Fusarium oxysporum (F221-B) as Biocontrol Agent against Plant Pathogenic Fungi in Vitro and in Hydroponics

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1 Plnt Protect. Sci. Vol. 53, 17, No. 2: Fusrium oxysporum (F221-B) s Biocontrol Agent ginst Plnt Pthogenic Fungi in Vitro nd in Hydroponics Titi Thongkmngm nd Tnimnun Jenksorn 1 Deprtment of Plnt Production Technology, Fculty of Agriculturl Technology, King Mongkut s Institute of Technology Ldkrbng, Bngkok, Thilnd *Corresponding uthor: kjtnimn@kmitl.c.th Abstrct Thongkmngm T., Jenksorn T. (17): Fusrium oxysporum (F221-B) s biocontrol gent ginst plnt pthogenic fungi in vitro nd in hydroponics. Plnt Protect. Sci., 53: Efficcy of non-pthogenic Fusrium oxysporum (F221-B) ws ssessed s possible biocontrol gent ginst fungl pthogens, nmely Curvulri lunt (C11, C12), F. semitectum (F113), F. oxysporum f.sp. lctuce (F221-R, F442-G), Rhizoctoni solni (R11, R12), Rhizoctoni sp. (R111, R112, R113) in vitro, while F221-B showed moderte bility to inhibit the mycelil growth of tested fungi bout 36 56%. Then, F221-B ws further evluted for its bility of controlling lettuce root rot nd wilt cused by F442-G in hydroponics. It ws reveled tht F221-B reduced disese incidence nd severity bout 6 8% compred to the inoculted control nd significntly promoted the growth of 3 lettuce vrieties. Interestingly, using only F221-B gve the significntly highest fresh weight (twice over the helthy control). Conclusively, this study provides n importnt suggestion for further development of F221-B since it showed the bility of biocontrol gent nd plnt growth promoting fungus. Keywords: Curvulri lunt (C11, C12); Fusrium semitectum (F113); F. oxysporum f.sp. lctuce (F221-R, F442-G); Rhizoctoni sp. (R111, R112, R113); R. solni (R11, R12); dul-culture test Biologicl control offers potentil lterntive control mesure in the re of plnt diseses nd becomes promising tool to use in griculturl production when it reduces the relese of polluting chemicl pesticides to the ecosystem (Kur et l. 1). At present, quite number of biocontrol gents (BCA) re found to be ble to mnge the plnt diseses effectively s well s they ll ecologiclly sound proof (Howell 3; Kur et l. 1; Kkvn et l. 13; Chen et l. 14; Kim et l. 14; Song et l. 14). Idelly, the ntgonist/bca must be ecologiclly fit to survive nd function with the prticulr condition of the ecosystem. Moreover, the ntgonist/bca must be present t n dequte level nd be cpble of effectively intercting with the pthogen or host plnt to provide cceptble disese control. The wilt nd root rot-inducing strins of Fusrium oxysporum cuse serious losses of mny economiclly importnt griculturl crops worldwide (Benhmou et l. 1989; Hervs et l. 1998; Pul et l. 1999; Kur 3; Kur et l. 1) including lettuce grown in hydroponics in Thilnd (Thongkmngm et l. 12). Effective control methods ginst the Fusrium disese of plnts in hydroponics re still limited in terms of n environmentlly sfe method. Biologicl control of Fusrium wilt nd root rot diseses hs been reported in numerous crops cultivted in the soil (Albouvette et l. 1993; Frvel et l. 3) such s tomto (Fuchs et l. 1997, 1999; Duijff et l. 1998; Lrkin & Frvel 1998), cucumber (Mndeel & Bker 1991), wtermelon (Lrkin et l. 1996), spinch (Ktsube & Alsk 1997), bsil (Minuto et l. 1997), chickpe (Hervs et l. 1998), nd rkkyo (Hond & Kwkub 1998) but not in crops grown in hydroponics especilly in the system without substrte. Non-pthogenic F. oxysporum F221-B recovered from roots of lettuce grown in hydroponics (Thongkmngm et l. 12) hs shown to significntly 85

2 Vol. 53, 17, No. 2: Plnt Protect. Sci. promote the growth nd yield of Cos, Green Ok, Red Ok, nd Butterhed lettuce, kle nd mung ben in hydroponics (Thongkmngm et l. 13). With the quite promising bility of F221-B, we therefore conducted the reserch to determine if (1) nonpthogenic F221-B could ct in vitro s potent BCA ginst 1 fungl plnt pthogens nd (2) F221-B cn potentilly protect three vrieties of hydroponiclly grown lettuce ginst the virulent isolte of F. oxysporum f.sp. lctuce (F442-G) (Thongkmngm et l. 12). Mteril nd Methods Fungl isoltes. Culture of F221-B ws obtined from the previous experiment (Thongkmngm et l. 12, 13). The test fungl pthogens were isolted from (lef, seed) nd hydroponiclly grown lettuce (Red Ok, Green Ok) showing disese symptoms from different plces (Tble 1) using tissue trnsplnting technique. Curvulri lunt (C11, C12), F. semitectum (F113), R. solni (R11, R12), Rhizoctoni sp. (R111, R112, R113) were obtined ccordingly, while the most virulent strins of F. oxysporum f.sp. lctuce (F221-R, F422-G) were obtined from the previous experiment (Thongkmngm et l. 12). Then, the fungi were trnsferred onto PDA slnts nd mintined s stock cultures for this experiment. In vitro ssessment of the ntgonistic ctivity of non-pthogenic F221-B ginst mycelil growth of 1 plnt pthogenic fungi by dul-culture test. F 221-B ws determined for its in vitro ntgonistic ctivity ginst ten phytopthogenic fungi by dulculture method (Dennis & Webster 1971) using potto dextrose gr (PDA). A mycelil disc (5 mm) obtined from the peripherl region of the colony 7-dy-old cultures of trgeted fungl pthogens (C11, C12, F113, F221-R, F422-G, R11, R12, R111, R112, nd R113) ws plced on PDA t the edge of ech plte, 1.5 cm from the periphery. Then, disc of mycelium of 5 mm in dimeter cut from the growing edge of the 7-dy-old culture of F221-B ws plced on ech plte, opposite to the inoculum of the pthogen, 1.5 cm from the periphery. A completely rndomised design (CRD) ws used with five replictes for ech isolte. In the control plte, sterile gr disc ws inoculted on the side opposite to the pthogen. All the inoculted pltes were incubted t room temperture (25 ± 2 C) for 9 dys until the control plte ws full. During the incubtion period, growth of pthogens ws dily mesured nd the percent growth inhibition (GI) ws clculted reltive to the control s follows: GI = ((D1 D2)/D1) 1 where: D1 dimeter of pthogen colony in control; D2 dimeter of pthogen colony in tretment Evlution of non-pthogenic F221-B for controlling Fusrium root rot of three lettuce vrieties grown in hydroponics. F 221-B ws further evluted for its biocontrol efficcy ginst Fusrium root rot nd wilt (cused by F422-G) of Butterhed, Cos, nd Red Ok lettuce grown in modified deep flow technique (DFT). Hydroponic cultivtion. Lettuce seeds were germinted in seedling try on moist sponge t room temperture. After 7 dys, seedlings were moved to new try with nutrient solution (EC = 1 ms/cm, ph ). Then, 14-dy-old seedlings were trnsplnted into mini DFT (plstic continer: cm) with nutrient solution (EC = 1.6 ms/cm, ph ) to be redy for being treted ccording to the tretments. Experiment. Within ech vriety of lettuce, four tretments were rrnged in CRD with five replictions of two plnts. Before trnsplnting the 14-dy-old lettuce seedlings into the DFT under outdoor conditions, seedling roots were inoculted with F221-B by dipping into spore suspension (1 1 8 spore/ml nd 5 ml per plnt). Three dys lter, the plnt roots were inoculted with spore suspension of the pthogen F422-G (1 1 6 spores/ml nd 5 ml per plnt), while sterile wter ws used for helthy control. Disese severity ws rted up to 6 levels ( helthy root, 1 reddish brown root, 2 reddish brown root tht hs become rotten, 3 rotten root, with slight wilting, 4 rotten root with severe wilting, 5 plnt tht hs become ded). Disese incidence (DI) nd disese index (DIn) were clculted s follows: % DI = (Number of plnts showing infected roots/ /Number of totl plnts) 1 % DIn = {[ (Number of plnts showing infected roots disese severity)]/(number of totl plnts the highest disese severity)} 1 The evlution ws done weekly on DI, DIn, nd plnt growth (lef number nd size, SPAD vlue, stem nd root fresh weight, stem nd root dry weight). Survivl of F221-B nd pthogen ws lso monitored 86

3 Plnt Protect. Sci. Vol. 53, 17, No. 2: Tble 1. Source, host, nd isoltion description of fungl pthogens used in this study No. Pthogen Source Host Isoltion description (symptom) 1 Curvulri lunt C11 Rtchburi (Thilnd) (seed) Seeds infected by dirty pnicle disese showing blck spot, smll brown lesion, prtil blckening nd covered by fungl spores 2 C. lunt C12 Bngkok (Thilnd) (seed) Undeveloped kernels with severe blck spots, brown lesions, prtil browning nd blckening 3 Fusrium semitectum F113 Rtchburi (Thilnd) (seed) Undeveloped kernels with brown lesion or blck spots covered with whitish spore mss 4 F. oxysporum f.sp. lctuce F221-R Smutprkrn (Thilnd) lettuce (red ok) Root rot, yellowing nd wilting of bsl leves s well s stunting nd plnt deth. The cortex of the crown nd upper root of infected plnts becme reddish brown nd decyed 5 F. oxysporum f.sp. lctuce F422-G Chonburi (Thilnd) lettuce (green ok) Disese symptoms included root rot, yellowing nd wilting of bsl leves s well s stunting nd plnt deth 6 R. solni R11 Angthong (Thilnd) (lef) Gryish lef lesions (1 3 cm) with red border 7 R. solni R12 Angthong (Thilnd) (lef) Blck nd brown lesions (1 5 cm) or stripes on young lef surrounded with red dots 8 Rhizoctoni sp. R111 Bngkok (Thilnd) (lef) Brownish ovl lesions (1 2 cm) on the lef sheths nd lef bldes surrounded with yellow re 9 Rhizoctoni sp. R112 Bngkok (Thilnd) (lef) Brownish ovl lesions (1 3 cm) on lef bldes with drkening t the center. 1 Rhizoctoni sp. R113 Nkornprthom (Thilnd) (lef) Brownish ovl lesions (1 1.5 cm) on lef sheths t the wter level in the pddy field from nutrient solution nd crop roots t hrvest by dilution pltes of selective medium. Dt were nlysed using nlysis of vrince (ANOVA). Tretment mens were compred using Duncn s Multiple Rnge Test (DMRT). Results In vitro ssessment of the ntgonistic ctivity of non-pthogenic F221-B ginst mycelil growth of ten plnt pthogenic fungi using dul-culture 87

4 Vol. 53, 17, No. 2: Plnt Protect. Sci. Tble 2. Assessment of the ntgonistic ctivity of F. oxysporum (F221-B) on colony growth of plnt pthogenic fungi using dul-culture ntgonistic test Pthogen Growth Inhibition (%) 1 Mechnism 2 3 DAI 5 DAI 7 DAI 9 DAI 3 DAI 5 DAI 9 DAI C A A A C A A A F A A A F221-R A A A F422-G A A A R A A A R A A A R C C C R A A C R A A A 1 growth inhibition over control (GI) = (D1 D2)/D1 1; DAI dys fter inocultion test. It reveled tht non-pthogenic F221-B significntly reduced the colony growth of ll tested fungl pthogens t three dys fter inocultion (DAI) (Tble 2). On DAI9, the inhibition percentge for C. lunt (C11 nd C12) ws 42.3 nd 36%, respectively; 42.5% for F. semitectum (F113); 42 nd 38.8% for F. oxysporum f.sp. lctuce (F221-R nd F422-G); 5.4 nd 35.9% for R. solni (R11 nd R12); 36.3, 43.4, nd 56.3% for Rhizoctoni spp. (R111, R112 nd R113). Considering the bove-mentioned inhibition percentges, F221-B showed moderte bility to inhibit ll fungl pthogens. Among the tested fungi, R113 nd R11 were the most susceptible nd reveled the highest percent inhibition of mycelil growth mounting to 5 nd 56%, respectively. Growth inhibition of ll tested fungi during in vitro interction with F221-B t DAI 3, 5, 7, nd 9 is shown in Figure 1. The min mechnism of F221-B ginst 3 DAI 5 DAI 7DAI 9 DAI C11 C12 F113 F21-R F422-G R11 R12 R111 R112 R113 BCA Dul Ptho- BCA Dul Ptho- BCA Dul Ptho- BCA Dul Pthogen gen gen gen Figure 1. Dul-culture ntgonistic test of F. oxysporum (F221-B) ginst 1 plnt pthogenic fungi t 3, 5, 7, nd 9 dys fter inocultion (DAI) 88

5 Plnt Protect. Sci. Vol. 53, 17, No. 2: Hyph Spore Figure 2. Morphologicl bnormlities of hyphe nd spores of plnt pthogenic fungi detected from the interction zone of dul-culture plte A = F. semitectum (F113), hyph: swollen, with short hyphl segment (yest-like cell); B = C. lunt (C11), hyph: deformities with globose form swelling; C = R. solni (R11), hyph: loss of structurl integrity showing deformities with shrivelling; D = C. lunt (C12), hyph: deformities with swelling nd curled; E = C. lunt (C11), spore: lysis, loss of cytoplsmic integrity; F = C. lunt (C12), spore: cytoplsm lekge out of spore; G = F. semitectum (F113), spore: ltertions with incresed size nd cogultion of the cytoplsm; H = F. oxysporum f.sp. lctuce (F422-G), spore: ltertions with incresed size nd cogultion of the cytoplsm the tested fungi ws ttributed to ntibiosis nd competition. Antibiosis of non-voltile metbolites ws more pronounced t the erly stge of incubtion (DAI 3) nd ws followed by competition tht could be involved in the ntgonistic process ginst both strins of C. lunt nd Rhizoctoni sp. R112. In the F221-B pthogen (R111) combintion, only competition ws noted from DAI 3 to DAI 9 bsed on microscopic observtions of hyphl interctions, no mycoprsitism ws detected. A cler zone of interction ws formed in the combintion of F221-B with 6 pthogens, nmely R11, R12, R113, F113, F221-R, nd F422-G. The presence of the inhibition zone without physicl contct between F221-B nd the pthogen colony t DAI 5-9 suggested the secretion of diffusible non-voltile inhibitory substnces. This ws confirmed by the detection of bnormlities of hyphe nd spores of the tested fungi (C11, C12, F113, F422-G, nd R11) from the interction zone or nerby re (Figure 2). Evlution of non-pthogenic F221-B for controlling Fusrium root rot of three lettuce vrieties grown in hydroponics. Neither roots of ll 3 vrieties of lettuces (Butterhed, Cos, nd Red Ok) inoculted with non-pthogenic F221-B lone nor roots of the (uninoculted) control showed ny disese symptoms throughout the tril (Figure 3). The roots inoculted with pthogenic F422-G lone were severely rotten with 1% disese incidence (DI) nd % disese index (DIn) t DAI 28 nd Cos ws shown to be the most susceptible vriety, wheres the tretment with F221-B significntly reduced the percentge of DIn of root rot with the level of control efficcy (over the inoculted control) of 64% for Butterhed, 85.3% for Cos, nd 7% for Red Ok (Figure 3). Interestingly, F221-B hd strong ntgonistic ctivity ginst Fusrium root rot especilly on Cos lettuce which ws the most susceptible vriety in our study (Figure 4). Considering plnt growth prmeters, results were in line with the disese control efficcy. Tht is, prt from hving the significnt control efficcy, the tretment with F221-B (with being chllenged with the pthogen) could significntly increse the plnt growth (e.g. lef number nd size, SPAD vlue, fresh nd dry weight of stem nd root) of tested lettuce over the inoculted control nd similrly in reltion to the helthy control t DAI 45 (Tble 3, 89

6 Vol. 53, 17, No. 2: Plnt Protect. Sci. Tble 3. Growth of 3 lettuce vrieties treted with non-pthogenic F. oxysporum (F221-B) nd pthogenic F. oxysporum (F422-G) in hydroponics Plnt growth prmeter Totl yield (g/plnt) lef number lef size (cm) chlorophyll (SPAD vlue) fresh weight dry weight 24 dys 31 dys 38 dys 45 dys 24 dys 31 dys 38 dys 45 dys 24 dys 31 dys 38 dys 45 dys stem root stem root Lettuce Tretment helthy control 4.2 b1 7.8 b 14.2 c 14.6 c 3.2 b 7.2 b 16.2 c 1.6 c 21.3 c 29.8 b 29.6 c 29.8 b 72.1 b 1.9 b 1.69 b.45 b F221-B 4.1 b F422-G 3.8 b 4.3 c 5.8 d 7.6 d 2.5 c 3.4 c 4.2 d 5.25 d 16.4 d 14.9 c 1.4 d 8.5 c 24.5 c 3.2 c.78 c.12 c Butterhed F221-B+F422-G b 16.9 b 16.1 b b 17.5 b 21.2 b 28.4 b 3.8 b 31.7 b b 2.3 b.54 b helthy control 4.4 b 7.5 b 12.4 c 16.3c 4.2 c 7.5 c 14.2 b b 24.5 c 28.5 b 29.8 b 3. b 11.7 c b.88 b F221-B F422-G 2.7 c 4.2 c 5.5 d 5.9 d 2.6 d 4.5 d 4.9 c 5.12 c 8.4 d 9.2 c 1.4c 1.5 c 7.2 d 2.5 b.3 c.1 c Cos F221-B+F422-G 4.5 b 8.5 b 15.4 b 18.3 b 5.1 b 9.8 b 15.3 b 19.2 b 29.6 b b 31.2 b b b 1. b helthy control 4.1 b 7.5 b 14.4 b 17.5 b 4.2 b 7.8c 12.2 c 16.5 c b 22.8c 24.5 b c.5 b c F221-B F422-G 3.1 c 3.8 c 4.1 d 4.8 c 3.3 c 4.1 d 4.3 d 4.5 d 1.5 b 11.9 c 14.2 d 1.5 c 37.6 d 4.4 c 1.1 b.18 d F221-B+F422-G 4.2 b 6.5 b 13.5 c 18.5 b 4.7 b 8.9 b 13.7 b 18.6 b b 24.5 b 25.2 b b 21.9 b b Red Ok Mens of five replictions in column within ech vriety of lettuce followed by common letter re not significntly different ccording to Duncn s Multiple Rnge Test (DMRT) t P =.5 Figure 4). Most striking nd interesting ws the finding tht tretment with only F221-B (in the bsence of the pthogen) gve the significntly highest fresh weight per plnt (115.9, 192.8, nd g for Butterhed, Cos, nd Red Ok, respectively) which were lmost twice over the helthy control (Figure 4). Aprt from disese incidence nd disese index, the survivls of biologicl control gent (BCA) nd pthogen were lso checked t hrvest from nutrient solution nd plnt roots in terms of colony forming units (CFU/ml). F422-G lone incresed its popultion compred to the originl mount while F221-B remined constnt throughout the tril. Survivls of F422-G in tretment treted with F221-B were lower thn those of the inoculted control (Tble 4). F221-B ws detected t hrvest in the rnge of CFU/ml. In the BCA-pthogen cotreted tretment, the mount of F422-G ws higher thn the originl mount but still lower thn tht in F422-G lone tretment. In ddition, the survivl of F221-B nd F422-G ws detected in roots nd nutrient solution (Tble 4). Discussion In this study, non-pthogenic F221-B, recovered from roots of hydroponiclly grown lettuce, ws ssessed for its possible ntgonistic ctivity ginst plnt pthogenic fungi in vitro nd in hydroponics. F221-B could considerbly inhibit the in vitro growth of 1 tested fungi nd exerted direct inhibitory effect e.g. ntibiosis nd competition. Unlike F221-B in our study, most of the non-pthogenic F. oxysporum strins which hve been so fr reported s promising BCA ginst fungl pthogens were isolted only from rhizosphere soils nd roots of plnts grown in suppressive soils (Benhmou et l. 2; Frvel et l. 3; Abeysinghe 6; Rodriguez et l. 6; Ptil et l. 11) but not from hydroponics. However, these results of the ntgonistic ctivity nd mechnisms of F221-B were still in line with those findings. Three non-pthogenic F. oxysporum isoltes from suppressive soil in Sri Lnk were in vitro proved to hve n ntgonistic potentil ginst pthogenic isoltes of F. oxysporum f.sp. rdicis-cucumerinum (Abeysinghe 6), while seven isoltes from rhizosphere soils of tomto from Krntk (Indi) were reported to hve n in vitro ntgonistic potentil ginst the tomto wilt pthogen (F. oxysporum f.sp. lycopersici) bout 24 4% inhibition with competition mechnism (Ptil et l. 11). F. oxysporum ginst plnt pthogens in dul-culture ssy provided the convincing evidence e.g. the non-pthogenic Fo47 exerted 9

7 Plnt Protect. Sci. Vol. 53, 17, No. 2: Tble 4. Survivl of non-pthogenic F. oxysporum (F221-B) nd pthogenic F. oxysporum (F422-G) in roots (R) nd nutrient solution (NS) t the end of the tril Lettuce Butter hed Cos Red ok Survivl of F. oxysporum (CFU/ml) Originl mount Tretment F221-B F422-G (spores/ml) R NS R NS F221-B F422-G F221-B+F422-G (3 1 5 ) + ( ) F221-B F422-G F221-B+F422-G (3 1 5 ) + ( ) F221-B F422-G F221-B+F422-G (3 1 5 ) + ( ) direct inhibitory effect on Pythium ultimum through combintion of ntibiosis nd mycoprsitism (Benhmou et l. 2) nd the non-pthogenic S6 hd mrked ntgonistic cpcity ginst the pthogen Sclerotini sclerotiorum with the responsible metbolite cyclosporine A (Rodriguez et l. 6). Disese incidence (%) Disese incidence (%) Disese incidence (%) Butter hed Cos Red ok b b 4 b b b b c c c c c c c c c c b c c b c c b c c b 7 DAI 14 DAI 21 DAI 28 DAI 7 DAI 14 DAI 21 DAI 28 DAI b b b c c c c c c c c 7 DAI 14 DAI 21 DAI 28 DAI b b b c c c c c c c c 7 DAI 14 DAI 21 DAI 28 DAI Disese index (%) Disese index (%) Disese index (%) 1 Butter hed Cos Red ok b b b b c c c c c c c c 7 DAI 14 DAI 21 DAI 28 DAI b b b b c c c c c c c c 7 DAI 14 DAI 21 DAI 28 DAI Figure 3. Effect of non-pthogenic F. oxysporum (F221-B) on disese development of Fusrium root rot nd wilt of lettuce grown in hydroponics helthy control; F221-B; F422-G; F221-B+F422G; DAI dys fter inocultion 91

8 Vol. 53, 17, No. 2: Plnt Protect. Sci. Red ok Cos Butter hed Helthy F221-B F422-G Stem F221-G+ F422-G Helthy F221-B F422-G Figure 4. Effect of non-pthogenic F. oxysporum (F221-B) nd pthogenic F. oxysporum (F422-G) on the growth (stem nd root) of 3 lettuce vrieties in hydroponics t 28 dys fter inocultion Root F221-G+ F422-G From our study, the differences in verge responses to ntgonism by F221-B between the different gener of pthogens imply tht genes of the pthogen might be involved in regulting the different levels of ntgonism. This confirmed the results of other studies reveling tht non-pthogenic isoltes from suppressive soils including Fo47 (Lemnceu et l. 1992, 1993) could differ in their efficcy s well s in their mechnisms (Nel et l. 6). Another interesting result of this study ws tht F221-B nd pthogenic F. oxysporum (F221-R, F422-G) reduced ech other s growth when grown together (Figure 1). This fungl ntgonism ppered to be minly ssocited with ntibiosis. The result ws in greement with Lemnceu et l. (1993), who demonstrted tht the sme pttern occurred in the co-inocultion experiment between pthogenic F. oxysporum (WCS816) nd non-pthogenic Fo47. In the present study, non-pthogenic F. oxysporum (F221-B) ws investigted s BCA ginst lettuce root rot nd wilt cused by F. oxysporum f.sp. lctuce F422-G in deep flow technique. F221-B ws found to effectively reduce Fusrium root rot nd wilt of 3 lettuce vrieties in the rnge of % s well s to promote their growth nd yield. So fr, mny reports hve provided such n evidence of successfully using mny strins of non-pthogenic F. oxysporum recovered from either disesed rhizosphere soil or suppressive soil for controlling soil-grown crops (Albouvette et l. 1993; Fuchs et l. 1997; Duijff et l. 1998; Frvel et l. 3) but not in hydroponics. Fo47 is the best known nd most effective nonpthogenic strin of F. oxysporum which ws isolted from soil nturlly suppressive to Fusrium wilt of tomto nd melon in Frnce (Albouvette 199; Albouvette & Couteudier 1992; Albouvette et l. 1993, 1996, 1998; Lrkin & Frvel 1999) nd hs been extensively studied for the control of Fusrium wilt disese of severl vegetbles nd flower crops such s tomto (Fuchs et l. 1997, 1999; Duijff et l. 1998), crntion (Lemncen et l. 1992, 1993), nd flx (Duijff et l. 1999). The result gives n importnt step for F221-B s biocontrol gent to be used in hydroponics. Few reports hve reveled the success of using non-pthogenic F. oxysporum for controlling plnts grown in hydroponics only with the rockwool substrte (Ghini et l. ; Minuto et l. 7; Horinouchi et l. 8, 1; Nhlkov et l. 8) which is different from this system. Interestingly, F221-B could survive pretty well in such system (in nutrient solution nd plnt root). This ws in line with 92

9 Plnt Protect. Sci. Vol. 53, 17, No. 2: two findings when one demonstrted the bility of non-pthogenic F. oxysporum (V2w2 nd III4w1) to penetrte the intct host tissue nd recolonise the host internlly upon inocultion in nutrient solution (Ppru et l. 6) nd the other reveled the cpbility of non-pthogenic F. oxysporum in colonising the lower prts of plntlets such s roots (Olivin & Albouvette 1999). The mechnism of ction ssocited with nonpthogenic F. oxysporum cn be divided into two brod ctegories: direct ntgonism of non-pthogenic strins to the pthogen nd indirect ntgonism medited through the host plnt such s induced resistnce (Fuchs et l. 1997). The sme non-pthogenic strin cn express severl modes of ction either simultneously or t different times. This is the cse of F221-B in which ntibiosis nd competition were involved nd could be observed in the in vitro experiment. However, its mechnism in hydroponics could not be concluded yet. Regrding this study, F221-B lso showed its remrkble bility s plnt growth-promoting fungus (PGPF) in ddition to its potentil in disese control. This confirmed the results of the previous work (Thongkmngm et l. 13) reporting its bility of significntly promoting the growth nd yield of Cos lettuce, Green Ok lettuce, Red Ok lettuce, Butterhed lettuce, kle, nd mung ben in hydroponics. Other strins of non-pthogenic Fusrium were reported s PGPF such s the isolte from rhizosphere soils of tomto in Indi (Ptil et l. 11), F. equiseti GF191 in Jpn (Horinouchi et l. 8, 1). Conclusion This study demonstrtes tht (i) non-pthogenic F. oxysporum (F221-B) (Thongkmngm et l. 12, 13) hs the in vitro ntgonistic bility ginst 1 plnt pthogenic fungi nd exerts direct inhibitory effect nmely ntibiosis nd competition, (ii) F221-B cn effectively reduce Fusrium root rot nd wilt of lettuce especilly on Cos, which is the most susceptible vriety with control efficcy of 85.3%, nd (iii) the newly described F221-B remrkbly promotes the growth nd yield of three lettuce vrieties grown in hydroponics twice over the helthy control especilly when it is treted in the bsence of the pthogen. Tken together, the competent non-pthogenic F. oxysporum (F221-B) hs gret potentil s promising biocontrol gent nd offers good prospect for integrted mngement of the root rot nd wilt disese of lettuce in hydroponics nd probbly for other crops s well. However, dditionl reserch on F221-B is still needed in severl res including: its colonistion pttern on plnt roots, its formultion nd pplictions; repetition of in vivo studies with other crops nd integrtion into production system; risk ssessment such s its behviour nd potentil impct on hydroponic ecosystem; nd probbly its genetic improvement. Acknowledgement. The uthors would like to thnk the Fculty of Agriculturl Technology, King Mongkut s Institute of Technology Ldkrbng for providing ll the fcilities both in lbortory nd in hydroponic greenhouse. References Abeysinghe S. (6): Biologicl control of Fusrium oxysporum f.sp. rdicis-cucumerinum, the cusl gent of root nd stem rot of Cucumis stivus by non-pthogenic Fusrium oxysporum. Ruhun Journl of Science, 1: Albouvette C. (199): Biologicl control of Fusrium wilts pthogen in suppressive soil. In: Hornby D. (ed.): Biologicl Control of Soil-borne Plnt Pthogens. Wllingford, CABI Publishing: Albouvette C., Couteudier Y. (1992): Biologicl control of Fusrium wilts with nonpthogenic Fusri. In: Tjmos E.C., Pvizs G.C., Cook R.J. (eds): Biologicl Control of Plnt Diseses. New York, Springer-US Plenum Press: Albouvette C., Lemnceu P., Steinberg C. (1993): Recent dvnce in biologicl control of Fusrium wilt. Pesticide Science, 37: Albouvette C., Lemnceu P., Steinberg C. (1996): Use of nonpthogenic Fusrium oxysporum nd fluorescent pseudomonds to control Fusrium wilts. In: Tng W., Cook R.J., Rovir A.D. (eds): Advnces in Biologicl Control of Plnt Diseses. Bejing, Chin Agriculturl University Press: Albouvette C., Steinberg C., Lemnceu P., Bkker P.A.H.M. (1998): Biologicl control of Fusrium wilt towrds the development of commercil products. In: Bolnd G.J., Kuykendll L.D. (eds): Plnt Microbe Interction nd Biologicl Control. New York, Mrcel Dekker: Benhmou N., Chrest P.M., Jrvis W.R. (1989): Biology nd host-prsite reltionships of Fusrium oxysporum. In: 93

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