EFFECT OF YEAST EXTRACT AND SUCCINIC ACID ON GROWTH AND S P O R U L A T I O N OF ALTERNARIA SPECIES by N. R. RAJDERKAR
Department o/ Botany, Marathwada University, Aurangabad (19.111.1965) In an earlier study it was shown that Alternaria solani (ELL. & Martin) JONES & GROUT. is able to use diverse types of nitrogen compounds. The ability to use ammonium, nitrate and organic nitrogen justifies its inclusion in the second group of the ~Robbinsclassification. The essentiality of nitrogen in the metabolism of fungi has long been recognised. The present paper deals with the nitrogen nutrition of Alternaria solani (ELL. & ~{ARTIN)JONES & GROUT. and A. nyctanthi DESHPANDE, K. ]3. & N. R. RAJDERKAR, spec. nov. and to investigate the effect of yeast extract and of succinic acid on the utilization of ammonium nitrogen during growth and sporulation. MATERIAL AND METHOD During the course of the studies 10 isolates of the genus Alternaria were treated in details. The specimens were collected from different hosts and the isolations, purification, etc. were done by the usual mycological laboratory method. Single spore cultures of A. solani (ELL. & MARTIN) JONES & GROUT. and A. nyctanthi DESHPANDE, K. ]3. & N. R. RAJDERKAR were used throughout this study. Cultures were grown on several media: Richard's medium, Coons medium, Dox's agar, Potatodextrose-agar(PDA), yeast extract with the usual formula glucose 1%, 1% peptone, 0.3% yeastreI, 2% agar and distilled water to make up 1 litre and yeast extract slightly modified. Both species of Alternaria, A. solani and A. nyctanthi showed good growth on PDA and yeast extract at 27 ° to 28 ° C. The general description of each isolate on different media was recorded after the dish was incubated at 26 ° to 28 ° C for 7 days. Even though the
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fungus made good growth on yeast extract and PAD at a particular range of temperature, it did not sporulate freely, unlike many species of Alternaria. The ability to sporulate diminished progressively with successive sub-culturing, but appeared to revive with the application of proper nitrogen sources and stimuli such as exposure to sunlight and freezing. The e//ect o/succinic acid on the fungus was pronounced. Organic acids, especially the four carbon dicarboxylic acids, are known to stimulate the utilization of ammonium nitrogen. Yeast extract also has marked effect on increased sporulation. Succinic acid was added to the basal medium together with the ammonium salts. Since poor sporulation occurred in the yeast extract medium when urea, asparagine glutamic acid, ammonium nitrate or ammonium sulphate was used as the source of nitrogen, the effect of addition of yeast extract was investigated. The resttlts are given below in Table I. TABLE I.
E[[ect o[ yeast extract on the growth and sporulation o~ Alternaria species after 10 days. Alternaria solani Compounds
Alternaria nyctanthi
Dry wt. mg
pH
Sporulation
Dry wt. mg
pH
Sporutation
48.3
5.4
poor
45.5
5.5
poor
52.2 55.0
6.8 6,5
poor fair
50.5 58.8
6.8 6.5
poor fair
yeast extract
65,5
6.4
good
66.8
6.4
good
Glutamic acid GIutamic acid and yeast extract A m m o n i u m nitrate Ammonium nitrate and yeast extract
68.5
6.6
good
68.5
6.5
good
72.6 45.5
6.8 4.8
good poor
70.0 45.8
6.5 4.8
good poor
55.5
5.4
good
52.4
5.2
good
50.9
5.2
good
50.5
5.4
good
Urea Urea and yeast extract Asparagine and
Aspa~agille and
Ammonium sulphate and yeast extract
It is evident fl'om the above results that yeast extract increases sporulation. Succinic acid was added to the basal medium together with the ammonium salts. Growth and sporulation were evaluated and cited in Table II. It is evident that in the presence of succinic acid good sporulation occurred with each of the ammonium salts and growth was enhanced. A chromatographic study of the assimilation rate of those amino acids which were present, either in tile free form or in the protein
E F F E C T OF Y E A S T AND SUCCINIC ACID ON A L T E R N A R I A
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TABLE II.
E//ect o/ succinic acid and ammonium salts on the growth and sporulation o/Alternaria sp. Alternaria solani
.Compounds
A m m o n i u m nitrate A m m o n i u m nitrate and succinic acid A m m o n i u m chloride A m m o n i u m chloride and succinic acid A m m o n i u m sulphate A m m o n i u m sulphate and succinic acid
Alternaria nyctanthi
Dry wt. mg
Sporulation
pH
Dry wt.
Sporulation
pH
45.6
poor
4.8
40.8
poor
4.8
65.5 55.6
good poor
6.4 4.8
60.8 52.0
good poor
6.4 4.6
78.5 48.5
good poor
6.8 4.2
75.0 48.5
good poor
6.8 4.2
85.8
good
6.8
85.5
good
7.0
complex; was under taken. Glutamic acid, aspartic acid, leucine etc. and their mixtures were used. The maximum growth was obtained on a mixture of the vaious amino acids. DISCUSSION
Alternaria solani (ELL. & MARTIN)JONES & GROUT. and A. myctauthi D E S H P A N D E K . B . & N . R . ]~AJDERKAR, these pathogens were isolated from the concerned host and grown on different culture media. The isolations, purification etc. were done b y the usual mycological laboratory methods. Fungi rarely grow appreciably on nitrites, however, there are conflicting opinions about whether the poor growth of an organism on ammonium nitrogen is a true character of the organism or is brought about by other factors. In the present investigation the yeast extract addition had a pronounced affect on growth and sporulation. Addition of succinic acid to ammonium sources induced good sporulation and growth.
Summary Alternaria solani and A. uyctanthi, these pathogens causing leafspot disease were able to metabolize a variety of nitrogen compounds when grown on different culture media. The amount of growth varied with the nitrogen source. Peptone produced the best zonation when added in definite proportion to the yeast extract medium. Ammonium compounds were found to be moderately effective for growth but poor for sporulation. The effect of adding succinic acid in media containing ammonium sources and the role of pH in the utilization of nitrite nitrogen was investigated. The fungus gave more vegetative growth on a mixture of aminoacids than in culture media in which the same amino acids were
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supplied singly to study the effect produced on growth and sporulation.
Acknowledgements Grateful thanks are due to Professor PAUL J. ALLEN of the University of Wisconsin and to Professor M. N. KAMAT, Head of the Mycology Section M.A.C.S., Poona for helpful suggestion and providing the necessary literature. The author is also grateful to Dr. L. B. KA]ALE for absorbing interest, to Mir IQBAL ALl BILGRAMI and to Marathwada University for laboratory facilities.
References LEwis, R. W. 1957. Amino acid nutrition of Al/ernaria solani. Phytopathology 47: 121. LILLY V. G. & BARNETT g . L. 1951. Physiology of Fungi. McGraw-Hill, New York. RANGASWAMI, G. & SAMBANDAM, C. ~ . 1960. Influence of the substrate on spore size of Alternaria melongenae. Phytopathology 50: 486. RAJDERKAR,~T. R. 1966. The influence of nitrogen nutrition on growth and sporulation of Alternaria solani (Ell. & Mart.) Jones & Grout. Mycopathol. et Mycol. appl. 29 : 55--58.
