The Establishment Of Rapid Detection Method For The Pathogen Of Hami Melon Fruit Blotch

Hami melon bacterial fruit blotch (HBFB), caused by Acidovorax avenae subsp. citrulli, is a new disease in our country, which damages the fruits and lowers their qualities. The disease severity is growing up in recently years, which threaten the productions of harm melon seriously. HBFB is a typically seedborne disease, and the important prevention and cure method is the quaratine of the seeds with bacteria. However, most kinds of detection methods are time-consuming, or in defect of low precision or low specificity. So in this study, a rapid and precise detection method was constructed for the detection of the bacteria in hami melon seeds.In this study, a culture medium named ASCM was confected in order to enrich and culture the HBFB bacteria exclusively while inhibiting growths of other kinds of bacteria. Serum anti-HBFB bacteria was made and used for indirect ELISA, which stimulated the detection of bacteria in seeds. A set of specific primers for BFB bacteria was chosen and used in PCR. The precision of bacteria detection by PCR was app. 3×105 CFU/ml. A practicable Taq Man probe was designed and composed and the precision of bacteria detection by real-time PCR was app. 3×104 CFU/ml. Then the immune-detection was combined with PCR by using the superparamagnetic beads to form IMS-PCR, which increase the precision of bacteria detection to app. 3×102 CFU/ml.In this study, the integrated method of bacterial detection , which was called BIO-IMS-(real-time)PCR, was formed by combining the bacterial isolation with MOPS, the culturing in ASCM, IMS and (real-time) PCR. One seed with bacteria in 1000 commercial seeds could be detected using this integrated method.