The concentration of the helminthosporal toxin in the crude culture filtrate for both the pathostrains were calculated based on the absorbance. The culture filtrate containing helminthosporal showed a maximum absorbance at 264 nm for G strain and 260 nm for I strain. This shift of wavelength by 2 to 6 nm from the standard 266 nm was likely due to impurity of the toxin preparation. Thus, our measurements were biased to some extent. In addition, unknown toxins might have been contained in our samples. Nevertheless based on the absorbance measurements, strain G had more toxin(s) with value of 1.6 x 10-4 moles/l in culture filtrate as compared to strain I with 1.1 x 10-4 moles/l. Pringle (1979) reported in barley plants that toxin production ability of H. sativum isolates varies widely and also varies in their disease development.
Screening by using intact leaves.
The reaction of pathogen and toxin(s) is, shown in Table
1 and Fig.1. WE 542 and PBW 226 showed
high to very high intensity of necrosis to pathogen and toxin of G
strain but little or no necrosis to both pathogen and toxin(s) of I
strain. PBW 154 showed moderate reaction and HD 2285 and Sonalika
moderate to little necrosis for G strain and little or no reaction
for I strain. Very little to no necrosis was seen for CPAN 3004, UP
2003 and UP 2121 for both G and I strains. Control treatment of water
did not show necrosis in any of the genotypes. Statistical comparison
of pathogen and toxin reactions for G and I strains were both found
to be non- signiflicant , indicating that pathogen and toxin(s) can
be equally used for screening.
Screening with detached leaves.
WH 542 and PBW 226 showed very high intensity of necrosis for both
pathogen and toxin(s) of G strain and very high to moderate intensity
of necrosis for I strain (Table 2). PBW 154
and Sonalika showed very high to high intensity of necrosis for G
strain and moderate to low intensity of necrosis for toxin(s) of G
strain, while no necrosis was seen for I strain. In the other
genotypes if intensity of necrosis was low for pathogen 'or toxin(s)
from G strain there was either little or no necrosis from I strain.
With control inoculations no necrosis was seen. Paired t-test
revealed nonsignificant differences between the pathogen and toxin
reactions for each pathostrain thus indicating similarity of
reaction.
Pathostrain G always showed more necrosis than I strain, indicating the variability in different strains for causing the virulent reaction. The production of toxic principles in culture filtrate of G strain was also higher than I strain. Vidhyasekaran et al. (1986) used detached leaf bioassay technique for comparison of pathogen and toxin reaction of H. oryzae against rice genotypes and found comparable reaction. They showed that toxin preparation obtained from diseased leaves induced characteristic brown spots surrounded by yellow halo symptoms. In tomato necrosis was never seen in resistant genotypes against AAL toxin of Alternaria, whereas very low concentration of toxin was sufficient to induce necrosis in leaves of susceptible genotypes (Hanneke et al. 1988).They reported that intact leaves and detached leaves of plants exhibited the same sensitivity to toxin. These studies suggest that toxin production ability varies in different pathostrains. In the present study one unit higher degree of necrosis was observed in the tests using detached leaves than in the test using intact leaves. But, generally these two tests exhibited similar levels of sensitivity. These intact and detached leaf bioassays can be performed for screening of genotypes against isolated toxin(s) from culture filtrate as effectively as pathogen inoculation.