Inter-laboratory optimization for the detection of Campylobacter jejuni in canine fecal samples using gyrA PCR

A 2016-2018 outbreak in the United States demonstrated the importance of pet dogs as a source of Campylobacter jejuni. Canine feces are not tested routinely for C. jejuni. We previously compared culture-based and molecular methods for detection of C. jejuni and found that a cpn60 real-time PCR (rtPCR) assay using DNA isolated directly from canine feces was more reliable than culture, with a limit of detection (LOD) of 320 cfu/g. Later, during an outbreak investigation, a gyrA rtPCR assay had a LOD of 40 cfu/g. In our inter-laboratory comparison exercise, 27 analysts at 19 laboratories successfully detected C. jejuni in canine feces using the gyrA rtPCR. Samples inoculated with 45,600 cfu/g were correctly identified as positive by 100% of analysts. Samples inoculated with 4,700, 460, or 200 cfu/g were correctly identified by 96%, 77%, and 40% of analysts, respectively. The addition of C. jejuni gyrA rtPCR to previous methods allows laboratories to rapidly identify C. jejuni-positive canine fecal samples at a lower LOD. Isolation of DNA directly from feces rather than from enrichment broth offers the advantage of fewer opportunities for cross-contamination during testing and decreased time between specimen collection and test completion. The updated method, with the addition of the gyrA rtPCR, offers increased sensitivity and will ultimately improve overall detection of C. jejuni in canine feces at veterinary diagnostic laboratories, further supporting canine and public health.