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Journal of Molecular Diagnostics 2007, Vol. 9, No. 5
Copyright © 2007 American Society for Investigative Pathology & Association for Molecular Pathology
DOI: 10.2353/jmoldx.2007.070064

Translational Genomics to Develop a Salmonella enterica Serovar Paratyphi A Multiplex Polymerase Chain Reaction Assay

Hong-Yu Ou*, Cindy Teh Shuan Ju{dagger}, Kwai-Lin Thong{dagger}, Norazah Ahmad{ddagger}, Zixin Deng*, Michael R. Barer§ and Kumar Rajakumar§

From the Laboratory of Microbial Metabolism and School of Life Science and Biotechnology, * Shanghai Jiaotong University, Shanghai, China; the Institute of Biological Sciences, {dagger} University of Malaya, Kuala Lumpur, Malaysia; the Institute for Medical Research, {ddagger} Kuala Lumpur, Malaysia; the Department of Infection, Immunity, and Inflammation, § Leicester Medical School, University of Leicester, Leicester, United Kingdom; and the Department of Clinical Microbiology, University Hospitals of Leicester NHS Trust, Leicester, United Kingdom

The use of pathogen genome sequence data for the control and management of infections remains an ongoing challenge. We describe a broadly applicable, web-enabled approach that can be used to develop bacteria-specific polymerase chain reaction (PCR) assays. Salmonella enterica Paratyphi A has emerged as a major cause of enteric fever in Asia. Culture-based diagnosis is slow and frequently negative in patients with suspected typhoid and paratyphoid fever, potentially compromising patient management and public health. We used the MobilomeFINDER web-server to perform in silico subtractive hybridization, thus identifying 43 protein-coding sequences (CDSs) that were present in two Paratyphi A strains but not in other sequenced Salmonella genomes. After exclusion of 29 CDSs found to be variably present in Paratyphi A strains by microarray hybridization and grouping of remaining CDSs by genomic location, four dispersed targets (stkF, spa2473, spa2539, hsdM) were used to develop a highly discriminatory multiplex PCR assay. All 52 Paratyphi A strains within the diverse panel investigated produced one of two pathognomonic four-band signatures. Given rapid and ongoing expansion of DNA and comparative genomics databases, our universally accessible web-server-supported do-it-yourself approach offers the potential to contribute significantly to the rapid development of species-, serovar-, or pathotype-specific PCR assays targeting pre-existing and emerging bacterial pathogens.




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W. W. Zhang and J. Versalovic
Expanding the Diagnostic Capabilities of Molecular Microbiology by Genomic Methods
J. Mol. Diagn., November 1, 2007; 9(5): 572 - 573.
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Copyright © 2007 by the American Society for Investigative Pathology and the Association for Molecular Pathology.