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PLANT CULTURE
Growth chamber
|
Container.......... |
Flats |
| Media............... |
Fine sand |
|
Seed Depth....... |
1.3 cm |
|
Temp/Light........ |
20ƒC; continuous light (approx. 100 @mol
m-2s-1) |
|
No. of Plants .... |
40 to 50 per replication |
| No.
of Reps....... |
4 minimum |
INOCULUM CULTURE AND PREPARATION
|
Source.............. |
Conidia (sporangia) from several
locations should be represented; conidia from field
plants usually are contaminated and germinate poorly; to
reduce contaminants add 50 mg
nystatin and 10 mg tetracycline/mL of inoculum (6); only conidia produced in
the lab should be used as inoculum for tests. |
| Storage............. |
Conidium viability declines rapidly if
harvest is delayed or if conidia are exposed to dry air
for more than a few minutes (4); however, a low
percentage of conidia will survive for a few weeks on
diseased seedlings stored at -20ƒC; conidia will remain
viable for many years in liquid nitrogen (1) |
| Production.........
|
Conidia form only during darkness at near
100% relative humidity; they will not form in free water
(4); to produce conidia place flats of infected plants (6
days after inoculation) into darkened, near-airtight
containers (we use plastic sweater boxes covered with
aluminum foil) about 16 hours before conidia are needed
for inoculum. |
| Preparation........ |
Remove flats from darkened containers,
immediately harvest plants, and place them into a jar
containing chlorine-free water (3); close the jar and
shake it vigorously to dislodge conidia; pour the spore
suspension through a tea strainer to remove plant debris;
adjust concentration
to at least 25,000 viable conidia per mL water and use
immediately.
|
INOCULATION
PROCEDURE
|
Inoculation ........ |
Spray suspension onto seedlings until a
drop forms between the cotyledons; to determine
viability, spray inoculum onto a slide, place on a filter
paper saturated with distilled water in a closed petri
dish, incubate in a dark area at 20ƒC, and 24 hours
later determine percent conidia with germ tubes |
SCHEDULE
The following schedule requires little attention on week
ends.
Day 1 (Th.) .....Plant seeds 1.3 cm deep in rows at least
2.5 cm apart in flats of fine sand
Days 3 to 6 .....Sprinkle water on flats twice daily to
settle the sand around the emerging seedlings
Day 5 (M 4pm) Induce sporulation on plants seeded the
previous week for inoculum production by placing them in
darkened containers
Day 6 (T 8am) Inoculate seedlings and place in darkened
containers
Day 7 (~ 8am) Remove flats from con~ainers and rogue any
plants that have emerged since inoculation
Days 8 to 12 .... Continue roguing newly emerged plants
Day 12 (M 4pm) Induce sporulation by placing flats of
plants into darkened containers
Day 13 (T 8am) Evaluate test
RATING
Evaluation of cultivars is based on the percentage of
resistant (symptomless) plants compared with the resistant check cultivar.
CHECK CULTIVARS
Expected symptomless plants (%)
| Resistant |
|
|
| Saranac** ... |
15-20 |
isolates I5 and 17 |
| |
50-60 |
isolate I8 |
| KS208(5)**... |
80-90 |
all isolates tested |
| Susceptible |
|
|
| Kanza** .... |
0-5 |
all isolates tested |
DISTRIBUTION AND SEVERITY OF
DOWNY MILDEW
Downy mildew, Peronospora trifoliorum de Bary
(Click on the map for an enlarged version See also the KEY)
SOURCE OF INOCULUM
Name ........Donald L. Stuteville
Address .....Department of Plant Pathology
Throckmorton Hall
Kansas State University
Manhattan, KS 66506-5502
Phone .............. 785-532-6176
SCIENTISTS WITH EXPERTISE
Name .........Donald L. Stuteville
Address ......Department of Plant Pathology
Throckmorton Hall
Kansas State University
Manhattan, KS 66506-5502
Phone ........785-532-6176
Name .........Daniel Z. Skinner
Address.......USDA/ARS
Dept. of Agronomy
Throckmorton Hall
Kansas State University
Manhattan, KS 66506-5501
Phone ........785-532-7247
CORRELATION TO FIELD
REACTION
Resistance among cultivars to the same race correlates
well, but the percentage of resistant (symptomless)
plants is typically much lower in the seedling test than
in the field.
RACES
Several races are known.
ALTERNATIVE METHODS
Downy mildew resistance of spaced plants in the field can
be evaluated following epiphytotics of downy mildew which
may occur during spring and fall (2):
1 Resistant ......No symptoms
2 Resistant ......Small, usually nonsporulating lesions
on one or two leaves
3 Susceptible ..Sporulating lesions on 10 to 25% of the
leaves
4 Susceptible .. General infection over the entire plant.
5 Susceptible .. Dead
Plants classified as 1 or 2 are considered resistant. Use
ASI or percentage of resistant plants to compare
cultivars.
REFERENCES
1. Bromfield, K.R., and C.G. Schmitt. 1967. Cryogenic
storage of conidia of Peronospora tabacina.
Phytopathology 57: 1133.
2. Elgin, J.H., JR., and D.K. Barnes, Eds. 1984. In:
Standard tests to characterize pest resistance in alfalfa
cultivars. USDA, ARS, Misc. Pub. No. 1434.
3. Fried, P.M., and D.L. Stuteville. 1975. Effect of
chlorine on Peronospora trifoliorum sporangial production
and germination. Phytopathology 65:929-930.
4. Fried, P.M., and D.L. Stuteville. 1977. Peronospora
trifoliorum sporangium development and effects of
humidity and light on discharge and germination.
Phytopathology 67:890-894.
5. Sorensen, E.L., D.L. Stuteville, and E.K. Horber.
1989. Registration of KS208 Alfalfa germplasm with
resistance to five diseases and three insects. Crop
Sci.29: 1094-1095.
6. Stuteville, D.L. 1977. Antibiotics that selectively
inhibit bacteria and fungi antagonistic to Peronospora
trifoliorum. Proc. Am. Phytopathol. Soc. 4:167-168
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