运用Real-time PCR建立即食虾中副溶血性弧菌分子预测模型

Development of a Molecular Predictive Model of Vibrio parahaemolyticus in Ready-to-eat Shrimps by Real-time PCR

运用Real-time PCR方法建立不同贮藏温度（4-30℃）下即食虾中副溶血性弧菌分子预测模型。将四株副溶血性弧菌混合接种于即食虾中,利用Real-time PCR定量4-30℃下虾中副溶血性弧菌数量,选用Gompertz模型拟合得到10-30℃下副溶血性弧菌最大比生长速率,并用Linear、Square Root和Ratkowsky模型拟合获得二级分子模型,然后对其验证;选用Weibull、Log-linear及Logistic模型拟合4℃及7℃下副溶血性弧菌的失活曲线。结果表明Gompertz模型R2均在0.97以上,可较好地拟合Real-time PCR定量的副溶血性弧菌生长数据;二级分子模型的R2均在0.94以上,准确因子（Af）和偏差因子（Bf）均在接受范围内,可较好地描绘即食虾中副溶血性弧菌的生长速率与贮藏温度之间的关系,其中Linear模型为最适二级模型;但Real-time PCR方法不适用于建立低温下副溶血性弧菌的失活模型。运用Real-time PCR方法可以建立可靠的一、二级分子预测模型,为实际样品中目标菌株的预测模型建立提供研究基础。

A molecular predictive model of Vibrio parahaemolyticus on ready-to-eat shrimps was developed and four Vibrio parahaemolyticus strains in ready-to-eat shrimps at 4-30 ℃ were quantitated by Real-time PCR method. Gompertz model was selected as the primary model, which was fitted to achieve maximum growth rate（MGR） for Vibrio parahaemolyticus on ready-to-eat shrimps at 10-30 ℃. The MGR was then used to establish secondary molecular models as linear model, square root model and Ratkowsky model, and these models were validated subsequently. Weibull model, Log-linear model, and Logistic model were applied to predict inactive curves of V. parahaemolyticus at 4 ℃ and 7 ℃, respectively. Results showed that the V. parahaemolyticus growth data could be well fitted by Gompertz model（R^2〉 0.96）. The R2 values of linear model, square root model and Ratkowsky model were all above 0.94, and the Af and Bf values were within the acceptable limit, indicating that the relationship between the growth rates of V. parahaemolyticus and storage temperatures could be well described by the models, especially the Linear model. However, Real-time PCR method could not be used to develop the inactive model of V. parahaemolyticus at 4 ℃ and 7 ℃. Accuracy molecular predictive models developed by real-time PCR method can aid to establish models of certain pathogens more accurately in the presence of other bacteria.