PROJECT RESULTS

Effects of Cropping Practices on Blackleg Disease of Canola

Applicant: 

Dr. Dilantha Fernando

Department of Plant Science

University of Manitoba

Winnipeg, Manitoba  R3T 2N2  Canada

Table of Contents:

• Effects of Rotation and Tillage on Blackleg Disease of Canola
• Persistence of Blackleg Pathogen in the Soil
• Diurnal Release of Ascospores and Spore Gradients

ARDI Project:

#98-242

Total Approved:

$45,453

Date Approved:

April 16, 1999

Project Status:

Completed January, 2003

Canola provides one of the world's major edible vegetable oils.  It belongs to the mustard family cruciferae, including the species Brassica napus L., Brassica rapa L., Brassica carinata L. and Brassica juncea (L.) Coss.  In Canada, the crop consists of spring-sown varieties of B. napus and B. rapa.  B. napus varieties are grown widely in Manitoba and Saskatchewan.

Blackleg disease, caused by Leptosphaeria maculans (Desm.) Ces. Et de Not. (Anamorph: Phoma lingam (Tode ex Fr.) Desm.) is a major disease in many areas where canola is grown.  L. maculans occurs in a weakly virulent form and a virulent form.  The virulent strain was first detected in 1975 in east central Saskatchewan.  Then the disease has spread rapidly to most canola growing areas of Saskatchewan and into Manitoba and Alberta, it is considered to be a serious threat to canola production in Canada.  L. maculans overwinters on canola stubble and produces spores in the spring; this becomes inoculum for infection of the new crop in the summer.  The cultural practices by farmers may increase disease inoculum, and a minimal level under any cultural practice may be enough to cause an epidemic if the environmental conditions favor the spores release and infection process.

The life cycle is well understood, but little is known about the epidemiology of the pathogen.  A handle on knowledge of the epidemiology will help understand good farming practices to avoid epidemics.  The goal of this project is to develop useful disease management strategies (rotation and tillage) based on the knowledge of the epidemiology of the pathogen.  Therefore, this project is broken up into several experiments to serve it:

1. Effects of rotation and tillage on blackleg disease of canola.
2. Persistence of blackleg pathogen in the soil.
3. Diurnal release of ascospores and spore gradients.

EFFECTS OF ROTATION AND TILLAGE ON BLACKLEG DISEASE OF CANOLA

Objective:

Rotation and tillage are important disease management methods.  This experiment was to examine the impact of rotation and tillage on blackleg spread and infection to canola.

Procedure and Project Activities:

This study was conducted at the Carman Research Station from 1999 to 2002.  Soil type was loam.  A split-spot design with three replications was used with conventional till (T) and zero till (Z) as main plots and canola (C) rotation with wheat (W) and flax (F) as subplots.  So there were eight treatments in total: CCCCZ, CCCCT, CWCCZ, CWCCT, CCWCZ, CCWCT, CWFCZ and CWFCT.  The experiment field size was 155 m x 81 m.  There were 24 plots; each plot size was 10 m x 4 m.  The distance was 15 m between plots and 10 m between a plot and the field border.  The canola variety was Westar (susceptible to blackleg disease); the wheat was Barrie and the flax was Norlin.  Border crop was sown to oats.  The canola was seed on May 30 each year; wheat and flax were seeded the same day in the second and third year respectively.  In 1999, 24 plots of canola plants were inoculated by spraying the blackleg (PG-2 isolate 86-12) pycnidiospore suspension at 1x10⁷ spores/ml at the 3-leaf stage of canola.

Measurements - 2001

In a 1 m x 1 m quadrant in the center of each canola plot, every other plant was assessed for leaf area infection at different growth stages, and at maturity for disease severity and incidence.

Measurements – 2002

Number of Lesions:  Zigzag-shape sampling method was used in each canola plot.  The zigzag line was composed of seven segment lines of equal length; the length of each segment line was 2.5 m.  The angle between two neighbor segment lines was 60°.  The starting point was 1.0 m and 0.6 m away from the long and short edge respectively.  Four canola plants were selected equidistance from each other along each segment line except the last segment line in which five plants were selected the same way.  There were 29 plants in total selected and measured for the number of lesions on each leaf of the plant selected at different growth stages of canola.

Percent Leaf Lesion Size over Leaf Size:  In a l m x l m quadrant in the center of each canola plot, all the plants with the No.4 leaves infected by blackleg were marked on July 7.  A digital picture was taken of the marked leaves.  A 20 cm distance between the leaf and the camera lens was kept each time.  The pictures were transferred into the computer, and the percent leaf lesion size was automatically calculated by the APS ASSESS program.

Stem Severity and Disease Incidence:  In a 9 m x 3 m rectangle in each canola plot with a 0.5 m margin, 10 lines were well distributed along the long edge with 1m between two neighbor lines.  At canola filling stage (seeds in pods turn yellow), 10 plants were evenly sampled along each line with about 0.3 m between two neighbor picked plants for the evaluation of stem severity and disease incidence.

Analyses

1. Number of infected plants:  Total number of infected plants at each measuring time.

2. Number of infected leaves per plant:  (total number of all infected leaves on all the infected plants) / (number of all the infected plants).

3. Number of lesions per plant:  (total number of all the lesions on one plant) / (number of all the infected plants).

4. Percent leaf lesion size:  APS ASSESS program.

5. Stem severity:  0 ~ 5 class system:  0 - no infection; 1 - lesion area is less than 25% of cross-section area of crown; 2 – 25 ~ 50%; 3 – 51 ~ 75%; 4 – 76 ~ 100%; 5 - plant dead.  Stem severity was evaluated as average stem severity over all the infected plants.

6. Disease incidence (%):  [(number of infected plants) / (total number of the sampled plants)] x 100%.

7. A LSD test at the 0.05 significant level was used for examining the differences among the treatments.

Results and Discussion:

Year 2001:

Figure 1.  Percent area covered by lesions on plants showing symptoms prior to July 10, 2001.

Figure 2.  Percent area covered by lesions on plants showing symptoms between July 10th to 13th, 2001.

Figure 3.  Stem severity of different treatments.

Figure 4.  Incidence of different treatments.

The results from year 2001 showed that the number and size of lesions on canola in the tilled canola-wheat-canola rotation (CWCT) was significantly lower than CCCT at all growth stages.  Similar significant results were obtained with CWCZ<CCCZ, CWCT<CWCZ and CCCT<CCCZ.  Stem severity and disease incidence at maturity were significantly lower when canola was rotated with wheat, and was grown on tilled plots.  Stem severity of plants in CWCT, CCCT, CWCZ and CCCZ were 2.9, 3.2, 3.4, and 3.6, respectively.  Disease incidence of plants in CWCT, CCCT, CWCZ and CCCZ were 49%, 57%. 63% and 81%.  The number of leaf lesions, their size, disease incidence and stem severity in the plant population suggests that appropriate rotation and tillage could significantly reduce blackleg of canola in farmers’ fields.  The effect of rotations with flax on blackleg disease in canola will be investigated.

Year 2002:

Significant Differences of Number of Lesions Among the Treatments

Different letters within a column indicate significant differences according to the Least Significant Difference (LSD) test (P=0.05).

Significant Differences of Percent Lesion Size Among the Treatments

Different letters within a column indicate significant differences according to the Least Significant Difference (LSD) test (P=0.05).

The results from year 2002 showed that there was no visible symptom showing up on leaves when canola was rotated with wheat and flax whether they were grown on plots tilled or not, or when canola was only rotated with wheat in the second or third year and they were grown on the tilled plots.  The number and percent lesion size in CCCCT was significantly lower than CWWCT and CCWCZ; CWCCZ and CCWCZ were individually significantly lower than CCCCZ.  There was no significant difference of number of lesions between CWCCZ and CCWCZ but percent lesion size.  Stem severity in 2002 and disease incidence in 2001 and 2002 were significantly lower when canola was rotated with wheat or/and flax, and were grown on tilled plots.  Disease incidence of plants in CWCT, CCCT, CWCZ and CCCZ were 48.9%, 57.2%, 63.7% and 80.6% respectively.  Stem severity of plants in CCCCT, CCWCZ, CWCCZ and CCCCZ were 1.34, 1.94, 2.01 and 3.07 respectively; there were significant differences of stem severity among CCWCT, CWFCZ, CWFCT and CWCCT, stem severity ranged from 1.04 to 1.18.  Disease incidence of plants in CCCCT, CCWCZ, CWCCZ and CCCCZ were 49.3%, 56.7%, 61.0% and 79.3% respectively; disease incidence of CCWCT, CWFCZ, CWFCT and CWCCT ranged from 16.3% to 21.5%.  The number and size of lesions, disease incidence and stem severity in the plant population suggests rotation and tillage could significantly reduce blackleg of canola in farmers’ fields.

PERSISTENCE OF BLACKLEG PATHOGEN IN THE SOIL

Objective:

To study the viability and virulence of the blackleg pathogen on lesions left on the soil surface or buried at different depths.

Procedure and Project Activities:

Materials - Canola - Westar; Blackleg-PG-2 isolate 86-12.

Treatments - Stubble showing typical blackleg lesions was collected and buried at the surface, 5 cm and 10 cm levels in three types of soil (clay, loam and sand soil) before the winter of 2001.  There were three replications in each type of soil plot.  In the spring of 2002, the samples were dug up on May 31, June 30 and July 28.  The stubble was washed under running cold water and the lesion area was cut into 1 cm pieces, which were plated on V-8 juice medium amended with Streptomycin Sulfate, Rose Bengal and CaC0₃ and measure the pathogen viability. 

Calculation of Pathogen Viability - (number of plates with spores available growing on them) / (number of total plates stubble was incubated on).

Results and Discussion:

With soil depth and canola growth stage, the viability of the blackleg pathogen significantly decreased.  On May 31 (about 7 months after burial) the viability at the depth of 0 cm, 5 cm and 10 cm of the sand soil was 77%, 57% and 37% respectively; on June 30 (8 months after) the viability was 60%, 47% and 27% respectively; and on July 28 (9 months after) it reduced to 57%, 37% and 23% respectively.  There were also significant differences among the different types of soil. On July 28, the viability at the three depths of the sand soil was 57%, 37% and 23%, and it was 47%, 27% and 10% respectively in the clay soil. This study initially presents that soil depth and type have reduced persistence of the blackleg pathogen from canola stubble. The risk of canola yield loss to the blackleg disease can be largely decreased by deeply ploughing after harvesting without lots of previous-year infected stubble brought out from soil. And this effect can be strengthened on clay soil.

Variability (%) of Blackleg Pathogen in Different Depths of Different Types of Soil

DIURNAL RELEASE OF ASCOSPORE AND SPORE GRADIENTS

Objective:

To correlate stage of crop development in the current year with ascospore discharge from the debris of the previous year's crop.  Understand the distance spores travel and how neighboring fields could be in danger of being infected.

Procedure and Project Activities:

Variety – Canola –Westar

Treatment - In year 2000, a 25x25m area was planted within the 45x126m block to the canola variety, Westar.  The 25x25m area was inoculated with a pycnidiospore suspension at 1x10⁷ spores/ml.  In year 2001, the entire 45x126m block was sown to Westar.  Rotorod impaction spore samplers were set up in the field in six directions NE, E, SE, SW, W and NW to catch blackleg ascospores.  A Campbell Scientific CR10 datalogger was programmed to run the rotorods for five minutes out of every hour.  Each rotorod was shifted every 24 hours and the spores on the rotorods were counted under the microscope.  Also, a seven-day Burkard spore trap was set up in the center of the block.  The tape was shifted every 7 days and divided into 7 fields equally, each field represents one day, spores were counted by each day.  Rainfall was collected from the weather station at the Carman Research Station.

In year 2001, the experiment was repeated in another field.  The rotorod sampler was set up in eight directions N, NE, E, SE, S, SW, W and NW.

Figure 5.  Schematic locations of spore traps in the field (2001).

Figure 6.  Schematic locations of spore traps in the field (2002).

Results and Discussion:

With the distance far from the inoculum's source, the concentration of spores released obviously decreased. Along the wind direction, the spores (ascospores and pycnidiospores) were released more than other directions. A few of spores were released beyond the cycle area of 25m radius. Ascospores were first released on June 21. Pycnidiospores were first trapped on June 30. The pycnidiospores released were more than the ascospores at the later stage of canola growth. Ascospores and pycnidiospores released decreased over the canola growth stage. In addition, the rainfall had significant effect on the spore release, the more the rainfall, the more the spores released. Pycnidiospores were released with raining, but ascospore release was found to have an about one day delay. However, this relationship was not always directly proportional. At the later stage, ascospores were released fewer, even though the rain.

Figure 7.  Spore release and rainfall (2001).

Figure 8.  Spore release and rainfall (2002).

Figure 9.  Diurnal spore release (2001).

Figure 10.  Diurnal spore release (2002).

Figure 11.  Ascospore release in SE direction (2001).

Figure 12.  Pycnodiospore release in SE direction (2001).

Figure 13.  Ascospore release in NW direction (2001).

Figure 14.  Pycnodiospore release in NW direction (2001).

Figure 15.  Ascospore release in SW direction (2001).

Figure 16.  Pycnodiospore release in SW direction (2001).

Figure 17.  Ascospore release in NE direction (2001).

Figure 18.  Pycnodiospore release in NE direction (2001).

Figure 19.  Ascospore release in E and W direction (2001).

Figure 20.  Pycnodiospore release in E and W direction (2001).

Figure 21.  Spore release at different distances from inoculum source (2002).

Figure 22.  Average spore release in different directions with northwest wind (NW) (2002).

Figure 23.  Average spore release in different directions with southeast wind (SE) (2002).

Figure 24.  Average spore release in different directions with northwest and northeast wind (NW and NE) (2002).

Figure 25.  Average spore release in different directions with northeast and southeast wind (NE and SE) (2002).

Figure 26.  Average spore release in different directions with southwest and southeast wind (SW and SE) (2002).

References:

X. Guo, W.G.D. Fernando and M. Entz, 2006. Dynamics of infection by Leptosphaeria maculans on canola (Brassica napus) as influenced by crop rotation and tillage.  Phytopathology and Plant Protection – DOI: 10.1080/PD-1-10.

X.W. Guo, W.G.D. Fernando, and M. Entz, 2005.  Effects of crop rotation and tillage on blackleg disease of canola. Can. J. Plant Pathol. 27:53-57.

Guo, X.W., and Fernando, W.G.D. 2005. Seasonal and Diurnal patterns of Spore Dispersal by Leptosphaeria maculans from canola stubble in relation to environmental conditions. Plant Disease 89:97-104.

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