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PCR revisited: A case for revalidation of PCR assays for microorganisms using identification of Campylobacter species as an exemplar


We re-examined the sensitivity and specificity of 31 PCR assays (including four commercially available and developed in-house methods) for the identification of Campylobacter species, with particular reference to taxa described since 2004, which are closely related to C. jejuni and C. coli, the pathogenic species of most interest. Each of the assays was used by at least one of the participating nine laboratories in eight countries. The sensitivity and specificity of these PCR assays examined varied considerably and ranged from 100% to 0% for sensitivity and 100% to 55% for specificity. None of the three assays examined for C. lari were successful in detecting all strains of this species, possibly reflecting its complex taxonomy. A number of assays for C. jejuni, C. coli, and a subgroup of enteropathogenic campylobacters, were found to yield false positive results for Campylobacter species described since PCR tests were reported, including C. cuniculorum, C. subantarcticus, C. peloridis and C. volucris. Our study supports the need for attention to detail in initial PCR assay design and evaluation, and also for on-going revalidation of laboratory assays to ensure that diagnoses are correct. Recommendations to guide the revalidation process are presented.

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