Abstract. Chikungunya virus (CHIKV) is an emerging alphaviral disease and a public health problem in South Asia including Nepal in recent years. In this study, sera were collected from patients presenting with fever, headache, muscular pain, fatigue, and joint pain of both upper and lower extremities. A total of 169 serum samples were tested for CHIKV and dengue virus (DENV) by using Immunoglobulin M (IgM) and Immunoglobulin G (IgG) antibody using enzymelinked immunosorbent assay (ELISA) method during August to November 2013. Results showed that 3.6% and 27.8% samples were positive for CHIKV and DENV IgM positive, respectively. Similarly, results of IgG showed 3.0% samples were positive for CHIKV IgG and 29.0% were for DENV IgG positive. Further, a 50% focal reduction neutralization test (FRNT 50 ) was performed to confirm the presence of CHIKV, which demonstrated that 8.9% of CHIKV IgM and/or IgG ELISA positive possessed neutralizing anti-CHIK antibodies. To our knowledge, this is the first report in which the presence of CHIKV is confirmed in Nepalese patients by FRNT 50 . Basic scientists and clinicians need to consider CHIKV as a differential diagnosis in febrile Nepalese patients, and policy makers should consider appropriate surveillance and actions for control strategies.Chikungunya virus (CHIKV) is a mosquito-borne febrile illness that is transmitted to humans through the bite of infected Aedes aegypti and Ae. albopictus mosquitoes.1 CHIKV belongs to the Alphavirus genus of the Togaviridae family whereas dengue viruses (DENVs) belong to genus Flavivirus of the family Flaviviridae. Both CHIKV and DENV are transmitted by the same mosquito vectors. The CHIKV was first isolated and characterized in humans and mosquitoes during an outbreak in Tanzania and Mozambique in 1955. 2 Afterwards, several outbreaks of CHIKV occurred and affected millions of people in eastern, southern, central Africa and Asia.3 CHIKV was introduced in the Americas in October 2013. As of April 2015, over 1,322,893 cases have been suspected and 30,309 confirmed to be CHIKV in the Americas. 4 It generally causes mild illness but sometimes can lead to severe and lifethreatening complications. The disease is characterized by an acute illness with fever, chills, headache, nausea, vomiting, joint pain with or without swelling, low back pain, and skin rash. DENV can progress to dengue hemorrhagic fever and dengue shock syndrome while CHIKV causes arthralgia, which may persist for months. 5 There is similarity in signs and symptoms of CHIK and DEN, which increases risks for misdiagnosis and underreporting of CHIKV infection in DEN-endemic areas. 5 The incubation period of CHIKV is usually 2-10 days, with constitutional symptoms lasting up to 7 days. The symptoms usually resolve within days to a few weeks; but in severe cases, these symptoms may last for months. Herein, we report a serological study of possible CHIKV infection with confirmation by 50% focal reduction neutralization test (FRNT 50 ) among febrile patients for the first time in N...