Abstract

Plasmids are components of many bacterial genomes. They enable the spread of a large pool of genetic information via lateral gene transfer. Many bacterial strains contain mega-sized replicons and these are particularly common in Alphaproteobacteria. Considerably less is known about smaller alphaproteobacterial plasmids. We analyzed the genomes of 14 such plasmids residing in 4 multireplicon carotenoid-producing strains of the genus Paracoccus (Alphaproteobacteria): P. aestuarii DSM 19484, P. haeundaensis LG P-21903, P. marcusii DSM 11574 and P. marcusii OS22. Comparative analyses revealed mosaic structures of the plasmids and recombinational shuffling of diverse genetic modules involved in (i) plasmid replication, (ii) stabilization (including toxin-antitoxin systems of the relBE/parDE, tad-ata, higBA, mazEF and toxBA families) and (iii) mobilization for conjugal transfer (encoding relaxases of the MobQ, MobP or MobV families). A common feature of the majority of the plasmids is the presence of AT-rich sequence islets (located downstream of exc1-like genes) containing genes, whose homologs are conserved in the chromosomes of many bacteria (encoding e.g. RelA/SpoT, SMC-like proteins and a retron-type reverse transcriptase). The results of this study have provided insight into the diversity and plasticity of plasmids of Paracoccus spp., and of the entire Alphaproteobacteria. Some of the identified plasmids contain replication systems not described previously in this class of bacteria. The composition of the plasmid genomes revealed frequent transfer of chromosomal genes into plasmids, which significantly enriches the pool of mobile DNA that can participate in lateral transfer. Many strains of Paracoccus spp. have great biotechnological potential, and the plasmid vectors constructed in this study will facilitate genetic studies of these bacteria.

Highlights

  • Bacterial plasmids have a modular structure: their genomes can be separated into several DNA cassettes encoding specific functions

  • As a result of this approach we have identified and analyzed (i) four related repABC as well as several pTAV3-type megaplasmids – both groups residing in P. versutus UW1 and four strains of Paracoccus pantotrophus [6,7], (ii) plasmid pALC1 of Paracoccus alcaliphilus JCM 7364, with an iteron-containing replication system [8], (iii) plasmid pMTH1 of Paracoccus methylutens DM12, whose genome is predominantly (80%) composed of transposable modules (TMos) [9], (iv) three plasmids of Paracoccus aminophilus JCM 7686, whose REP modules were used for the construction of versatile DIY cassettes [10,11], as well as (v) plasmid pWKS1 of P. pantotrophus DSM 11072 – the smallest replicon identified so far in Paracoccus spp. [12]

  • Four of the strains (P. aestuarii DSM 19484, P. haeundaensis LG P-21903, P. marcusii DSM 11574 and P. marcusii OS22 – all able to produce beta-carotenoid pigments) contained numerous smaller replicons ranging in size from approx. 2.5 kb to 85 kb

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Summary

Introduction

Bacterial plasmids have a modular structure: their genomes can be separated into several DNA cassettes encoding specific functions. Many plasmids are giant molecules that can even exceed the size of some bacterial chromosomes Such mega-sized replicons (megaplasmids) are common in Alphaproteobacteria. Analysis of the genomic data collected by The National Center for Biotechnology Information (NCBI) revealed that the sequenced genomes of 240 alphaproteobacterial strains include a total of 315 plasmids. Twenty six of these strains are multi-plasmid containing, with at least five extrachromosomal replicons. DNA into bacterial cells and plasmid stability assay. Stationary-phase cultures of plasmid-containing strains were diluted in fresh medium without antibiotic selection and cultivated for approximately 10, 20 and 30 generations. Plasmid retention was determined from the percentage of kanamycin-resistant colonies

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