Biomedical Engineering Reference
In-Depth Information
Desm., which teleomorph is Mycosphaerella graminicola (Fuckel) J. Schröt.
(Palmer and Skiner 2002).
The defeat of the S. nodorum fungus becomes visible in the stage of
seedlings, due to reduced growth and formation of brown nodes on the
fi rst leaves of seedling. The highest pathogen activity develops on adult
wheat plants. During initial periods the infection is extremely diffi cult to
identify. This is due to the fact that the pathogen is hemibiotroph and such
symptom as necrotic on leaf is similar to that of other leaf blotch diseases.
In this regard, diagnostics of this disease is a very urgent problem for the
choice of fungicide and time of its application. Now one of the most specifi c
methods of diagnostics of infection diseases of plants, animals and humans
is polymerase chain reaction (PCR) (Atkins and Clark 2004, Kolesov et al.
2004, Manuku 2004, Eibel et al. 2005, Lievens et al. 2006, Ryazantsev et al.
2009). It allows us to determine with high accuracy the presence of organisms
of any pathogen in wheat tissues, including the fungus S. nodorum (Beck
and Ligon, 1995 (U.S. Patent 5,955,274 (1999), Pfi rter et al. 1999, Gubis et
al. 2005, Abramova et al. 2008, Oliver et al. 2008).
Since pathogens enzyme activity, such as chitin deacetylase ( CDA ) is
closely related to their aggressiveness (Levis et al. 2001, Hu et al. 2007),
probably PCR method can be used to determine the presence of fungus in
tissues S. nodorum and to study its aggressiveness degree. In this regard,
the purpose of this work is to use a PCR method of diagnostic of S. nodorum
fungus in infected wheat plants tissues and determine the aggressiveness
degree of strains of this fungus with primers on CDA gene.
METHODS
The experiments were conducted using strains 4VD, 6VD and 9MN of
the fungus S. nodorum , which were kindly provided by the Institute
of Experimental Botany V.F. Kuprevich National academy of sciences
of Belorussia and the strain Bas1, selected from local South-Ural
populations of the fungus. Wheat seedlings Triticum aestivum L. grade
Zhnitsa susceptible to pathogen Septoria glume blotch S. nodorum, were
also used.
Wheat seeds were sterilized with 70% ethanol for 3 minutes, washed
with sterile water and germinated on fi lter paper. Infection of plants
was carried out on the seventh day. Inoculated leaves were kept at room
conditions for 10 hr. First completely unfolded leaves were cut, placed in
Petri dishes and covered with cottonwool wetted with benzimidazole.
Infection was carried out by spraying the leaves with a suspension of spores
at concentration of 10 6 /ml. Part of the leaves, after preliminary washing
under running tap water, was collected for analysis of expression CDA gene
and some were left for monitoring the disease progression.
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