E. D. Mytsa, L.Yu. Kokaeva, S.N. Elansky
Oomycete Phytophthora infestans (Mont) de Bary causes late blight, a dangerous disease of potatoes and tomatoes. One of the main sources of primary inoculum P. infestans are thick-walled resting reproductive structures - oospores.
Hybrid oospores formed as a result of crossing genetically different parental strains contribute to an increase in genotypic diversity in the population, as a result of which the process of adaptation of strains to new varieties and applied fungicides is accelerated.
Oospore formation P. infestans in the field was noted in many countries of the world: Russia (Smirnov et al., 1999), Norway (Hermansen et al., 2002), Sweden (Strömberg et al., 2001), the Netherlands (Kessel et al., 2002) and others regions. Oospores are able to survive for more than 2 years in the soil in a viable state (Bødker et al., 2006) and cause infection of plants after overwintering (Ulanova et al., 2010; Lehtinen et al., 2002).
Earlier it was shown that the leaves of plants of different tiers of the potato bush have differences in resistance to late blight; it increases from the lower leaves to the upper ones. This trend does not depend on leaf age, plant age, and potato variety (Vesper et al., 2003). At the same time, we were unable to find data on differences in the intensity of oospore formation in the leaves of different layers of the potato bush in the literature. However, if such differences exist, then they should be taken into account when taking leaf samples for the analysis of oospore formation and when interpreting the results obtained by other authors. The aim of this work was to study the formation of oospores in the detached leaves of different layers of a potato plant, placed in the same conditions in humid chambers.
Materials and methods
We used 5 isolates P. infestans different types of mating isolated from affected samples brought from Moscow, Ryazan and Leningrad regions. Of these, 3 pairs of strains of different mating types were selected, giving abundant oospores when tested in agar oat medium.
For testing, we used virus-free potato plants of the following varieties grown in a greenhouse (in a peat substrate): early Sandrin, Zorachka, Uladar, Osiris, mid-early Ilyinsky, mid-season Yanka.
Simple leaves from different levels of the potato plant were selected for analysis. Three leaves were selected from the lower level (4 lower compound leaves), three from the top (3-4 compound leaves on top), and three leaves from the center of the bush. Simple leaves were weighed, photographed on coordinate paper (to calculate the volume and area), and then placed upside down on the surface of sterile water, poured 25 ml in Petri dishes. Then each leaf was infected with one drop of a mixed suspension of zoosporangia isolates A1 and A2 of the mating type. For three leaves collected from each level, an inoculum of three different pairs of isolates was used. One leaf was infected with an inoculum mixture of one pair of isolates.
To prepare a suspension of zoosporangia, isolates of different mating types were grown on agar oat medium for 7 days, after which the zoosporangia were washed off with sterile distilled water. The inoculum concentration was 5-7 zoospores in the field of view of the microscope at 80x magnification. Each leaf was infected with one drop of a mixed suspension of zoosporangia isolates A1 and A2 mating type.
In all experiments, 3 potato leaves were used per variant. After incubation for 20 days at 18 ° C, each leaf was homogenized in a mortar with 2 ml of distilled water. From the resulting suspension, 3 samples were taken, from which preparations for microscopy were prepared.
In each variant, 180 fields of vision were examined, after which the number of oospores was recalculated per 1 mm2 sheet surface. The counting results for each variant were averaged.
To calculate the confidence interval (μ) for a significance level of 0,05, the following formula was used
where s is the standard deviation, n is the number of measurements, t is the t-test constant for a significance level of 0,05. All calculations were performed in Excel (Microsoft Office package).
Results and discussion
Inoculation of leaves collected from different layers of the potato bush and placed in the same conditions in humid chambers revealed differences in the intensity of oospore formation. In all studied varieties, the maximum number of oospores was formed during inoculation of leaves collected from the lower and middle tiers of the bush. No significant differences were found in the formation of oospores in the lower and middle leaves.
The minimum number of oospores was formed in the leaves of the upper layer (Fig. 1).
In the leaves of the lower and middle tiers, the maximum number of oospores was formed in the varieties Sandrin, Ilyinsky, Zorachka, Uladar and Osiris. They were formed somewhat less intensively in the leaves of the Yanka variety. The maximum number of oospores in the leaves of the upper layer was observed in the leaves of the Uladar variety, then Zorachka, Osiris, Yanka, Ilyinsky, Sandrin went in decreasing order, but not all differences were statistically significant.
Fig. 1. Formation of oospores in the leaves of different layers of the potato bush. The lower ones are the leaves of the three lower levels of the bush, the upper ones are 3-4 complex leaves from the top of the plant, the middle ones are from the middle of the bush.
Error bars show the confidence interval for a significance level of 0,05.
In the field, microclimatic conditions also contribute to the formation of oospores in the lower and middle leaves of the bush: higher humidity, reduced solar insolation, and daily temperature fluctuations (Harrison, 1992). Top spraying results in less pesticides in the bottom and center of the bush. To reduce the formation of oospores, technologies should be used that allow successfully delivering fungicides to the lower and middle parts of the bush. Good results can be obtained by using turbulent air sprayers and systemic fungicides that can spread to untreated parts of the bush.
This work was supported by the Russian Science Foundation (project No. 14-50-00029).
The article was published in the journal "Potato Protection" (No. 2, 2015)