AbstractPurpose To analyze the etiological distribution of long‐lasting and severe anterior uveitis based on the analysis of the aqueous humor.Methods Retrospective study of patients presenting with atypical and severe anterior uveitis and managed in a tertiary referral centre, between January 2008 and December 2009. Patients with panuveitis, intermediate and posterior uveitis and typical B27‐associated uveitis or herpes zoster ophthalmicus were excluded. Patients were divided into three groups : atypical viral uveitis (group 1), typical herpetic uveitis (group 2), Fuchs uveitis (group 3). An anterior chamber paracentesis was performed in all cases.Results The medical files of 153 patients were analysed. The mean age was 49 years (range 8‐83 Years), the M/F sex ratio was 84/69. Uveitis was unilateral in 86.5% of cases. Group 1 included 42 patients with sarcoidosis (13 cases), tuberculosis (7 cases), endophthalmitis (6 cases), spondylarthropathy (10 cases), syphilis (2 cases), neoplasia (2 cases) and multiple sclerosis (2 cases). Group 2 included 83 cases. A viral agent was definitely identified in 33 cases (39.8%). The distribution of herpes viruses was the following : CMV (19 cases), HSV‐1 (8 cases) and VZV (6 cases). The yield of anterior chamber paracentesis is increased by repetition of the procedure (up to 77.4% for CMV). The PCR‐negative patients in Group 2 responded well to a specific antiviral treatment. None of the patients with Fuchs uveitis (Group 3) had a positive PCR for CMV.Conclusion Most cases of atypical anterior uveitis, especially when unilateral, are viral‐induced. Molecular tools applied to ocular fluids may confirm a viral infection in atypical cases of anterior uveitis, leading to a specific antiviral therapy.