PCR-DGGE技术分析不同包装条件下鱼肉表面优势菌的菌群变化

PCR-DGGE Analysis of Dominant Bacterial Populations on the Surface of Grass Carp Meat under Different Packaging Conditions

采用聚合酶链式反应-变性梯度凝胶电泳(polymerase chain reaction-deformation gradient gel electrophoresis analysis,PCR-DGGE)技术研究3种包装(空气、真空、气调)结合钴60辐照技术对鱼肉表面优势菌菌群的影响,分别选择6种辐照剂量(0、2、4、6、8、10 kGy)照射4℃贮藏18 d后的鱼肉表面优势腐败菌群进行研究。方法:采集第18天不同包装不同剂量的草鱼肉样品,洗脱鱼肉表面菌;提取DNA,扩增16S rDNA的V3可变区,用PCRDGGE技术鉴定优势菌,割胶回收测序。结果表明:不同包装鱼肉表面优势菌群数量和种类不同,在样品中共同的优势腐败菌为假单胞菌,在空气包装中优势腐败菌是假单胞菌、乳酸菌和沙雷菌,沙雷菌在辐照剂量高于8 kGy时被抑制。在真空包装中优势腐败菌是假单胞菌、沙雷菌、热死环丝菌和耶尔森菌,沙雷菌辐照剂量高于6 kGy时被抑制,而耶尔森菌在2 kGy以上即被抑制,热死环丝菌在6、8 kGy辐照条件下才出现,其具有很高的辐照抗性。在气调包装中优势腐败菌是假单胞菌和乳酸菌,但乳酸菌仅在气调包装2 kGy辐照和气调包装8 kGy两种辐照包装条件下是优势腐败菌。结果表明,PCR-DGGE技术可以很好地鉴别鱼肉表面优势腐败菌。

Changes in dominant bacterial populations(DBP) on the surface of grass carp meat packaged under three different atmospheric conditions(air, vacuum and modified atmosphere packaging environment) and then irradiated by Co-60 gamma ray at six different doses(0, 2, 4, 6, 8, and 10 kGy) after subsequent storage at 4 ℃ for 18 days were investigated by polymerase chain reaction-denaturing gradient gel electrophoresis analysis(PCR-DGGE). The bacteria on the surface of fish meat samples were washed and collected, and their DNA was then extracted. The V3 variable region of 16 S rDNA was amplified with PCR. Identification of DBP was performed with DGGE. The amounts and species composition of DBP varied with the type of packaging. Pseudomonas sp. was dominant on all fish samples. Lactic acid bacteria and Serratia sp. were also dominant on air-packaged samples, and Serratia sp. was inhibited when the irradiation rose was more than 8 kGy. Pseudomonas sp., Serratia sp., Brochothrix thermosphact, and Yersinia sp. were identified as the dominant bacteria on vacuum-packaged samples; Serratia sp. was inhibited when the irradiation dose was above 6 kGy, Yersinia sp. could be inhibited even when the irradiation dose exceeded 2 kGy, and B. thermosphact exhibited high irradiation resistance for its occurrence even at doses of 6 and 8 kGy. For modified atmosphere packaged fish, the DPB were Pseudomonas sp. andLactobacillus sp.; however, lactic aci bacteria were dominant only when the irradiation dose was 2 or 8 kGy. In conclusion, PCR-DGGE provides a good way to analyze dominant bacterial populations on the surface of grass carp meat.