Detection of histamine-producing bacteria using polymerase chain reaction techniques and DNA probes

Abstract

The polymerase chain reaction technique was used to enable the detection of DNA specific amplicons, corresponding to gene fragments coding for histidine decarboxylase present in the histidine decarboxylating bacteria frequently found in canned fish. The level of histamine in several fish product samples was quantified by HPLC and the DNA of the samples containing histamine was isolated and amplified with hdcA histidine-decarboxylase gene-specific primers. The primer sets used, CL1/CL2, JV16HC/JV17HC and CL1/JV17HC, amplified 150 bp, 370 bp and 500 bp products respectively. Non-expected fragments (100 bp, 150 bp and 250 bp) were also amplified by JV16HC/JV17HC. Some amplified fragments were sequenced automatically, presenting high homologies with the Clostridium perfringens hdcA gene. A DNA probe from a C. perfringens pure strain was hybridized with the DNA fragments amplified from the contaminated samples. The hybridization blots were then detected by colorimetry, revealing that the samples were contaminated by C. perfringens.