In 2008 alone, 85,400 cases of acute flaccid paralysis (AFP) were reported from the countries with active poliovirus surveillance programs, of which less than 1,731 (2.03%) were found to be associated with poliovirus infection. Using a metagenomic approach we have discovered a group of highly divergent picornaviruses in children suffering from non-polio AFP. The complete genomes of six new viruses were sequenced and phylogenetic analysis indicated them to be sufficiently divergent from all currently known picornaviruses to form a new genus in the Picornaviridae family. In order to begin testing whether these new viruses, named Human Cosavirus (HCoSV), were a significant pathogen associated with non-polio AFP we tested stool samples from 160 non-polio AFP cases using specific PCR primers. HCoSV RNA was detected in stool samples of 107 AFP cases. High prevalence and high viral load of HCoSV variants in stool samples of multiple AFP children leads us to hypothesize that HCoSV is a human pathogen associated with AFP in children. Initial phylogenetic analysis of HCoSV variants showed the existence of four different viral species within this new Picornaviridae genus. The level of genetic divergence between species of genus Cosavirus was similar to that between species of genus Enterovirus. Further information regarding HCoSV genetic diversity and epidemiology is now required to clarify its pathogenic role in AFP and, if confirmed, for developing strategies to prevent its transmission and disease. AIM 1 will determine the prevalence of different HCoSV genotypes in samples from a larger number of AFP cases plus demographically matched healthy controls to test whether there is a significant association with AFP, followed by full genome genetic characterization of the most divergent HCoSV variants. AIM 2 will be the development of a serological assay for HCoSV using virions produced by cell culture or viral capsids derived from baculovirus/insect cell expression. AIM 3 will determine the prevalence of seroreactivity to HCoSV in AFP cases, in healthy controls and in the general population. This exploratory study of a new human picornavirus genus that we have recently characterized and detected at high frequency in developing countries will therefore provide the foundation for possible steps to relieve the still high health burden imposed by infantile paralytic diseases. Our long-term goal is to define the pathogenic potential of HCoSV and to prevent new infections.