Variations in Antioxidase Activities and MDA Content in Potato Tubers Infected by Fusarium trichothecioides

Fenglan LI,Yao FU,Qiang YUAN,Wenhe LU,Yongqing XU,Rongmei LIU,Baozhong HU,Fuwei CHEN,Yuanyuan XU,Yanzhong FENG

1.College of Life Science,Northeast Agricultural University,Harbin 150030,China;2.Heilongjiang Academy of Agricultural Sciences,Harbin 150086,China

Potato,the main crop for food,vegetable,animal feed and industrial raw materials,has high yield,wide adaptability,high economic efficiency and abundant nutrients,and has contributed much to the food supply and safety in China’s underdeveloped areas.It also plays an important role in post-disaster relief,adjustment of agricultural structure and regional distribution[1].In recent years,potato has been planted in an increasing scale,and has become China’s forth most important food crop.China now is the largest potato planting country all over the world,producing 1/4 of world’s potato[2].In Northern China,both commercial and seed potatoes are mainly stored in cellars in winters,while the inappropriate storage conditions often result in some disease to potatoes,such as late blight,ring rot,dry rot,black heartand softrot,severely affecting potato yield and quality,as well as potato production and sales in the next year[5].Various diseases of potato emerge with the planting area increasing,and now have become the main factor limiting the development of potato industry.Among them,Fusariumdry rot is one of the main potato diseases at storage period,and causes 60% loss of potato stored in cellars[6-7].

Most previous studies about potato dry rot focused on the isolation and identification of pathogens and disease prevention,but the interaction mechanism between pathogens and potato is still unclear[8].The genetic characteristics,variations in physiological and biochemical properties of potato tubers duringF.trichothecioidesinfection have been rarely re-ported so far[9-11].At present,there are only some simple chemical methods forthe prevention and controlofFusariumdry rot in potato,and all the methods can only reduce the damages.However,they can not avoid the incidence of the disease,but also causes serious environmental pollution.So far,researchers have not found an effective method for the prevention and control of potato dry rot,because of the lack of theoretical basis.The interaction mechanism between potato andF.trichothecioidesis the basis to select an effective way to prevent and control potato dry rot[12-14].

Potato dry rot is caused by various pathogens of genusFusarium,and those pathogens can grow in a wide range of temperature 5-30℃and humidity.High temperature and humidity are most conducive to dry rot incidence,particularly under poor storage and ventilation conditions[15].It is reported that the occurrence ofFusariumdry rot is more frequent in earlymaturing potato varieties than in latematuring varieties[16].According to statistics,there are six species or variants causing potato dry rot in Heilongjiang Province,they areFusariumsambucinum,F.avenaceum,F.trichothecioides,F.solanivar.coeruleum,Fusarium solani,F.sporatrioidesandF.culmorum,respectively.Among them,F.sambucinum,F.avenaceumandF.trichothecioidesare more pathogenic[17-18].In this study,the activities of main antioxidases and the content of malondialdehyde(MDA)in potato tubers infected withF.trichothecioideswere measured,to determine the roles of the defense enzymes duringF.trichothecioidesinfection,and to provide theoretical basis for the scientific control ofFusariumdiseases,and the breeding of new potato varieties with broad-spectrum disease resistance.

Materials and Methods

Materials

The detoxified tubers of two major potato varieties in Heilongjiang Province Keshan 1(resistant toFusarium trichothecioides),Atlantic(susceptible toF.trichothecioides)were used as the host materials.F.trichothecioidesWollenw was preserved in PDA medium in our laboratory.

Methods

The solid culture medium ofF.trichothecioidesWollenw was punched with a hole punch to obtain 1 cm disks.Potato tubers of Atlantic and Keshan 1 were surface sterilized,peeled,and sliced into pieces 5 mm in thickness and 3 cm in diameter.Then,pathogen disks were inoculated into the center of each potato pieces in dishes,and cultured at 25℃.1.0 g of the samples were collected 3,6,12,24,48 and 72 h later,wrapped with foil and immediately placed in liquid nitrogen.Three repetitions were set for each sampling.After that,all the samples were measured through enzymeprotein assay for target physiological and biochemical indices[19].

Results and Analysis

Variations in soluble protein content during F.trichothecioides infection

The tubers of Atlantic and Keshan 1 were infected byF.trichothecioides.During this process,the soluble protein content in the tubers was measured at different time.As shown in Fig.1,the soluble protein content in Atlantic changed over time afterFusariuminfection,peaked and exceeded the uninfected control at hour 9,and was below the control at other time periods.The soluble protein content in Keshan 1 tubers infected byF.trichothecioideschanged slightly compared with the uninfected control,and it was 43% higher than that of the uninfected at only hour 9.It could be concluded from Fig.1 that the soluble protein content in the infected tubers of the two potato varieties changed in a similar pattern,lower than that of the uninfected at most time.In addition,the soluble protein content of Atlantic which is susceptible toF.trichothecioides,was higher than that of Keshan 1,which is resistant toF.trichothecioides.The results suggested that the soluble protein content may be negatively correlated to the resistance of potato varieties toF.trichothecioides.

Variations of SOD activity during F.trichothecioides infection

As shown in Fig.2,the SOD activity of both potato varieties decreased 3 h after infection and then rose sharply.The SOD activity in infected Keshan 1 peaked at hour 6,that in infected Atlantic peaked at hour 9.From hour 9 to 48,the SOD activity in infected Keshan 1 which is resistant toF.trichothecioidesincreased again,while that in infected Atlantic which is susceptible toF.trichothecioideskept decreasing.The results indicated that the increasing SOD activity in infected potato tubers had a certain relationship to the resistance of potato varieties toF.trichothecioides.

Variation of POD activity during F.trichothecioides infection

As shown in Fig.3,the POD activity in infected Atlantic and Keshan 1 tubers decreased within the first 6 h,while that in Atlantic changed more significantly.6 h after infection,the POD activity in both potato varieties began to increase.In addition,the POD activity in Keshan 1 which is resistant toF.trichothecioideswas always higher than that in Atlantic which is susceptible toF.trichothecioideswithin the 12 h after infection.The results proved that the high POD activity is closely related to the strong resistance of potato varieties toF.trichothecioides.

Variation of malondialdehyde(MDA)content during F.trichothecioides infection

As shown in Fig.4,compared with the control uninfected,the MDA content in infected Keshan 1 tubers decreased slightly from the beginning,and greatly at hour 24;the MDA content in infected Atlantic tubers decreased significantly after infection,and more significantly at hour 12.A large number of antioxidants were generated after fungal infection,which might be the reason causing the decrease in MDA content.

Discussion

Reactive oxygen species(ROS)is a very important part in plant defense response,as it can trigger programmed cell death(PCD),drive the oxidative crosslinking of cell wall proteins,so that the pathogens can be confined to the suicidal host cells,thereby slowing down pathogen invasion[20].A free radical reaction may be triggered in the plants under stress.The study by Huet al.[21]revealed that the SOD activity inAcer truncatumunder simulated drought stress decreased at first and then gradually increased to normallevel.Previous studies have shown that the SOD activity in the plants resistant to drought,frost,high salinity and toxics is significantly higher,indicating that high SOD activity is related to plant’s resistance to stresses,facilitating the selection of new varieties with strong resistance and high economic benefits[22].Peroxidase (POD),a ubiquitous and active enzyme in plants,is closely related to plant metabolism and resistance[23-24].It has a dual role:it has a protective role at early period of aging and response to stress,and then it is involved in the generation of reactive oxygen species at late period of aging and response to stress.It is a product of plant aging,and even can be used as an index of senescence[25].Malondialdehyde(MDA)is a most commonly used index for the measurement of membrane lipid peroxidation[26].The damages caused by stress or aging to plants are closely related to reactive oxygen-induced membrane lipid peroxidation.Diene conjugates,lipid peroxides,MDA and ethylene,etc.are the products of membrane lipid peroxidation,among which,MDA is one of the most important one.So MDA content can be used as an index for measuring the degree of membrane lipid peroxidation,thereby indirectly determining the damages in membrane system and plant resistance to stresses.

In this experiment,we found that the soluble protein content inF.trichothecioidesinfected Atlantic and Keshan 1 tubers were lower than that of the uninfected;the soluble protein content of infected Atlantic which is susceptible toF.trichothecioideswas higher than that of Keshan 1,which is resistant toF.trichothecioides,indicating that soluble protein content is negatively correlated to the resistance of potato varieties toF.trichothecioides.From 9 to 48 h afterF.trichothecioidesinfection,the SOD activity of Keshan 1 began to increase,while that of Atlantic kept decreasing,suggesting that SOD activity has a certain relationship with the resistance of potato varieties toF.trichothecioides.The POD activity in both potato varieties changed significantly afterF.trichothecioidesinfection,proving that it is closely related to the resistance of potato varieties toF.trichothecioides.We also found that the MDA content in potato tubers also significantly reduced a certain time period afterF.trichothecioidesinfection,which indicated that MDA content may have a certain role in disease resistance of potato.

The defense enzymes play a crucial role in potato’s resistance toFusariumdry rot.The activities of related defense enzymes inF.trichothecioides-resistant potato varieties increased at varying degrees afterF.trichothecioidesinfection,indicating that these enzymes are related to potato’s resistance toF.trichothecioides.This paper illustrates the physiological basis of potato’s resistance according to the changes in antioxidase activity and provides a theoretical basis for the selection of disease-resistant potato varieties.

[1]XIE JH(謝建華).Potato production situationanddevelopmentcountermeasures in China(我國(guó)馬鈴薯生產(chǎn)現(xiàn)狀及發(fā)展對(duì)策)[J].China Agricultural Technology Extension(中國(guó)農(nóng)技推廣),2007,(5):4-7.

[2]SUN DS(孫東升),LIU GH(劉合光).The analysis and prospect outlook of China's potato industry(我國(guó)馬鈴薯產(chǎn)業(yè)發(fā)展現(xiàn)狀及前景展望)[J].Agricultural Outlook(農(nóng)業(yè)展望),2009,(3):25-28.

[3]SONG BF(宋伯符).Promising potato industry in China(充滿朝陽(yáng)的中國(guó)馬鈴薯產(chǎn)業(yè))[A].Potato industry and high technologies(馬鈴薯產(chǎn)業(yè)與高新技術(shù))[C].Harbin:Harbin Engineering University Press(哈爾濱:哈爾濱工程大學(xué)出版社),2002:185-188.

[4]QU DY(屈冬玉),XIE KY(謝開云),JIN LP(金黎平),et al.Development of pota-to industry and food security in China(中國(guó)馬鈴薯產(chǎn)業(yè)發(fā)展與食物安全)[J].Scientia Agricultura Sinica(中國(guó)農(nóng)業(yè)科學(xué)),2005,(2):258-262.

[5]TIAN F(田芳).Occurrence and control of diseases of potato in cellars in Northern China(北方地區(qū)馬鈴薯窖藏病害的發(fā)生及防治)[J].Guide of Sci-tech Magazine(科技致富向?qū)?,2012,(27):378-402.

[6]CARNEGIE SF,RUTHVEN AD,LINDSAY DA,et al.Effects of fungicides applied to seed potato at harvest or after grading on fungal storage diseases and plant development[J].Ann Appl Biol,1990,116:61-72.

[7]ZHAO SS(趙生山),NIU LH(牛樂(lè)華).Survey on potato disease and chemical control during storage(馬鈴薯貯藏期病害調(diào)查及藥劑防治研究)[J].Information of Agricultural Science and Technology(農(nóng)業(yè)科技與信息),2008,(11):44-46.

[8]PENG XW(彭學(xué)文),ZHU WH(朱杰華).Fungal diseases and distribution of in potato in Hebei Province(河北省馬鈴薯真菌病害種類及分布)[J].Chinese Potato Journal(中國(guó)馬鈴薯),2008,22(1):31-33.

[9]LIU CH(劉程惠),HU WZ(胡文忠).Physio-biochemical changes of freshcut potato at different storage temperatures(不同貯藏溫度下鮮切馬鈴薯的生理生化變化)[J].Food&Machinery(食品與機(jī)械),2008,24(2):38-42.

[10]LI GL(李國(guó)龍),WU HX(吳海霞),WEN L(溫麗).Progress in physiological and molecular mechanism of drought resistant in crop(作物抗旱生理與分子作用機(jī)制研究進(jìn)展)[J].Chinese Agricultural Science Bulletin(中國(guó)農(nóng)學(xué)通報(bào)),2010,40(23):185-191.

[11]CHEN Q,SUN HS,YANG JL,et al.Inheritance and molecular mapping of new genes conferring late blight and CPB resistance in Mexican wild potato speciesSolanum pinnatisectum[J].Agricultural Science & Technology,2009(5):257-261.

[12]ZHANG RJ,CHEN YJ,MENG Ml,et al.Regulation of nitrogen on potato under NaCl stress[J].Agricultural Science&Technology,2010,11(11-12):65-67.

[13]BI Y,LI YC,GE YH.Induced resistance in postharvest fruits and vegetables by chemicals and its mechanism[J].Stewart Postharvest Review,2007,(6):1-7.

[14]WU B(吳斌),WANG FL(王建富),SUN RL(孫瑞林),et al.Potato pest occurrence and control in Taixing City(泰興市馬鈴薯病蟲害發(fā)生特點(diǎn)與防治技術(shù))[J].Modern Agricultural Sciences and Technology(現(xiàn)代農(nóng)業(yè)科技),2011,(14):179-180.

[15]THEV DJ,LIAO XL(廖曉蘭).Effects of temperature on dry occurrence in potatoes inoculated with differentFusariumspecies(溫度對(duì)接種不同鐮刀菌的馬鈴薯干腐病發(fā)展的影響)[J].Rain Fed Crops(國(guó)外農(nóng)學(xué):雜糧作物),1991(6):30-33.

[16]HOU ZY(侯忠艷).Occurrence and control of Fusarium dry rot in potato(馬鈴薯干腐病的發(fā)生與防治)[J].Modern Agricultural Sciences and Technology(現(xiàn)代農(nóng)業(yè)科技),2012,(10):173-179.

[17]MIN FX(閔凡祥),WANG XD(王曉丹),HU LS(胡林雙),et al.Identification of species and pathogenicity ofthe Fusarium on potato in Heilongjiang Province(黑龍江省馬鈴薯干腐病菌種類鑒定及致病性)[J].Plant Protection(植物保護(hù)),2010,(4):112-115.

[18]LI FL(李鳳蘭),JIANG XF(蔣先鋒),SHI LJ(史麗娟),et al.Isolation and identification of two pathogenic stains associated with potato dry rot(兩株馬鈴薯干腐病病原菌的分離和鑒定)[J].Crops(作物雜志),2013,4:125-127.

[19]WANG QM(汪俏梅),GUO DP(郭得平),KYIKYI WIN.Effect of 1-MCP on storage life,quality and antioxidant enzyme activities of broccoli(1-MCP對(duì)青花菜貯藏壽命、品質(zhì)和抗氧化酶活性的影響)[J].Journal of Zhejiang University(Agriculture&Life Sciences)(園藝學(xué)報(bào):農(nóng)業(yè)與生命科學(xué)版),2004,31(2):205-209.

[20]BRISSON LF,TENHAKEN R,LAMB CJ.Function of oxidative cross-linking of cell wall structural proteins in plant disease resistance[J].Plant Cell,1994,6:1703-1712.

[21]HU JJ(胡景江),LIU ST(劉樹滔),PAN JR(潘劍茹).Research progress on technologies of SOD property improvement(SOD特性改善技術(shù)研究進(jìn)展)[J].Strait Pharmaceutical Journal(海峽藥學(xué)),2009,21(2):11-14.

[22]LI JH(李金華),WANG F(王豐),LIAO YL(廖亦龍).Advance on seed vigor physiological-biochemical and genetic mechanisms in rice(水稻種子活力的生理生化及遺傳研究)[J].Molecular Plant Breeding(分子植物育種),2009,7(4):772-777.

[23]WANG JY(王晶英),ZHAO YS(趙雨森),WANG Z(王臻),et al.Effect of drought stress on physiologic and biochemical characteristic of Populus alba×Populus berolinensis(干旱脅迫對(duì)銀中楊生理生化特性的影響)[J].Journal of Soil and Water Conservation(水土保持學(xué)報(bào)),2006,20(1):197-200.

[24]JIAO LL(繳麗莉),NI ZY(倪志云),LU BS(路丙社),et al.Effects of low temperature on cold-resistance of David maple(低溫脅迫對(duì)青榨槭幼樹抗寒指標(biāo)的影響)[J].Journal of Agricultural University of Hebei(河北農(nóng)業(yè)大學(xué)學(xué)報(bào)),2006,29(4):44-47.

[25]ZHANG SN(張蜀寧),ZHANG ZC(張振超),ZHANG HL(張紅亮),et al.Growth,physiological and biochemical characteristics of non-heading Chinese cabbage under low temperature stress(低溫脅迫對(duì)不同倍性不結(jié)球白菜生長(zhǎng)及生理生化特征的影響)[J].Acta Botanica Boreali-Occidentalia Sinica(西北植物學(xué)報(bào)),2008,28(1):109-112.

[26]ZHAO SJ(趙世杰),XU ZC(許長(zhǎng)成),ZOU Q(鄒琦),et al.Improved method for determining malondialdehyde content in plant tissues(植物組織中丙二醛測(cè)定方法的改進(jìn))[J].Plant Physiology Communications(植物生理學(xué)通訊),1994,(3):207-210.