PET瓶装无气天然矿泉水中污染菌及其系统进化分析

Case Analysis of Contaminated Bottled Natural Mineral Water:Isolation and Phylogenetic Analysis of Contaminant Bacteria

目的:了解某一批次聚对苯二甲酸乙二醇酯(polyethylene terephthalate,PET)瓶装无气天然矿泉水产生异味的缘由。方法:首先采用膜过滤结合平板菌落计数法分离和计数污染菌,16S r RNA序列分析鉴定其为假单胞菌属(Pseudomonas spp.),再采用假单胞菌属特异性rpo B引物进行梯度聚合酶链式反应优化扩增条件,产物进行测序和序列分析将污染菌鉴定到种。结果:这批产品的主要污染菌是一种2009年从南极分离鉴定的假单胞菌属新种P.extremaustralis。反接实验证实该菌能在天然矿泉水中生长繁殖,并产生刺激性异味。结论:P.extremaustralis部分细菌能够透过两级串联0.2μm过滤器,耐受中、低压紫外杀菌工艺,耐低温和低氧压,能在无任何添加的PET瓶装无气天然矿泉水中生长并产生异味。

Objective：To figure out the cause of the occurrence of unpleasant odor in one batch of polyethylene terephthalate（PET）bottled natural mineral water.Methods：Firstly,we counted and isolated strains from the samples by membrane filtration and culture on plate count agar（PCA）.Secondly,the main spoilage bacteria were identified as Pseudomonas spp.by 16 S r RNA analysis.Then gradient PCR was used to optimize the PCR conditions for amplification of rpo B gene.The PCR amplified rpo B sequence was analyzed.Results：The main pollutant bacteria in this batch of bottled drinking water was found to belong to a new species of P.extremaustralis,which was firstly isolated from a temporary pond in Antarctica in 2009.The rpo B sequence analysis showed that P.extremaustralis spp.was responsible for the occurrence of off-flavor in natural mineral water.When being inoculated back into normal products,it grew well and stink.Conclusion：Some strains of P.extremaustralis spp.have the capability of passing through double 0.2 μm-pore-size filters,tolerating ultraviolet sterilization,growing in PET bottled natural mineral water at low temperature and low oxygen content,and giving out an unpleasant odor.