a single application made5days before harvest.Furthermore,early applications (90and30days before harvest)of calcium chloride provided better control offield rots,mainly sour rots.It is presumed that an early application of calcium chloride favoured the penetration of calcium through the skin and that a second application increased the calcium level inside the fruit,result-ing in higher levels of protection compared to treatments made just a few days before harvest.Our data are in accor-dance with previous reports on pre-harvest application of calcium chloride.Pre-harvest applications of calcium chlo-ride increased the Ca2+content of sweet cherries and apples (Brown et al.,1996),and table grape berries(Miceli et al.,。