Soil Biol. Biochem. Vol. 13, pp. 75 10 76 0 Pergamon Press Ltd 1981. Printed in Great Britain

SHORT COMMUNICATION

Volatile and non-volatile metabolites of actinomycetes and the growth of some litter decomposing fungi

BHARAT RAI, ATUL K. SRIVASTAVA and D. B. SINGH Department of Botany, Banaras Hindu University, Varanasi 221005, India

(Accepted 25 July 1980)

The role of actinomycetes in organic matter decomposition and their possible contribution to microbial antagonism regulating the composition of the soil microbial com- munity have been extensively reviewed (Lacey, 1973; Goodfellow and Cross, 1974; Alexander, 1976). Volatile and non-volatile antibiotic substances produced by actino- mycetes in cultures inhibit the growth of many pathogenic soil fungi (Batra and Bajaj, 1966, 1967; Whaley and Boyle, 1967; Hora and Baker, 1970, 1972; K. P. Singh, unpub- lished Ph.D. thesis, Banaras Hindu University, 1970; Hora et al., 1971; D. Singh, unpublished Ph.D. thesis, Banaras Hindu University, 1971; Fries, 1973; Gupta and Tandon, 1977). Little is known about the effect of these substances on fungi involved in litter decomposition on the forest floor. We investigated the effect of volatile and non-volatile metabolites of six dominant soil and litter isolates of acti- nomycetes on the growth behaviour of six dominant litter- decomposing fungi.

Six actinomycetes viz., Micromonospora elongata Krassil- nikov, M. parva Jensen, Streptomyces armillatus Mancy- Courtillet and Pinnert-Sindico, S. griseolus (Waksman) Waksman and Henrici, S. griseus Waksman and Henrici and S. rochei Berger et al., were selected for our study as they were dominant in all isolations from forest soil and litter. The dominant litter-decomposing fungi selected as test organisms were Acremonium furcatum (F. et V. Moreau) ex W. Gams, Aspergillus niger van Tieghem, C/a- dosporium cladosporioides (Fresen.) de Vries, Curvularia lunata var. aeria (Batista, Lima & Vasconcelos) M. B. Ellis, Penicillium rubum St011 and Trichoderma harzianum Rifai.

To study the effect of possible volatile substances test actinomycetes were cultured separately in five replicated Petri dishes (9 cm dia) containing 20 ml Jensen’s medium (dextrose 2.0 g, Casein 0.2 g dissolved in 10 ml 0.1 N NaOH, K2HP04 0.5 g, MgS04, 7Hz0 0.2 g, FeCl, trace, Bacto- Difco agar-agar 15.0 g, distilled water 1000 ml, pH 6.5 and mycostatin 25 pg ml-’ after autoclaving). The Petri dishes were incubated at 30 k 2°C for 15 days after which the cover was replaced by a bottom plate containing Czapek- Dox + 0.05’4 yeast extract agar medium (20 ml per plate) preinoculated with the test fungus. The two plates were taped together with Sellotape to seal the dishes air tight. The covers of control plates which had not been inoculated with actinomycetes were also replaced in the same way. Colony diameters of the test fungi were measured (mm) after 3 days at 25°C in diffused light (fluorescent tubes).

To determine the effect of non-volatile substances, the actinomycetes were cultured individually in 250 ml conical flasks each containing 1OOml sterile Jensen’s liquid medium; cultures were incubated at 30 f 2°C for 15 days. Thereafter, cultures were filtered through a bacterial filter (Seitz filter). Twenty ml filtrate of each isolate was poured into a conical flask containing 80ml melted and cooled (35°C) Czapek-Dox + 0.05% yeast extract agar medium maintaining the treatment at 20% concentration, shaken well and poured into sterile Petri dishes. The same amount of sterile Jensen’s liquid medium was added to Czapek-

Dox + 0.05% yeast extract agar medium as a control. Agar blocks 5 mm dia. of the test fungi, cut from the margins of actively-growing colonies, were placed centrally in all the treated as well as control plates and incubated at 25°C for 3 days and the radial growth measured.

A 5% inhibition or stimulation of growth was considered to be due to variation in growth of the test fungi in colony diameter in control plates. Any variation greater than this was thought to be caused by volatile and non-volatile sub- stances of antinomycetes tested.

Results show that the volatile and non-volatile metab- olites of the six test actinomycetes were more or less inhibi- tory to the growth of A. niger, P. rubrum and T. .harzianum and stimulatory to C. lunata var. aeria (Table 1).

The volatile metabolites of both Micromonospora sp. did not affect any of the test fungi except A. niger where growth was inhibited slightly by M. parva. Volatile metab- olites of the four species of Streptomyces slightly inhibited the mycelial growth of Acremonium furcatum, A. niger, Cladosporium cladosporioides, P. rubrum and ‘I: harzianum. However, S. griseolus produced maximum inhibition of ?Y harziunum (Table 1). The growth inhibition or stimulation of the fungi was presumably caused by the production of volatile substances (McCain, 1966; Batra and Bajaj, 1966, 1967; Whaley and Boyle, 1967; Gerber, 1968; Hora and Baker, 1970; Hora et al., 1971), each species did not respond in a uniform manner and each fungus shows variable sensitivities to the products of the different actinomycetes.

Non-volatile metabolites of M. parvu and M. elongate inhibited the growth of A. niger, P. rubrum and ‘I: harzio- num but stimulated the growth of A. furcatum, C. cladospor- ioides and C. lunatn var. aeria. The minimum (17%) and the maximum (30%) inhibition was observed in the case of A. niger and 7: harzianum respectively whereas the minimum (10%) and the maximum (79%) stimulation in case of C. cladosporioides and A. furcatum respectively and were caused by the metabolites of M. parva. Non-volatile metab- olites of all the four test species of Streptomyces caused inhibition in the growth of A. niger, C. cladosporioides, P. rubrum and 17: harzianum. However, the growth of A. furca- turn was inhibited only by S. armillatus and S. griseolus. The minimum growth inhibition (6%) was for C. cladospor- ioides caused by the metabolite of S. armillatus and maxi- mum (42%) for Z harzianum due to S. griseolus. Growth of C. lunata var. aeria was stimulated slightly due to metab- olites of all the four Streptomyces spp. Growth of A. furca- turn was significantly stimulated (80%) due to the metab- olite of S. rochei.

The growth behaviour of the test fungi on the nutrient medium supplemented with the metabolites of different species of actinomycetes was not similar. The degree of effect might vary according to the nature, quality and quantity of the antibiotics or growth-promoting substances produced by actinomycetes. However, relatively low inhi- bition and rather higher stimulation in the growth of test fungi on nutrient media supplemented with non-volatile

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metabolites might be partly due to a masking effect pro- duced by sufficient nutrients in the supplemented medium. The exact nature of antibiotic and growth-promoting sub- stances produced by the actinomycetes has not yet been determined and needs investigation. Our results suggest Streptomyces spp to be more active antagonists than Mic- romonospora spp.

Acknowledgemenrs-We thank the Head of the Depart- ment of Botany for providing laboratory facilities. AKS and DBS thank the University Grants Commission and Council of Scientific and Industrial Research, New Delhi respectively for financial assistance.

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