Cyclospora cayetanensis is a foodborne protozoan parasite that causes the human diarrheal disease cyclosporiasis. Recently, the US FDA developed a modified real-time PCR method based on a specific mitochondrial target gene (Mit1C) to detect C. cayetanensis in fresh produce. The method was validated by single laboratory validation (SLV) studies in Romaine lettuce, cilantro, and raspberries. The present study aimed to evaluate the performance of the new real-time Mit1C (Mit1C qPCR) method by comparing it with the current BAM
- Chapter 19b qPCR (18S qPCR) as the reference method for the detection of the protozoan parasite C. cayetanensis in fresh produce in a multi-laboratory validation (MLV) setting with the participation of 13 collaborating laboratories. Each laboratory analyzed twenty-four blind-coded Romaine lettuce DNA test samples that included: two unseeded samples, three samples seeded with five oocysts, and one sample seeded with 200 oocysts in the first round and five unseeded samples, eight samples seeded with five oocysts, and five samples seeded with 200 oocysts in the second round. The overall detection rates across laboratories for Romaine lettuce samples inoculated with 200 and 5 oocysts and un-inoculated samples were 100% (78/78), 69.23% (99/143), and 1.1% (1/91), respectively, for Mit1C qPCR, and 100% (78/78), 61.54% (88/143) and 0% (0/91), respectively, for 18S qPCR. The relative level of detection (RLOD = LOD