Cucurbit Genetics Cooperative Report 4:17-19 (article 8) 1981
Pathogenicity of Isolates of Didymella bryoniae
and Reisolation of the Fungus Out of Symptomless Plants
A. P. M. den Nijs and A. C. van der Giessen
Institute
for Horticultural Plant Breeding (IVT), P. O. B. 16, Wageningen,
The Netherlands
Five different isolates of Didymella bryoniae
(Auersw.) Rehm, causal fungus of fruit and stem rot, were
separately used to inoculate three cucumber genotypes with
different levels of resistance, to detect possible differences
in pathogenicity. Three tests were carried out, the first
in 1978 and the last two in 1980. The isolates had been
collected from 1974 to 1977 by Van Steekelenburg (Research
Institute for Plant Protection) and were subcultured manifold
on malt extract agar (1). The plant material consisted of
two IVT breeding lines with different levels of resistance
against fruit and stem rot and were derived from cv. Rheinische
Vorgebirge (2). Susceptible check was cv. Levo. The experimental
design was a split block with eight replications of two
plants per treatment randomly arranged in two plastic tents.
The plants were inoculated in the potted plant stage, as
described earlier (1). We inoculated the plants with each
isolate separately, while shielding the remainder of the
plants and taking care to keep the treatments apart.
Average visual disease ratings according to the scale in
(1) of all 16 plants per treatment are in Table 1. These
are the results of one test only, inoculated October 10,
1980 and evaluated October 20, 1980. The data from the other
two tests are similar. The ratings in the water-control
treatment may reflect damage from water-logging due to the
extremely high relative humidity under the tents. ANOVA
of all the data revealed that the apparent resistance level
of the three genotypes differed significantly over all isolates
(F2/14 = 23.97**). The isolates were also significantly
different (F3/35 = 29.02**). Separate analyses per host
genotype established that on line A, isolate M77-3 was more
pathogenic than all others with M74-3 as second most pathogenic,
both significantly differing from the water-control. On
line B both isolates were equally pathogenic and both differed
significantly from the others. On 'Levo' their pathogenicity
was similar, and all isolates induced significantly more
symptoms than the water-control. Yet there was no significant
interaction between isolates and host genotypes (F10/70 = 1.94), so from this test we cannot conclude that physiological
races of D. bryoniae exists, although differences
in overall pathogenicity were clear. The results of the
other two experiments pointed in the same direction. There
was a noteworthy large difference in the coefficient of
variation of the two lines (67%) in comparison with that
of cv. Levo (26%). The lines appear to be segregating. There
was almost complete concordance of the ranking of all five
isolates on susceptible cv. Levo in all three tests. Apparently
no relative change in pathogenicity has occurred in these
isolates for over two years, despite frequent subculturing
under blacklight and the fortuitous selection for in
vitro sporulation (1).
The value of the resistance found in the potted plant stage
depends on how well it holds out in plants during the cropping
period. Initial results of correlation studies with related
breeding lines were not reassuring (3). We have investigated
whether symptomless plants in our tests were indeed free
of the fungus following 4-5 weeks of recovery after the
test. Therefore, we attempted to reisolate the fungus from
pieces (length 0.5-1 cm) of disinfected internodes cultured
according to (1). We also applied the same technique to
plants with mild, moderate and severe symptoms in the potted
plant stage, but at the time of dissection, all plants were
without symptoms. The plants had been inoculated with very
pathogenic isolate M77-3 or with M74-4. Fungal growth was
assessed following five days of incubation of the stem pieces.
From almost all of 67 dissected plants the fungus was
reisolated from the hypocotyl. From less that half of the
plants we also obtained fungal colonies from higher internodes,
often from the first one or from the top of the plant (9th to 15th internode). Only one to three internodes per
plant yielded positive reisolation results. A summary is
in Table 2. The most resistant line contained most plants
from which the fungus was reisolated. From plants that were
severely diseased in the potted plant test we obtained less
often fungal growth than from moderately or slightly attacked
plants. From one-third of the plants that did not show symptoms
in the potted plant test, we were able to reisolate Didymella,
so these plants apparently carried it without visible damage.
The results of both isolates differed slightly.
These preliminary data caution against overreliance on
the potted plant test. Other reisolation attempts out of
inoculated plants of up to four months of age indicate that
the fungus may be present in the plants for a long time
unnoticed, and possibly break out in disease when the environment
and condition of the plants are right.
Table 1. Average disease rating of five isolates of Didymella bryoniae on two lines and cv. Levo in a potted plant test.
Isolate |
Line A |
Line B |
Levo |
Mean |
M74-2 |
0.61z |
0.60 |
1.76 |
0.99 |
M74-3 |
1.60 |
1.30 |
2.30 |
1.73 |
M74-4 |
1.08 |
0.43 |
2.05 |
1.19 |
M75-3 |
1.20 |
0.59 |
1.51 |
1.10 |
M77-3 |
2.29 |
1.28 |
2.33 |
1.97 |
Mean |
1.36 |
0.84 |
1.99 |
1.40 |
Water |
0.16 |
0.06 |
0.39 |
0.20 |
z Average of 8 plots of 2 plants.
Table 2. Number of plants from which Didymella bryoniae was reisolated, as a fraction of the number of plants tested (plants were inoculated in a potted plant test 6 weeks before dissection).z
Line |
Isolate |
0 |
1 |
2 |
3 |
Mean |
A |
M77-3 |
- |
3/3 |
- |
1/8 |
4/11 |
|
M74-4 |
2/6 |
- |
1/1 |
0/2 |
3/9 |
B |
M77-3 |
1/2 |
3/5 |
8/8 |
- |
12/15 |
|
M74-4 |
2/8 |
0/1 |
0/1 |
- |
2/10 |
Levo |
M77-3 |
- |
0/1 |
3/5 |
0/5 |
3/11 |
|
M74-4 |
- |
1/2 |
1/5 |
1/4 |
3/11 |
|
Mean |
5/16 |
7/12 |
13/20 |
2/9 |
27/67 |
z Only internodes above hypocotyl considered.
Literature Cited
- Giessen, A. C. van der and A. P. M. den Nijs. 1981.
In vitro growth and sporulation of Didymella
bryoniae and a glasshouse method for screening for
resistance. Cucurbit Genetics Coop. Rpt. 4:6-8.
- Meer, Q. P. van der, J. L. van Bennekom and A. C. van
der Giessen. 1978. Gummy stem blight resistance of cucumbers
(Cucumis sativus L.). Euphytica 27:861-864.
- Steekelenburg, N. A. M. van. 1981. Comparison of inoculation
methods with Didymella bryoniae on Cucumis
sativus L. Euphytica (accepted).
