Background With declining vectorial transmission, Chagas disease predominantly affects adults nowadays. The efficacy of nifurtimox in the chronic phase in adult patients is poorly known, particularly in regions where there is no risk of reinfection. Recommendations for treatment outcome assessment rely on serological follow-up. We evaluated the serological and parasitological response to nifurtimox in a cohort of adult patients three years post-treatment in Switzerland. Methods Patients treated with nifurtimox in 2008 during a cross-sectional study in Geneva, Switzerland, were contacted for follow-up in 2011. Two ELISAs and a rapid immunochromatographic test were used to test 2008 and 2011 serum samples simultaneously. In addition, conventional and real-time PCR were performed on 2011 samples. Results Thirty-seven (84.1%) of 44 eligible patients, predominantly female, middle-aged, Bolivians at the indeterminate stage, were enrolled. All 2011 ELISA and immunochromatographic tests were positive. Twenty-eight (75.7%) patients presented a lower optical density (OD) in 2011 compared to 2008. This OD difference was significant in both commercial (P < 0.001) and in-house (P = 0.002) ELISAs. Agreement between the two ELISAs was low (Kappa = 0.469). All patients had negative conventional PCR results but one (2.7%) was positive with real-time PCR. Conclusion Our results highlight the inadequacy of serology for assessing response in adults, three years after treatment. In our cohort, 97.3% had results that could either indicate treatment failure or persistant humoral response despite treatment. The lack of accurate early post-treatment tests of cure prevents appropriate patients information and councelling. New follow-up tests are needed to assess treatments efficacy given the large adult population in need of antiparasitic therapy.