In spite of the increasing number of BHR plasmids identified, complete sequences of BHR plasmids available in GenBank are still not sufficient for systematically analyzing their genetic diversity. With the development of next-generation sequencing methods, such as 454 pyrosequencing and Illumina high-throughput sequencing technology, more complete sequences of BHR plasmids have been added to the data pool in recent years, most of which were identified as IncP-1 plasmids ( Sen et al., 2012 Brown et al., 2013). To the best of our knowledge, no more than 15 BHR plasmids had been fully sequenced until 2006. Despite the general agreement on the importance of BHR plasmids in the adaptive evolution of bacteria, the BHR plasmids being identified and completely sequenced are still few, initially limited by the high sequencing cost of first generation (Sanger) sequencing technology. Other plasmid regions are comprised of various “accessory” genes conferring important benefits to the host, including resistance to antibiotics ( Rhodes et al., 2004), resistance to heavy metals ( Schneiker et al., 2001), catabolic functions ( Ono et al., 2007), and virulence determinants ( Schlüter et al., 2008), etc.Ĭonjugative gene transfer mediated by BHR plasmids is generally believed to be a common and widespread mechanism for the transfer of genes across a broad phylogenetic range of bacteria ( Top and Springael, 2003 Van der Auwera et al., 2009), and plays a crucial role in the adaptation of bacteria to environmental challenges and spread of antibiotic resistance ( Jechalke et al., 2013). The “plasmid backbone” genes encode proteins involved in the replication, maintenance, control and conjugative transfer of the BHR plasmid. The BHR plasmids typically have mosaic genomes including two distinct regions ( Thomas, 2000). The broad-host-range (BHR) plasmids have been defined as those plasmids that can self-transfer themselves and can stably replicate and maintain in bacterial species from at least two subgroups within the Proteobacteria (e.g., between α- and β- Proteobacteria) ( Szpirer et al., 1999 Sen et al., 2011). They are important members of the mobile gene pool, and are among the most important contributors to horizontal gene transfer between bacteria ( Frost et al., 2005). Plasmids are extra-chromosomal self-replicating DNA elements within the microorganisms ( Mela et al., 2008). Our study increases the available collection of complete genome sequences of BHR plasmids, and since pSFA231 is the only characterized PromA plasmid from China, our findings also enhance our understanding of the genetic diversity of this plasmid group in different parts of the world. Alternatively, they may also be accessory genes that were first acquired and then stayed as the plasmid diverged. Interestingly, a cluster of hypothetical orfs located between parA and traA of pSFA231 shows high similarity with the corresponding regions on pMOL98, pIPO2T, and pTer331, suggesting these hypothetical orfs may represent “essential” plasmid backbone genes for the PromA-β subgroup. We propose to divide the PromA group into two subgroups, PromA-α (pMRAD02, pSB102) and PromA-β (pMOL98, pIPO2T, pSFA231, pTer331), based on the splits network analysis of the RepA protein. Nevertheless, phylogenetic divergence was found in specific gene products. Further comparative genomic analysis shows that pSFA231 shares the common backbone regions with the other PromA plasmids, i.e., genes involved in replication, maintenance and control, and conjugative transfer. Phylogenetic analysis grouped pSFA231 into the newly defined PromA plasmid family, which currently includes five members. Based on its complete sequence the plasmid has a size of 41.5 kb and codes for 50 putative open reading frames (orfs), 29 of which represent genes involved in replication, partitioning and transfer functions of the plasmid.
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