Assessing genetic diversity in plasmids from Escherichia coli and Salmonella enterica using a mixed‐plasmid microarray

Aims:  To compare genetic composition of plasmids using microarrays composed of randomly selected fragments of plasmid DNA. Methods and Results:  Separate shotgun libraries were constructed from plasmid DNA pooled from Escherichia coli and Salmonella enterica. Cloned fragments were used as probes fo...

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Bibliographic Details
Published in:Journal of applied microbiology 2006-01, Vol.100 (1), p.15-28
Main Authors: Call, D.R., Kang, M.‐S., Daniels, J., Besser, T.E.
Format: Article
Language:English
Subjects:
beta-Lactamases - genetics
Biodiversity
Biological and medical sciences
blaCMY‐2
Cephalosporin Resistance - genetics
comparative genomics
DNA fingerprint
DNA, Bacterial - genetics
Escherichia coli
Escherichia coli - genetics
Fundamental and applied biological sciences. Psychology
Gene Library
Genes, Bacterial - genetics
Microbiology
mosaic
Nucleic Acid Hybridization - methods
Oligonucleotide Array Sequence Analysis - methods
Plasmids - genetics
Reproducibility of Results
Salmonella
Salmonella enterica
Salmonella enterica - genetics
Sequence Analysis, DNA - methods
transposase
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Summary:Aims:  To compare genetic composition of plasmids using microarrays composed of randomly selected fragments of plasmid DNA. Methods and Results:  Separate shotgun libraries were constructed from plasmid DNA pooled from Escherichia coli and Salmonella enterica. Cloned fragments were used as probes for microarrays. Plasmid targets were labelled, hybridized overnight, and bound targets were imaged after enzymatic signal amplification. Control hybridizations demonstrated significantly higher signal when probes and targets shared >95% sequence identity. Diagnostic sensitivity and specificity of the assay was 95 and 99%, respectively. Cluster analysis showed close matches for replicate experiments with a high correlation between replicates (r = 0·91) compared with the correlation for nonreplicates (r = 0·09). Analysis of hybridization data from 43 plasmids generated five distinct clusters, two for known serovar‐specific plasmids, one for enterohemorrhagic E. coli plasmids, and two for plasmids harboring a recently disseminated antibiotic resistance gene (blaCMY‐2). Conclusion:  Mixed‐plasmid microarrays are suitable for comparing genetic content of wild‐type plasmids and hybridization results from this study suggest several novel hypotheses about plasmid gene exchange between E. coli and S. enterica. Significance and Impact of Study:  Mixed‐plasmid microarrays permit rapid, low cost analysis and comparison of many plasmids. This ability is critical to understanding the source, fate, and transport of plasmids amongst commensal and pathogenic bacteria.
ISSN:1364-5072
1365-2672
DOI:10.1111/j.1365-2672.2005.02775.x