The New Zealand Institute for Plant and Food Research Limited, Auckland, New Zealand.
Institute of Food, Nutrition and Human Health, Massey University, Palmerston North, New Zealand.
Appl Environ Microbiol. 2021 Apr 27;87(10). doi: 10.1128/AEM.02349-20.
is a ubiquitous foodborne pathogen that results in a high rate of mortality in sensitive and immunocompromised people. Contamination of food with is thought to occur during food processing, most often as a result of the pathogen producing a biofilm that persists in the environment and acting as the source for subsequent dispersal of cells onto food. A survey of seafood-processing plants in New Zealand identified the persistent strain 15G01, which has a high capacity to form biofilms. In this study, a transposon library of 15G01 was screened for mutants with altered biofilm formation, assessed by a crystal violet assay, to identify genes involved in biofilm formation. This screen identified 36 transposants that showed a significant change in biofilm formation compared to the wild type. The insertion sites were in 27 genes, 20 of which led to decreased biofilm formation and seven to an increase. Two insertions were in intergenic regions. Annotation of the genes suggested that they are involved in diverse cellular processes, including stress response, autolysis, transporter systems, and cell wall/membrane synthesis. Analysis of the biofilms produced by the transposants using scanning electron microscopy and fluorescence microscopy showed notable differences in the structure of the biofilms compared to the wild type. In particular, inactivation of and produced coccoid-shaped cells and elongated cells in long chains, respectively, and the mutant produced a unique biofilm with a sandwich structure which was reversed to the wild-type level upon magnesium addition. The transposant was successfully complemented with the wild-type gene, whereas the phenotypes were not or only partially restored for the remaining mutants. The major source of contamination of food with is thought to be due to biofilm formation and/or persistence in food-processing plants. By establishing as a biofilm, cells become harder to eradicate due to their increased resistance to environmental threats. Understanding the genes involved in biofilm formation and their influence on biofilm structure will help identify new ways to eliminate harmful biofilms in food processing environments. To date, multiple genes have been identified as being involved in biofilm formation by ; however, the exact mechanism remains unclear. This study identified four genes associated with biofilm formation by a persistent strain. Extensive microscopic analysis illustrated the effect of the disruption of , , , and and the influence of magnesium on the biofilm structure. The results strongly suggest an involvement in biofilm formation for the four genes and provide a basis for further studies to analyze gene regulation to assess the specific role of these biofilm-associated genes.
是一种普遍存在的食源性病原体，在敏感和免疫功能低下的人群中导致高死亡率。人们认为，食品受到污染是在食品加工过程中发生的，最常见的原因是病原体产生了生物膜，这种生物膜在环境中持续存在，并成为随后细胞散布到食物上的来源。对新西兰海产品加工厂的一项调查发现了具有高生物膜形成能力的持久性菌株 15G01。在这项研究中，对 15G01 的转座子文库进行了筛选，以寻找通过结晶紫测定法评估的生物膜形成发生改变的突变体，从而鉴定参与生物膜形成的基因。该筛选确定了 36 个转座子，与野生型相比，它们的生物膜形成发生了显著变化。插入位点位于 27 个基因中，其中 20 个导致生物膜形成减少，7 个导致生物膜形成增加。两个插入位于基因间区域。基因注释表明，它们参与了多种细胞过程，包括应激反应、自溶、转运系统和细胞壁/膜合成。使用扫描电子显微镜和荧光显微镜分析转座子产生的生物膜表明，与野生型相比，生物膜的结构有明显差异。特别是，和的失活分别导致球菌形状的细胞和长链中的长形细胞，而突变体产生具有独特三明治结构的生物膜，在添加镁后恢复到野生型水平。成功地用野生型基因对 转座子进行了互补，而其余突变体的表型则没有或仅部分恢复。人们认为，食品受到污染的主要原因是生物膜的形成和/或在食品加工工厂中的持续存在。由于其对环境威胁的抵抗力增强，成为生物膜后，细胞更难被根除。了解生物膜形成涉及的基因及其对生物膜结构的影响将有助于确定消除食品加工环境中有害生物膜的新方法。迄今为止，已经确定了多个与生物膜形成有关的基因；然而，确切的机制仍不清楚。本研究确定了与持久性菌株生物膜形成有关的四个基因。广泛的显微镜分析说明了破坏、、和以及镁对生物膜结构的影响。结果强烈表明这四个基因参与了生物膜的形成，并为进一步研究分析基因调控以评估这些生物膜相关基因的具体作用提供了基础。
