Sorghum Diseases in India

host genotype. Copious amounts of bacterial exudate
are usually apparent on both leaf surfaces.
Widespread dissemination of the bacteria is
probably attributable to seed or infested debris.
Entry into the plant is possibly through wounds,
stomata, or hydathodes. Tolerance or susceptibility
of the germplasm tested is listed in
Table 2.
Restriction endonuclease analysis of the
genome of Xanthomonas campestris pv
holcicola
Xanthomonas campestris pv holcicola cannot be
differentiated from other pathovars of X. campestris
by physiological or biochemical tests. Serological
probes are useful, but unless monoclonal
antibodies are used, the serological
probes often cross-react with other pathovars.
Development of monoclonals is expensive, a
constraint that prohibits their use with pathogens
of limited economic importance. Restriction
endonuclease analysis (REA) has accurately
differentiated other X campestris pathovars
(Leach et al. 1987; Lazo et al. 1987). Plant assays
require several weeks to complete; DNA restriction
analyses of many isolates can be completed
within a week. These features of the REA procedure
make it attractive for specific purposes.
REA is based on the identification of specific
DNA fragmentation patterns. The number and
locations of endonuclease restriction sites along
the DNA strand are unique for each genome.
Separation of the fragments by digestion with
specific endonucleases on agarnose gels reveals
fragment-size classes unique for each genome.
Such unique fragment classes form the specific
restriction patterns, or fingerprints, of individual
isolates.
To identify unique DNA banding patterns
which correlate with the pathovar holcicola and
differentiate it from all other pathovars of
X. campestris, X. campestris pv holcicola isolates
from USA (Kansas, Nebraska, and Texas), Mexico,
Lesotho (Africa), and Australia were screened
by restriction analysis of their DNA. The DNA
restriction patterns were compared with those of
24 X. campestris pathovars. Total DNA was extracted
(Shepard and Polisky 1979, pp. 503-506)
and digested to completion with various restriction
enzymes (Maniatis et al. 1982). The DNA
fragments were separated by electrophoresis in
0.75% agarose gels and stained with ethidium
bromide. X. campestris pv holcicola was then
characterized by visual assessment of unique
fragment subsets (or bands) within the total genomic
profile.
An EcoR1 restriction pattern, consisting of a
broad space (spanning about 4.8 to 5.0 kb) and a
pair of bands at about 3.7 and 3.9 kb, differentiates
X. campestris pv holcicola from the 24 different
pathovars of X. campestris tested (for example, see
Fig. 9). Fifteen X. campestris pv holcicola isolates
were screened; the pattern was consistent in all
isolates (Fig. 10). While isolates of other pathovars
may have bands in the same positions,
not one had the complete pattern. Thus the subset
pattern was unique to X.campestris pv
holcicola and REA is useful in confirming identity
of this pathovar.
Figure 9. Xanthomonas campestris pv
holcicola DNA Eco R1 restriction pattern. Pattern
which differentiates X, c. pv holcicola
from other pathovars is shown by arrows.
Lane 1,1 kb ladder size marker; 2-5, X. c. pv
translucens; 6-7, pv secalis; 8-9, pv undulosa;
10-11, pv oryzicola; 12, pv phleipratensis; 13,
pv graminis; 14, pv holcicola; 15-19, pv oryzae.
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