摘要

通过与作为可动员质粒的pBBR1MCS-2和作为受体的假单胞菌（Pseudomonas resinovorans）的三亲交配，从环境样品中外源捕获新的自身可传递的质粒。从厌氧甲烷发酵厂的颗粒和牛粪中成功获得了总共272个受体作为质粒宿主候选物。测定了6个质粒的全部核苷酸序列，包括一个IncP-1质粒（pSN1104-59），四个PromA样质粒（pSN1104-11，pSN1104-34，pSN0729-62和pSN0729-70），以及一个新的质粒。 （pSN1216-29），其先前未确定其不相容性组。在这些质粒中未发现先前已知的抗生素抗性基因。深入的系统发育分析表明，PromA样质粒属于PromA亚群（命名为PromAγ和PromAδ），与先前提出的亚群PromAα和PromAβ不同。通过与其他PromA质粒的比较，将24个基因鉴定为骨架基因。 pSN1216-29的核苷酸序列与临床分离株中的核苷酸序列具有高度同一性。从编码复制起始蛋白和oriV的repA成功构建了pSN1216-29的微型复制子。发现所有捕获的质粒具有广泛的宿主范围，并且可以转移到不同类别的变形菌门中并在其中复制。值得注意的是，pSN1216-29的repA和oriV显示出与pSRC119-A / C的两个复制系统之一的高度相似性，pSRC119-A / C被称为具有在沙门氏菌血清型Senftenberg中发现的多药抗性基因的质粒。我们的研究结果表明，这些“神秘”但广泛宿主范围的质粒对于将几种基因作为“载体”在自然环境中的更广泛的细菌中传播可能是重要的。

Novel self-transmissible plasmids were exogenously captured from environmental samples by triparental matings with pBBR1MCS-2 as a mobilizable plasmid and Pseudomonas resinovorans as a recipient. A total of 272 recipients were successfully obtained as plasmid host candidates from granules of an anaerobic methane fermentation plant and from cow manure. The whole nucleotide sequences of six plasmids were determined, including one IncP-1 plasmid (pSN1104-59), four PromA-like plasmids (pSN1104-11, pSN1104-34, pSN0729-62, and pSN0729-70), and one novel plasmid (pSN1216-29), whose incompatibility group has not been previously identified. No previously known antibiotic resistance genes were found in these plasmids. In-depth phylogenetic analyses showed that the PromA-like plasmids belong to subgroups of PromA (designated as PromAγ and PromAδ) different from previously proposed subgroups PromAα and PromAβ. Twenty-four genes were identified as backbone genes by comparisons with other PromA plasmids. The nucleotide sequences of pSN1216-29 share high identity with those found in clinical isolates. A minireplicon of pSN1216-29 was successfully constructed from repA encoding a replication initiation protein and oriV. All the captured plasmids were found to have a broad host range and could be transferred to and replicated in different classes of Proteobacteria. Notably, repA and oriV of pSN1216-29 showed high similarity with one of two replication systems of pSRC119-A/C, known as a plasmid with multidrug resistance genes found in Salmonella enterica serovar Senftenberg. Our findings suggest that these "cryptic" but broad-host-range plasmids may be important for spreading several genes as "vehicles" in a wider range of bacteria in natural environments.

https://www.ncbi.nlm.nih.gov/pubmed/30459733