Comparative Analysis of Two Real-Time PCR Techniques for the Detection of Congenital Syphilis in Umbilical Cord Blood

Background: Syphilis remains a relevant public health problem and continues to be associated with adverse perinatal outcomes. In newborns, conventional serological tests are often difficult to interpret due to maternal antibody transfer and the immaturity of the neonatal immune response. These limitations have led to increasing interest in molecular approaches as complementary diagnostic tools. Methods: This pilot study compared two real-time PCR methodologies for detecting Treponema pallidum DNA in umbilical cord blood: a SYBR Green-based qPCR targeting the polA gene and a TaqMan probe-based qPCR targeting the TpN47 gene. Thirty samples from neonates born to mothers with suspected gestational syphilis were analyzed under standardized conditions. Analytical performance and reproducibility were evaluated before application to clinical samples. Results: Both assays showed stable amplification and acceptable reproducibility. During standardization, the TaqMan assay reached a lower detection limit. However, when applied to clinical samples, the SYBR Green approach identified more positive cases (10/30 vs. 4/30), highlighting differences between analytical performance and behavior in clinical samples. Conclusions: Both methods were suitable for molecular detection of T. pallidum. However, their performance differed in neonatal samples, indicating that analytical sensitivity alone may not predict clinical detection capacity. Molecular testing in cord blood may provide complementary information when serological results are inconclusive.