Zeng Ximin, Ardeshna Devarshi, Lin Jun
Department of Animal Science, The University of Tennessee, Knoxville, Tennessee, USA.
Department of Animal Science, The University of Tennessee, Knoxville, Tennessee, USA
Appl Environ Microbiol. 2015 Jul;81(13):4546-52. doi: 10.1128/AEM.00346-15. Epub 2015 Apr 24.
Campylobacter jejuni, the leading bacterial cause of human gastroenteritis in the United States, displays significant strain diversity due to horizontal gene transfer. Conjugation is an important horizontal gene transfer mechanism contributing to the evolution of bacterial pathogenesis and antimicrobial resistance. It has been observed that heat shock could increase transformation efficiency in some bacteria. In this study, the effect of heat shock on C. jejuni conjugation efficiency and the underlying mechanisms were examined. With a modified Escherichia coli donor strain, different C. jejuni recipient strains displayed significant variation in conjugation efficiency ranging from 6.2 × 10(-8) to 6.0 × 10(-3) CFU per recipient cell. Despite reduced viability, heat shock of standard C. jejuni NCTC 11168 and 81-176 strains (e.g., 48 to 54°C for 30 to 60 min) could dramatically enhance C. jejuni conjugation efficiency up to 1,000-fold. The phenotype of the heat shock-enhanced conjugation in C. jejuni recipient cells could be sustained for at least 9 h. Filtered supernatant from the heat shock-treated C. jejuni cells could not enhance conjugation efficiency, which suggests that the enhanced conjugation efficiency is independent of secreted substances. Mutagenesis analysis indicated that the clustered regularly interspaced short palindromic repeats system and the selected restriction-modification systems (Cj0030/Cj0031, Cj0139/Cj0140, Cj0690c, and HsdR) were dispensable for heat shock-enhanced conjugation in C. jejuni. Taking all results together, this study demonstrated a heat shock-enhanced conjugation efficiency in standard C. jejuni strains, leading to an optimized conjugation protocol for molecular manipulation of this organism. The findings from this study also represent a significant step toward elucidation of the molecular mechanism of conjugative gene transfer in C. jejuni.
空肠弯曲菌是美国人类肠胃炎的主要细菌性病因，由于水平基因转移而表现出显著的菌株多样性。接合是一种重要的水平基因转移机制，有助于细菌致病性和抗菌药物耐药性的进化。据观察，热休克可提高某些细菌的转化效率。在本研究中，检测了热休克对空肠弯曲菌接合效率的影响及其潜在机制。使用改良的大肠杆菌供体菌株，不同的空肠弯曲菌受体菌株在接合效率上表现出显著差异，范围为每个受体细胞6.2×10^(-8)至6.0×10^(-3) CFU。尽管活力降低，但标准空肠弯曲菌NCTC 11168和81 - 176菌株的热休克（例如，48至54°C处理30至60分钟）可显著提高空肠弯曲菌的接合效率，最高可达1000倍。空肠弯曲菌受体细胞中热休克增强的接合表型可维持至少9小时。热休克处理的空肠弯曲菌细胞的过滤上清液不能提高接合效率，这表明增强的接合效率与分泌物质无关。诱变分析表明，成簇规律间隔短回文重复序列系统和选定的限制修饰系统（Cj0030/Cj0031、Cj0139/Cj0140、Cj0690c和HsdR）对于空肠弯曲菌热休克增强的接合是可有可无的。综合所有结果，本研究证明了标准空肠弯曲菌菌株中热休克增强的接合效率，从而为该生物体的分子操作制定了优化的接合方案。本研究的结果也代表了在阐明空肠弯曲菌接合基因转移分子机制方面迈出的重要一步。
