Real-time PCR assays for genotyping of Cryptococcus gattii in North America
Author
Kelley, ErinDriebe, Elizabeth
Etienne, Kizee
Brandt, Mary
Schupp, James
Gillece, John
Trujillo, Jesse
Lockhart, Shawn
Deak, Eszter
Keim, Paul
Engelthaler, David
Affiliation
The Translational Genomics Research Institute, 3051 W. Shamrell Blvd. Ste. 106, Flagstaff, AZ 86001, USACenters for Disease Control and Prevention, Atlanta, GA, USA
Department of Pathology and Laboratory Medicine, David Geffen School of Medicine at UCLA, Los Angeles, CA, USA
Center for Microbial Genetics and Genomics, Northern Arizona University, Flagstaff, AZ, USA
Issue Date
2014
Metadata
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BioMed CentralCitation
Kelley et al. BMC Microbiology 2014, 14:125 http://www.biomedcentral.com/1471-2180/14/125Journal
BMC MicrobiologyRights
© 2014 Kelley et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0).Collection Information
This item is part of the UA Faculty Publications collection. For more information this item or other items in the UA Campus Repository, contact the University of Arizona Libraries at repository@u.library.arizona.edu.Abstract
BACKGROUND:Cryptococcus gattii has been the cause of an ongoing outbreak starting in 1999 on Vancouver Island, British Columbia and spreading to mainland Canada and the US Pacific Northwest. In the course of the outbreak, C. gattii has been identified outside of its previously documented climate, habitat, and host disease. Genotyping of C. gattii is essential to understand the ecological and geographical expansion of this emerging pathogen.METHODS:We developed and validated a mismatch amplification mutation assay (MAMA) real-time PCR panel for genotyping C. gattii molecular types VGI-VGIV and VGII subtypes a,b,c. Subtype assays were designed based on whole-genome sequence of 20 C. gattii strains. Publically available multilocus sequence typing (MLST) data from a study of 202 strains was used for the molecular type (VGI-VGIV) assay design. All assays were validated across DNA from 112 strains of diverse international origin and sample types, including animal, environmental and human.RESULTS:Validation revealed each assay on the panel is 100% sensitive, specific and concordant with MLST. The assay panel can detect down to 0.5 picograms of template DNA.CONCLUSIONS:The (MAMA) real-time PCR panel for C. gattii accurately typed a collection of 112 diverse strains and demonstrated high sensitivity. This is a time and cost efficient method of genotyping C. gattii best suited for application in large-scale epidemiological studies.EISSN
1471-2180Version
Final published versionAdditional Links
http://www.biomedcentral.com/1471-2180/14/125ae974a485f413a2113503eed53cd6c53
10.1186/1471-2180-14-125
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Except where otherwise noted, this item's license is described as © 2014 Kelley et al.; licensee BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0).