Molecular Characterization of Plasmids Harbored by Actinomycetes Isolated From the Great Salt Plains of Oklahoma Using PFGE and Next Generation Whole Genome Sequencing

Abstract

One of the unique features of actinomycetes, especially the genus Streptomyces, is the presence of linear plasmids. These range in size from 12 to 600 kb, and are often termed mega-plasmids. While many of the genes involved in secondary metabolite production reside in clusters on the chromosome, several studies have identified biosynthetic clusters on large linear plasmids that produce important secondary metabolites, including antibiotics. In this study, Pulse Field Gel Electrophoresis (PFGE) was used to screen 176 actinomycete isolates for the presence of plasmids; these bacterial strains were previously isolated from the Great Salt Plains of Oklahoma. Seventy-eight of the 176 actinomycete isolates (44%) contained plasmids. Several strains contained more than one plasmid, accounting for a total of 109 plasmids. Ten isolates showed extrachromosomal DNA larger than 200 kb, thus falling into the category of mega-plasmids. A subset of plasmids from 55 isolates was treated with S1 nuclease to determine topology; all plasmids examined appeared to be linear and ranged from ~55 to 400 kb. Eleven isolates were chosen for Whole Genome Next Generation Sequencing. From the 11 sequenced isolates, seven plasmids were partially assembled. While the majority of the genes identified on the plasmids coded for hypothetical proteins, others coded for general functions, stress response, and antibiotic and heavy metal resistance. Draft genome sequences of two mega-plasmid-bearing Streptomyces sp. strains, BF-3 and 4F, revealed the presence of genes involved in antibiotic production, antibiotic, and heavy metal resistance, osmoregulation, and stress response, which likely facilitate their survival in this extreme halophilic environment. To our knowledge, this is the first study to explore plasmids harbored by actinomycetes isolated from the Great Salt Plains of Oklahoma.

First Report of the Local Spread of Vancomycin-Resistant Enterococci Ascribed to the Interspecies Transmission of a vanA Gene Cluster-Carrying Linear Plasmid

“…2; see also Fig. S3) (19). In contrast to the 22 isolates that seemed to carry the same linear plasmid, the size of each plasmid of KUHS4, KUHS20, and KUHS22 was observed to be larger and not consistent with that of each of the others; however, these plasmids also migrated into the PFGE gel without S1 nuclease treatment, suggesting that their topology was also linear (Fig.…”

Novel Multidrug-Resistant Enterococcal Mobile Linear Plasmid pELF1 Encoding vanA and vanM Gene Clusters From a Japanese Vancomycin-Resistant Enterococci Isolate

“…This plays an important role in the horizontal spread of vancomycin resistance. Among mobile genetic elements, a “plasmid” is originally considered an extrachromosomal genetic element with circular topology (Cornell et al, 2018). In contrast, linear DNA elements have already been reported in various prokaryotes and eukaryotes (Hinnebusch and Tilly, 1993; Cornell et al, 2018).…”

Molecular characterization of covalently closed circular plasmid DNA in local Streptomyces strains isolated from soil with detection of antibiotic production and antibacterial activity

“…Since a Streptomyces strain contains about 20 different gene clusters for the production of antibiotics, they form an enormous pool of antibiotic resistance genes in the soil, which can be passed to another bacteria by horizontal gene transfer (Muth, 2012) . The genes of Streptomyces involved in secondary metabolite production be located as clusters on the chromosome, also, several studies have recognized biosynthetic clusters on the linear plasmids that produce important secondary metabolites (Cornell et al, 2018) .…”