DETECTION OF AEROMONAS-SALMONICIDA FROM FISH BY USING POLYMERASE CHAIN-REACTION AMPLIFICATION OF THE VIRULENCE SURFACE ARRAY PROTEIN GENE

被引:88
作者
GUSTAFSON, CE
THOMAS, CJ
TRUST, TJ
机构
[1] UNIV VICTORIA, DEPT BIOCHEM & MICROBIOL, VICTORIA V8W 3P6, BC, CANADA
[2] UNIV VICTORIA, CANADIAN BACTERIAL DIS NETWORK, VICTORIA V8W 3P6, BC, CANADA
[3] UNIV ADELAIDE, DEPT MICROBIOL & IMMUNOL, ADELAIDE, SA 5001, AUSTRALIA
关键词
D O I
10.1128/AEM.58.12.3816-3825.1992
中图分类号
Q81 [生物工程学(生物技术)]; Q93 [微生物学];
学科分类号
071005 ; 0836 ; 090102 ; 100705 ;
摘要
A DNA-based assay was developed to detect Aeromonas salmonicida from infected fish by analyzing tissues, feces, and the tank water in which the infected fish were held. This analysis was done both by direct detection from samples and after a bacterial outgrowth step. Polymerase chain reaction (PCR) amplification of a 421-bp sequence from the 3' region of the surface array protein gene (vapA) of A. salmonicida provided a specific and sensitive method for the detection and identification of this important fish pathogen. The sensitivity of PCR detection of A. salmonicida directly from tissues was less than 10 CFU/mg. Furthermore, a detection level of 5 fg, equivalent to approximately 1 cell, was obtained by using purified chromosomal DNA as the template. This highly reproducible assay, which requires 45 min to complete, is therefore sensitive enough to be used as a noninvasive method for monitoring fish populations for the presence of carrier fish. Because the surface protein array (A-layer) is a virulence factor of A. salmonicida, PCR analysis with oligonucleotide primers directed at vapA can also be used to provide information on the potential virulence of a strain.
引用
收藏
页码:3816 / 3825
页数:10
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