Feline coronavirus (FCoV) is endemic in feral cat populations and cat colonies, frequently preceding outbreaks of fatal feline infectious peritonitis (FIP). FCoV exhibits 2 biotypes: the pathogenic disease and a benign infection with feline enteric coronavirus (FECV). Uncertainty remains regarding whether genetically distinctive avirulent and virulent forms coexist or whether an avirulent form mutates in vivo, causing FIP. To resolve these alternative hypotheses, we isolated viral sequences from FCoV-infected clinically healthy and sick cats (8 FIP cases and 48 FECV-asymptomatic animals); 735 sequences from 4 gene segments were generated and subjected to phylogenetic analyses. Viral sequences from healthy cats were distinct from sick cats on the basis of genetic distances observed in the membrane and nonstructural protein 7b genes. These data demonstrate distinctive circulating virulent and avirulent strains in natural populations. In addition, 5 membrane protein amino acid residues with functional potential differentiated healthy cats from cats with FIP. These findings may have potential as diagnostic markers for virulent FIP-associated FCoV.