Some Chemical Aspects of Plant Disease Resistance - American

2 Some Chemical Aspects of Plant Disease Resistance R. L. Wain

Downloaded by UNIV LAVAL on July 11, 2016 | http://pubs.acs.org Publication Date: January 16, 1986 | doi: 10.1021/bk-1986-0296.ch002

Department of Chemistry, University of Kent, Canterbury, England

Growing plants are exposed to pressures of environment and disease, and without built-in defense mechanisms, they could not survive. They protect them selves against stresses such as drought by producing a chemical which reduces both the rate of growth and leaf transpiration. Plants can also exert chemical defenses against fungal pathogens. Some of the chemi cals which operate occur normally in plants; others build up in the tissues following an attempted invasion by the fungus. Like chlorophyll, all these chemicals have arisen over thousands of years of plant develop ment, and unlike manmade systemic fungicides, they never lose their effectiveness. Their presence in plants helps to explain the wide-spread natural resistance shown towards disease. Isolation and iden tification of these naturally occurring fungicides is now a routine procedure and such chemicals from a resistant plant can sometimes be used to protect a susceptible species from disease attack. Whereas animals can take shelter, plants remain i n the same place, no matter how unfavourable environmental conditions may be. It i s not surprising then that throughout their evolutionary history, plants have developed their chemistry to protect themselves against environmental stresses such as drought, waterlogged s o i l , and s o i l s a l i n i t y . There i s now evidence that they do this by bioeynthes l s l n g r e l a t i v e l y large amounts of the endogenous plant-growth hor mone i n h i b i t o r a b s c i s i c a c i d . This then r e s t r i c t s the a c t i v i t i e s of

CH CH

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CHO Abscisic acid

0097-6156/86/0296-Ό016S06.00/0 © 1986 American Chemical Society

Green and Hedin; Natural Resistance of Plants to Pests ACS Symposium Series; American Chemical Society: Washington, DC, 1986.

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the growth hormones which are responsible for promoting extension growth and c e l l d i v i s i o n i n plants. In this way, growth may v i r t u a l l y cease, thereby conserving the plant's energy during the c r i t i c a l stress period. The marked increase i n the a b s c i s i c acid l e v e l which can occur during water stress was f i r s t observed by Wright i n my laboratory (l_ 2)« In a t y p i c a l experiment, the a b s c i s i c acid l e v e l i n the leaves of a tomato plant was shown to Increase over f i f t y f o l d when water was withheld for three days. It was l a t e r shown elsewhere that not only does this a b s c i s i c acid r e s t r i c t the plant's growth but Its presence In the leaves leads to a closure of the stoma so that water lose by t r a n s p i r a t i o n Is decreased (3). Turning now to plant diseases, we know that although a l l plants are exposed to a very wide range of p o t e n t i a l l y pathogenic fungi, they are completely resistant to most of them. Indeed It Is true that In general, under natural conditions, resistance i s the r u l e . There are many ways i n which a plant can r e s i s t fungal i n f e c t i o n , but i t i s now well established that amongst these, chemical defense mechanisms are Important. That f u n g i c i d a l compounds are present i n the washings of healthy leaves of a number of plant species was demonstrated by Topps and myself (4_). Sakural, also working i n my laboratory, has shown that an a c i d i c f u n g i c i d a l substance, not yet i d e n t i f i e d , can be obtained from the washings of healthy leaves of V i c i a faba. Such compounds then could be a factor i n determining whether the spores of some fungi can germinate on the leaf surface. A well recognised defense mechanism against a p o t e n t i a l pathogen i s the plant c u t i c u l a r layer which may provide a b a r r i e r to i n f e c t i o n to c e r t a i n fungi. I f , however, a germinating fungal spore does penetrate into the plant c e l l s , It must f i n d suitable n u t r i t i o n a l and other conditions for successful establishment. In some cases, t h i s establishment may f a l l owing to the rapid death of the host tissue c e l l s at the s i t e of i n f e c t i o n so that here, hypers e n s i t i v i t y of the host plant could be a primary cause of resistance. Research in recent years has shown that protective chemicals i n plants can operate i n disease resistance. Some of these chemicals occur i n disease-free plants, but f u n g i c i d a l compounds can build up i n tissues i n response to attempted i n f e c t i o n . When this occurs, these defense chemicals are known as phytoalexins. Papers to be presented at this conference by Cutler et_ a_l., Harborne, Bailey, B e l l et^ a l . and Gustine w i l l deal with recent developments i n research on phytoalexins. E a r l i e r research on phenolic substances i n r e l a t i o n to plant disease resistance was concerned not only with the possible protect i v e e f f e c t s of preformed phenolic compounds, but also whether these compounds are mobilized or their synthesis i s promoted at the s i t e of i n f e c t i o n . Another aspect which has received attention i s that the phenolic compound might be liberated from i t s glycoside or sugar ester at the i n f e c t i o n s i t e i n response to i n f e c t i o n (5_)» Chemical changes In Infected tissue due to the a c t i v i t y of polyphenoloxidases and peroxidases, leading to the production of quinones and other f u n g i c i d a l compounds, have also been Investigated (6_). The p o s s i b i l i t y that f u n g i c i d a l action may a r i s e from the l i b e r a t i o n of an aglycone from i t s glycoside within the tissues of the fungus provides an approach to systemic fungicides which i s f


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being followed i n my laboratory. Well known synthetic fungicides, not themselves systemic, are converted to their more water soluble β-D-glueosides which are more able to move within the plant. L i b e r a t i o n and metabolism of the sugar from the glucoside by the plant i s thought to be unlikely since adequate carbohydrate i s a v a i l a b l e from the phytosynthetic pathway. The pathogen, however, which i s e n t i r e l y dependent on supplied carbohydrate, might well adapt i t s e l f to hydrolyze the glycoside, thereby releasing the f u n g i c i d a l aglycone. The l i t e r a t u r e on phytoalexins i s now very extensive as Dr. Harborne w i l l r e l a t e i n his paper. These fungitoxic substances which are produced i n plants i n response to attempted fungal inva sion are thought to arise from an i n t e r a c t i o n between s p e c i f i c meta b o l i c systems of the host and the fungus. But although phytoalexins are synthesised and accumulated by plant tissues i n response to the presence of fungi, phytoalexln formation can be stimulated by other means. Thus, chemical, mechanical and heat treatment, and anaerobic storage of host plant tissues (7_,8) have a l l been shown to promote the synthesis of phytoalexins. A l l these "stress" factors then, both b i o l o g i c a l and non-biological, can I n i t i a t e the changes i n host plant metabolism upon which phytoalexln formation depends. This suggests that the production of phytoalexln might arise from a s h i f t i n an already e x i s t i n g biosynthetic mechanism; indeed, the i n i t i a t i o n of a completely new synthetic pathway involving s p e c i f i c enzymes i s most u n l i k e l y as i t would require the operation of new genes. It would be l o g i c a l , therefore, to expect traces of the phy t o a l e x l n to occur In the normal, uninfected plant together with substances which can y i e l d the phytoalexln i n response to the a c t i vation or suppression of enzyme reactions which are already established within the host. Albersheim has developed a concept of phytoalexln production being promoted by s p e c i f i c " e l i c i t o r s " - that i s , by molecules pro duced by certain fungi, which can sometimes be found i n fungal culture media (9). Such e l i c i t o r s have been Isolated and t h e i r role i n the defensive process has been investigated (10). The resistance of plants towards disease may well be related to phytoalexln formation, but whatever be the precise function of these compounds, they can only represent one of the complex of factors which operate i n disease resistance and immunity. Any protective substances present i n the healthy plant must also be important i n t h i s connection. A good example of such a substance i s provided by our discovery that healthy seedlings of broad bean ( V i c i a faba) contain a potent antifungal chemical to which we have given the name Wyerone. Our f i r s t i n d i c a t i o n that Wyerone occurs i n broad bean plants arose from an observation that fungal growth on nutrient sugar was

Wyerone



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i n h i b i t e d by the presence of segments of the stem or root tissue (11)· The f u n g i c i d a l compound present i n healthy seedlings was i s o lated, i t s structure and s t e r i c configuration were established and the substance (Wyerone) was also synthesised (12,13). Wyerone was found to show a wide f u n g i c i d a l spectrum when tested against phytopathogens i n spore germination tests (14). It showed high i n v i t r o a c t i v i t y , p a r t i c u l a r l y against A l t e r n a r i a b r a s 8 i c i c o l a ( E D 5 0 3 ppm). However, i t was some 40 times less active against B o t r y t i s cinerea ( E D 5 0 125 ppm). This i s of considerable i n t e r e s t since A. b r a s s i c i c o l a does not a f f e c t broad bean plants whereas B o t r y t i s spp. are the pathogens which cause the well known chocolate spot disease. Wyerone levels i n V i c i a faba have been shown to increase i n response to attempted fungal invasion so i n this sense, Wyerone, although i t occurs i n healthy tissues must also be considered as a phytoalexln. Wyerone acid has been i s o l a t e d from bean infected tissues (15) and this f u n g i c i d a l acid ( i n which the COOCH3 group of Wyerone has been hydrolysed to - C 0 0 H ) must also be considered as a substance which protects against disease. The detection and i s o l a t i o n of phytoalexins and other naturally occurring fungicides has been greatly s i m p l i f i e d using the technique whereby plant tissue extracts are subjected to t h i n layer chromatography a f t e r which the developed plate i s sprayed with spores of an appropriate fungus suspended i n a nutrient s o l u t i o n . On incubating the plates, the darkly pigmented fungus grows over the plate except i n the areas where the antifungal compounds are situated. Using this technique, we found another source of naturally occurring fungicides. This came from a speculation on why plant roots growing i n s o i l usually remain healthy although they are exposed to a wide range of bacteria and fungi which can readily destroy dead root t i s s u e . Such considerations led us to investigate the chemicals exuded by pea and bean seedling roots into the water i n which the roots were growing. It was found i n both cases that antifungal compounds were present (16). Thus i t i s clear that the l i v i n g root operates to defend i t s e l f i n the h o s t i l e environment of the s o i l . Amongst the known phytoalexins, one finds considerable variations i n chemical structure and some of these naturally occurring fungicides taken from one plant have been shown to protect other plants against fungal attack. This has been shown for example with the solanaceoue phytoalexln c a p s i d i o l which i s highly active against Phytophthora infestans. When the compound was applied to tomato plants at 100 ppm p r i o r to inoculation with P^. i n f estans, i t provided almost complete protection ( 1 7 ) . Another example of the use of a naturally occurring fungicide i n plant protection i s provided by 'sclareol' - an epimeric mixture of s c l a r e o l and 13-episclareol. We have shown that this mixture of diterpenes which occurs on the surface of healthy leaves of tobacco (Nicotiniana glutlnosa) w i l l prevent the germination of rust spores at low concentrations (18). This finding led us to examine i t s use as a rust fungicide. It was then found that when i t was applied as a spray to broad bean and dwarf bean at 100 ppm, i t provided almost complete protection against certain rust diseases (19). These examples i l l u s t r a t e possible a g r i c u l t u r a l uses for these organic fungicides which occur naturally within the plant kingdom. Such compounds can often be synthesised as has been done, for


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example, with p i s a t i n (20) Wyerone (13) r i s h i t i n (21) vignafuran (22) and orchinol (23). Furthermore, the approach can also be used to prepare analogues closely related to the naturally occurring fungicide. Not only might these be of a g r i c u l t u r a l s i g n i f i c a n c e , but they might be of value i n c o n t r o l l i n g fungal pathogens of man and animals. Fungal infections of the eye, f o r example, although not common are extremely d i f f i c u l t to treat with e x i s t i n g fungicides because these chemicals are often i r r i t a n t or toxic to the délicates of the eye. Such cases of keratomycosis might however be more succ e s s f u l l y treated with naturally occurring fungicides and these p o s s i b i l i t i e s are now being investigated.


Literature Cited 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. 12. 13. 14. 15. 16. 17. 18. 19. 20. 21.

Wright, S. T. C. Planta, 1969, 86, 10. Wright, S. T. C.; Hiron, R. W. P. Nature, Lond., 1969, 224, 719. Jones, R. J.; Mansfield, T. A. J. Exp. Bot., 1970, 21, 714. Topps, J. Α.; Wain, R. L. Nature, London, 1957, 179, 652. Flood, A. E.; Kirkham, D. S. In Phenolics in Plants in Health and Disease; J. B. Pridham ed.; Pergamon Press, Oxford, 1960. Goodman, R. N.; Kiraly, Z.; Zaitlin, M. Biochemistry and Physiology of Infectious Plant Disease; van Nostrand Co., Princeton N.J., 1967, p. 187. Cruickshank, I. A. M.; Perrin, D. R. Aus. J. Biol. Sci., 1963, 16, 111. Perrin, D. R.; Cruickshank, I. A. M. Aus. J. Biol. Sci., 1965, 18, 803. Ayers, A. R.; Ebel, J.; Valent, Β.; Albersheim, P. Plant Physiol., 1976, 37, 751; 760; 766. Ebel, J.; Ayers, A. R.; Albersheim, P. Plant Physiol., 1976, 37, 775. Spencer, D. M.; Topps, J. H.; Wain, R. L. Nature, Lond., 1957, 179, 651. Fawcett, C. H.; Spencer, D. M.; Wain, R. L.; Jones, Sir Ewart; Le Quan, M.; Page, C. B.; Thaller, V. Chem. Comm., 1965, 1, 422. Fawcett, C. H.; Spencer, D. M.; Wain, R. L.; Fallis, A. G.; Jones, Sir Ewart; Le Quan, M.; Page, C. B.; Teller, V.; Shubrook, D. C.; Witham, P. M. J. Chem. Soc., 1968, p. 2455. Fawcett, C. H.; Spencer, D. M.; Wain, R. L. Neth. J. Plant Path., 1969, 75, 72. Deverall, B. J. Proc. Symp. Phytochem. Soc., 1971, p. 217. Burden, R. S.; Rogers, P. M.; Wain, R. L. Ann. Appl. Biol., 1974, 78, 59. Ward, E. W. B.; Unwin, C. H.; Stoessel, A. Phytopath., 1975, 65, 168. Bailey, J. Α.; Vincent, G. G.; Burden, R. S. J. Gen. Microbiol., 1974, 85, 57. Bailey, J. Α.; Carter, G. Α.; Burden, R. S.; Wain, R. L. Nature, Lond., 1975, 255, 328. Bevan, C. W. L.; Birch, A. J.; Moore, B.; Mukerjee, S. K. J. Chem. Soc., 1964, p. 5991. Katsui, N.; Matsunaga, Α.; Imazumi, K.; Matsume, T.; Tomiyama, K. Tetrahedron Lett., 1971, 983.


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Preston, N. W.; Chamberlain, K.; Skipp, R. A. Phytochem., 1975, 14, 1983. 23. Stoessel, Α.; Rock, G. L.; Fisch, M. H. Chem. Ind., 1974, p. 703. 1985


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Some Chemical Aspects of Plant Disease Resistance - American

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