Purified RNA from five lentogenic Newcastle disease virus (NDV) strains (B1, England-F, Nebraska, Queensland V4, and Ulster) was subjected to oligonucleotide fingerprinting analysis. This technique could resolve over 100 oligonucleotides per virus strain, of which 50 to 80 were unique. Although the fingerprints generated from these viral RNAs exhibited similar distribution patterns of non-unique RNA fragments, all could be distinguished from one another on the basis of the migration of unique RNA fragments. It is concluded that oligonucleotide mapping is a useful method of unambiguously identifying individual lentogenic strains of NDV and that this technique may serve as a means of characterizing highly pathogenic NDV isolates.