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2 Egypt. J. Phytopathol., Vol. 32, No. 1-2, pp. 11-21 (2004) Preliminary Evaluation of Salicylic Acid and Acetylsalicylic Acid Efficacy for Controlling Root Rot Disease of Lupin under Greenhouse Conditions Nehal S. El-Mougy Plant Pathol. Dept., National Res. Centre, Giza, Egypt. S alicylic acid (SA) and acetylsalicylic acid (ASA) in addition to Rizolex-T were evaluated as seed dressing or soil drench against lupin root rot pathogens under greenhouse conditions. No significant reduction in seed germination was observed when lupin seeds were treated with SA and ASA up to 2 and 3 g/kg, respectively. Raising the applied dosages has reflected negatively on seed germination. SA, ASA and Rizolex-T as well as seed dressing or soil drench at the rate of 2, 3 and 3 g/kg, respectively, have significantly reduced the percentage of root rot incidence at both pre-, and post-emergence stages of lupin plants growth comparing with untreated control in artificially infested soil with Fusarium solani, Rhizoctonia solani and Sclerotium rolfsii. Seed dressing showed higher significant reduction in disease incidence than soil drench treatment. Acetylsalicylic acid as seed dressing or soil drench showed superior effect on root rot incidence followed by SA and Rizolex-T, respectively. The percentage of disease infection at these treatments where 4.0 & 12.5%; 12.0 & 22.7% and 24.0 & 31.6% at pre-emergence stage, while the recorded infection at post-emergence stage were 16.0 & 19.6%; 16.0 & 23.8% and 20.0 & 25.0% for seed and soil treatments, respectively. Furthermore, all the applied treatments significantly increased the chitinase activity in treated lupin plants grown in soil infested with F. solani, R. solani and S. rolfsii. Lupin plants treated with SA and ASA showed more activity in chitinase than Rizolex-T treatment. It is thus suggested that the usage of SA and ASA as seed dressing or soil drench could be considered as fungicide alternatives for controlling such soilborne diseases. Key words: Acetylsalicylic acid, disease control, F. solani, R. solani, root rot, S. rolfsii and salicylic acid. Root rot diseases caused by several soilborne fungal pathogens are widespread and serious in many crops cultivated in different soil types. Lupin (Lupinus termis Forsk) is subjected to attack by many pathogenic organisms wherever the crop is grown. Several root rot pathogens such as Rhizoctonia solani, Sclerotium rolfsii and Fusarium solani are reported to attack lupin roots and stem base causing serious losses in seed germination and plant stand as well (Fahim et al., 1983). An investigation for controlling such diseases is considered important, especially in view of their wide prevalence in Egypt. Several attempts to control root rot diseases could be accepted. However, fungicides are considered one of several factors involving in environmental pollution, in spite of their satisfactory results in 12 NEHAL S. EL-MOUGY the control of plant diseases. In addition, control of disease with fungicides has proven very difficult, and almost all fungicides are effective only at phytotoxic levels (Jarvis, 1988). On the other hand, the application of biological control using antagonistic microorganisms proved to be successful for controlling various plant diseases in many countries. At the same time, it still not easy and costly in application, however it can serve as a better control measure under greenhouse conditions (Sivan and Chet, 1986). Recently, the growing concern over the use of pesticides to human health and environment has brought increasing interest in the use of alternatives characterized with negative impact on the environment. Therefore, present activity focuses on finding compounds that are safe to human and environment. An alternative to fungicidal application, it may be possible to utilize a scheme of inducible plant defenses which provides protection against a broad spectrum of disease–causing organisms. Among synthetic inducers, salicylic acid (SA) and acetylsalicylic acid (ASA) have been found to be active as antimicrobial agents in various trials as disease resistance inducers. These have been reported for inducing resistance against several plant pathogens, i.e. TMV (Marrero et al., 1990) bacterial soft (El-Sayed, 1996) bacterial wilt (Abdel-Said et al., 1996) as well as soilborne fungal root rot and wilt diseases (Chen-Chunquan et al., 1999 and Mandavia et al., 2000), in addition to fungal foliar diseases (Srinivas et al., 1997). Moreover, few attempts proved the direct inhibitor effect of SA and ASA on the growth of phytopathogenic microorganisms (Dwivedi, 1990; Guo et al., 1993; Srinivas et al., 1997; Matthew and Alexander, 1999; Mandavia et al., 2000 and El-Mougy, 2002). The purpose of the present study aimed to evaluate the efficacy of salicylic acid and acetylsalicylic acid applied as seed or soil treatments for controlling root rot disease of lupin under greenhouse conditions. Materials and Methods Root rot pathogens: An aggressive isolate of Fusarium solani, Rhizoctonia solani and Sclerotium rolfsii, obtained from Plant Pathol. Dept., National Res. Centre (NRC), were used in the present study. These fungal isolates proved to be highly pathogenic to induce root rot disease on various plants in previous works (El-Kazzaz and El-Mougy, 2001 and Abdel-Kader et al., 2002). In vitro assay: The influence of SA and ASA as seed dressing or soil drench on lupin seeds germination was evaluated under in vitro conditions. a) Seed dressing: Lupin seeds (cv. Giza 2) were surface disinfected by immersing in sodium hypochlorite (2%) for 2 min, and washed several times with sterilized water, then dried between two sterilized layers of filter paper. The disinfected lupin seeds were coated with either SA or ASA at the rate of 1, 2, 3, 4 and 5 g/kg seeds. Seed dressing was carried out by applying the tested SA Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) PRELIMINARY EVALUATION OF SALYCYLIC ACID …. 13 and ASA individually to the moistened seeds in polyethylene bags and shaked well to ensure even distribution of the added chemicals. The treated lupin seeds were sown in 2x2 cm cells in polystyrene foam trays with 1x1 cm bottom holes for drainage and root emergence. Mixture of autoclaved peat-moss and sand (1:1, v/v) was used for sowing. b) Soil drench: Salicylic acid and acetylsalicylic acid were individually mixed thoroughly at different rates, i.e. 1, 2, 3, 4 and 5 g/kg of soil mixture. The drenched peat-mosssand soil was filled into polystyrene foam trays, then sown with disinfected lupin seeds. The fungicide Rizolex-T at the recommended dose (3 g/kg) was applied either as seed or soil treatment as stated before, in addition to disinfected lupin seeds sown in autoclaved peat-moss-sand soil were used as comparison treatments. Seventy two lupin seeds were sown for each particular treatment. All polystyrene foam trays were kept under laboratory conditions (18-21°C) and irrigated every two days for three successive weeks, then the percentage of germinated lupin seeds was calculated. Greenhouse experiment: Effect of SA and ASA on root rot disease incidence of lupin was studied. Pot experiment was carried out in the greenhouse of Plant Pathology Dept., NRC. The above mentioned lupin root rot pathogens were used in the present experiment. Loamy soil was artificially infested individually (at the rate of 5% w/w) with the inoculum of each fungus tested which previously grown for two weeks on sand barley medium (1 : 1, w/w and 40% water) at 25±2ºC. A set of varied infested soils were filled in plastic pots (20 cm in diameter) and sown, with relevant to the specific treatment, with disinfected lupin seeds dressed individually with SA and ASA at the rate of 2 or 3 g/kg seeds, respectively. Another set of varied infested soil was thoroughly mixed individually with SA and ASA at the rate of 2 or 3 g / kg soil weight, then filled in plastic pots (20-cmdiameter) and sown with disinfected lupin seeds. The fungicide Rizolex-T was applied to artificially infested soils, at the recommended dose, as seed dressing or soil drench at the rate of 3 g/kg seeds or soil, and used for comparison treatment. A set of disinfected lupin seeds were sown in artificially infested soils and served as general check treatment. Five lupin seeds (cv. Giza 2) were sown in each pot and five replicated pots were used for each particular treatment. Percentage of root rot disease incidence was calculated as pre- and postemergence damping-off after 20 and 40 days of sowing date, respectively. Determination of chitinase activity: The effect of usage the inducers SA and ASA as seed or soil treatment in addition to untreated control on chitinase activity was determined in lupin plants as follows: Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) 14 NEHAL S. EL-MOUGY (a) Extraction of enzyme: Plant tissues (g), relevant to each particular treatment, were homogenized in 0.2 M Tris HCL buffer (pH 7.8) containing 14mM -mercaptoethanol at the rate of 1 : 3 (w/v). The homogenate was centrifuged at 3000 rpm for 15 min. The supernatant was used to determine enzyme activity (Tuzum et al., 1989). (b) Chitinase assay: The substrate, colloidal chitin was prepared from chitin powder according to the method described by Ried and Ogryd-Ziak (1981). Twenty five grams of chitin was milled, suspended in 250ml of 85% phosphoric acid and stored at 4ºC for 24 hr, then blended in 2 liters of distilled water using a waring blender and the suspension was centrifuged. The washing procedure was repeated twice. In the final wash, the colloidal chitin suspension was adjusted to pH 7.0 with (1 N) NaOH, separated by centrifugation, then stored at 4ºC. Determination of the enzyme activity was carried out according to the method of Monreal and Reese (1969), 1 ml of 1% colloidal chitin in 0.05 M citrate phosphate buffer (pH 6.6) in test tubes, 1 ml of enzyme extract was added and mixed by shaking. Tubes were kept in a water bath at 37ºC for 60 min, then cooled and centrifuged before assaying. Reducing sugar was determined in 1 ml of the supernatant by dinitrosalicylic acid (DNS). Optical density was determined at 540nm. Chitinase activity was expressed as mM N-acetylglucosamine equivalent released / gram fresh weight tissue / 60 min. Statistical analysis: Tukey test for multiple comparisons among means was utilized (Neler et al., 1985). Results and Discussion In vitro assay: The effect of SA and ASA application as seed or soil treatment on lupin seed germination was tested under in vitro conditions. Data presented in Table (1) show that usage of salicylic acid and acetylsalicylic acid as seed dressing or soil drench up to 2g/kg had no inhibitory effect on lupin seed germination. Raising the used dosage more than 2 or 3 g/kg for SA and ASA, respectively caused significant reduction in the percentage of lupin seed germination, while no inhibitory effect was observed with Rizolex-T when applied at the recommended dose (3g/kg) as seed dressing or soil drench treatment. In this regard, many investigators recorded the effect of SA and ASA on seed germination. Sushma-Negi et al. (2001) studied the effect of different concentrations of SA and ASA on seed germination of soybean. They found that percentage of seed germination was decreased with the increasing level used of SA. Zhang-ShiGong et al. (1999) stated that wheat seeds treated with 1 g/l salicylic acid or 2 g/l aspirin increased their germination rate. They added that the addition of SA and ASA not only increase germination rate but also increase germination and activities of alphaamylase and proteinase in endosperm and their contents of soluble sugars, protein and free amino acids. Moreover, Elangovan et al. (1995) reported that treatment of seeds Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) PRELIMINARY EVALUATION OF SALYCYLIC ACID …. 15 Table 1. In vitro effect of Salicylic acid (SA) and Acetylsalicylic acid (ASA) on lupin (cv. Giza 2) seed germination Tested dose SA ASA (g/kg) Seed dressing Soil drench Seed dressing Soil drench 1 98.6 a* 97.2 a 98.6 a 98.6 a 2 95.8 a 94.4 a 97.2 a 95.8 a 3 84.7 b 81.4 b 95.8 a 94.4 a 4 81.9 b 79.2 b 83.3 b 83.3 b 5 77.8 b 76.4 b 80.5 b 80.5 b Comparison treatment: 98.6 a Rizolex-T seed dressing 98.6 a Soil drench (3g/kg) 97.2 a Check treatment * Percentage of lupin seed germination. Figures with the same letters are not significantly different (P= 0.05). with 1 and 2 mM salicylic acid induced 25% and 16% higher in seed germination, respectively, of Phaseolus aureus (Vigna radiata), while treatment with 5 mM SA inhibited the growth of seedlings. These reports are in agreement with the present data that concentrations of SA and ASA positively affect seed germination up to certain concentration, then inhibition in germination rate starts. On the other hand, several workers recorded the positive effect on germination of seeds treated with different concentrations of various fungicides (Siddique et al., 1996; Shalaby, 1997 and Vyakarnahal et al., 2000). Greenhouse experiment: The efficacy of salicylic acid and acetylsalicylic acid in addition to Rizolex-T as seed dressing or soil drench on lupin root rot incidence was evaluated in pots experiment using soil artificially infested with the disease agents under greenhouse conditions. Data in Table (2) reveal that all the tested treatments have significantly reduced the percentage of root rot incidence at both pre- and post-emergence stages of lupin plant growth comparing with the check treatment. The highest percentage of root rot infection was observed in lupin plants grown in infested soil with R. solani followed by these grown soil infested with S. rolfsii and F. solani. The corresponding disease percentages were 64.0 & 55.5%; 56.0 & 54.5% and 36.0 & 43.8% at pre- and postemergence stages of plant growth, respectively. Data also show that all treatments varied in their effect on disease incidence. Seed treatments showed higher significant reduction on disease incidence than soil treatments. In infested soil with F. solani, ASA as seed dressing or soil drench showed superior effect on disease incidence followed by SA and Rizolex-T, respectively. Disease incidence (%), for these treatments, recorded 4.0 & 12.5%; 12.0 & 22.7% and 24.0 & 31.6% at pre-emergence stage for seed and soil treatments, respectively. Meanwhile, at post-emergence stage the recorded root rot Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) NEHAL S. EL-MOUGY 16 Table 2. Percentage of lupin root rot incidence in response to seed dressing or soil drench with SA, ASA and Rizolex-T under greenhouse conditions Tested fungus Disease incidence (%) Treatment Check* SA Seed F. solani Soil ASA Seed Soil Pre-emergence 36.0 b** 12.0 e 22.7 d 4.0 f 12.5 e Post-emergence 43.8 a 16.0 e 23.8 d 16.0 e 19.6 d R. solani Pre-emergence 64.0 a 28.0 c 27.7 c 24.0 d 31.3 b Post-emergence 55.5 a 32.0 b 29.4 c 28.0 c 33.3 b S. rolfsii Pre-emergence 56.0 a 20.0 d 25.0 c 16.0 e 28.6 c Post-emergence 54.5 a 24.0 d 26.3 c 20.0 d 30.0 b * Lupin plants grown in artificially infested soil with root rot pathogens. ** Figures with the same letters are not significantly different (P= 0.05). Rizolex-T Seed Soil 24.0 d 20.0 d 8.0 f 16.0 e 16.0 e 20.0 d 31.6 b 25.0 c 26.0 c 28.5 c 28.5 c 30.0 b infection was 16.0 & 19.6%; 16.0 & 23.8% and 20.0 & 25.0% for the same treatment, in a respective order. Furthermore, in infested soil with R. solani and S. rolfsii another feature of tested treatments on lupin root rot incidence was observed. Rizolex-T occupied the first order for reduction of root rot incidence at both damping-off stages followed by ASA and SA treatments especially when used for seed dressing. It is interesting to note that, more reduction in disease incidence was observed at pre-emergence stage of plant growth than at post-emergence. This observation was also recorded by Abdel-Kader (1997) who reported that Rizolex-T as seed dressing decreased bean root rot incidence to a lower extent at postemergence stage in comparison with the biocide treatments. He concluded that this may be due to the expected degradation of fungicide when introduced into the soil and exposed to the environmental conditions. This conclusion is in harmony with the present results that treated lupin seeds with SA; ASA or Rizolex-T provide such protection to seed bed region against soilborne pathogens reflected on the observed lower disease incidence at pre-emergence stage before exposure to degradation factors. The efficacy of SA and ASA for decreasing disease incidence on various plants was recorded by many investigators. Palva et al. (1994) reported that the addition of SA to the growth medium of tobacco seedlings made them almost fully resistant to infection by soft rot bacteria. They stated that the molecular mechanism of the SA-induced resistance to Erwinia carotovora subsp. carotovora appears to involve inhibition of plant cell wall-degrading enzymes secreted by the pathogen. Okey and Sreenivasan (1996) recorded that seed treatment, plant spray and soil drench with SA reduced infection with Phytophthora palmivora of cacao. They add that these results indicated that SA induces systemic resistance in cacao and should be considered as a potential chemical for control of the pathogen. Moreover, Voget and Buchenauer (1997) reported that both SA and ASA reduced the percentage of cucumber damping off when applied as soil drench. Also, Wisniewska and Chelkowski (1999) confirmed the above mentioned reports. They stated that the application of SA as seed treatment at different concentrations from 1-10 g/l reduced the Fusarium blight disease on cucumber seedlings as 38 up to 66%. Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) PRELIMINARY EVALUATION OF SALYCYLIC ACID …. 17 These results are in a harmony with those obtained in the present study, as SA and ASA, either used as seed dressing or soil drench, had the ability to reduce lupin root rot incidence. The recorded effect might be attributed to the act of SA and ASA as plant defense inducers or to their direct effect on soilborne plant pathogens as stated by El-Mougy (2002). Dressing lupin seed or soil drench with Sa, ASA and Rizolex-T were applied to evaluate their effect on chitinase activity in lupin plants grown in soil artificially infested with root rot pathogens. Data in Table (3) indicate that all treatments have significantly increased the chitinase activity. Lupin seeds treated with SA and ASA showed more activity of chitinase enzyme than Rizolex-T treatment. In this concern, seeds treated with ASA showed superior effect followed by SA and Rizolex-T, in a respective order. These treatments increased the chitinase activity up to 8.1, 150.0 and 142.3% and to 56.9, 94.8 and 67.7% as well as to 8.1, 93.5 and 15.2% in lupin plants grown in infested soil with F. solani, R. solani and S. rolfsii, respectively. Table 3. Chitinase activity in lupin plants treated with SA, ASA and Rizolex-T as seed dressing or soil drench under greenhouse conditions Tested Fungus Chitinase measure Treatment Check * SA Seed Soil F. solani Activity 0.86a** 1.35 c 1.21 c Increase (%) -----56.9 40.6 R. solani Activity 0.75a 1.52 c 1.10 c Increase (%) -----94.8 41.0 S. rolfsii Activity 0.59a 0.99 ab 0.71 ab Increase (%) -----67.7 20.3 ASA Seed Soil 1.79 c 1.11 c 108.1 29.0 1.95 c 1.07 c 150.0 37.1 1.43 c 1.08 c 142.3 83.0 Rizolex-T Seed Soil 0.93 ab 0.89 ab 8.1 3.4 1.52 c 1.01 c 93.5 29.4 0.68 ab 0.77 ab 15.2 30.5 * Lupin plants grown in artificial infested soil with root rot pathogens. ** Figures with the same letters are not significantly different (P= 0.05). In case of soil drench, the highest increase in chitinase activity was observed in plants grown in soil infested with F. solani, R. solani and S. rolfsii then drenched with SA (being 40.6, 41.0 and 20.0 %, respectively) as more than in check treatment. These figures followed by 29.0, 37.1, 83.0% and 3.4, 29.4, 30.5% increase in chitinase activity resulted for treatments of ASA and Rizolex-T, respectively. Meanwhile, the lowest chitinase activities (being 0.86, 0.75 and 0.59%) was recorded in untreated lupin plants grown in infested soil with the same pathogens, respectively. In this regard, many investigators reported that induced resistance in various plants is associated with enhancing the activities of chitinase and ß-1,3-glucanase which hydrolyse hyphal cell wall of fungi (Matta et al., 1988). Also, Sathiyabama and Balasubramanian (1999) reported that prior treatment of groundnut leaves with salicylic acid showed a reduction in the number of rust pustules caused by Puccinia Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) 18 NEHAL S. EL-MOUGY arachidis. They concluded that enhanced activities of intercellular chitinase and glucanase of treated leaves might be associated with induced resistance responses in treated groundnut with SA against rust disease. Promising applicable technique could be suggested in the light of the results obtained in the present study. The usage of salicylic acid and acetylsalicylic acid as seed dressing or soil drench might be considered as safe, cheap and easily applied method for controlling soilborne plant pathogens taking into consideration the avoidance of environmental pollution. References Abdel-Kader, M.M. 1997. Field application of Trichoderma harzianum as biocide for control bean root rot disease. Egypt. J. Phytopathol., 25: 19-25. Abdel-Kader, M.M.; El-Mougy, Nehal S. and Ashour, A.M.A. 2002. Suppression of root rot incidence in faba bean fields by using certain isolates of Trichoderma. Egypt. J. Phytopathol., 30 (2): 15-25. Abdel-Said, W.M.; Abdel-Ghfar, N.Y. and Shehata, S.A.M. 1996 . Application of salicylic and aspirin for induction of resistance to tomato plants against bacterial wilt and its effect on endogenous hormones. Ann. Agric . Sci . Ain Shames Univ., 41: 1007-1020. Chen-Chunquan, S.; Belanger, R.R.; Benhamou, N.; Paulitz, T.C. and Chen, C.Q. 1999. Role of salicylic acid in systemic resistance induced by Pseudomonas spp. against Pythium aphanidermatum in cucumber roots. European J. Plant Pathol., 105: 477-486. Dwivedi, S.K. 1990. Antifungal activity of some phenolic compounds on Fusarium oxysporum f.sp. psidi causing guava wilt. Hindustan Antibiotics Bull., 32: 33-35. Elangovan, N.; Stevens, T. and Kalaichelvan, P.T. 1995. Effect of salicylic acid on Phaseolus aureus seed germination. J. of Ecotoxicol. and Environmental Monitoring, 5: 67-70. El-Kazzaz, A.A. and El-Mougy, Nehal S. 2001. Inheritance of Disease Resistance in Cucumber Plants to Root Rot Caused by Fusarium solani Using Tissue Culture Techniques, Egypt. J. Phytopathol., 29 (2): 57-68. El-Mougy, Nehal S. 2002. In vitro studies on antimicrobial activity of salicylic acid and acetylsalicylic acid as pesticidal alternatives against some soilborne plant pathogens. Egypt. J. Phytopathol., 30: 41-55. El-Sayed, W.M. 1996. Induction resistance to bacterial soft rot disease of potato tubers by application of acetylsalicylic acid (Aspirin). Ann. Agric. Sci., Ain Shams Univ., 41: 993-1006. Fahim, M.M.; Sahab, A.S.; Osman, A.R. and Abdel-Kader, M.M. 1983. Studies on some soilborne fungi attacking lupin plant. Egypt. J. Phytopathol., 15: 17-26. Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) PRELIMINARY EVALUATION OF SALYCYLIC ACID …. 19 Guo, D.C.; Wang, Q. F.; Yan, S.Z. and Dai, K.S. 1993. A study on a new kind of fungicide. Zhiweiling Scientia Agricultura Sinica, 26: 63-68. Jarvis, W.R.1988. Fusarium crown root rot of tomatoes. Phytoprotection, 69: 49-64. Mandavia, M.K.; Khan, N.A.; Gajera, H.P.; Andaria, J.H. and Parameswaram, M. 2000. Inhibitory effects of phenolic compounds on fungal metabolism in hostpathogen interactions in Fusarium wilt of cumin. Allelopathy J., 7: 85-92. Marrero, N.; Fernandez, T.; Caballero, O.; Rivero, M. and Lopez, M. 1990. Induction of antiviral factors in kidney beans var. Bolita 42. Ciencias- de- la Agricultura, 43: 17-21. Matta, A.; Abattista, Gentile I. and Ferraris, L. 1988. Stimulation of 1,3-glucanase and chitinase by stress that induce resistance to Fusarium wilt in tomato. Phytopathol. Medit., 27: 45-50. Matthew, E.S. and Alexander, J.E. 1999. Salicylic acid induces resistance to Alternaria solani in hydroponically grown tomato. Phytopathology, 89: 722-727. Monreal, J. and Reese, E.T. 1969. The chitinase of Servatia marcescens. Canad. J. Microbiol., 15: 689-696. Neler, J.; Wassermann, W. and Kutner, M.H. 1985. Applied Linear Statistical Models. Regression, Analysis of Variance and Experimental Design. 2nd Ed. Richard, D. (ed.). Irwin Inc., Homewood, Illinois, USA. Okey, E.N. and Sreenivasan, T.N. 1996. Salicylic acid: a factor in systemic resistance of cacao to Phytophthora palmivora. Brighton Crop Protect. Conf., Pests and Diseases, 3:955-960. Palva, T.K.; Hurtig, M.; Saindrenan, P. and Palva, E.T. 1994. Salicylic acid induced resistance to Erwinia carotovora subsp. carotovora in tobacco. Molecular Plant Microbe Interactions, 7: 356-363. Ried, J.D. and Ogryd-Ziak, D.M. 1981. Chitinase over producing mutant of Servatia marcescens. Appl. Environ. Microbiol., 41: 664-669. Sathiyabama, M. and Balasubramanian, R. 1999. Treatment of groundnut leaves with salicylic acid controls the development of rust disease caused by Puccinia arachidis Speg. Zeitschrift fur Pflanzenkrankheiten und Pflanzenschutz, 106: 166-173. Shalaby, S.I.M. 1997. Effect of fungicidal treatment of sesame seeds on root rot infection, plant growth and chemical components. Bull. Fac. Agric. Cairo Univ., 48: 397-411. Siddique, Z.S.; Soaliha-Ahmed and Ahmed, S. 1996. 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Induced systemic resistance to blue mould: Early induction and accumulation of -1,2–glucanase, chitinase and other pathogenesis proteins (b-proteins) in immunized tobacco. Phytopathology, 79: 979-983. Voget, W. and Buchenauer, H. 1997. Enhancement of biological control by combination of antagonistic fluorescent pseudomonas strains and resistance inducers against damping off and powdery mildew in cucumber. Zeitschrift fur Pflanzenkrankheiten und Pflanzenschutz, 104: 272-280. Vyakarnahal, B.S.; Shekhargouda, M. and Prabhakar, A.S. 2000. Efficacy of halogens, plant products and fungicides on storage potentiality of sunflower restorer lines. Karnataka J. Agric. Sci., 13: 36-45. Wisniewska, H. and Chelkowski, J. 1999. Influence of exogenic salicylic acid on Fusarium seedling blight reduction in barley. Acta Physiologiae Plantarum, 21: 63-66. Zhang-ShiGong; Gao-JiYin; Song-TingZhi; Zhang, S.G.; Gao, J.Y. and Song, J.Z. 1999. Effect of salicylic acid and aspirin on wheat seed germination under salt stress. Plant Physiol. Communications, 35:29-32. (Received 11/02/2004; in revised form 14/04/2004) Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004) PRELIMINARY EVALUATION OF SALYCYLIC ACID …. . - - – - . .. . 21 / / - . .. % : % % - . .. % % % . .. . Egypt. J. Phytopathol., Vol. 32, No, 1-2 (2004)