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Decomposition of cellulose by three pea pathogens in pure culture
[ 113 ] Trans. Br. mycol, Soc. 62 (1), 1I3-II7 (1974) Printed in Great Britain
DECOMPOSITION OF CELLULOSE BY THREE PEA PATHOGENS IN PURE CULTURE By P. S. RATTAN
Department of Biological Sciences, University of Exeter Ascochyta pisi (Lib.), Mycosphaerella pinodes (Berk. & Blox.) Vestergr., Phoma medicaginis var. pinodella (L. K. Jones) Boerema, which cause leaf and pod spot, and foot rots of pea grow on filter paper as a sole carbon source in pure culture. Different isolates of the same pathogen utilized cellulose at different rates and decomposition was affected by the amount of nitrogen in the culture. These results have relevance to studies of the saprophytic survival of these organisms.
Wehlburg (1932) reported that Ascochyta pisi did not utilize shredded filter, whilst Baumann (1953) reported that Mycosphaerella pinodes could grow on filter paper soaked in Czapek-Dox solution. Heath & Wood (1971) found evidence that M. pinodes secreted enzymes (in vivo) that attacked native cellulose (cotton muslin), whereas A. pisi did not. The present investigation reports the relative abilities of the three pea pathogens to utilize filter-paper cellulose as a sole carbon source. The bearing of the results on the saprophytic survival of the fungi is discussed. MATERIALS AND METHODS
The isolates of the pathogens used in the present investigation were:
Ascochyta pisi (Lib.) PS 17. Isolated from seed of pea cv. Giant Stride. PS 38. Isolated from seed of pea cv. Thomas Laxton.
Mycosphaerella pinodes (Berk. & Blox.) Vestergr. PS ro. Isolated from seed of pea cv. Pauli. PS 56. Isolated from stem of pea by Dr G. H. Boerema, Ministerie van Landbouw en Visserij, Plantenziektenkundige Dienst, Wageningen, The Netherlands.
Phoma medicaginis var. pinodella (L. K. Jones) Boerema ( =. Ascochyta pinodella L. K. Jones). PS27. Isolated from seed of pea cv.Jade. PS59. Isolated from seed of pea cv. Sharpe's 99 Canner. PS7I. Isolated from pea by W.]. Rennie, Department of Agriculture and Fisheries for Scotland, East Craigs, Edinburgh 12. The method used was that of Garrett (1962, 1966). Wads of ten Whatman no. 3 filter paper (extra thick) of 7 em diameter were put into 8
MYC
62
114
Transactions British Mycological Society Table I. Percentage loss in dry weight with time, offilterpaper inoculated with M. pinodes, A. pisi, and P. medicaginis var. pinodella Period of incubation (weeks)
,
Pathogen (isolate no.) M, pinodes (PS 10)
A, pisi (PS 38) P. medicaginis var, pinodella (PS 71)
6
4 3'3 ±O'S 5.6 ±o'4 lS'2 ±1'1
40'S ±0'7 7'0 ±1'1 15'1 ±o'3
8 8,6 ±1'2 8'9 ±1'6 16,6 ±o'3
10
12
10'1 ±O'9 8'S ±o'7 17'7 ± 1'1
lO'7 ±0·8 8,6 ±1'2 18'0 ±O'2
2S0 ml flasks, and moistened with 20 ml of mineral solution containing NaNOs in variable quantities, KH 2P0 4, 1'0 g; MgS04.7H20, o'S g; FeS04.7H20, 0'01 g; ZnS04.7H20, 0'01 g; CUS04' SH 20, o'ooS g; made up to I 1 with distilled water. Since all the three pathogens were known (Bajan, 1968) to utilize sodium nitrate equally well, it was selected as the nitrogen source for the culture solution. The flasks were autoclaved at 121°C for 30 min. Each wad was inoculated with a 10 mm agar disk from a 3-day-old mycelial culture, on pea seed decoction glucose agar. Inoculated and uninoculated flasks were incubated at 22'So for the periods stated. After incubation, the wads of filter paper were removed from the flasks, dried to a constant weight at 80° for 24 h and the ovendried weight recorded. The original air-dried weights were converted to oven-dried weights by determining both the air-dried and oven-dried weights of the uninoculated wads, and the mean losses in dry weights were calculated. These losses represent only that portion of cellulose actually respired by the pathogens and do not include the further portion converted into fungal substance. RESULTS
Experiment I. One isolate of each pathogen was used. All the three pathogens invaded the filter paper. P. medicaginis var. pinodella fully colonized the wads within 3 weeks and also grew to the bottom of the wads. A. pisi and M, pinodes grew more slowly. However, the filter paper was covered by both the pathogens within 4-5 weeks. In this experiment, sodium nitrate was supplied at the rate of 5 gil. Four replicates per pathogen were used at each sampling. The results (Table I) show clearly that the three pathogens are capable of utilizing filter paper as a sole carbon source in pure culture. The degradation process by M. pinodes was slow, but it continued progressively over a period of 12 weeks (limit of the experiment), whereas in the case of A. pisi it ceased after 8 weeks. P. medicaginis var. pinodella was very active in the first half of the incubation period, but then the degradation of cellulose was slower than by M. pinodes. M. pinodes and P. medicaginis var. pinodella also formed abundant chlamydospores on the filter paper. Experiment 2. This experiment was carried out to compare the rate of
115
Cellulose decomposition by pea pathogens. P. S. Rattan
Table 2. Percentage loss in dry weight of filter paper inoculated with different isolates of M. pinodes, A. pisi and P. medicaginis var. pinodella Pathogen
Isolate no,
M,pinodes
PSIO PSS6 PSI7 PS3S PS27 PS59 PS71
A, pisi
r. medicaginis
var, pinodella
Least significant differences at P" 0'05
Percentage loss
Mean
6'7 ± a's"'''} S·6±o'g··· 7'2 ± 1'2· 7'8±o'7· g'O± I'O••• } S'2+ 1'0··· 1I'3±1·1·"
7'6± 1'0
}
= 0'45,
P" 0'01
7'5±0'3 9'5± 1'3···
= 0'60, P"· 0'001 = 0·7g.
Table 3. Percentage loss in dry weight offilter paper inoculated with M. pinodes, A. pisi and P. medicaginis var. pin odella at different levels of nitrogen Pathogen (isolate no.) M.pinodes(PSIO) A.pisi(PS38) P.medicaginisvar. pinodella (PS71)
5 gIl NaNO a (full nitrogen) 6·8+1·6 6'7±I'o 16'0±O'4
2'5 gIl NaNO a 1'25 gIl NaNO a (half nitrogen) (quarter nitrogen) 5'7±o'9 5,S±0·6 g'o±o'6
Least significant differences at P 0'05
= 0,83,
3'9±o'6 3'7±O'6 4'9±o'8 P 0'01
=
1'10.
decomposition of filter paper by different isolates of the three pathogens. Sodium nitrate was supplied at 2'5 gil. Eight replicate flasks were used and loss in dry weight was recorded after 8 weeks. The results (Table 2) indicate that different isolates of the same pathogen decomposed cellulose at different rates. The difference between the mean dry-weight loss caused by isolates of P. medicaginis var. pinodella was significantly higher (P > 0'001) than that of M. pinodes or A. pisi. Experiment 3. This was designed to measure the effect of nitrogen level on cellulose decomposition, Using the same isolates as in Expt I, NaNGa was supplied at 5'0 gil (full strength), 2'5 gil, and 1'25 gil, and flasks were incubated for 8 weeks at 22'5°, In the full-strength and half-strength series the growth was similar but in quarter-strength it was less in M. pinodes and A. pisi. The lowest filter paper was not colonized fully in quarterstrength series by anyone of the three pathogens. The results (Table 3) show that percentage loss in dry weight of filter paper inoculated with each pathogen decreased with the reduction in nitrogen content of the culture. For M. pinodes dry-weight losses were in the ratio of 1'00:0.84:0'57, for A. pisi 1'00:0'88:0'56. For P. medicaginis var. pinodella the ratio was 1'00:0'56:0'31. For each pathogen, the differences in means of the three nitrogen series were statistically significant at the I % level except that in A. pisi, the difference between full-strength and half-strength nitrogen was significant at the 5 % level only. The differences between the dry-weight losses caused by M. pinodes and A. pisi were not significant for the three levels of nitrogen but were significant in the case of P. medicaginis var. pinodella at the I % level. The percentage loss in dry weight (full-strength nitrogen) after an 8'2
116
Transactions British Mycological Society
8-week period was less than the one previously recorded (Table I) for the same period. It was probably due to a faulty incubator (loose fitting glass door). The evaporation in this incubator was more than the one previously used, and the contents gradually dried up, affecting the growth of the pathogens. DISCUSSION
These experiments show that A. pisi, M. pinodes and P. medicaginis var. pinodella can decompose native cellulose in the form of filter paper. Wehlburg's (1932) negative results can perhaps be attributed either to the level of inoculum used by him or to the level of nitrogen used in the nutrient solution or even the source of nitrogen (no details of Wehlburg's experimental data were available). Garrett (1962) stated that it is possible that the level of inoculum potential required for the initiation of cellulose decomposition varies with different species offungi. It was noticed in the present investigation that the pathogens made abundant growth on the agar block before colonizing the filter paper. Arakawa (1934) found that the process of decomposition of cellulose is influenced by several factors including the kind of cellulose, the species of fungus and composition of the culture medium. Some sugars stimulate the decomposition of filter paper. Perhaps the results of Heath & Wood (1971) can be explained on the same basis. The only other report on cellulose decomposition by Ascochyta is by Hancock & Millar (1965), who found that Ascochyta imperfecta Peck, the cause of black stem of alfalfa (now considered a synonym of P. medicaginis var. medicaginis by Boerema, Dorenbosch & Leffring, 1965), can make abundant growth on filter paper. The results of these experiments indicate that the three pathogens have more or less equal ability to decompose cellulose in pure culture and utilize the breakdown products nutritionally and thus possibly they have the same ability for saprophytic survival. The three pathogens are able to persist and overwinter in pea refuse (Jones, 1927; Linford & Sprague, 1927; Hare & Walker, 1944). The actual period of survival in soil depends upon the soil conditions and competition with other organisms, though this competition is not so severe as that involved in competitive colonization of dead tissues (Garrett, 1963). A cellulose decomposing fungus may possess a high degree of saprophytic ability for exploitation of cellulose component of plant tissue, yet in actual competition may obtain no more than a negligible share of the simpler carbon compounds (Garrett, 1963). Dickinson & Sheridan (1968) concluded from their studies on survival of A. pisi and M. pinodes in unsterilized soil that M. pinodes and A. pisi could be classified as moderately successful and poor saprophytes respectively. Wallen & Jeun (1968) reported that Ascochyta pinodes (M. pinodes) and Ascochyta pinodella (== P. medicaginis var. pinodella) can survive longer than A. pisi because of their ability to tolerate a wide range of temperature, their formation of chlamydospores and their antagonism towards A. pisi. They thought it possible that other fungi and micro-organisms exert a strong antagonistic action on A. pisi in natural soil.
Cellulose decomposition by pea pathogens. P. S. Rattan
117
This work formed part of a Ph.D. thesis submitted to the University of Exeter. I am grateful to Dr S. A. J. Tarr for help and encouragement, and to the University of Exeter for the financial support. REFERENCES
ARAKAWA, S. (1934). The influence of sugars on the cellulose decomposition by the soil fungi. Transactions of the Tottori Society of Agricultural Science 5, 27-35. (Abstr. in Review of Applied Mycology 14 (1935), 332.) BAJAN, C. (1968). Investigations on the pathogenicity and biology of three species of genus Ascochyta occurring on pea (Pisum sativum L.). Acta Agrobotanica lU, 17-74. BAUMANN, G. (1953). Untersuchungen zur Biologie von Mycosphaerella pinodes (Berk, et Blox.) Stone. Kiihn-Archiv 67,305-383. BOEREMA, G. H., DORENBOSCH MARIA M.]. & LEFFRING, L. ([965). A comparative study of the black stem fungi on lucerne and red clover and the footrot fungus on pea. Netherlands Journal of Plant Pathology 71, 79-89. DICK.l~SO~, C. H. & SHERIDAN,].]. ([968). Studies on survival of Mycosphaerella pinodes and Ascochyta pisi. Annals of AppliedBiology 62, 474-483. GARRETT, S. D. (1962). Decomposition of cellulose in soil by Rhizoctonia solani, Transactions of the British Mycological Society 45, [15-120. GARRETT, S. D. ([963). Soilfungi and soilfertility. Oxford: Pergamon Press. GARRETT, S. D. (1966). Cellulose decomposing ability of some cereal foot rot fungi in relation to their saprophytic survival. Transactions of the British Mycological Society 49,57--68. HANCOCK, ]. G. & MILLAR, R. L. ([965)' Association of cellulolytic, proteolytic and xylolytic enzymes with southern anthracnose, spring black stem, and Stemphylium leaf spot of alfalfa. Phytopathology 55, 356-360. HARE, W. W. & WALKER, ]. C. (1944). Ascochyta diseases of canning pea. Research Bulletin Wisconsin Agricultural Experiment Station no. 150, p. 31. HEATH, M. C. & WOOD, R. K. S. (1971). Role of cell-wall degrading enzymes in the development of leaf spots caused by Ascochyta pisi and Mycosphaerella pinades. Annals of Botany (New Series) 35,451-474. JONES, L. K. (1927). Studies on the nature and control of blight, leaf and pod spot and foot rot of peas caused by some of the species of Ascochyta. Bulletin of the New York (Geneva) Agricultural Experimental Station no. 547, p. 46. LINFORD, M. B. & SPRAGUE, R. (1927). Species of Ascochyta parasitic on pea. Phytopathology 17, 38 1-397. RATTAN, P. S. ([97[). Studies on the diseases of pea caused by Ascochyta pisi, Mycosphaerellapinodes and Phoma medicaginis var. pinodella. Ph.D. thesis, University of Exeter. WALLEN, V. R. & ]EUN,]. (1968). Factors limiting survival of Ascochyta spp. on peas in soil. Canadian Journal of Botany 46, 1279-1286. WEHLBTJRG, C. (1932). [Investigations of pea anthracnose. Thesis, University of Utrecht (Hollandia-Drukkerij, Baarn)']. (Abstr. in Review of Applied Mycology 12 (1933),483.)
(Accepted for publication
20
April 1973)
DECOMPOSITION OF CELLULOSE BY THREE PEA PATHOGENS IN PURE CULTURE By P. S. RATTAN
Department of Biological Sciences, University of Exeter Ascochyta pisi (Lib.), Mycosphaerella pinodes (Berk. & Blox.) Vestergr., Phoma medicaginis var. pinodella (L. K. Jones) Boerema, which cause leaf and pod spot, and foot rots of pea grow on filter paper as a sole carbon source in pure culture. Different isolates of the same pathogen utilized cellulose at different rates and decomposition was affected by the amount of nitrogen in the culture. These results have relevance to studies of the saprophytic survival of these organisms.
Wehlburg (1932) reported that Ascochyta pisi did not utilize shredded filter, whilst Baumann (1953) reported that Mycosphaerella pinodes could grow on filter paper soaked in Czapek-Dox solution. Heath & Wood (1971) found evidence that M. pinodes secreted enzymes (in vivo) that attacked native cellulose (cotton muslin), whereas A. pisi did not. The present investigation reports the relative abilities of the three pea pathogens to utilize filter-paper cellulose as a sole carbon source. The bearing of the results on the saprophytic survival of the fungi is discussed. MATERIALS AND METHODS
The isolates of the pathogens used in the present investigation were:
Ascochyta pisi (Lib.) PS 17. Isolated from seed of pea cv. Giant Stride. PS 38. Isolated from seed of pea cv. Thomas Laxton.
Mycosphaerella pinodes (Berk. & Blox.) Vestergr. PS ro. Isolated from seed of pea cv. Pauli. PS 56. Isolated from stem of pea by Dr G. H. Boerema, Ministerie van Landbouw en Visserij, Plantenziektenkundige Dienst, Wageningen, The Netherlands.
Phoma medicaginis var. pinodella (L. K. Jones) Boerema ( =. Ascochyta pinodella L. K. Jones). PS27. Isolated from seed of pea cv.Jade. PS59. Isolated from seed of pea cv. Sharpe's 99 Canner. PS7I. Isolated from pea by W.]. Rennie, Department of Agriculture and Fisheries for Scotland, East Craigs, Edinburgh 12. The method used was that of Garrett (1962, 1966). Wads of ten Whatman no. 3 filter paper (extra thick) of 7 em diameter were put into 8
MYC
62
114
Transactions British Mycological Society Table I. Percentage loss in dry weight with time, offilterpaper inoculated with M. pinodes, A. pisi, and P. medicaginis var. pinodella Period of incubation (weeks)
,
Pathogen (isolate no.) M, pinodes (PS 10)
A, pisi (PS 38) P. medicaginis var, pinodella (PS 71)
6
4 3'3 ±O'S 5.6 ±o'4 lS'2 ±1'1
40'S ±0'7 7'0 ±1'1 15'1 ±o'3
8 8,6 ±1'2 8'9 ±1'6 16,6 ±o'3
10
12
10'1 ±O'9 8'S ±o'7 17'7 ± 1'1
lO'7 ±0·8 8,6 ±1'2 18'0 ±O'2
2S0 ml flasks, and moistened with 20 ml of mineral solution containing NaNOs in variable quantities, KH 2P0 4, 1'0 g; MgS04.7H20, o'S g; FeS04.7H20, 0'01 g; ZnS04.7H20, 0'01 g; CUS04' SH 20, o'ooS g; made up to I 1 with distilled water. Since all the three pathogens were known (Bajan, 1968) to utilize sodium nitrate equally well, it was selected as the nitrogen source for the culture solution. The flasks were autoclaved at 121°C for 30 min. Each wad was inoculated with a 10 mm agar disk from a 3-day-old mycelial culture, on pea seed decoction glucose agar. Inoculated and uninoculated flasks were incubated at 22'So for the periods stated. After incubation, the wads of filter paper were removed from the flasks, dried to a constant weight at 80° for 24 h and the ovendried weight recorded. The original air-dried weights were converted to oven-dried weights by determining both the air-dried and oven-dried weights of the uninoculated wads, and the mean losses in dry weights were calculated. These losses represent only that portion of cellulose actually respired by the pathogens and do not include the further portion converted into fungal substance. RESULTS
Experiment I. One isolate of each pathogen was used. All the three pathogens invaded the filter paper. P. medicaginis var. pinodella fully colonized the wads within 3 weeks and also grew to the bottom of the wads. A. pisi and M, pinodes grew more slowly. However, the filter paper was covered by both the pathogens within 4-5 weeks. In this experiment, sodium nitrate was supplied at the rate of 5 gil. Four replicates per pathogen were used at each sampling. The results (Table I) show clearly that the three pathogens are capable of utilizing filter paper as a sole carbon source in pure culture. The degradation process by M. pinodes was slow, but it continued progressively over a period of 12 weeks (limit of the experiment), whereas in the case of A. pisi it ceased after 8 weeks. P. medicaginis var. pinodella was very active in the first half of the incubation period, but then the degradation of cellulose was slower than by M. pinodes. M. pinodes and P. medicaginis var. pinodella also formed abundant chlamydospores on the filter paper. Experiment 2. This experiment was carried out to compare the rate of
115
Cellulose decomposition by pea pathogens. P. S. Rattan
Table 2. Percentage loss in dry weight of filter paper inoculated with different isolates of M. pinodes, A. pisi and P. medicaginis var. pinodella Pathogen
Isolate no,
M,pinodes
PSIO PSS6 PSI7 PS3S PS27 PS59 PS71
A, pisi
r. medicaginis
var, pinodella
Least significant differences at P" 0'05
Percentage loss
Mean
6'7 ± a's"'''} S·6±o'g··· 7'2 ± 1'2· 7'8±o'7· g'O± I'O••• } S'2+ 1'0··· 1I'3±1·1·"
7'6± 1'0
}
= 0'45,
P" 0'01
7'5±0'3 9'5± 1'3···
= 0'60, P"· 0'001 = 0·7g.
Table 3. Percentage loss in dry weight offilter paper inoculated with M. pinodes, A. pisi and P. medicaginis var. pin odella at different levels of nitrogen Pathogen (isolate no.) M.pinodes(PSIO) A.pisi(PS38) P.medicaginisvar. pinodella (PS71)
5 gIl NaNO a (full nitrogen) 6·8+1·6 6'7±I'o 16'0±O'4
2'5 gIl NaNO a 1'25 gIl NaNO a (half nitrogen) (quarter nitrogen) 5'7±o'9 5,S±0·6 g'o±o'6
Least significant differences at P 0'05
= 0,83,
3'9±o'6 3'7±O'6 4'9±o'8 P 0'01
=
1'10.
decomposition of filter paper by different isolates of the three pathogens. Sodium nitrate was supplied at 2'5 gil. Eight replicate flasks were used and loss in dry weight was recorded after 8 weeks. The results (Table 2) indicate that different isolates of the same pathogen decomposed cellulose at different rates. The difference between the mean dry-weight loss caused by isolates of P. medicaginis var. pinodella was significantly higher (P > 0'001) than that of M. pinodes or A. pisi. Experiment 3. This was designed to measure the effect of nitrogen level on cellulose decomposition, Using the same isolates as in Expt I, NaNGa was supplied at 5'0 gil (full strength), 2'5 gil, and 1'25 gil, and flasks were incubated for 8 weeks at 22'5°, In the full-strength and half-strength series the growth was similar but in quarter-strength it was less in M. pinodes and A. pisi. The lowest filter paper was not colonized fully in quarterstrength series by anyone of the three pathogens. The results (Table 3) show that percentage loss in dry weight of filter paper inoculated with each pathogen decreased with the reduction in nitrogen content of the culture. For M. pinodes dry-weight losses were in the ratio of 1'00:0.84:0'57, for A. pisi 1'00:0'88:0'56. For P. medicaginis var. pinodella the ratio was 1'00:0'56:0'31. For each pathogen, the differences in means of the three nitrogen series were statistically significant at the I % level except that in A. pisi, the difference between full-strength and half-strength nitrogen was significant at the 5 % level only. The differences between the dry-weight losses caused by M. pinodes and A. pisi were not significant for the three levels of nitrogen but were significant in the case of P. medicaginis var. pinodella at the I % level. The percentage loss in dry weight (full-strength nitrogen) after an 8'2
116
Transactions British Mycological Society
8-week period was less than the one previously recorded (Table I) for the same period. It was probably due to a faulty incubator (loose fitting glass door). The evaporation in this incubator was more than the one previously used, and the contents gradually dried up, affecting the growth of the pathogens. DISCUSSION
These experiments show that A. pisi, M. pinodes and P. medicaginis var. pinodella can decompose native cellulose in the form of filter paper. Wehlburg's (1932) negative results can perhaps be attributed either to the level of inoculum used by him or to the level of nitrogen used in the nutrient solution or even the source of nitrogen (no details of Wehlburg's experimental data were available). Garrett (1962) stated that it is possible that the level of inoculum potential required for the initiation of cellulose decomposition varies with different species offungi. It was noticed in the present investigation that the pathogens made abundant growth on the agar block before colonizing the filter paper. Arakawa (1934) found that the process of decomposition of cellulose is influenced by several factors including the kind of cellulose, the species of fungus and composition of the culture medium. Some sugars stimulate the decomposition of filter paper. Perhaps the results of Heath & Wood (1971) can be explained on the same basis. The only other report on cellulose decomposition by Ascochyta is by Hancock & Millar (1965), who found that Ascochyta imperfecta Peck, the cause of black stem of alfalfa (now considered a synonym of P. medicaginis var. medicaginis by Boerema, Dorenbosch & Leffring, 1965), can make abundant growth on filter paper. The results of these experiments indicate that the three pathogens have more or less equal ability to decompose cellulose in pure culture and utilize the breakdown products nutritionally and thus possibly they have the same ability for saprophytic survival. The three pathogens are able to persist and overwinter in pea refuse (Jones, 1927; Linford & Sprague, 1927; Hare & Walker, 1944). The actual period of survival in soil depends upon the soil conditions and competition with other organisms, though this competition is not so severe as that involved in competitive colonization of dead tissues (Garrett, 1963). A cellulose decomposing fungus may possess a high degree of saprophytic ability for exploitation of cellulose component of plant tissue, yet in actual competition may obtain no more than a negligible share of the simpler carbon compounds (Garrett, 1963). Dickinson & Sheridan (1968) concluded from their studies on survival of A. pisi and M. pinodes in unsterilized soil that M. pinodes and A. pisi could be classified as moderately successful and poor saprophytes respectively. Wallen & Jeun (1968) reported that Ascochyta pinodes (M. pinodes) and Ascochyta pinodella (== P. medicaginis var. pinodella) can survive longer than A. pisi because of their ability to tolerate a wide range of temperature, their formation of chlamydospores and their antagonism towards A. pisi. They thought it possible that other fungi and micro-organisms exert a strong antagonistic action on A. pisi in natural soil.
Cellulose decomposition by pea pathogens. P. S. Rattan
117
This work formed part of a Ph.D. thesis submitted to the University of Exeter. I am grateful to Dr S. A. J. Tarr for help and encouragement, and to the University of Exeter for the financial support. REFERENCES
ARAKAWA, S. (1934). The influence of sugars on the cellulose decomposition by the soil fungi. Transactions of the Tottori Society of Agricultural Science 5, 27-35. (Abstr. in Review of Applied Mycology 14 (1935), 332.) BAJAN, C. (1968). Investigations on the pathogenicity and biology of three species of genus Ascochyta occurring on pea (Pisum sativum L.). Acta Agrobotanica lU, 17-74. BAUMANN, G. (1953). Untersuchungen zur Biologie von Mycosphaerella pinodes (Berk, et Blox.) Stone. Kiihn-Archiv 67,305-383. BOEREMA, G. H., DORENBOSCH MARIA M.]. & LEFFRING, L. ([965). A comparative study of the black stem fungi on lucerne and red clover and the footrot fungus on pea. Netherlands Journal of Plant Pathology 71, 79-89. DICK.l~SO~, C. H. & SHERIDAN,].]. ([968). Studies on survival of Mycosphaerella pinodes and Ascochyta pisi. Annals of AppliedBiology 62, 474-483. GARRETT, S. D. (1962). Decomposition of cellulose in soil by Rhizoctonia solani, Transactions of the British Mycological Society 45, [15-120. GARRETT, S. D. ([963). Soilfungi and soilfertility. Oxford: Pergamon Press. GARRETT, S. D. (1966). Cellulose decomposing ability of some cereal foot rot fungi in relation to their saprophytic survival. Transactions of the British Mycological Society 49,57--68. HANCOCK, ]. G. & MILLAR, R. L. ([965)' Association of cellulolytic, proteolytic and xylolytic enzymes with southern anthracnose, spring black stem, and Stemphylium leaf spot of alfalfa. Phytopathology 55, 356-360. HARE, W. W. & WALKER, ]. C. (1944). Ascochyta diseases of canning pea. Research Bulletin Wisconsin Agricultural Experiment Station no. 150, p. 31. HEATH, M. C. & WOOD, R. K. S. (1971). Role of cell-wall degrading enzymes in the development of leaf spots caused by Ascochyta pisi and Mycosphaerella pinades. Annals of Botany (New Series) 35,451-474. JONES, L. K. (1927). Studies on the nature and control of blight, leaf and pod spot and foot rot of peas caused by some of the species of Ascochyta. Bulletin of the New York (Geneva) Agricultural Experimental Station no. 547, p. 46. LINFORD, M. B. & SPRAGUE, R. (1927). Species of Ascochyta parasitic on pea. Phytopathology 17, 38 1-397. RATTAN, P. S. ([97[). Studies on the diseases of pea caused by Ascochyta pisi, Mycosphaerellapinodes and Phoma medicaginis var. pinodella. Ph.D. thesis, University of Exeter. WALLEN, V. R. & ]EUN,]. (1968). Factors limiting survival of Ascochyta spp. on peas in soil. Canadian Journal of Botany 46, 1279-1286. WEHLBTJRG, C. (1932). [Investigations of pea anthracnose. Thesis, University of Utrecht (Hollandia-Drukkerij, Baarn)']. (Abstr. in Review of Applied Mycology 12 (1933),483.)
(Accepted for publication
20
April 1973)