Immunohistochemical- and PCR-based assay for the reproducible, routine detection of erythrovirus B19 in thyroid...

It is generally accepted that thyroid follicle cells are at least semi-permissive for erythrovirus B19 (EVB19). Thus, various laboratory techniques have been successfully used to detect EVB19 in the thyroid gland, including polymerase chain reaction (PCR), immunohistochemistry, and in situ hybridization. However, the detection of EVB19 within the thyroid gland is problematic, and none of the detection protocols in the literature have been unequivocally validated. This multidisciplinary study in which 32 thyroidectomy subjects undergoing thyroidectomy in a French University hospital were prospectively recruited was performed over a period of 3 years. Prior to surgery, all the subjects were assayed for blood levels of anti-EVB19 antibodies and (using a quantitative PCR [qPCR] assay) EVB19 itself. A qPCR assay for EVB19 and an immunohistochemical assay (based on polyclonal anti-VP2 antibodies) were performed on the thyroidectomy samples. None of the subjects had an acute EVB19 infection. A viral load was detected in two serum samples and six thyroid biopsies. Three subjects had both a positive immunohistochemical assay and a positive qPCR assay for the thyroid tissue. It is noteworthy that the thyroid immunohistochemical and qPCR assays were negative in the two patients with detectable serum loads of EVB19. In conclusion, EVB19 can be detected in thyroid follicle cells by using immunohistochemical and qPCR assays. Ideally, patients should be tested with both PCR and immunohistochemical assays, in order to unequivocally confirm or rule out the presence of EVB19 in the thyroid gland. The present protocol must now be validated in larger series--notably with respect to its reliability and in order to determine qPCR positivity thresholds for application in future large-scale studies.