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ARS Home » Northeast Area » Beltsville, Maryland (BARC) » Beltsville Agricultural Research Center » Environmental Microbial & Food Safety Laboratory » Research » Publications at this Location » Publication #340902

Research Project: Zoonotic Parasites Affecting Food Animals, Food Safety, and Public Health

Location: Environmental Microbial & Food Safety Laboratory

Title: Interlaboratory validation of an improved U.S. Food and Drug Administration method for detection of Cyclospora cayetanensis in produce using TaqMan real-time PCR

Author
item Murphy, Helen - Us Food & Drug Administration (FDA)
item Cinar, Hediye Nese - Us Food & Drug Administration (FDA)
item Gopinath, Gopal - Us Food & Drug Administration (FDA)
item Im, Annhee - Us Food & Drug Administration (FDA)
item Noe, Kathy - Us Food & Drug Administration (FDA)
item Chatman, Lacresha - Us Food & Drug Administration (FDA)
item Miranda, Nancy - Us Food & Drug Administration (FDA)
item Wetherington, June - Us Food & Drug Administration (FDA)
item Neal-mckinney, Jason - Us Food & Drug Administration (FDA)
item Piresc, Gabrielle - Us Food & Drug Administration (FDA)
item Sachs, Elizabeth - Us Food & Drug Administration (FDA)
item Stanya, Kristopher - Us Food & Drug Administration (FDA)
item Jognson, Cynthia - Us Food & Drug Administration (FDA)
item Nascimiento, Fernanda - Centers For Disease Control And Prevention (CDCP) - United States
item Santin-duran, Monica
item Molokin, Aleksey
item Samadpour, Mansour - National Institute Of Environmental Health Sciences (NIEHS, NIH)
item Janagama, Harish - National Institute Of Environmental Health Sciences (NIEHS, NIH)
item Kahler, Amy - Centers For Disease Control And Prevention (CDCP) - United States
item Miller, Candace - Centers For Disease Control And Prevention (CDCP) - United States
item Da Silva, Alexandre - Us Food & Drug Administration (FDA)

Submitted to: International Journal of Food Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 8/8/2017
Publication Date: 8/24/2017
Citation: Murphy, H.R., Cinar, H., Gopinath, G., Im, A., Noe, K.E., Chatman, L.D., Miranda, N.E., Wetherington, J.H., Neal-Mckinney, J., Piresc, G.S., Sachs, E., Stanya, K.J., Jognson, C.L., Nascimiento, F., Santin, M., Molokin, A., Samadpour, M., Janagama, H., Kahler, A., Miller, C., Da Silva, A.J. 2017. Interlaboratory validation of an improved U.S. Food and Drug Administration method for detection of Cyclospora cayetanensis in produce using TaqMan real-time PCR. International Journal of Food Microbiology. 69:170-178.

Interpretive Summary: Cyclospora cayetanensis is a protozoan parasite that has emerged worldwide as an agent of diarrheal disease in humans called cyclosporiasis, which can be transmitted by consumption of contaminated food or water. In developed countries foodborne transmission of C. cayetanensis has become a major public health concern and is generally associated with contaminated fresh produce. In the U.S, foodborne outbreaks and sporadic cases of cyclosporiasis have occurred linked to imported fruit and vegetables, including basil, snow peas, berries, cilantro, and mixed salads. A collaborative validation study was performed to evaluate the performance of a new U.S. Food and Drug Administration method developed for detection of C. cayetanensis on cilantro and raspberries. The method includes a sample preparation step followed by a molecular detection assay that provides species specific identification. Five laboratories blindly analyzed a total of 320 samples consisting of 25 g of cilantro or 50 g of raspberries that were either uninoculated or artificially contaminated with C. cayetanensis oocysts. Reproducibility between laboratories and analysts was high and the method was shown to be an effective analytical tool for detection of C. cayetanensis in produce. This method will facilitate studies to understand the prevalence of C. cayetanensis in fresh produce and the environment and identify the necessary preventative control measures to reduce foodborne exposure to C. cayetanensis.

Technical Abstract: A collaborative validation study was performed to evaluate the performance of a new U.S. Food and Drug Administration method developed for detection of the protozoan parasite, Cyclospora cayetanensis, on cilantro and raspberries. The method includes a sample preparation step in which oocysts are recovered from produce using an enhanced produce washing solution containing 0.1% Alconox and a commercially available method to disrupt the C. cayetanensis oocysts and extract DNA. A TaqMan real-time PCR assay targeting the C. cayetanensis 18S rDNA gene with an internal amplification control to monitor PCR inhibition provides species specific identification. Five laboratories blindly analyzed a total of 320 samples consisting of 25 g of cilantro or 50 g of raspberries that were either uninoculated or artificially contaminated with C. cayetanensis oocysts. Detection rates for cilantro inoculated with 200, 10, and 5 oocysts, were 100%, 80%, and 31%, respectively. For raspberries, the detection rates for samples inoculated with 200, 10, and 5 oocysts were 100%, 90% and 50%, respectively. All uninoculated samples, DNA blank extracts, and no-template PCR controls were negative. Reproducibility between laboratories and analysts was high and the method was shown to be an effective analytical tool for detection of C. cayetanensis in produce.