A novel picornavirus genome was sequenced, showing 42.6%, 35.2%, and 44.6% of deduced amino acid identities corresponding to the P1, P2, and P3 regions, respectively, of the Aichi virus. Divergent strains of this new virus, which we named salivirus, were detected in 18 stool samples from Nigeria, Tunisia, Nepal, and the United States. A statistical association was seen between virus shedding and unexplained cases of gastroenteritis in Nepal (P ؍ 0.0056). Viruses with approximately 90% nucleotide similarity, named klassevirus, were also recently reported in three cases of unexplained diarrhea from the United States and Australia and in sewage from Spain, reflecting a global distribution and supporting a pathogenic role for this new group of picornaviruses.The falling cost of DNA sequencing has led to a recent surge in human and animal virus discoveries (1-3, 5-12, 14, 16-17, 19-24, 27, 30, 31, 33, 39, 43, 44). While the pathogenicity of some newly characterized human viruses has been demonstrated, it remains unknown or controversial for other viruses, which may be commensal or pathogenic in only a very small fraction of infections (25,32,40,42,45). Genetic characterization of previously unknown viruses allows the rapid design of nucleic acid tests needed to determine their association with different medical conditions, their presence in different populations, and the design of antibody tests for determining seroprevalence (25, 28, 34, 35, 47).Using sequence-independent PCR amplification, pyrosequencing, and sequence similarity searches (46) (see the text in the supplemental material), we analyzed the virus sequences present in 95 stool samples from Nigerian children suffering from nonpolio acute flaccid paralysis (AFP). Sequences derived from a 10-month-old female child exhibiting right-side asymmetric sudden flaccid paralysis (patient no. NG-J1) formed a 6,981-bp contig consisting of 2,903 individual sequence reads, which was distantly related to sequence of the Aichi virus species in the Kobuvirus genus of the Picornaviridae family (48, 49). Similar sequences were also observed in a second, 24-month-old patient with right-side asymmetric sudden flaccid paralysis (patient no. NG-F1). Gaps between sequenced viral fragments were connected by nested reverse transcription-PCR (RT-PCR), while the 5Ј and 3Ј extremity sequences were acquired using primers designed over conserved regions of bovine, porcine, and human kobuviruses. We temporarily named these viruses saliviruses (stool Aichi-like viruses).The resulting salivirus genome, NG-J1, was 7,124 bp in length with a GC content of 57%, excluding a poly(A) tail. NG-J1 contained a large open reading frame of 7,125 bp encoding a putative polyprotein precursor of 2,374 amino acids (aa), a 5Ј untranslated region (UTR) of 709 bp, and a 3ЈUTR of 148 bp (Fig. 1).NG-J1 and NG-F1 were highly similar, with nucleotide similarities of 94% and 95% in the P1 and P3 regions, respectively. Salivirus NG-J1 had approximately 90% nucleotide similarity to the recently described klasse...