Postgraduate
student Andrusyk I. M.
Candidate
of Agricultural Sciences Antipov I. M.
National University of Life and Environmental Sciences
of Ukraine, Ukraine
Testing of virus diseases of the sugar beet plants in
agrocenoses of Ukraine
The
viral infections of the sugar beet plants are monitored and tasted in several
farms of Ternopil region of Ukraine. The
infection of beet mosaic virus (BtMV) and beet yellow
virus (BYV) are detected by the methods of visual diagnostics,
biological testing and PCR.
To
prevent the spread of this infection and reduce disease level to economically
acceptable, all stages of the process must be followed by the rules of prophylaxis,
moreover the seed has to be free from any diseases. However, the information
about the spreading and species diversity of viruses which infect sugar beets
on the territory of Ternopil district is insufficient. The study of the etiology
of diseases will allow to develop some effective methods of protecting plants
and preserve the quality of harvest and raw materials.
During
the summer of 2013, we had been conducting inspection of sugar beet crops in Ternopil district
(Ternopil region., v. Osipovka, v. Volya; Pidvolochisskiy region, v. Kamyanka; Zalishchyky region., v.
Novosylka). Such observations shown us uneven spread of these
viruses in all inspected areas.
Evaluation
infestation of plants was performed by the method of visual diagnostics and
plant material samples were taken for further laboratory investigations, furthermore
weeds with the symptoms of virus diseases were selected on the sugar beet
fields. During the inspection, symptoms had been noticed as the light painted
spots of different form and size, which form a marble or mosaic picture. Yellow or orange spots with vague contours appear on the leaves at the defeat of plants by beet yellow virus
(BYV). Yellowing
starts from the top of the list. Some necrotic spots appear on the yellowed leaf and the leaf
becomes thicker and wavy.
As the diagnosis of diseases of plants by
only visual signs are not always accurate, this method can be useful only if
the symptoms of virus lesion are characteristic to it. To confirm the previous
conclusion on viral lesion, together with external review it is necessary to
apply additional methods. One of them is biological testing.
The
method of inoculation by juice of diseased plants is suitable for transmission
to healthy plants only for those viruses which can spread through
contact-mechanical method [2]. The leaves with the exact symptoms were used for
making juice. Finely cut leaves were ground in a mortar with a small amount of
water and silicon carbide. Than the juice was squeezed from that mixture
through a double layer of cheesecloth and smeared to a healthy leaf by spatula.
The appearing of symptoms was observed after some while.
Infectious
nature of noticed diseases was identified by using established biological
tests. The juice of diseased plants of sugar beet had been used as infectious
material. The infectious material was
crushed and ground in a mortar with the addition of 0.1 M phosphate buffer at a
ratio of 1: 2. The resulted homogenate was filtered [2].
To
avoid the inactivation of viruses in the environment for extracting, the reducing
agent was added. The inoculation of virus-containing material was made by
putting carbide powder on plants-indicators leaves. An excess of inoculum was
washed with water, and then plant indicators were kept in a shaded place during
the day [3].
Biological
testing performed by mechanical inoculation of plant indicators. Such indicators of plant were used: Tetragonia
expansa Murr., Chenopodium amaranticolor C. et R., Chenopodium quinoa Willd.,
Chenopodium album L., Amaranthus retroflexus L., Nicotiana tabacum L., Cucumis
sativus L., Phaseolus vulgaris L.
The
properties of viruses in infectious
juice was determined by conventional methods using as indicator plants
Chenopodium amaranticolor for beet mosaic virus and Chenopodium quinoa for beet
yellow virus. Systemic reaction in the form of a yellow-green mosaic leaf plate
and yellowing appeared at 15th day. After 21 days the symptoms of viral lesions
with the enlightenment of veins on the young leaves appeared in plants
Chenopodium amaranticolor. Leaves changed color to light-yellow[1].
Contamination
of sugar beet by viral pathogens in accordance with the identified symptoms was
checked using PCR. The advantages of this method were identified by extraordinary precision experiment, a
minimum number of plant samples and virtually absolute determination of the
virus in the sample.
As
a result, a test system was created for the diagnosis and identification BtMV
and BYV by PCR.
Consequently
after analyzing sampled sugar beet in
the Ternopil region by using PCR, the presence of virus disease was shown. Beet
mosaic virus was found on the fields of such villages as Volya in Ternopil region., Kamyanka in Pidvolochysky
region and Novosylka, Zalischyky region. Beet yellow virus was detected on the
fields of Osipovka village, Ternopil region
and Kamyanka village, Pidvolochysky region.
Therefore,
the results obtained on isolation and identification of phytoviruses in
combination with classic methods of viral diagnosis allowed to draw a clear
phytosanitary control of sugar beet plants in Ternopil. Infection of sugar beet
by BtMV and BYV in the field revealed by the results of visually identify of
symptoms, typical for viral diseases, biological testing juice of affected
samples on plants-indicators of virus diseases. The selection of healthy
planting material and early detection using PCR is the key to identify diseased
plants, which consequently leads to higher yields of sugar beet.
Literature:
1. Andrusyk I. PCR diagnostic and identification of beet yellow virus / I.Andusyk,
I.Antipov, A.Kyrychenko // Bulletin of Sumy National Agriculture
University. Serie “Agronomics and biology". V. 11 (26), 2013 : scientific magazine. – Sumy : SNAU, 2013. – P. 42.
2. Melnychuk M. Diagnostics of virus infections of sugar
beet in agrocoenoses of Ukraine / M. Melnychuk, S. Smirnova, E. Kachura, A. Smirnova // Collection of scientific works [Institute for Sugar Beet UAAN]. – 2003. – V. 5. – P. 320-325.
3. Polyschuk V. Manual of practical lessons to the course "General
virology" / V.Polyschuk, Y.Budzanyvska, T.Shevchenko. – Kiev: Phytosocial
center, 2005. – P. 129-133
4. Markoulatos P.
Multiplex polymerase chain reaction: a practical approach / P. Markoulatos,
N.Siafakas, M.Moncany // J. Clin. Laborat. Analysis. – 2002. – V. 16. – P.
47–51.