Author:
Mitchell Marilyn,Dizon Dominic,Libke Robert,Peterson Michael,Slater David,Dhillon Akashdeep
Abstract
Rapid real-time PCR (RT-PCR) can be performed in a community hospital setting to identifyCoccidioidesspecies using the new Becton Dickinson molecular instrument BD Max. Following sample preparation, DNA extraction and PCR were performed on the BD Max using the BD Max extraction kit ExK-DNA-1 test strip and a master mix prepared by BioGX (Birmingham, AL). Sample preparation took 2 h, and testing on the BD Max took an additional 2 h. Method sensitivity and specificity were evaluated along with the limits of detection to confirm that this convenient method would provide medically useful information. Using serial dilutions, the lower limit of detection was determined to be 1 CFU/μl. Testing with this method was validated using samples from various body sites, including bronchial alveolar lavage (BAL) fluid; sputum and lung tissue samples; and pleural and spinal fluids. Safety protocols were established, and specimen preparation processes were developed for the various types of specimens. The range for the cycle threshold (CT) indicating adequate fluorescent signal to signify a positive result was established along with the acceptable range for the internal standard. Positive controls run with each batch were prepared by spiking a pooled BAL fluid specimen with a known dilution ofCoccidioides immitisorganism. Our experience with testing >330 patient samples shows that clinically relevant information can be available within 4 h using an RT-PCR method on the BD Max to identifyCoccidioidesspp., with sensitivity equivalent to culture.
Publisher
American Society for Microbiology
Reference14 articles.
1. New perspectives on coccidioidomycosis;Proc Am Thorac Soc,2010
2. Increase in reported coccidioidomycosis–United States, 1998–2011;Centers for Disease Control and Prevention (CDC);MMWR Morb Mortal Wkly Rep,2013
3. Recent advances in our understanding of the environmental, epidemiological, immunological, and clinical dimensions of coccidioidomycosis;Clin Microbiol Rev,2013
4. Centers for Disease Control and Prevention. 2014. Valley fever (coccidioidomycosis) statistics. http://www.cdc.gov/fungal/diseases/coccidioidomycosis/statistics.html.
5. Coccidioidomycosis as a common cause of community-acquired pneumonia;Emerg Infect Dis,2006
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