Urine as a complementary specimen for RT-qPCR detection of Oropouche virus.

Oropouche fever is an emerging arboviral disease in South America, for which reliable molecular diagnostic specimens throughout the course of infection are essential for surveillance and clinical management. This study aimed to compare the diagnostic performance of RT-qPCR in paired urine and serum samples and to evaluate temporal trends in cycle threshold (Ct) values according to time since symptom onset. A retrospective analysis was conducted on 41 paired serum–urine samples (one pair per patient) collected within 15 days after symptom onset. RT-qPCR results were classified as detectable or undetectable, with Ct values recorded when available. Positivity rates were compared using McNemar’s test, inter-matrix agreement was assessed with Cohen’s kappa, and paired Ct values were analyzed using the Wilcoxon signed-rank test. Temporal trends were evaluated according to clinical phases defined by time since symptom onset: acute phase (0–7 days) and early convalescent phase (8–14 days), and univariable logistic regression was used to assess the effect of time on detection probability. Urine samples showed a significantly higher positivity rate (75.6%, 31/41) than serum samples (43.9%, 18/41; McNemar p = 0.037), with poor agreement between matrices (Cohen’s kappa ≈ − 0.52). While serum Ct values increased over time), indicating declining detectability, urine Ct values remained showed no significant temporal trend throughout the two clinical phases. No significant association was observed between days since symptom onset and urine detection probability within 15 days. These findings suggest that urine may serve as a complementary specimen for molecular detection of Oropouche virus, particularly when serum testing occurs later in the course of infection.