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Pathogenic variation in Septoria nodorum (Berk.) Berk. in relation to organ specificity, apparent photosynthetic rate and yield of wheat
Physiological
Plant Pathology
(1973) 3, 187-194
Pathogenic variation in Septoria nodorum (Berk.) Berk. in relation to organ specificity, apparent photosynthetic rate and yield of wheat? J. M. KRUPINSKY,$ A.L.
SCHAREN
Plant Science Research Division, Agricultural Research Service, United States Debartment of Agriculture, Plant Industry Station, Beltsville, Maryland
J. A.
and
Agronomy
20705,
U.S.A.
SCHILLINGER~
Department,
University of Maryland,
College
Park, Maryland
(Accepted for publication
Sepember
20740,
U.S.A.
1972)
Cultures of Seporia nodorum that varied in morphology were isolated from wheat plants that had been artificially inoculated with a composite culture. Other culture types were isolated from wheat leaves naturally infected in the field. Variation in pathogenicity of selected cultures was measured by symptom development, by apparent photosynthetic rates and by grain yield. Fungus variants that caused differences in photosynthesis, which were statistically significant at the 1 y0 level, were considered different in pathogenicity. Photosynthetic differences invariably were followed by yield differences. The role of environmental variation vs. pathogen variation was discussed.
INTRODUCTION Septoria nodorum (Berk.) Berk. (Leptosphaeria nodorum Mtiller) causes the glume blotch disease of wheat, Triticum aestiuum L. A major problem encountered in screening for resistance to S. nodorum has been the variability in disease intensity and yield loss due to environmental influences [Z, 81. Variation in the pathogen has been studied in pure culture [I, 3, 41, and in combination with wheat hosts [I, 41. The investigations cited above tended to show that environmental variation was the greatest deterrent to progress in selecting resistant host lines in the field. The present investigation was designed to study quantitatively the variation in pathogenicity of selected cultures of the fungus under rigidly controlled environmental conditions. It was hoped that some relative evaluation of the contribution of genetic variation in the fungus pathogen versus the variability caused by environmental factors could t Cooperative investigations of the National Agricultural Research Center, Agricultural Research Service, United States Department of Agriculture, and the University of Maryland Agricultural Experiment Station, Journal Paper No. A1745. This work is part of a thesis submitted by the senior author in partial fulf?lment of the requirements for the M.S. degree. f: Agricultural Research Technician and Research Plant Pathologist, respectively. Present address: USDA, ARS, Department of Botany, Montana State University, Bozeman, Montana 59715. f Associate Professor. 13
188
J. M. Krupinsky,
A. L. Scharen
and
J. A. Schillinger
be made as a result of these studies. In addition to visible symptoms and yields, apparent photosynthetic rates (APR) were measured to determine whether metabolic effects of the disease could be detected before visible symptoms of infection appeared, and as an additional measure of culture virulence. Comparisons of visible symptoms, APR and yield of grain permitted conclusions to be made concerning correlation of the aforementioned factors, and how they affect a search for genetic resistance to S. nodorum. MATERIALS
AND
METHODS
Cultures with maximum potential variability in morphology and pathogenicity were sought for our experiments. Since previous reports [Z, 41 had shown that a mixture of cultures collected in the geographic area of interest gave the most consistently useful results in inoculation experiments, we began with a composite collection of S. nodorum from seven locations. Wheat plants were inoculated with the composite, and after symptoms developed, isolations were made from infected leaves. Newly isolated cultures of S. nodorum that were derived from single spores always produced additional pycnidia and spores; however, indiscriminate mycelial transfers resulted in cultures that tended rapidly toward nonsporulating, mycelial types [a]. An extension of the single-spore isolation was the development of a mass spore transfer (MST) technique in which a mass of spores exuding from a pycnidium was picked up on a sterile needle or loop and transferred. MST were used as a rapid means of transferring cultures, while maintaining their sporulating ability and associated virulence. Three basic types of cultures were isolated that varied in production of pycnidia and spores, rate of mycelial growth and mycelial color. These were subsequently tested for virulence and effect on CO, exchange in wheat. For comparison, some cultures were isolated in the more traditional way by making a single-spore culture from a single plant at a single location. Two cultures of this type were used: N-40 from “Lerma Rojo 64-A” growing at Stoneville, Mississippi and N-42 from “Purplestraw” at Tifton, Georgia. These were maintained in culture by selective mass transfers of the “wild type”. Gas exchange analysis was done using an infrared CO, analyzer (IRGA). The intact head, peduncle and flag leaf of wheat, Wisconsin selection, C.I. 12632, were inserted into a Plexiglass air-sealed assimilation chamber [IO], and the APR was measured. A reading of APR was made prior to inoculation, then 14 daily measurements of photosynthesis in the head, peduncle and flag leaf were obtained to determine the effect of each S. nodorum isolate on APR [5]. All plants were grown in controlled environment chambers. The chambers had a 12-h photoperiod and the temperature was maintained at 2 1 “C during the day and 18 “C during the night. The light intensity was maintained at 70 + 5 x IO4 erg/ cm2 per s. For each study, six pots were selected that contained three plants which were just beginning to extrude anthers. The three treatments in each experiment included an uninoculated control head and flag leaf (treatment 1) plus two heads and flag leaves (treatments 2 and 3) that were inoculated with different culture types of S. nodorum. Details of inoculation are listed in Table 1. In the split plot design used for the IRGA studies, days were the whole unit treatments, and the three individual treatments were the subunits. An analysis of
Pathogenic
variation
in S.
189
nodorum
variance was run on each individual study, for treatment 1 us. the average of treatments
and orthogonal comparisons were made 2 and 3, and treatment 2 vs. 3 [9].
TABLE Isolates, Experiments and treatments Al 2 3 Bl
their description
and inoculation
Culture
type
Control Pycnidia Pink mycelia; pycnidia
2 3
Control Pycnidia Yellow mycelia; pycnidia
2 3
Control Pycnidia Pink mycelia; pycnidia
Cl
Dl 2 3 El 2 3 Fl 2 3
procedures
1 used in studying
Source of isolate
the pathogenic@ Concentration (spores and mycelia/ml)
of S. nodorum Incubation (‘4
Composite
15 x 10s
64
Composite
14x
106
64
Composite
12 x 10s
48
Composite
13 x 106
48
Composite
14 x 10s
48
Composite
13 x 106
48
Control Pink mycelia; no pycnidia Pycnidia
N-40 N-40
12 x 10s 14x 10s
64 64
Control White mycelia; no pycnidia Pycnidia
N-42 N-42
Control Gray-white mycelia; no pycnidia Pycnidia
N-40 N-40
8x 8x
lo6 lo6
72 72
12 x 10s 13 x 10s
72 72
RESULTS Decreases in APR were noted when heads and peduncles of test wheat were inoculated with selected cultures of 5’. nodorum (Fig. 1). Significant differences (1%) were observed in all studies and at all time intervals when the uninoculated control, treatment 1, was compared with the average of inoculated treatments 2 and 3. A sharp initial decrease in APR during the first 24 h after inoculation was seen in every experiment. At that time, no symptoms, or a few tiny chlorotic flecks were all that could be seen. Obvious visible symptoms of disease did not develop until 4 or 5 days after inoculation. Isolates were considered to be different from each other in pathogenicity only if they caused differences in APR significant at the 1 oh level. The dark areas in Figs 1 and 2 visualize the significant differences in the pathogenicity of isolates. When measuring the APR of the heads and peduncles in experiment A, a pycnidial isolate with white mycelium [Fig. 1 (a) 21 was less pathogenic than a pink mycelial isolate [Fig. 1 (a) 31. These isolates were also significantly different from one another in experiment C [Fig. 1 (c)l. Cultures used in treatments 2 and 3 of experiment B
190
J. M. Krupinsky,
A. L. Scharen
and
J. A. Schillinger
similarly reduced APR [Fig. 1(b)]. I n experiment D inoculation with a pycnidial isolate [Fig. 1(d) 31 re d uced APR more than similar inoculation with a pink mycelial isolate [Fig. l(d) 21. In experiment E, a white mycelial isolate [Fig. 1(e) 21 and a pycnidial isolate [Fig. 1(e) 31 had differential effects on the APR of the heads and peduncles for the entire period after inoculation. In experiment F a gray-white mycelial isolate [Fig. 1 (f) 21 reduced APR more than a pycnidial isolate [Fig. l(f) 31 during the first 8 days; however, after the twelfth day, the effect of the pycnidial isolate became greater.
(b)
0
11
0’
11
2
10
4
10
”
I””
6
8
IO
I
I
12 14 0 Days after inoculation
I
2
I
I
4
,
I
6
I
I
8
I
11
10
11
1
12
14
FIG. 1. Apparent photosynthetic rates of heads and peduncles of C.I. 12632 when inoculated at preanthesis stage of growth with different isolates of S. nodorum. Dark areas show significant differences ( 1%) in the pathogenicity of isolates : (a) pycnidial US. mycelial-pycnidial type; (b) pycnidial US. mycelial-pycnidial type; (c) pycnidial US. mycelial-pycnidial type; (d) mycelial US. pycnidial type; (e) mycelial US. pycnidial type and (f) mycelial US. pycnidial type. Legend: (o), control, treatment 1; (o), treatment 2; (0), treatment 3.
Pathogenic
variation
191
in S. nodorum
Flag leaves were less affected by S. nodorum inoculation than heads and peduncles (Fig. 2). In experiments A, D and F the APR of the control was significantly different from one inoculated treatment or the means of the two inoculated treatments. No significant differences were found in experiments B, C and E. The length of incubation time (48 h) was associated with poor S. nodorum infection in experiments B and C. In experiment E, the uninoculated flag leaves of control plants deteriorated for unknown reasons, resulting in the APR of the controls being lower than the APR of the infected for four consecutive days. In experiment A, a pycnidial isolate [Fig. 2(a) 21 reduced APR more than a pink mycelial isolate [Fig. 2(a) 31 and in
I’ll
I’
I
’
I
/
I
i
1’1’1
I
1 I
8
IO
(b)
II o Days after
FIG. 2. Apparent
I 2
/
I 4
/
6
I
I I2
inoculation
photosynthetic rates of flag leaves of C.I. 12632 when inoculated at preanthesis stage of growth with different isolates of S. nodomm. Dark areas show significant differences (1%) in the pathogenicity of isolates: (a) pycnidial VS. mycelial-pycnidial type; (b) pycnidial us. mycelial-pycnidial type; (c) pycnidial US. mycelial-pycnidial type; (d) myceiial US. pycnidial type; (e) mycelial US. pycnidial type and (f) mycelial US. pycnidial type. Legend as for Fig. 1.
I
" I 14
192
J. M. Krupinsky,
A. L. Scharen
and J. A. Schillinger
experiment D, a pycnidial isolate [Fig. 2(d) 31 was more effective in reducing APR than a pink mycelial isolate [Fig. 2(d) 21. Finally, a pycnidial isolate [Fig. 2 (f )] reduced APR more than a gray-white mycelial isolate [Fig. 2(f) 21 in experiment F. Other experiments [Fig. 2(b) (c) and (e)] detected no differences between treatments 2 and 3. The APR during the 15-day period of the experiments were averaged, and percentage of reduction was calculated for treatments 2 and 3 (Table 2). All isolates, excepting one, caused a greater decrease in APR on the heads and peduncles than on the flag leaves. S. nodorum infection caused an average 43% loss in APR on heads and peduncles and a 21 O/Oloss on the flag leaves. The decrease in APR of the heads and peduncles ranged from 26 to 55%, while the loss on the flag leaves ranged from 5 to 47%. When the APR of the infected heads, peduncles and flag leaves were added together, fungus infection caused an overall loss in APR of 3 1%. The overall range was 17 to 50%. TABLE
The average apparent Experiment study
photosynthetic
Treatmenta
A
B
C
D
E
F
rate for
Heads and pedunclesb
Reduction (%)
1 2 3
2.60 1.46 1.18
43.8 54.6
1 2 3
3.13 1.90 1.78
:
of plant parts
with S.
Flag leavesb
nodorum Reduction (%)
3.11
1.65 1.91
46.9 38.6
39.3 43.1
2.34 2.22 2.01
5.1 14.1
3
3.23 2.11 l-83
34.7 43.3
3-28 3.07 3.00
6.4 8.5
1 2 3
3.41 2.06 1.52
39.6 55.4
3.21 2.71 1.77
15.6 44.7
1 2 3
2.89 1.77 2.13
38.8 26.3
2.37 2.13 2.21
10.1 6.8
:
3.88 2.07 2.18
46.6 43.8
3.37 2.79 2.57
17.2 23.7
3 Total Total
2
I5 days after inonclation
for 1 for 2 and 3 2
a 1 = Uninoculated control. 2 and b APR = mg (CO&/organ per h.
19.14 IO.99
3 = Different
17.68 14.02
42.6
isolates
of S. nodomm.
The 1000 kernel weight and the grain weight per head were measured (Table 3). The loss in grain weight per head of inoculated plants ranged from 8 to 56%, and the overall loss averaged 31%. These variations in yield loss also demonstrate the variability in pathogenicity of S. nodorum.
Pathogenic
variation in S. nodorum
193 TABLE
Average yield
Study
of C.I.
12632
inoculated 1000 kernel weight (g)
Treatment
at preanthesis
3 stage of growth
Reduction (%I
with isolates of S. nodorum
Grain weight per h=d k)
Reduction (%I
1 2 3
33.04 20.15 20.12
39-l 39-2
1.19 0.66 0.71
44.6 40.6
1 2 3
35.34 24.20 27.58
31.6 22.0
1.48 1.00 l-11
32.5 25-l
C
: 3
32-99 25.28 26.90
23.4 18.5
1.50 1.16 l-18
22.7 21.1
D
1 2 3
31.30 2944 16.38
6.0 47.7
1.29 l-20 0.56
7.5 56.3
1 2 3
27Ti 21.17 20.79
23.2 24.6
1.06 0.79 0.84
26.0 21.3
1 2 3
33.97 23.69 17.93
30.3 47.3
1.26 0.89 0.68
29.0 45.9
A
B
E
F
Total Total
for 1 for 2 and 2
194.22 3
136.83
7.80 29.6
5.41
30.8
DISCUSSION
As reported previously [5-71, APR in wheat was reduced by 5’. nodorum infection. In the case of heads and peduncles, it was possible to distinguish the control from the infected material by means of gas exchange analysis from the first through the fourteenth day after inoculation. Decreases in APR were evident earlier than previously reported [5]. This is probably due to inoculation with increased numbers of spores, since greater reductions in yield have been associated with higher spore concentrations [2]. As in an earlier study [5j, the decrease in APR was not as evident with the flag leaves as with the head and peduncle. Under the experimental conditions used, a minimum of 64 h incubation was needed for flag leaves to become well infected. The selected mycelial types of S. nodorum or the mycelial type with pycnidia were sometimes more pathogenic than the typical pycnidial sorts. When the APR of the heads and peduncles were studied, the mycelial type and the mycelial type with pycnidia were more pathogenic than the pycnidial cultures. When flag leaves were inoculated and studied, the typical pycnidial type was more pathogenic than the selected mycelial type. Thus, the pycnidial type is more pathogenic on the flag leaves, indicating again the variation in pathogenicity among the isolates as well as variations due to the part of the plant infected. Variation in pathogenicity between different types of S. nodorum has been demonstrated. Considerable differences in APR, yield and symptomology were found
J. M. Krupinsky,
194
A. L. Scharen
and J. A. Schillinger
when inoculations with differing isolates of S. nodorum were made, even though environmental conditions were constant. We have shown, therefore, that the genetic propensities of the fungus population do indeed influence disease development and ultimate losses from S. nodorum infection even though stringent environmental conditions are required for any disease at all to develop. These environmental conditions include rainfall which is essential for the spread of the pathogen, and high humidity which is necessary for infection of the host and development of the pathogen [8]. To put it another way, when inoculum of S. nodorum is present and the host is in a receptive condition, then the environmental conditions control the initiation of disease development. If the environment is favorable, then the amount and intensity of disease development depends upon the pathogenicity of the fungus population. This situation is the one that should be attained in any search for resistance to S. nodorum under artificial conditions. REFERENCES A. (1968). Priifimg der Pathogen&t einiger St%mme von Septoria nodomm Berk. Phytopathologische ,+itschrift 62, 190-194. BR~NNIMANN, A. (1968). Zur Kenntnis von Septoria nodorum Berk., dem Erreger der Spelzenbraune und einer Blattdtire des Weizens. Phytofiathogix~ zeitschrift 61, 101-146. KIETREIBER, M. (1966). Atypical symptoms of S$toria nodorum on wheat seedlings. The behavior of the cultivar. Probus. Proceedines of the In&national Seed Test& Association 31, 179-186. SCHAREN, A. L. & KRUPINSKY, J. My (l!970). Cultural and inocula%on studies of Septoria nodoncm, cause of glume blotch of wheat. Phytopathology 60, 1480-1485. SCHAREN, A. L. & KRUPINSKY, J. M. (1969). Effect of Septia nodorum infection on CO, absorption and yield of wheat. Phytopathology 59, 1298-1301. SCHAREN, A. L. (1968). CO, absorption in four wheat varieties and differential responses to infection by S&to& nod&k. Phy&patholoD 58, 887 (Abstr.). SCHAREN. A. L. & TAYLOR. T. M. (1968). CO. assimilation and vield of Little Club wheat infected by Septoria nodo&.” Phytopkolo~ 58, G7-451. SCHAREN, A. L. (1964). Environmental influences on development of glume blotch in wheat. Phytofiathology 54, 300-303. McGraw-Hill, New STEEL, R. G. D. & TORRIE, J. H. (1960). P rink pl es and Procedures of Statistics. York. WOLF, D. D., PEARCE, R. B., CARLSON, G. E. & LEE, D. R. (1969). Measuring photosynthesis of attached leaves with air seal chambers. Crop Science 9, 24-27.
1. BRBNNIMANN, 2.
3. 4. 5. 6. 7. 8. 9. 10.
Plant Pathology
(1973) 3, 187-194
Pathogenic variation in Septoria nodorum (Berk.) Berk. in relation to organ specificity, apparent photosynthetic rate and yield of wheat? J. M. KRUPINSKY,$ A.L.
SCHAREN
Plant Science Research Division, Agricultural Research Service, United States Debartment of Agriculture, Plant Industry Station, Beltsville, Maryland
J. A.
and
Agronomy
20705,
U.S.A.
SCHILLINGER~
Department,
University of Maryland,
College
Park, Maryland
(Accepted for publication
Sepember
20740,
U.S.A.
1972)
Cultures of Seporia nodorum that varied in morphology were isolated from wheat plants that had been artificially inoculated with a composite culture. Other culture types were isolated from wheat leaves naturally infected in the field. Variation in pathogenicity of selected cultures was measured by symptom development, by apparent photosynthetic rates and by grain yield. Fungus variants that caused differences in photosynthesis, which were statistically significant at the 1 y0 level, were considered different in pathogenicity. Photosynthetic differences invariably were followed by yield differences. The role of environmental variation vs. pathogen variation was discussed.
INTRODUCTION Septoria nodorum (Berk.) Berk. (Leptosphaeria nodorum Mtiller) causes the glume blotch disease of wheat, Triticum aestiuum L. A major problem encountered in screening for resistance to S. nodorum has been the variability in disease intensity and yield loss due to environmental influences [Z, 81. Variation in the pathogen has been studied in pure culture [I, 3, 41, and in combination with wheat hosts [I, 41. The investigations cited above tended to show that environmental variation was the greatest deterrent to progress in selecting resistant host lines in the field. The present investigation was designed to study quantitatively the variation in pathogenicity of selected cultures of the fungus under rigidly controlled environmental conditions. It was hoped that some relative evaluation of the contribution of genetic variation in the fungus pathogen versus the variability caused by environmental factors could t Cooperative investigations of the National Agricultural Research Center, Agricultural Research Service, United States Department of Agriculture, and the University of Maryland Agricultural Experiment Station, Journal Paper No. A1745. This work is part of a thesis submitted by the senior author in partial fulf?lment of the requirements for the M.S. degree. f: Agricultural Research Technician and Research Plant Pathologist, respectively. Present address: USDA, ARS, Department of Botany, Montana State University, Bozeman, Montana 59715. f Associate Professor. 13
188
J. M. Krupinsky,
A. L. Scharen
and
J. A. Schillinger
be made as a result of these studies. In addition to visible symptoms and yields, apparent photosynthetic rates (APR) were measured to determine whether metabolic effects of the disease could be detected before visible symptoms of infection appeared, and as an additional measure of culture virulence. Comparisons of visible symptoms, APR and yield of grain permitted conclusions to be made concerning correlation of the aforementioned factors, and how they affect a search for genetic resistance to S. nodorum. MATERIALS
AND
METHODS
Cultures with maximum potential variability in morphology and pathogenicity were sought for our experiments. Since previous reports [Z, 41 had shown that a mixture of cultures collected in the geographic area of interest gave the most consistently useful results in inoculation experiments, we began with a composite collection of S. nodorum from seven locations. Wheat plants were inoculated with the composite, and after symptoms developed, isolations were made from infected leaves. Newly isolated cultures of S. nodorum that were derived from single spores always produced additional pycnidia and spores; however, indiscriminate mycelial transfers resulted in cultures that tended rapidly toward nonsporulating, mycelial types [a]. An extension of the single-spore isolation was the development of a mass spore transfer (MST) technique in which a mass of spores exuding from a pycnidium was picked up on a sterile needle or loop and transferred. MST were used as a rapid means of transferring cultures, while maintaining their sporulating ability and associated virulence. Three basic types of cultures were isolated that varied in production of pycnidia and spores, rate of mycelial growth and mycelial color. These were subsequently tested for virulence and effect on CO, exchange in wheat. For comparison, some cultures were isolated in the more traditional way by making a single-spore culture from a single plant at a single location. Two cultures of this type were used: N-40 from “Lerma Rojo 64-A” growing at Stoneville, Mississippi and N-42 from “Purplestraw” at Tifton, Georgia. These were maintained in culture by selective mass transfers of the “wild type”. Gas exchange analysis was done using an infrared CO, analyzer (IRGA). The intact head, peduncle and flag leaf of wheat, Wisconsin selection, C.I. 12632, were inserted into a Plexiglass air-sealed assimilation chamber [IO], and the APR was measured. A reading of APR was made prior to inoculation, then 14 daily measurements of photosynthesis in the head, peduncle and flag leaf were obtained to determine the effect of each S. nodorum isolate on APR [5]. All plants were grown in controlled environment chambers. The chambers had a 12-h photoperiod and the temperature was maintained at 2 1 “C during the day and 18 “C during the night. The light intensity was maintained at 70 + 5 x IO4 erg/ cm2 per s. For each study, six pots were selected that contained three plants which were just beginning to extrude anthers. The three treatments in each experiment included an uninoculated control head and flag leaf (treatment 1) plus two heads and flag leaves (treatments 2 and 3) that were inoculated with different culture types of S. nodorum. Details of inoculation are listed in Table 1. In the split plot design used for the IRGA studies, days were the whole unit treatments, and the three individual treatments were the subunits. An analysis of
Pathogenic
variation
in S.
189
nodorum
variance was run on each individual study, for treatment 1 us. the average of treatments
and orthogonal comparisons were made 2 and 3, and treatment 2 vs. 3 [9].
TABLE Isolates, Experiments and treatments Al 2 3 Bl
their description
and inoculation
Culture
type
Control Pycnidia Pink mycelia; pycnidia
2 3
Control Pycnidia Yellow mycelia; pycnidia
2 3
Control Pycnidia Pink mycelia; pycnidia
Cl
Dl 2 3 El 2 3 Fl 2 3
procedures
1 used in studying
Source of isolate
the pathogenic@ Concentration (spores and mycelia/ml)
of S. nodorum Incubation (‘4
Composite
15 x 10s
64
Composite
14x
106
64
Composite
12 x 10s
48
Composite
13 x 106
48
Composite
14 x 10s
48
Composite
13 x 106
48
Control Pink mycelia; no pycnidia Pycnidia
N-40 N-40
12 x 10s 14x 10s
64 64
Control White mycelia; no pycnidia Pycnidia
N-42 N-42
Control Gray-white mycelia; no pycnidia Pycnidia
N-40 N-40
8x 8x
lo6 lo6
72 72
12 x 10s 13 x 10s
72 72
RESULTS Decreases in APR were noted when heads and peduncles of test wheat were inoculated with selected cultures of 5’. nodorum (Fig. 1). Significant differences (1%) were observed in all studies and at all time intervals when the uninoculated control, treatment 1, was compared with the average of inoculated treatments 2 and 3. A sharp initial decrease in APR during the first 24 h after inoculation was seen in every experiment. At that time, no symptoms, or a few tiny chlorotic flecks were all that could be seen. Obvious visible symptoms of disease did not develop until 4 or 5 days after inoculation. Isolates were considered to be different from each other in pathogenicity only if they caused differences in APR significant at the 1 oh level. The dark areas in Figs 1 and 2 visualize the significant differences in the pathogenicity of isolates. When measuring the APR of the heads and peduncles in experiment A, a pycnidial isolate with white mycelium [Fig. 1 (a) 21 was less pathogenic than a pink mycelial isolate [Fig. 1 (a) 31. These isolates were also significantly different from one another in experiment C [Fig. 1 (c)l. Cultures used in treatments 2 and 3 of experiment B
190
J. M. Krupinsky,
A. L. Scharen
and
J. A. Schillinger
similarly reduced APR [Fig. 1(b)]. I n experiment D inoculation with a pycnidial isolate [Fig. 1(d) 31 re d uced APR more than similar inoculation with a pink mycelial isolate [Fig. l(d) 21. In experiment E, a white mycelial isolate [Fig. 1(e) 21 and a pycnidial isolate [Fig. 1(e) 31 had differential effects on the APR of the heads and peduncles for the entire period after inoculation. In experiment F a gray-white mycelial isolate [Fig. 1 (f) 21 reduced APR more than a pycnidial isolate [Fig. l(f) 31 during the first 8 days; however, after the twelfth day, the effect of the pycnidial isolate became greater.
(b)
0
11
0’
11
2
10
4
10
”
I””
6
8
IO
I
I
12 14 0 Days after inoculation
I
2
I
I
4
,
I
6
I
I
8
I
11
10
11
1
12
14
FIG. 1. Apparent photosynthetic rates of heads and peduncles of C.I. 12632 when inoculated at preanthesis stage of growth with different isolates of S. nodorum. Dark areas show significant differences ( 1%) in the pathogenicity of isolates : (a) pycnidial US. mycelial-pycnidial type; (b) pycnidial US. mycelial-pycnidial type; (c) pycnidial US. mycelial-pycnidial type; (d) mycelial US. pycnidial type; (e) mycelial US. pycnidial type and (f) mycelial US. pycnidial type. Legend: (o), control, treatment 1; (o), treatment 2; (0), treatment 3.
Pathogenic
variation
191
in S. nodorum
Flag leaves were less affected by S. nodorum inoculation than heads and peduncles (Fig. 2). In experiments A, D and F the APR of the control was significantly different from one inoculated treatment or the means of the two inoculated treatments. No significant differences were found in experiments B, C and E. The length of incubation time (48 h) was associated with poor S. nodorum infection in experiments B and C. In experiment E, the uninoculated flag leaves of control plants deteriorated for unknown reasons, resulting in the APR of the controls being lower than the APR of the infected for four consecutive days. In experiment A, a pycnidial isolate [Fig. 2(a) 21 reduced APR more than a pink mycelial isolate [Fig. 2(a) 31 and in
I’ll
I’
I
’
I
/
I
i
1’1’1
I
1 I
8
IO
(b)
II o Days after
FIG. 2. Apparent
I 2
/
I 4
/
6
I
I I2
inoculation
photosynthetic rates of flag leaves of C.I. 12632 when inoculated at preanthesis stage of growth with different isolates of S. nodomm. Dark areas show significant differences (1%) in the pathogenicity of isolates: (a) pycnidial VS. mycelial-pycnidial type; (b) pycnidial us. mycelial-pycnidial type; (c) pycnidial US. mycelial-pycnidial type; (d) myceiial US. pycnidial type; (e) mycelial US. pycnidial type and (f) mycelial US. pycnidial type. Legend as for Fig. 1.
I
" I 14
192
J. M. Krupinsky,
A. L. Scharen
and J. A. Schillinger
experiment D, a pycnidial isolate [Fig. 2(d) 31 was more effective in reducing APR than a pink mycelial isolate [Fig. 2(d) 21. Finally, a pycnidial isolate [Fig. 2 (f )] reduced APR more than a gray-white mycelial isolate [Fig. 2(f) 21 in experiment F. Other experiments [Fig. 2(b) (c) and (e)] detected no differences between treatments 2 and 3. The APR during the 15-day period of the experiments were averaged, and percentage of reduction was calculated for treatments 2 and 3 (Table 2). All isolates, excepting one, caused a greater decrease in APR on the heads and peduncles than on the flag leaves. S. nodorum infection caused an average 43% loss in APR on heads and peduncles and a 21 O/Oloss on the flag leaves. The decrease in APR of the heads and peduncles ranged from 26 to 55%, while the loss on the flag leaves ranged from 5 to 47%. When the APR of the infected heads, peduncles and flag leaves were added together, fungus infection caused an overall loss in APR of 3 1%. The overall range was 17 to 50%. TABLE
The average apparent Experiment study
photosynthetic
Treatmenta
A
B
C
D
E
F
rate for
Heads and pedunclesb
Reduction (%)
1 2 3
2.60 1.46 1.18
43.8 54.6
1 2 3
3.13 1.90 1.78
:
of plant parts
with S.
Flag leavesb
nodorum Reduction (%)
3.11
1.65 1.91
46.9 38.6
39.3 43.1
2.34 2.22 2.01
5.1 14.1
3
3.23 2.11 l-83
34.7 43.3
3-28 3.07 3.00
6.4 8.5
1 2 3
3.41 2.06 1.52
39.6 55.4
3.21 2.71 1.77
15.6 44.7
1 2 3
2.89 1.77 2.13
38.8 26.3
2.37 2.13 2.21
10.1 6.8
:
3.88 2.07 2.18
46.6 43.8
3.37 2.79 2.57
17.2 23.7
3 Total Total
2
I5 days after inonclation
for 1 for 2 and 3 2
a 1 = Uninoculated control. 2 and b APR = mg (CO&/organ per h.
19.14 IO.99
3 = Different
17.68 14.02
42.6
isolates
of S. nodomm.
The 1000 kernel weight and the grain weight per head were measured (Table 3). The loss in grain weight per head of inoculated plants ranged from 8 to 56%, and the overall loss averaged 31%. These variations in yield loss also demonstrate the variability in pathogenicity of S. nodorum.
Pathogenic
variation in S. nodorum
193 TABLE
Average yield
Study
of C.I.
12632
inoculated 1000 kernel weight (g)
Treatment
at preanthesis
3 stage of growth
Reduction (%I
with isolates of S. nodorum
Grain weight per h=d k)
Reduction (%I
1 2 3
33.04 20.15 20.12
39-l 39-2
1.19 0.66 0.71
44.6 40.6
1 2 3
35.34 24.20 27.58
31.6 22.0
1.48 1.00 l-11
32.5 25-l
C
: 3
32-99 25.28 26.90
23.4 18.5
1.50 1.16 l-18
22.7 21.1
D
1 2 3
31.30 2944 16.38
6.0 47.7
1.29 l-20 0.56
7.5 56.3
1 2 3
27Ti 21.17 20.79
23.2 24.6
1.06 0.79 0.84
26.0 21.3
1 2 3
33.97 23.69 17.93
30.3 47.3
1.26 0.89 0.68
29.0 45.9
A
B
E
F
Total Total
for 1 for 2 and 2
194.22 3
136.83
7.80 29.6
5.41
30.8
DISCUSSION
As reported previously [5-71, APR in wheat was reduced by 5’. nodorum infection. In the case of heads and peduncles, it was possible to distinguish the control from the infected material by means of gas exchange analysis from the first through the fourteenth day after inoculation. Decreases in APR were evident earlier than previously reported [5]. This is probably due to inoculation with increased numbers of spores, since greater reductions in yield have been associated with higher spore concentrations [2]. As in an earlier study [5j, the decrease in APR was not as evident with the flag leaves as with the head and peduncle. Under the experimental conditions used, a minimum of 64 h incubation was needed for flag leaves to become well infected. The selected mycelial types of S. nodorum or the mycelial type with pycnidia were sometimes more pathogenic than the typical pycnidial sorts. When the APR of the heads and peduncles were studied, the mycelial type and the mycelial type with pycnidia were more pathogenic than the pycnidial cultures. When flag leaves were inoculated and studied, the typical pycnidial type was more pathogenic than the selected mycelial type. Thus, the pycnidial type is more pathogenic on the flag leaves, indicating again the variation in pathogenicity among the isolates as well as variations due to the part of the plant infected. Variation in pathogenicity between different types of S. nodorum has been demonstrated. Considerable differences in APR, yield and symptomology were found
J. M. Krupinsky,
194
A. L. Scharen
and J. A. Schillinger
when inoculations with differing isolates of S. nodorum were made, even though environmental conditions were constant. We have shown, therefore, that the genetic propensities of the fungus population do indeed influence disease development and ultimate losses from S. nodorum infection even though stringent environmental conditions are required for any disease at all to develop. These environmental conditions include rainfall which is essential for the spread of the pathogen, and high humidity which is necessary for infection of the host and development of the pathogen [8]. To put it another way, when inoculum of S. nodorum is present and the host is in a receptive condition, then the environmental conditions control the initiation of disease development. If the environment is favorable, then the amount and intensity of disease development depends upon the pathogenicity of the fungus population. This situation is the one that should be attained in any search for resistance to S. nodorum under artificial conditions. REFERENCES A. (1968). Priifimg der Pathogen&t einiger St%mme von Septoria nodomm Berk. Phytopathologische ,+itschrift 62, 190-194. BR~NNIMANN, A. (1968). Zur Kenntnis von Septoria nodorum Berk., dem Erreger der Spelzenbraune und einer Blattdtire des Weizens. Phytofiathogix~ zeitschrift 61, 101-146. KIETREIBER, M. (1966). Atypical symptoms of S$toria nodorum on wheat seedlings. The behavior of the cultivar. Probus. Proceedines of the In&national Seed Test& Association 31, 179-186. SCHAREN, A. L. & KRUPINSKY, J. My (l!970). Cultural and inocula%on studies of Septoria nodoncm, cause of glume blotch of wheat. Phytopathology 60, 1480-1485. SCHAREN, A. L. & KRUPINSKY, J. M. (1969). Effect of Septia nodorum infection on CO, absorption and yield of wheat. Phytopathology 59, 1298-1301. SCHAREN, A. L. (1968). CO, absorption in four wheat varieties and differential responses to infection by S&to& nod&k. Phy&patholoD 58, 887 (Abstr.). SCHAREN. A. L. & TAYLOR. T. M. (1968). CO. assimilation and vield of Little Club wheat infected by Septoria nodo&.” Phytopkolo~ 58, G7-451. SCHAREN, A. L. (1964). Environmental influences on development of glume blotch in wheat. Phytofiathology 54, 300-303. McGraw-Hill, New STEEL, R. G. D. & TORRIE, J. H. (1960). P rink pl es and Procedures of Statistics. York. WOLF, D. D., PEARCE, R. B., CARLSON, G. E. & LEE, D. R. (1969). Measuring photosynthesis of attached leaves with air seal chambers. Crop Science 9, 24-27.
1. BRBNNIMANN, 2.
3. 4. 5. 6. 7. 8. 9. 10.