Plant and Soil 8 7 , 4 3 5 - 4 3 8 (1985). Ms. 5865 �9 1985 Martinus Ni/hoffPublishers, Dordrecht. Printed in the Netherlands.

Biocontrol of Macrophomina root.rot disease of jute by an

antagonistic organism, Aspergillus versicolor

D. BHATTACHARYYA, S. BASU, J. P. CHATTAPADHYAY and S. K. BOSE Department of Biochemistry, University College of Science, 35, Ballygunge Circular Road, Calcutta 700 019, India

Received 20 March 1984. Revised November 1984

Key words Antagonist Aspergillus versicolor Biocontrol Enti-soil Macrophomina phaseolina

Summary Aspergillus versicolor was grown in soil-compost medium at pH 4.0 for 10 days under diffused light to get A. versicolor-grown soil which was found to control experimental infection in jute by Macrophomina phaseolina to the extent of 56% in pot-culture experiment.

Introduction

Many pathogens like Rhizoctonia solanL Macrophomina phaseolina, Pythium spp. etc. live in soil and may be responsible for a soil-borne infection. Various attempts have been made in the recent past to combat soil-borne infection by applying microorganisms instead of microbial products. Bacillus subtilis protected barley from seedling blight caused by Helmin- thosporium sativum in the field ~ and Chaetomium globosum protected oats from victoriae blight caused by H. victoriae 11 . Thirumalachar and O'Brien reported the control of charcoal- rot of potato caused by Macrophomina phaseolina and Botryodiplodia solani tuberosi by applying antagonistic bacteria 1~ Antibiotics might be responsible for these instances of bio- control, although their production in soil could not be easily demonstrated 2,3,6. The effec- tiveness of organic amendments has been observed in controlling root-diseases including root- rot of strawberry with soybean residues *,~2. Phymatotrichum root-rot of cotton with organic manure s and Fusarium root-rot of bean with straws of high C:N ratio 9. Biological control of soil-borne infection of jute has received much less attention than biocontrol of other soil- borne infections. The present paper aims at exploring the possibilities of biological control of root-rot disease of jute caused by Macrophomina phaseolina.

Materials and methods

Organisms Aspergillus versicolor, producer of mycoversilin, was isolated in this laboratory 7. A strain

of Macrophomina phaseolina (Maubl.) Ashby was obtained through the courtesy of Jute Agricultural Research Institute, Barrackpore, West Bengal, India.

Jute seed Jute seeds of cultivar JRO-632 which are susceptible to Macrophomina phaseolina causing

root-rot and stem-rot diseases of jute, were collected from National Seeds Corporation of India.

Organic manure Organic manure was collected from Globe Nursery, Calcutta, India.

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Table 1. Effect of compost on the biological control of root-rot disease of jute seedlings in soil preinoculated with A. versicolor

Concentration Disease index* Inhibition of compost pH of infection (%) unadjusted Control Experimental (%)

12.5 5.3 2.34 • 0.026 1.40 • 0.041"* 39.99 10.0 5.5-5.6 2.60 -+ 0.088 1.22 • 0.051"* 52.72

7.5 6.0 2.39 • 0.068 1.27 • 0.093** 46.28 0.0 6.8 3 3 0

* Results are expressed as mean -+ SEM of five different experiments. ** Significantly different from control, p < 0.001.

Soil Soil was collected from Kalinarayanpur, a jute growing area of West Bengal, India.

Preparation of soil-compost medium The soil compost medium contained per 100 ml. Eighty gms of soil and compost in variable

proportions as indicated in the text. The pH of the medium was recorded or adjusted as per requirements and then autoclaved at 121~ for 1 h.

Experiment on biological con trol The soil compost medium was inoculated with the antagonist. The inoculum consisted

of spores harvested from Sabouraud's agar slants and freed from mycelial fragments by cotton filtration. The medium so inoculated was incubated at 25 • 2~ for 10 days, then taken out and dried over aluminium trays to get a semisolid cake which was then infested by spreading on the soil surface sclerotial plus mycellal suspension of Macrophomina phaseolina. The in- festing suspension so used was obtained by culturing the pathogen in potato-dextrose-broth for 4 - 5 days at 27 -+ 2~ to obtain a fungal biomass which was ground with the same volume of sterile water in a Waring blender for 30 sec. Two days' old jute seedlings were planted in porcelain pots containing soil and grown for 15 days under greenhouse condition. The seedlings were then removed from the pots and the roots washed in running water to remove adhering soil particles. Extent of infection by M. phaseolina was {hen determined. Control experiments were run side by side.

Determination of disease index The disease index used in this case was a modification of Cralley's system of disease

measurement. The extent of infection by M. phaseolina was indicated by the presence of dark brown lesion and also by the presence of micro-sclerotia of the fungus on root-systems. Healthy and infected seedlings were divided into four groups as follows:

1. Healthy seedlings = 0 - No root-rot symptom 2. Slightly infected = 1 - Slight root-rot symptom

seedlings 3. Heavily infected = 2 - Heavy root-rot symptom

seedlings 4. Dead seedlings = 3

Disease index was calculated as follows:

D.I. = O(Hn) + 1 (S n) + 2 (H n*) + 3 (D n) Total number of seedlings examined

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where H n = Number of healthy seedlings S n = Number of slightly infected seedlings H n* = Number of heavily infected seedlings D n = Number of dead seedlings

Results and discussion

Effect of compost on the biological control of root-rot disease of jute seedlings in soil preinoculated with A. versicolor

It appears from the results (Table 1) that the inhibition of infection was maximum i.e., 52.72 + 2.507 at 10% compost and this increased value of inhibition was significantly different from other values as confirmed by statistical analysis. However, the disease indices in case of A. versicolor inoculated and non-inoculated soil was 3 in both the cases where the soil was not enriched with compost. Thus the presence of organic matter particularly amino acids of com- post including aspartic acid which was reported to support maximum production of myco- versilin 8 was responsible for the increased value of inhibition of infection.

Effect o flight on the biological control of root-rot disease of lute seedlings in soil preinoeulated with A. versicolor

The disease indices for A. versieolor inoculated and non-inoculated soil exposed to strong light were 1.73 + 0.0307 and 2.2 + 0.0209 respectively. So, inhibition of infection was 21.07 + 1.238% in the presence of strong light. The disease indices for A. versicolor inoculated and non-inoculated soil exposed to diffused light were 1.19 +- 0.062 and 2.47 + 0.077 respec- tively. So, inhibition of infection was 51.8 + 2.146% under exposure to diffused light. So, diffused light is superior in this respect.

Effect of pH of the growth medium of A. versicolor on effectiveness of disease inhibition

The disease indices in A. versicolor inoculated soil whose pH was initially adjusted to 4.5, 4.0 and 3.5 were 1.27 + 0.019, 1.07 + 0.052 and 1.41 +- 0.03 and the corresponding figures for non-inoculated control were 2.42 + 0.03, 2.46 + 0.107 and 2.38 _+ 0.026 respectively. So, the infection was inhibited to the extent of 47.37 +- 1.282, 56.65 _+ 0.916 and 40.72 + 1.485% (as confirmed statistically not shown in the table) for pH values 4.5, 4.0 and 3.5 respectively, the opt imum pH for inhibition being 4.0. It might be that mycoversilin pro- duction which was reported to be maximum at pH 4.0 might account for effective biological control at that pH 8.

Effect of incubation period of A. versicolor for the effectiveness of disease inhibition

The disease indices in A. versicolor inoculated soil which was incubated for varying periods viz. 8, 10, and 12 days were 1.39 + 0.04, 1.1 + 0.054 and 1.27 + 0.02 and the corresponding figures for non-inoculated control were 2.44 + 0.06, 2.52 + 0.081 and 2.49 + 0.055 respec- tively. So, infection was inhibited to the extent of 42.76 + 1.013, 56.39 + 1.691 and 48.404 +_ 0.845% for the incubation periods 8, 10 and 12 days respectively, the opt imum period of incubation being 10 days. Normally 8 - 9 days of incubation were required for opt imum pro- duction in a glucose-peptone broth s which in case of soil compost medium was extended to 10 days probably due to nutritional deficiency of this medium.

Effect of elimination of drying of A. versicolor-grown soil compost medium on the effectiveness of disease inhibition

The disease index was 1.404 + 0.03 for A. versicolor-grown soil-compost medium whose moisture content was initially kept sufficiently low to avoid drying for subsequent use in biological control experiments and the corresponding figure for non-inoculated control was

438 SHORT COMMUNICATION

2.448 -+ 0.055. So, the infection was inhibited to the extent of 42.56 _+ 1.654% as against 53.23 • 0.599% when soil compost medium contained enough water to be dried for subsequent use. It might be that mycoversilin production was not favoured in soil until it was diluted.

Thus under an optimal combination of these parameters the biological control of experi- mental infection to the extent of 56% appears feasible.

Acknowledgement Thanks are due to Indian Council of Agricultural Research, Krishi Bhavan, New Delhi, India, for financial assistance.

References

1 Anwar A A 1949 Phytopathology 39, 1005-1019. 2 Brian P Wet al. 1951 Nature London 167 ,347-349 . 3 Gottlieb D e t al. 1952 Phytopathology 42 ,493 - 496 . 4 Hildebrand A A and West P M 1941 Can. J. Res. (C) 19 ,183-198 . 5 King C J e t al. 1934 J. Agric. Research 49, 1093-1107. 6 Martin M and Gottlieb D 1955 Phytopathology 45 ,407 -408 . 7 Samanta A K e t al. 1983 Ind. J. Exptl. Biol. 21 ,577-578 . 8 Samanta A K and Bose S K 1984 J. Antibiotics 37 ,728-732 . 9 Snyder W C et al. 1959 Phytopathology 49 ,755 - 756 .

10 Thirumalachar M J and O'Brien M J 1977 P1. Dis. Reptr. 61 ,543 -546 . 11 Tveit M and Moore M S 1954 Phytopathology 44 ,686 -689 . 12 West P M and Hildebrand A A 1941 Can. J. Res. (C) 19 ,199-210 .