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Effect of light quality on growth and sporulation in Verticillium agaricinum
[ Trans. Br. mycol. Soc. 72 (1) 145-146 (1979)
145
]
Printedin Great Britain
EFFECT OF LIGHT QUALITY ON GROWTH AND SPORULATION IN VERTICILLIUM AGARICINUM By MOHAMED OSMAN AND L. R. G. VALADON Department of Botany, Royal Holloway College, University of London, Egham Hill, Egham, Surrey TW20 oEX, England Growth and sporulation of Verticillium agaricinum were studied under various wavelengths between 300 and 750 nm. The fungus produced spores under all wavelengths tested as well as in the dark. Of all lights used only actinic blue 320-450 nm (max 370 nm) markedly increased sporulation and retarded growth. The whole process can be considered as lightvariable and not light-dependent. Mycochrome which may be involved in sporulation in certain fungi is not operative in this system. Micro-organisms respond to light in different ways: by changed growth, spore production, and pigment formation (Carlile, 1965, 1970). Aspergillus fiavus (Muntanjola-Cvetkovic & Neskovic, 1968) and Trichoderma viride (Gressel & Galun, 1967) were found to produce abundant conidia only after exposure to light. Hill (1976) found that light induces conidiation and inhibits growth in Aspergillus ornatus. Several early observations, reviewed by Smith (1936), Page (1952), Snyder & Hansen (1941) and Harter (1939), confirmed that light retards mycelial growth. Zentmyer & Ribeiro (1977) found that Phytophthora cinnamomi produced sporangia at all wavelengths and intensities tested, and in darkness, and concluded that sporangial production is light-variable. Tan & Epton (1973) found that sporulation of Botrytis cinerea was stimulated by near-u.v. light and was slightly affected by infrared, red and yellow light, but blue and green were ineffective. They observed also that continuous black light retarded linear growth. Carotenogenesis in Verticillium agaricinum is photo-dependent and near-u.v. light is very effective. The near-u.v. light effect is reversed by blue light but the photoreceptor is not mycochrome (Osman & Valadon, 1979). Furthermore, this fungus does not behave in the same way as other fungi in response to light (Valadon & Mummery, 1973; Mummery & Valadon, 1973). The object of the present paper is to study the effect of light on growth and sporulation in V. agaricinum and to compare it with other fungi. MATERIALS AND METHODS
The fungus V. agaricinum (Lk) Cda was used in this study. Malt extract-agar medium (20 ml) was poured into Petri dishes (8-5 em diam).
Spore suspension was prepared from 7-day cultures of the fungus. A loopful was placed in the centre of each dish, which was then incubated in the dark and under different lights for 5 days at 24-25 DC. The lights used were white, black, actinic blue, blue, yellow and red (Table 1). Linear growth of the developed colonies was measured. The spores were counted using a haemocytometer. The light intensity was 95 flW cm- 2 for white and actinic blue light, and 10 flW em -2 for the other lights, between 300 and 750 nm. RESULTS AND DISCUSSION
Linear growth was markedly affected by actinic blue (near-u.v.) light (Table 1) and the colony appeared to be wrinkled and very deep-orange in colour. Carotenoids are greatly increased under near-u.v, light (Osman & Valadon, 1978) and this was reflected by the appearance of the cultures. Linear growth (6 em) under black light was also inhibited compared with cultures in the dark (8-2 em), and it is also known that carotenoid production is more than doubled under black light compared to white light (Osman & Valadon, 1978). Growth under all other lights used did not differ significantly from dark controls. The colonies under black light appeared flat and were lightorange in colour; under white light the appearance was again flat and slightly orange coloured; whilst under all other lights and in the dark there was no trace of colouration and the cultures were all white and fluffy. The spores under near-u.v, light were variable in size and shape in comparison with those grown in the dark (Osman & Valadon, 1979). As can be seen from Table 1, the average number of spores was 6-8 x 105 ml - 1 which was significantly different from dark-grown spores or
0007-1536/79/2828-483° $01.00 © 1979 The British Mycological Society
Photobiology of Verticillium Table
1.
Effect of light quality on linear growth, sporulation and appearance of colonies of V. agaricinum
Light quality Dark White Black Actinic blue (near-u.v.) Blue Yellow Red
Transmittance (nm) Visible spectrum 300-420 (peak 355) 320-450 (peak 370) 380-5 25 (peak 475) 545-620 (peak 570) 625 and above
spores under any other kind of light. Although there is an increase of approximately 40 % in number of spores under near-u.v., on the whole sporulation in V. agaricinum is not photo-dependent, but continuous near-u. v, light retards growth and increases sporulation. Black light, on the other hand, inhibits linear growth but does not affect sporulation, so we are in disagreement with a number of reviewers in that light does not inhibit growth in V. agaricinum. Kumagai et al. (1976) have proposed the term mycochrome for their photoreceptor system, in which near-u.v. caused sporulation, and this is reversed by blue light. They proposed the action of mycochrome as follows: B
mycochrome (MB ) ~ mycochrome (M:01uv) N-UV Irradiation with near-u.v. light will convert the M N U V form to the M B form which is active in conidiation. In our case near-u.v. increased sporulation by about 40 % but blue light had no effect, i.e. blue light cannot reverse the effect of near-u.v. light. We have already shown that mycochrome cannot operate in V. agaricinum (Osman & Valadon, 1979) and that the pigment system must be new. This conclusion is reinforced by the results above. Since there is a high irradiance reaction involved in carotenogenesis in this fungus (Osman & Valadon, 1979), phytochrome must also be involved. REFERENCES CARLILE, M. J. (1965). The photobiology of fungi. Annual Review of Plant Physiology 16, 175-202. CARLILE, M. J. (1970). The photoresponses offungi. In Photobiology of Micro-organisms, pp. 309-344. New York: Wiley-Interscience. GRESSEL, J. & GALUN, E. (1967). Morphogenesis in Trichoderma: Photoinduction and RNA. Developmental Biology 15, 575-598.
Linear growth (ern)
Spore Number ml ' ! x 10- 5
8'2 8'0 6'0 1'0 8'1 7'9 8'1
4'5 5'2 5'5 6·8 5'4 4'9 5'4
Character of Colonies Fluffy and white Flat and slightly orange Flat and light-orange Wrinkled and deep-orange Fluffy and white Fluffy and white Fluffy and white
HARTER, L. L. (1939). Influence of light on the length of the conidia in certain species of Fusarium. American Journal of Botany 26, 234-243. HILL, E. P. (1976). Effect of light on growth and sporulation of Aspergillus ornatus, Journal of General Microbiology 95, 39-44· KUMAGAI, T., YOSHIOKA, N. & aDA, Y. (1976). Further studies on the blue and near ultraviolet reversible photoreaction with an intracellular particulate fraction of the fungus, Alternaria tomato. Biochimica et Biophysica Acta 421, 133-140. MUMMERY, R. S. & VALADON, L. R. G. (1973). The effect of antimycin A on carotenogenesis in Verticillium agaricinum. Planta (Berlin) 109, 353-356. MUNTANJOLA-CVETKOVIC, M. 1. & NESKOVIC, M. (1968). Studies on the effect of some substances as possible substitutes of light in the sporulation of Aspergillus flaous 28-A. Bulletin de l'Institut et du Jardin Botanique de l'Unioersite de Belgrade 3, 35-41. OSMAN, M. & VALADON. L. R. G. (1978). Effect oflight quality on the photoinduction of carotenoid synthesis in Verticillium agaricinum. Microbios 18, 229-234. OSMAN, M. & VALADON, L. R. G. (1979). Studies on the near u.v. effect on carotenogenesis in Verticillium agaricinum. Microbios. (In the press.) PAGE, R. M. (1952). The effect of nutrition on growth and sporulation of Pilobolus. American Journal of Botany 39, 731-739. SMITH, E. C. (1936). In Biological Effects of Radiation (ed. B. M. Dugger), pp. 889-918. New York: McGraw-Hill. SNYDER, W. C. & HANSEN, H. N. (1941). The effect of light on taxonomic characters in Fusarium. Mycologia 33, 580-591. TAN, K. K. & EpToN, H. A. S. (1973). Effect of light on the growth and sporulation of Botrytis cinerea. Transactions of the British Mycological Society 61, 147-157. VALADON, L. R. G. & MUMMERY, R. S. (1973). Effect of certain inhibitors of carotenogenesis in Verticillium agaricinum in response to nicotine and to CPTA. Microbios 10A, 97-104. ZENTMYER, G. A. & RIBEIRO, O. K. (1977). The effect of visible and near-visible radiation on sporangium production by Phytophthora cinnamomi. Phytopathology 67, 91-95.
(Accepted for publication 30 June 1978)
145
]
Printedin Great Britain
EFFECT OF LIGHT QUALITY ON GROWTH AND SPORULATION IN VERTICILLIUM AGARICINUM By MOHAMED OSMAN AND L. R. G. VALADON Department of Botany, Royal Holloway College, University of London, Egham Hill, Egham, Surrey TW20 oEX, England Growth and sporulation of Verticillium agaricinum were studied under various wavelengths between 300 and 750 nm. The fungus produced spores under all wavelengths tested as well as in the dark. Of all lights used only actinic blue 320-450 nm (max 370 nm) markedly increased sporulation and retarded growth. The whole process can be considered as lightvariable and not light-dependent. Mycochrome which may be involved in sporulation in certain fungi is not operative in this system. Micro-organisms respond to light in different ways: by changed growth, spore production, and pigment formation (Carlile, 1965, 1970). Aspergillus fiavus (Muntanjola-Cvetkovic & Neskovic, 1968) and Trichoderma viride (Gressel & Galun, 1967) were found to produce abundant conidia only after exposure to light. Hill (1976) found that light induces conidiation and inhibits growth in Aspergillus ornatus. Several early observations, reviewed by Smith (1936), Page (1952), Snyder & Hansen (1941) and Harter (1939), confirmed that light retards mycelial growth. Zentmyer & Ribeiro (1977) found that Phytophthora cinnamomi produced sporangia at all wavelengths and intensities tested, and in darkness, and concluded that sporangial production is light-variable. Tan & Epton (1973) found that sporulation of Botrytis cinerea was stimulated by near-u.v. light and was slightly affected by infrared, red and yellow light, but blue and green were ineffective. They observed also that continuous black light retarded linear growth. Carotenogenesis in Verticillium agaricinum is photo-dependent and near-u.v. light is very effective. The near-u.v. light effect is reversed by blue light but the photoreceptor is not mycochrome (Osman & Valadon, 1979). Furthermore, this fungus does not behave in the same way as other fungi in response to light (Valadon & Mummery, 1973; Mummery & Valadon, 1973). The object of the present paper is to study the effect of light on growth and sporulation in V. agaricinum and to compare it with other fungi. MATERIALS AND METHODS
The fungus V. agaricinum (Lk) Cda was used in this study. Malt extract-agar medium (20 ml) was poured into Petri dishes (8-5 em diam).
Spore suspension was prepared from 7-day cultures of the fungus. A loopful was placed in the centre of each dish, which was then incubated in the dark and under different lights for 5 days at 24-25 DC. The lights used were white, black, actinic blue, blue, yellow and red (Table 1). Linear growth of the developed colonies was measured. The spores were counted using a haemocytometer. The light intensity was 95 flW cm- 2 for white and actinic blue light, and 10 flW em -2 for the other lights, between 300 and 750 nm. RESULTS AND DISCUSSION
Linear growth was markedly affected by actinic blue (near-u.v.) light (Table 1) and the colony appeared to be wrinkled and very deep-orange in colour. Carotenoids are greatly increased under near-u.v, light (Osman & Valadon, 1978) and this was reflected by the appearance of the cultures. Linear growth (6 em) under black light was also inhibited compared with cultures in the dark (8-2 em), and it is also known that carotenoid production is more than doubled under black light compared to white light (Osman & Valadon, 1978). Growth under all other lights used did not differ significantly from dark controls. The colonies under black light appeared flat and were lightorange in colour; under white light the appearance was again flat and slightly orange coloured; whilst under all other lights and in the dark there was no trace of colouration and the cultures were all white and fluffy. The spores under near-u.v, light were variable in size and shape in comparison with those grown in the dark (Osman & Valadon, 1979). As can be seen from Table 1, the average number of spores was 6-8 x 105 ml - 1 which was significantly different from dark-grown spores or
0007-1536/79/2828-483° $01.00 © 1979 The British Mycological Society
Photobiology of Verticillium Table
1.
Effect of light quality on linear growth, sporulation and appearance of colonies of V. agaricinum
Light quality Dark White Black Actinic blue (near-u.v.) Blue Yellow Red
Transmittance (nm) Visible spectrum 300-420 (peak 355) 320-450 (peak 370) 380-5 25 (peak 475) 545-620 (peak 570) 625 and above
spores under any other kind of light. Although there is an increase of approximately 40 % in number of spores under near-u.v., on the whole sporulation in V. agaricinum is not photo-dependent, but continuous near-u. v, light retards growth and increases sporulation. Black light, on the other hand, inhibits linear growth but does not affect sporulation, so we are in disagreement with a number of reviewers in that light does not inhibit growth in V. agaricinum. Kumagai et al. (1976) have proposed the term mycochrome for their photoreceptor system, in which near-u.v. caused sporulation, and this is reversed by blue light. They proposed the action of mycochrome as follows: B
mycochrome (MB ) ~ mycochrome (M:01uv) N-UV Irradiation with near-u.v. light will convert the M N U V form to the M B form which is active in conidiation. In our case near-u.v. increased sporulation by about 40 % but blue light had no effect, i.e. blue light cannot reverse the effect of near-u.v. light. We have already shown that mycochrome cannot operate in V. agaricinum (Osman & Valadon, 1979) and that the pigment system must be new. This conclusion is reinforced by the results above. Since there is a high irradiance reaction involved in carotenogenesis in this fungus (Osman & Valadon, 1979), phytochrome must also be involved. REFERENCES CARLILE, M. J. (1965). The photobiology of fungi. Annual Review of Plant Physiology 16, 175-202. CARLILE, M. J. (1970). The photoresponses offungi. In Photobiology of Micro-organisms, pp. 309-344. New York: Wiley-Interscience. GRESSEL, J. & GALUN, E. (1967). Morphogenesis in Trichoderma: Photoinduction and RNA. Developmental Biology 15, 575-598.
Linear growth (ern)
Spore Number ml ' ! x 10- 5
8'2 8'0 6'0 1'0 8'1 7'9 8'1
4'5 5'2 5'5 6·8 5'4 4'9 5'4
Character of Colonies Fluffy and white Flat and slightly orange Flat and light-orange Wrinkled and deep-orange Fluffy and white Fluffy and white Fluffy and white
HARTER, L. L. (1939). Influence of light on the length of the conidia in certain species of Fusarium. American Journal of Botany 26, 234-243. HILL, E. P. (1976). Effect of light on growth and sporulation of Aspergillus ornatus, Journal of General Microbiology 95, 39-44· KUMAGAI, T., YOSHIOKA, N. & aDA, Y. (1976). Further studies on the blue and near ultraviolet reversible photoreaction with an intracellular particulate fraction of the fungus, Alternaria tomato. Biochimica et Biophysica Acta 421, 133-140. MUMMERY, R. S. & VALADON, L. R. G. (1973). The effect of antimycin A on carotenogenesis in Verticillium agaricinum. Planta (Berlin) 109, 353-356. MUNTANJOLA-CVETKOVIC, M. 1. & NESKOVIC, M. (1968). Studies on the effect of some substances as possible substitutes of light in the sporulation of Aspergillus flaous 28-A. Bulletin de l'Institut et du Jardin Botanique de l'Unioersite de Belgrade 3, 35-41. OSMAN, M. & VALADON. L. R. G. (1978). Effect oflight quality on the photoinduction of carotenoid synthesis in Verticillium agaricinum. Microbios 18, 229-234. OSMAN, M. & VALADON, L. R. G. (1979). Studies on the near u.v. effect on carotenogenesis in Verticillium agaricinum. Microbios. (In the press.) PAGE, R. M. (1952). The effect of nutrition on growth and sporulation of Pilobolus. American Journal of Botany 39, 731-739. SMITH, E. C. (1936). In Biological Effects of Radiation (ed. B. M. Dugger), pp. 889-918. New York: McGraw-Hill. SNYDER, W. C. & HANSEN, H. N. (1941). The effect of light on taxonomic characters in Fusarium. Mycologia 33, 580-591. TAN, K. K. & EpToN, H. A. S. (1973). Effect of light on the growth and sporulation of Botrytis cinerea. Transactions of the British Mycological Society 61, 147-157. VALADON, L. R. G. & MUMMERY, R. S. (1973). Effect of certain inhibitors of carotenogenesis in Verticillium agaricinum in response to nicotine and to CPTA. Microbios 10A, 97-104. ZENTMYER, G. A. & RIBEIRO, O. K. (1977). The effect of visible and near-visible radiation on sporangium production by Phytophthora cinnamomi. Phytopathology 67, 91-95.
(Accepted for publication 30 June 1978)