Background Chikungunya is a widely distributed, re-emerging tropical disease caused by the chikungunya virus (CHIKV). Little is known about the duration for which CHIK RNA are detectable in bodily fluids, especially genital secretions, and current evidence is based on small series or case reports. An understanding of viral dynamics across different body compartments can inform diagnostic testing algorithms and public health prevention interventions. Methodology A prospective cohort study was conducted to assess the presence and duration of detectable levels of CHIKV RNA in blood, urine, saliva, semen, and vaginal secretions. Men and women (� 18 years) with a positive reverse transcriptase-polymerase chain reaction (RT-PCR) test for CHIKV in the acute phase (1–14 days) of the disease were included. After enrollment, clinical data and samples were collected every 15 days over the first 2 months, and a final collection was performed 3 months after recruitment. The Kaplan–Meier interval-censoring method and the parametric Weibull model were fitted to estimate the median time of viral persistence until the lack of CHIKV RNA detection among all body fluids. Punctual estimates of the median time of CHIKV RNA persistence for each fluid were estimated using a 95% confidence interval (CI). Results From April to December 2019, 170 participants were screened. Of these, 152 (100 women) were enrolled in the study. The median and interquartile range (IQR) ages for men and women were 39.3 (IQR: 26.9, 50.7) and 43.5 (IQR: 33.8, 53.6) years, respectively. CHIKV RNA was detected in 80.3% (122/152) of serum samples, 23.0% (35/152) of urine samples, 30.3% (46/152) of saliva samples, 14.3% (6/42) of semen samples, and 20.2% (20/99) of vaginal secretion samples. The median time until the loss of CHIKV RNA detection was 19.6 days (95% CI, 17.5–21.7) in serum, 25.3 days (95% CI, 17.8–32.8) in urine, 23.1 days (95% CI, 17.9–28.4) in saliva, and 25.8 days (95% CI, 20.6–31.1) in vaginal secretion. The number of semen samples available was too small to make statistical estimates, but a last positive sample was obtained from a participant 56 days after the onset of symptoms. Conclusions CHIKV RNA could be detected in all bodily fluids studied, including genital secretions during the acute and convalescent phases and additional studies on viral infectivity in semen and vaginal secretions are warranted.