The genetic variation and evolutionary relationships amongst the five serotypes of bluetongue virus (BTV) endemic to the United States were investigated by oligonucleotide fingerprint analysis. The viruses analyzed include prototype viruses of the five U.S. serotypes, and 32 viruses isolated from domestic and wild ruminants from the U.S. in the years 1979-1981. With the exception of serotype 2, most genes encoding the viral core and non-structural proteins were demonstrated to be highly conserved both within and between serotypes and some also appear to have reassorted in nature. Gene segments 2 and 6, which encode the outer capsid proteins VP2 and VP5 respectively, were more variable and were not consistently linked as serotype determination was dependent solely on gene segment 2. Gene segment 2 was the most variable gene between serotypes, but it was highly conserved within serotypes and stable over time. This suggests that the emergence of new BTV serotypes, which would require the stable incorporation of numerous mutations, must be a very slow process. Fingerprint comparisons further suggested that BTV serotypes 10, 11, 13 and 17 have evolved together in the U.S. over a considerable period of time, whereas serotype 2, which is genetically distinct, has evolved elsewhere and is most likely a recent introduction to North America.