Moschetti G, Blaiotta G, Villani F, Coppola S
Dipartimento di Scienza degli Alimenti, Sezione di Microbiologia Agraria, Alimentare ed Ambientale e di Igiene, Università degli Studi di Napoli Federico II, Portici, Italy.
Int J Food Microbiol. 2001 Jan 22;63(1-2):109-16. doi: 10.1016/s0168-1605(00)00411-6.
In this work we studied using different molecular methods the population dynamics of nisin-producing organisms and the persistence of such organisms within a complex ecosystem, 'Fior di latte' cheese, a traditional high-moisture pasta filata cheese. Using the primers targeting the eubacterial 16S-23S rRNA spacer region, together with those amplifying the nisA or nisZ gene, we were able to provide a rapid species identification of the isolates. Inhibitors of Lactococcus lactis subsp. lactis DSM 20481T used as indicator occurred during the whole process of cheese manufacture as a significant part of lactic microflora; however, only 12 among 109 isolates of bacteriocin producers were nisin producers. Amplification of the nisA or nisZ gene, using DNA extracted directly from dairy samples as templates, showed that the nisin structural gene was detected during cheese-making from milk samples up to the end of curd ripening but not in the final cheese. In order to monitor nisin-producing strains during cheese manufacturing, the 12 Lactococcus lactis nis+ strains were analysed by low frequency restriction fragment and PFGE. Nine isolates among the 12 nisin-producers exhibited an unique and distinct DNA banding pattern and are considered to be genetically diverse. The other three isolates from curd after ripening showed the same restriction pattern and could be the same strain. In fact, it was also isolated 2 months after the first analysis of cheese-making of 'Fior di latte'.
在这项工作中，我们使用不同的分子方法研究了产乳链菌肽的微生物的种群动态以及这些微生物在复杂生态系统——“菲奥里·迪拉泰”奶酪（一种传统的高水分拉伸凝乳奶酪）中的持久性。使用靶向真细菌16S - 23S rRNA间隔区的引物，以及扩增nisA或nisZ基因的引物，我们能够对分离株进行快速的物种鉴定。乳酸乳球菌乳酸亚种DSM 20481T用作指示菌，在奶酪制造的整个过程中作为乳酸微生物群的重要组成部分出现；然而，在109株细菌素产生菌分离株中，只有12株是产乳链菌肽的。以直接从乳制品样品中提取的DNA为模板扩增nisA或nisZ基因，结果表明在奶酪制作过程中，从牛奶样品到凝乳成熟结束都能检测到乳链菌肽结构基因，但在最终的奶酪中未检测到。为了在奶酪制造过程中监测产乳链菌肽的菌株，对12株乳酸乳球菌nis +菌株进行了低频限制性片段分析和脉冲场凝胶电泳分析。12株产乳链菌肽的分离株中有9株表现出独特且不同的DNA条带模式，被认为在遗传上是多样的。另外三株成熟后凝乳中的分离株显示出相同的限制性模式，可能是同一菌株。事实上，在首次分析“菲奥里·迪拉泰”奶酪制作后的2个月也分离到了该菌株。
