Degenerate oligonucleotide primers were synthesized to amplify nucleotide sequences from portions of the fusion protein and matrix protein genes of Newcastle disease virus (NDV) genomic RNA that could be used diagnostically. These primers were used in a single-tube reverse transcription PCR of NDV genomic RNA coupled to direct nucleotide sequencing of the amplified product to characterize more than 30 NDV isolates. In agreement with previous reports, differences in the fusion protein cleavage sequence that correlated genotypically with virulence among various NDV pathotypes were detected. By using sequences generated from the matrix protein gene coding for the nuclear localization signal, lentogenic viruses were again grouped phylogenetically separate from other pathotypes. These techniques were applied to compare neurotropic velogenic viruses isolated from an outbreak of Newcastle disease in cormorants and turkeys. Cormorant NDV isolates and an NDV isolate from an infected turkey flock in North Dakota had the fusion protein cleavage sequence 109 SRGRRQKRFVG 119. The R-for-G substitution at position 110 may be unique for the cormoranttype isolates. Although the amino acid sequences from the fusion protein cleavage site were identical, nucleotide sequence data correlate the outbreak in turkeys to a cormorant virus isolate from Minnesota and not to a cormorant virus isolate from Michigan. On the basis of sequence information, the cormorant isolates are virulent viruses related to isolates of psittacine origin, possibly genotypically distinct from other velogenic NDV isolates. These techniques can be used reliably for Newcastle disease epidemiology and for prediction of pathotypes of NDV isolates without traditional live-bird inoculations.

Nine Newcastle disease virus (NDV) isolates obtained from Puerto Rico, Georgia, Alabama, Mississippi, and Texas were analyzed for in vivo pathogenicity, biological properties (hemagglutination of mammalian erythrocytes), and for sequence variation at the amino acid and sense RNA level of the fusion protein cleavage site. Intracerebral pathogenicity index values ranged from 0 to 0.3 and the intravenous pathogenicity index obtained for all isolates was 0. Four isolates hemagglutinated bovine erythrocytes, whereas no hemagglutination was observed using equine erythrocytes. By direct sequencing of reverse transcription polymerase chain reaction products, all the isolates had a predicted fusion cleavage sequence comparable to lentogenic NDV strains. Based on nucleotide sequence, the viruses could be grouped phylogenetically with the B1 vaccine-type virus. However, nucleotide sequences were not 100% similar to the B1 or La Sota NDV strains, indicating that minor genetic heterogeneity occurs among lentogenic field isolates of NDV.

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The infectious bronchitis virus (IBV) spike glycoprotein S1 subunit is required to initiate infection and contains virus-neutralizing and serotype-specific epitope(s). Reported are the S1 gene nucleotide and predicted amino acid sequences for the Florida 18288 strain and isolates GA-92, CV-56b, CV-9437, CV-1686, and 1013. These sequences were compared with previously published gene sequences of IBV strains, and phylogenetic relationships are reported. The S1 amino acid sequence of Florida 18288 was 94.9% similar to the Connecticut strain, and GA-92 was 92.8% similar to the Arkansas 99 strain. S1 amino acid sequences of the California variants, CV-56b, CV-9437, and CV-1686, were 97.6-99.3% similar to one another and only 76.6%-76.8% similar to the Arkansas-type strains. Isolate 1013, also from California, was 84.0% similar to Ark DPI and 77.9% similar to CV-56b. When comparing 19 viruses isolated from the United States, sequence variations were observed between amino acids 55-96, 115-149, 255-309, and 378-395. Similar regions are reported to be involved in virus-neutralizing and/or serotype-specific epitopes. These data demonstrate that variant IBV strains continue to emerge, and unique variants may circulate among poultry in geographically isolated areas.