Agriculture Reference
In-Depth Information
well developed to bear its characteristic reproductive structures. The microscopic
examination
in vitro
of pure cultures after isolation may also involve coaxing the
pathogen into producing any identifiable sexual or asexual reproductive structures.
Such manipulation is neither rapid nor completely dependable and requires some
experience of mycological techniques. Isolation would also be made easier and more
successful if a wider range of standard media were developed.
Frequently, isolating the pathogen responsible for a disease from the host,
regardless of whether it is a fungus, bacterium, virus or another agent possibly as yet
unknown, is not straightforward, particularly when the exact region of infection is
not clearly defined. In this case, the whole plant must be thoroughly examined for
the pathogen, including roots plus attached soil, as well as the aerial parts of the
plant. When there are no localized symptoms but just a general malaise, where the
pathogen could be present in the roots, stem or leaves but could result in similar host
reactions (such as during wilting due to root rot, vascular blockage or leaf infection),
it would be very helpful if there was a rapid non-specific scanning system to locate
the presence of the pathogen. Similar problems in detecting breast cancer in humans
have been tackled by screening patients with infrared detectors.
In general, traditional methods still require more experience on the part of an
investigator than do tests based on differences in nucleic acids and immunology,
where knowledge is increasingly being replaced by expensive equipment and
reagents. At present there is often a hierarchy of methods. Visual inspection is
unavoidable and can often give an instantaneous diagnosis when symptoms clearly
conform to a well known syndrome or when signs of infection are revealed. When
they are not, the next traditional route to identification in the laboratory is the
incubation of the specimen to allow the pathogen to develop sufficiently to be
identified and then, if necessary, isolated into culture. Biochemical and/or
immunological tests may then follow. Immunoassays have been revolutionized by
ELISA, which make them routinely completed within hours rather than the days or
even weeks required if isolation proved necessary. The problem of non-specific
binding has often meant that rapid diagnosis by monoclonal antibody ELISA has
been substituted by PCR (Martin
et al.,
1992a,b). Nonetheless, techniques based on
PCR and other methods that involve matching nucleic acid from the pathogen with
known sequences have been less suited for field use as they have not yet been
adapted to be as portable as many techniques based on immunology.
Traditional methods of diagnosis by experts, probably with increasing help from
electronic aids, will doubtless survive and even thrive but farmers and growers are
increasingly choosing to detect and monitor low but treatable levels of disease on
the spot under field conditions with a wide selection of relatively inexpensive
diagnostic kits that are simple to use (Klausner, 1987; Miller and Martin 1988;
Miller
et al.,
1988, 1990; MacAskill, 1989). Immunology has already provided
cheap kits for the pesticide industry that are sensitive enough for lower numbers of
pathogens than previously to be detected, and hence treated with less fungicide.
Each year, more immunological and nucleic acid hybridization techniques are being
developed for the rapid detection of many of those plant pathogens that cannot be
easily identified by other routine ways. Even pathogens that cause diseases with
variable or latent symptoms on the host plant and those with an indistinct structure
