Sorghum Diseases in India

observed. Testing for potential vectors is a critical
step, especially if the virus is not sapinoculable.
The following means of transmission
should be tested: grafting, arthropod, nematode,
fungus, parasitic seed plant (dodder), seed, and
pollen. Following identification of a vector, the
mechanism of transmission must be ascertained.
The next identification step is usually the establishment
of a host range, using both dicotyledonous
and monocotyledonous test plants. In
this phase the identification of diagnostic, propagation,
and assay host species is desirable for
identifying, multiplying, and quantifying the virus,
respectively (Hamilton et al. 1981).
At this point, one can deviate somewhat from
empirical procedures and conduct serological
tests (Von Wechmar et al. 1983) without purification
of the virus, provided antisera to known
viruses are available. Selecting the antisera is a
crucial step, as the selection must include as
many and the types needed to identify the unknown
virus and determine its interrelationships
with known virus groups, viruses, and
virus strains. If the serological test is positive to
a known virus, additional testing can identify
similarities to or deviations from the type virus.
If the serology tests are negative, two quick tests
may identify the unknown. One is the quick-dip
test, designed to demonstrate the presence of
rod-shaped virus particles as well as their relative
lengths and morphologies (rigid, flexuous,
or bacilliform) by electron microscopy (EM).
However, the test is not reliable for small polyhedral
viruses, because of confusion with ribosomes.
If the quick-dip test can determine
morphology of the virus particle, this alone will
serve to eliminate many virus groups.
Immunosorbent electron microscopy (ISEM)
provides both serological and particle morphology
data (Derrick and Brlansky 1976). In this
procedure, antiserum-treated grids are floated
on a drop of infected crude sap, washed,
stained, and viewed by EM. If the antiserum is
homologous, the virus particles will adhere to
the grid and their morphology can be noted
(Derrick 1975; Giorda et al. 1986a). In virus identification,
the greater the number of criteria used
and tests applied, the greater are the chances of
positive identification of a known virus, or
placement of an unknown virus in an established
virus group. If serology, quick-dip, and
ISEM are negative, the next step is to check
for possible viral inclusions (Christie and
Edwardson 1977). This involves staining and examination
by light or electron microscope. Viral
inclusion may be specific for a single virus, or
general for a particular virus group. Detection of
ultramicroscopic inclusions, such as pinwheels,
requires electron microscopy.
Purification and Molecular
Characteristics
Study of the ultrastructuxe of infected host tissue
is required before purification. Such study will
confirm presence or absence of inclusions, types
of tissue infected, location of virus in the host
cell, structural or cellular changes in the host,
and morphology of the virus particle (Kurstak
1981). At this point, if evidence is insufficient or
the virus appears to be one not yet described,
purification is necessary. Purification schemes
are available for viruses of all types (Giorda and
Toler 1986a; Giorda et al 1987; Langham 1986;
Van Regenmortel 1982a). Preliminary information
on nonspecific criteria such as thermal inactivation
point, dilution-end point, effect of pH,
longevity in vitro, and the effect of diethyl ether
is helpful (Noordam 1973). After purification,
the sedimentation coefficient, molecular weight,
isoelectric point, extinction coefficient, 260/280
ratio of unfractionated and separated components,
and buoyant density in CsCl or Cs2SO4
can be measured (Matthews 1981; Noordam
1973). Purity is critical. If artifacts produced by
nonviral material remain, the results are likely to
be misread. Also, one must determine if more
than a single component is present in the virus
preparation. Analytical uitracentrifugation and
Schlieren optics are employed to separate viral
components sedimenting at different rates. If
purified preparations contain more than one
component, each component can be categorized
by number or name (Trautman and Hamilton
1972).
Molecular characterization is another step in
the identification of viruses. Protein is separated
from the nucleic acid and the molecular weight
determined. Next, the number of protein species
in the particles are counted and described by
size and number (Langham 1986; Smith 1984).
Terminal amino acids are identified, and then
the entire amino acid composition is sequenced
(Langham 1986). The presence of viral encoded
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