PCR扩增特异性16SrDNA和溶血素基因检测致病性嗜水气单胞菌

PCR detection of pathogenic Aeromonas hydrophila by specific 16S rDNA and aerolysin gene

根据已发表的气单胞菌16SrDNA基因序列及气单胞菌气溶素(aerolysin)基因序列,设计了2对引物,建立了检测致病性嗜水气单胞菌的PCR方法。通过对12株气单胞菌的检测,发现16SrDNA引物具有高度的特异性,仅对嗜水气单胞菌扩增阳性。而Aero基因引物检测结果与采用生物学方法(鲜血平板法)检测的结果符合率为97.2%,且具有高度的敏感性,可检测最低1fg的模板。将16SrDNA与Aero基因结合PCR方法检测致病性嗜水气单胞菌与用致病性嗜水气单胞菌检测试剂盒的符合率为94.4%。该方法的建立为致病性嗜水气单胞菌的检测提供了一种简便、快速的途径,是一种比较实用的致病性嗜水气单胞菌的检测方法。

Based on the published 16S rDNA gene sequence of Aeromonas spp. and aerolysin gene sequence of Aeromonas hydrophila, the synthetic oligonucleotide primers were used in a polymerase chain reaction (PCR) technique to detect the gene for specific 16S rDNA and aerolysin of pathogenic A. hydrophila. A. hydrophila can be clearly discriminated from the other Aeromonas species by 16S rDNA gene PCR, the detection limit for the aerolysin gene by PCR amplification was 1fg DNA. 36 strains were tested for pathogenic A. hydrophila by PCR method and pathogenic aeromonads diagnosis kit and their coincident rate was 94.4%. The PCR can clearly identify A. hydrophila from Aeromonas species, and can identify aerolysin-producing strain of A. hydrophila. In conclusion, this PCR-based method is rapid, sensitive and specific for the detection of pathogenic A. hydrophila, and it is a practical method for pathogenic A. hydrophila detection.