Avian nephritis virus (ANV), which belongs to the Astroviridae family, has been associated with acute nephritis in chickens. Cases of ANV infection have been recorded in Japan and in several European countries. However, related studies have never been performed in China. Thus, this study isolated ANV in Chinese chicken flocks. ANV RNA was detected by reverse transcription–PCR in stool samples collected from healthy layer chickens in the Sichuan Province of China in 2009. Of the 192 stool specimens collected, 32.3% (62/192) were positive for ANV infection. The whole genome of ANV-Sichuan54, the first representative Chinese strain, was 6941 nucleotides in length, including the 5′ untranslated region, three open reading frames (ORFs), a 3′ UTR, and a poly-(A) tail. Comparative and phylogenetic analyses based on partial RNA-dependent RNA polymerase (ORF1b) demonstrated that the majority of ANV investigations were more closely related to the U.S. ANV strain (DQ324827–324836) than to the G-4260 (AB033998).