Detection And Identification Of Pantoea Stewartii Subsp.Stewartii Real-time Fluorescent PCR

Pantoea stewartii subsp. stewartii is a very important quarantine bacterium on the A1 list of China , which has not been reported in China yet. The traditional methods for detection of Pantoea stewartii subsp. stewartii were isolating culture, physiological and biochemical identification, bacteriophage specifity and serological test.These methods are time-consuming, low efficiency and sensitivity and these limits make it difficulit to identify species of the bacteria and unfit for the port identification. So it is significance to establish a rapid and accurate detection method of Pantoea stewartii subsp. stewartii.A group-specific probe was designed based on 16S rDNA of Pantoea and it was used to detect 3 species of Pantoea ,3 species of Erwinia , 4 species of other species of bacteria and 5 species of Phytoplasmas. The results show that no signal was detected for other bactseria and Phytoplasmas except Pantoea stewartii. subsp. stewartii. The absolute sensitivity is 2.53 ×106 positive molecules and the relative sensitivity was 107CFU/mL. The sensitivity of colony's suspension was 10 CFU/mL . That is to say, if reactionary system had 2 live bacteria, real-time fluorescent PCR can detect it.Few contamination will occur because the whole detection process is finished in the contained tubes.The method is more specific than normal PCR,TaqMan probe designed in the study can detect Pantoea stewartii subsp. stewartii directly during the PCR amplification.