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We developed a polyclonal antibody based-ELISA system to monitor inocula accurately and rapidly before onset of anthracnose on soybean sprouts. Titer of mouse antisera against conidia of Colletotrichum gloeosporioides, determined by indirect ELISA, was high enough to be detectable up to ${\times}$25,600 dilutions. Both PAb1 and PAb2 had the highest level of reactivity to Colletotrichum gloeosporioides. Absorbance readings exceeded 0.15. Sensitivity of PAb to C. gloeosporioides was precise enough to detect spore concentration as low as 500 conidia/well by indirect ELISA. Both antibodies are very sensitive and highly specific to the target pathogen Colletotrichum gloeosporioides, apparently discriminating other unrelated pathogen, or epiphytes. This kit fulfills the requirements far detecting inocula before infection and onset of anthracnose. Our ELISA system should also be feasible to detect C. acutatum (Mungbean sprouts rot) and G. cingulata (C. gleosporioides), (apple, pepper). It was remarkable that absorbance value was not reduced even after 4 consecutive washings (Fig.4), suggesting that antigenic determinants are on the surface of conidia. Antigenic determinant was characterized by heating and enzyme treatment: Both PAb1 and PAb2 bind to protein epitope that does not contain residue of amino acid, arginine, and Iysine, even though more work needs to be done.

참고문헌 (10)

  1. Clark, M. F. 1981. Immunosorbent assays in plant pathology. Annu. Rev. Phytopathol. 19:83- I06 
  2. Jung, Y. K. and Kim, S. H. 2004. Isolation and identification of pathogenic and epiphytic microorganisms from soybean sprouts and control of soybean sprout rot. B.S. Thesis, Gyeongsang National University 
  3. Kim, D. K., Lee, S. C., Kang, J. H. and Kim, H. K. 2003. Colletotrichum Disease of Mungbean Sprout by Colletotrichum acutatum. Plant Pathol. 19:203-204 
  4. Sundaram, S., Plasencia, J. and Banttari, E. E. 1991. Enzymelinked immunosorbent assay for detection of verticillium spp. Using antisera produced to V. dahliae from potato. Phytopathology 8I: 1485- I489 
  5. Velicheti, R. K., Lamison, C., Brill, L. M. and Sinclair, l. B. 1993. Immunodetection of Phomopsis species in asymptomatic soybean plants. Plant Dis. 77:70-73 
  6. Lee, J. H., Han, K. S., Lee, S. C., Shim, C. K., Bae, D. W. and Kim, H. K. 2004. Early detection of epiphytic anthracnose inoculum on phyllosphere of Diospyros kaki var. domestica . Plant Pathol. J 20:(in press) 
  7. Kang, J. H. and Song, G A. 2002. Clean soybean sprouts produced by using light and seed floating on water and its production model. Patent number: 0379839, Korean Patent Administration 
  8. Somai, B. M. and Keinath, A. P. 2002. Development of PCRELISA for detection and differentiation ofDidymella blyoniae from related phoma species. Plant Dis. 86:7 I0-7 I6 
  9. Meyer, U. M., Spotts, R. A. and Dewey, F. M. 2000. Detection and quantification of Botrytis cinerea by ELISA in pear stems during cold storage. Plant Dis. 84: I099- I103 
  10. Singh, O., Trevors, C. M., Boer, S. H. and lanse, J. D. 2000. Fimbrial- specific monoclonal antibody-based ELISA for Europiean potato strains of Erwinia ehrysanthemi and comparison to PCR. Plant Dis. 84:443-448 

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