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United States Department of Agriculture

Agricultural Research Service

Research Project: MOLECULAR CHARACTERIZATION OF PATHOGENS AND THEIR RESPONSES TO ENVIRONMENTAL FACTORS

Location: Molecular Characterization of Foodborne Pathogens

Title: Typing and characterization of ColE1-like plasmids conferring kanamycin resistance in Salmonella enterica serotypes

Authors
item Chen, Chin-Yi
item Lindsey, Rebecca
item Strobaugh, Terence
item Frye, Jonathan
item Cray, Paula
item Meinersmann, Richard

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: February 21, 2009
Publication Date: May 17, 2009
Citation: Chen, C., Lindsey, R.L., Strobaugh Jr, T.P., Frye, J.G., Cray, P.J., Meinersmann, R.J. 2009. Typing and characterization of ColE1-like plasmids conferring kanamycin resistance in Salmonella enterica serotypes. Meeting Abstract. Poster Z048.

Technical Abstract: Background: Multi-antibiotic resistant Salmonella enterica serotypes are increasing in prevalence and concern in human and animal health. Many strains carry resistance determinants on plasmids; current practices focus heavily on large plasmids and the role small plasmids play in resistance gene transfer are largely unknown. Previous studies in our labs found that some isolates harbor ColE1-like plasmids that carry the aph gene responsible for kanamycin-resistance (KAN-R) phenotypes. Methods: 97 KAN-R Salmonella isolates from a NARMS study between 2005 and 2007 were screened by PCR using a ColE1 primer set. DNA was isolated and transformed into E. coli DH5alpha for plasmid propagation and further characterization. Plasmid DNA from DH5alpha was purified and the nucleotide sequences were determined from representative clones using ABI BigDye Terminator and custom primers. Sequences were assembled using Sequencher software and nucleotide and amino acids sequence similarity was compared to the NCBI GenBank using BLAST. Results: Using the ColE1 typing primers, 26 of the 97 KAN-R isolates screened were positive (27%). Restriction mapping revealed three major plasmid types consisting of 3 or more isolates, and 3 other patterns representing only a single isolate. Nucleotide sequencing results showed that 2 of the plasmid types were conserved with previously sequenced Salmonella plasmids carrying aph(3’)-I gene, pU302S and pSN11/00Kan, respectively. The third major type showed limited sequence similarity to other ColE1-like plasmids found in GenBank. Conclusion: The ColE1 typing primers successfully identified known plasmid types as well as new types. These and other ColE1 typing primers will be added to the widely used plasmid replicon typing primer sets for routine screening, and will also be added to a microarray probe set for characterizing antibiotic-resistance genes/plasmids. Due to high copy number and mobility, the ColE1-like plasmids may play a critical role in transmission of antibiotic resistance genes among enteric pathogens and these findings warrant a close monitoring of this plasmid incompatibility group.

Last Modified: 4/17/2014
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