Virulence markers to distinguish high from low virulence bovine viral diarrhea virus genotype 2 isolates have not been previously reported. The objective of this study was to identify virulence markers by evaluating the primary and secondary structures of the 5'-untranslated region of low and high virulence bovine viral diarrhea virus genotype 2 isolates. The nucleotide sequences of the entire 5'-untranslated region mRNA of eight bovine viral diarrhea virus genotype 2 isolates, four of high virulence and four of low virulence, and two genotype 1 reference isolates were determined using a polymerase chain reaction and a 5' Rapid Amplification of cDNA Ends System. Two nucleotide substitutions were identified in the internal ribosomal entry site that distinguished the high virulence from the low virulence genotype 2 isolates. The low virulence isolates had a cytosine at position 219, whereas the high virulence isolates had a uracil. At position 278, a uracil or cytosine was found in the low and high virulence groups, respectively. The substituted bases are virulence markers that were used to identify bovine viral diarrhea virus genotype 2 isolates of high virulence.
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