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Title: An oligonucleotide microarray to characterize multidrug resistant plasmids

item Lindsey, Rebecca
item Frye, Jonathan
item Cray, Paula
item Welch, Timothy - Tim
item Meinersmann, Richard - Rick

Submitted to: Journal of Microbiological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/28/2010
Publication Date: 2/4/2010
Citation: Lindsey, R.L., Frye, J.G., Cray, P.J., Welch, T.J., Meinersmann, R.J. 2010. An oligonucleotide microarray to characterize multidrug resistant plasmids. Journal of Microbiological Methods. 81(2):96-100.

Interpretive Summary: Salmonella are food borne bacterial pathogens that can result in serious illness in humans and animals. Salmonella may also be acquired by the oral ingestion of contaminated water. Approximately 600 people die each year in the US after being infected. Salmonella carries assorted plasmids which are pieces of deoxyribonucleic acid (DNA) that can be easily transferred from one bacterial strain to another. There is a lot of interest in characterizing and tracking plasmids because they may carry genes or sequences which benefit survival of the bacterium. Some of these genes may confer resistance to antimicrobials. While treatment of salmonellosis is not typically recommended, in cases where it is necessary, resistance to antimicrobials, and in particular multiple antimicrobials, may compromise outcome. Inc A/C and H1 type plasmids are of interest because they are associated with multidrug resistance. Hybridization microarrays are a tool that is superior to PCR for analysis of genetic content of complex plasmids as well as genomes. Due to their high density, microarrays have the ability to interrogate simultaneously a much larger number of genes in a single assay. Detecting the gene content of the plasmids can be used to determine the minimum core content needed for transmission and also to reconstruct a phylogeny. Phylogeny is useful for demonstrating patterns of transmission and determining the rate of transmission between potential host bacteria. Thus we undertook to create a tool, hybridization arrays, to evaluate the gene content of Inc A/C and H1 plasmids. These data are useful for physicians, veterinarians, scientists and epidemiologists as they develop models to better understand the evolution and transmission of antimicrobial resistance.

Technical Abstract: Bacteria plasmids are fragments of extra-chromosomal double stranded deoxyribonucleic acid (DNA) that can contain a variety of genes beneficial to the host organism like antibiotic drug resistance. Many of the Enterobacteriaceae carry multiple drug resistance (MDR) genes on large plasmids of replicon type Inc A/C and Inc H1. It is important to understand the transmission of these MDR plasmids because the genes they carry can affect the outcome of antimicrobial therapy. The aim was to design a microarray with oligonucleotide probes for every gene in the six Inc A/C and one Inc H1 plasmids of interest while representing all redundant sequences only once. The microarray is printed in triplicate with 493 unique oligonucleotide probes 70 nucleotides in length. Salmonella enterica and Escherichia coli control strains and test isolates (both native and transformed into a known E. coli background strain) were hybridized to the plasmid microarray. This hybridization arrays presents a rapid and cost effective method for high-density screening of isolates to evaluate the gene content of Inc A/C and H1 plasmids.