Complete genome sequence of highly virulent neurotropic Newcastle disease virus strain Texas GB

Newcastle disease virus (NDV) strain Texas GB is a highly virulent neurotropic virus that is used as a standard vaccine challenge virus in the U.S. In this study, the complete genome sequence of strain Texas GB was determined and compared with the complete genome sequences of other NDV strains. The genome is 15,186 nucleotides (nt) long and consists of six genes in the order of 3′leader-N-P-M-F-HN-L-5′trailer. The genome contains a 55-nt leader sequence at the 3′ end and a 114-nt trailer sequence at the 5′ end. The intergenic sequences are 2, 1, 1, 31, and 47 nt between N/P, P/M, M/F, F/HN, and HN/L genes, respectively. The putative cleavage site of fusion protein showed amino acid sequence of R-R-Q-K-R↓F in position 112 to 117, which corresponds to those of virulent NDV strains. The phylogenetic analysis showed that strain Texas GB is closely related to the neurovirulent mesogenic strain Beaudette C (BC) and to NDV viruses isolated in China and Egypt than to other strains of NDV.     

Complete genome sequence and molecular characterization of thermostable Newcastle disease virus strain TS09-C

A thermostable avirulent Newcastle disease virus (NDV) strain TS09-C was developed by serial passage of V4 strain in BHK-21 cells. The complete genome sequence of strain TS09-C was determined and compared with the sequences of other NDV isolates representing different thermostable phenotypes. The TS09-C genome was 15,186 nucleotides long and consisted of eight genes in the order of 3′-NP-P/V/W-M-F-HN-L-5′. High levels of nucleotide and amino acid sequence identity existed among the thermostable NDV isolates. Thermostable strains TS09-C, V4, I-2, and Ulster all belonged to genotype I. The F protein cleavage site of strain TS09-C was ¹¹²G-K-Q-R–R-L¹¹⁷, with an isolated basic amino acid and a pair of contiguous basic amino acids, differing from that of its parental V4 strain (¹¹²G-K-Q-G-R-L¹¹⁷). Our characterization of the complete genome of thermostable avirulent NDV strain TS09-C may facilitate the development of a thermostable NDV reverse genetic system and further study of the mechanism of thermostability of NDV.     

Characterization of a recombinant Newcastle disease virus vaccine strain

A recombinant La Sota strain (KBNP-C4152R2L) in which fusion (F) and hemagglutinin-neuraminidase (HN) genes were replaced with those of a contemporary genotype VIId virus, KBNP-4152, has been developed. To attenuate the virulence of the recombinant strain, the F cleavage motif was mutated from ₁₁₂RRQKR₁₁₆ to ₁₁₂GRQAR₁₁₆, and to reduce pathogenic instability, a codon which does not allow changes to basic amino acids by single point mutation was inserted at codon 115. In addition a six-nucleotide sequence was inserted into the intergenic region between matrix protein and F genes for attenuation without breaking the “rule-of-six.” The HN protein length was increased from 571 to 577 as a marker. Serological tests revealed that the antigenicity of KBNP-C4152R2L was similar to that of KBNP-4152 but distinct from that of the La Sota strain. KBNP-C4152R2L was avirulent (intracerebral pathogenicity index, 0.0; mean death time, >168 h) and stable in pathogenicity through in vivo passages. The killed oil emulsion of and live KBNP-C4152R2L were completely protective against mortality and egg drop caused by virulent strains, and KBNP-C4152R2L was applicable to in ovo vaccination. Therefore, KBNP-C4152R2L is a promising vaccine strain and viral vector in terms of antigenicity, productivity, safety, and pathogenic stability.     

Characterization of complete genome sequence of genotype VI and VII velogenic Newcastle disease virus from Japan

The complete genome sequences of three strains of Newcastle disease virus (NDV) isolated from vaccinated commercial layer flocks in Japan in the span of three decades were characterized. All strains had genome lengths of 15,192 nucleotides consisting of six genes in the order of 3′-NP–P/V/W–M–F–HN–L-5′. The general genomic characteristics of the Japanese field strains were consistent with previously characterized class II NDV, except for those belonging to early genotypes (genotype I–IV), which lack the six nucleotide insertion at nucleotide positions 1,648–1,653 of the nucleoprotein (NP) gene. Phylogenetic analysis showed that the Japanese strains could be classified into genotypes VIc and VIIe using the complete genome sequence and the complete coding sequence of the fusion (F) gene according to the unified NDV classification system. Characterization of functional domains and neutralizing epitopes of the F and hemagglutinin-neuraminidase (HN) proteins of Japanese field strains revealed a total of 31 amino acid substitutions, as compared to vaccine strains Ishii and B1, which were widely used in Japan. Although virus neutralization (VN) test showed that poor flock immunity due to vaccination failure or partial and non-uniform immunization maybe the major factors involved in the mechanism of breakthrough infection of the Japanese field strains, approximately two to threefold decrease in the VN titers of the field NDV strains possessing a point mutation (E347K or E347G) at the linear epitope of the HN protein was observed, as compared to vaccine strain B1 and field strain 2440/69, which lack the point mutation. This study may be a useful reference in characterizing future ND outbreaks in vaccinated chickens and as a genetic map for future investigations regarding vaccine designs, reverse genetics systems, and development of molecular diagnostic tools to prevent future ND outbreaks in vaccinated poultry flocks.     

Complete genome sequence of a velogenic Newcastle disease virus isolated in Mexico

In Mexico, the number of cases of the highly virulent Newcastle disease virus is increasing. In 2005, an outbreak of Newcastle disease occurred on an egg laying hen farm in the state of Puebla despite vaccination with the LaSota strain. Farmers experienced a major drop in egg production as a consequence of a field challenge virus. In this study, we characterize the virus, APMV1/chicken/Mexico/P05/2005, responsible for the outbreak. The virus is categorized as a velogenic virus with an intracranial pathogenicity index of 1.99 and a chicken embryo mean death time of 36?h. The complete genome length of the virus was sequenced as consisting of 15,192?bp. In addition, phylogenetic analysis classified the virus as a member of the class II, genotype V. The highly pathogenic nature of the virus has been linked to the amino acid sequence at the fusion protein cleavage site, which contains multiple basic amino acids (RRQKR↓F).     

Complete genome sequence of a virulent Newcastle disease virus isolated from an outbreak in chickens in Egypt

The complete genome sequence of a virulent Newcastle disease virus (NDV) isolated from chickens in Egypt was determined and compared to the sequence of NDV strains isolated from different parts of the world. The genome is 15,186 nucleotides (nt) long and consists of 6 genes in the order of 3′-N-P-M-F-HN-L-5′. The genome contains a 55-nt leader region at the 3′ end and a 114-nt trailer region at the 5′ end. Interestingly, the phylogenetic analysis showed that strain Egypt is closely related with the NDV strains isolated in China. In addition, the sequence of the fusion protein cleavage site of strain Egypt was identical to that of the NDV strain recently isolated in Mali. Determination of complete genome sequences of additional NDV strains from Africa is necessary to understand the epidemiology of currently circulating viruses in Africa.     

Sequence variation in the Newcastle disease virus genome

Full-length genome sequences of five virulent and five avirulent strains of Newcastle disease virus isolated between 1998 and 2002 in Victoria and New South Wales, Australia were determined. Comparisons between these strains revealed that coding sequence variability in the haemagglutinin-neuraminidase (HN), matrix (M) and phosphoprotein (P) gene sequences appeared to be more variable than in the fusion (F), nucleocapsid (N) and RNA dependent-RNA replicase (L) genes. Sequence analysis of a number of other isolates made during the recent virulent NDV outbreaks, also identified the presence of a number of variants with altered F gene cleavage sites, which resulted in altered biological properties of those viruses. Quasispecies analysis of a number of field isolates indicated the presence of virulent virus in one particular isolate. Gene sequence analysis of the progenitor virus isolated in 1998 showed very little sequence variation when compared to that of a progenitor-like virus isolated in 2001, demonstrating that in the field, viral genome sequence variation appears to be biologically restricted to that of a consensus sequence.     

Genomic sequence of an antigenic variant Newcastle disease virus isolated in Korea

In Korea, extensive Newcastle disease (ND) vaccine programs have been implemented, but ND outbreaks continue to occur occasionally, even in well-vaccinated farms. KBNP-4152 is a virulent ND virus, which has been isolated from vaccinated chickens in Korea. In this study, we conducted a comparison of the antigenicity of KBNP-4152 with that of a vaccine strain, La Sota, via virus-neutralization (VN) and cross haemagglutination-inhibition (HI) tests, and analyzed the genomic sequences. The antigenicity of KBNP-4152 was distinct from La Sota, and the expected genome size was 15,192 nt, as was the case with other recent virulent ND viruses analyzed. Based on the partial F gene, the strain was phylogenetically classified into the VIId genotype, but was distinct from other VII viruses due to amino acid changes at (E347K) and proximal to (M354K), the major linear epitope of HN, as well as relatively low amino acid similarity of the V protein, and a truncated W protein (203 aa vs. 227 aa). Therefore, KBNP-4152 is unique among genotype VII.     

Nucleotide sequence of the envelope protein genes of a highly virulent, neurotropic strain of Newcastle disease virus

The envelope glycoproteins of Newcastle disease virus (NDV), hemagglutinin-neuraminidase (HN) and fusion (F) proteins, play important roles in determining the host immune response and the virulence of that particular virus strain. The complete nucleotide sequence of the HN and F genes of a highly neurovirulent strain of NDV (Texas G. B., 1948) was determined in an effort to study the molecular basis of this strain's neurotropic properties. Comparison of the predicted amino acid sequences for the HN and F among the American NDV strains revealed that the Texas G. B. and Beaudette C envelope genes are closely related to each other and are less closely related to the avirulent B1 Hitchner strain. We have found 11 amino acid changes in the predicted HN protein between the Beaudette C and Texas G. B. strain but only 2 conservative amino acid changes (amino acids 11 and 197) in the F protein between these two strains. Although the virulence of NDV strains has been related to sequences at the cleavage site of F0, the property of neurovirulence cannot depend solely upon these sequences because there are no sequence differences between the Beaudette C and Texas G. B. strains. We suggest that the neurovirulence phenotype could be due to the molecular properties of the HN protein; however, we cannot exclude the possibility that the two conservative amino acid differences between the two F proteins could also play a role in determining the phenotypic differences between these two virus strains.     

Haemagglutinating activity of the lentogenic Newcastle disease virus strain MET95

Using the rapid glass plate method, the Newcastle disease virus strain MET95 showed much weaker haemagglutination (HA) activity for chicken erythrocytes than 69 other Newcastle disease viruses, including 56 field strains isolated from chickens reared in Japan between 1988 and 2001. Using erythrocytes from other avian species, only the MET95 strain failed to show HA activity for erythrocytes from ducks, geese or pigeons. The haemagglutinin-neuraminidase protein of the MET95 strain was shown to have unique substitutions of isoleucine for thereonine and leucine at amino acide residues 216 and 552. It is suggested that these two substitutions might relate to the unique HA activity of the MET95 strain. This HA activity may be a useful marker for this strain.     

The complete genome sequence of the Tanzanian strain of Cassava brown streak virus and comparison with the Ugandan strain sequence

The complete genome sequence for an isolate of the Ugandan and Tanzanian strain types of Cassava brown streak virus have been determined using the novel approach of non-directed next generation sequencing. Comparison of the genome sequences revealed that CBSV is highly heterogeneous at the isolate level as well as the strain level. The isolate of the Ugandan strain was found to have a genome 9,070 nucleotides long coding for a polypeptide with 2,902 amino acid residues. The isolate of the Tanzanian strain was 9,008 nucleotides long and coded for a polypeptide with 2,916 amino acid residues. Nucleotide identity between the isolates across the genome was 76%, with protein encoding regions 57–77% and individual proteins had 65–91% amino acid similarity. In addition between the two strains four protein products (PIPO, CI, NIa-Vpg and coat protein) varied in size and an unusual HAM1-like protein, whilst of identical nucleotide length, was found to have the lowest homology. The implication of diversity of CBSV is discussed in the context of speciation, evolution, development of diagnostics, and breeding for resistance.     

Complete genome characterisation of a Newcastle disease virus isolated during an outbreak in Sweden in 1997

The complete genome sequence of a Newcastle disease virus (NDV) isolated from a chicken in Sweden was determined and compared with other NDV sequences. The isolate was shown to belong to genotype VIIb, which arose in the Far East and spread around the world during the 1990s. It had a length of 15,192 bases and consisted of six genes in the order 3′-NP-P-M-F-HN-L-5′. The F protein cleavage site was 112-RRQRRF-117, corresponding to that of a virulent pathotype.     

Some properties of an avirulent Newcastle disease virus

Summary Newcastle disease virus was isolated from healthy birds using chick kidney cultures. The virus strain involved was exceptionally attenuated in character and appeared to be transmitted by the enteric route. It caused no clinical illness even in day old chicks and did not regularly kill embryonated eggs. The virus is relatively thermostable and its haemagglutinin is exceptionally stable at 56 C. This easily observed property may be useful in genetic studies of the virus.     

Immunization by aerosol with La Sota strain of Newcastle disease virus

The respiratory symptoms observed after aerosol vaccination with the La Sota strain of Newcastle disease virus were eliminated by mixing the anti-inflammatory compound benzydamine with the vaccine prior to aerosol administration. Benzydamine had no effect on the vaccine virus or on the development of haemagglutination-inhibiting antibodies.     

The genome nucleotide sequence of a contemporary wild strain of measles virus and its comparison with the classical Edmonston strain genome

The only complete genome nucleotide sequences of measles virus (MeV) reported to date have been for the Edmonston (Ed) strain and derivatives, which were isolated decades ago, passaged extensively under laboratory conditions, and appeared to be nonpathogenic. Partial sequencing of many other strains has identified >/=15 genotypes. Most recent isolates, including those typically pathogenic, belong to genotypes distinct from the Edmonston type. Therefore, the sequence of Ed and related strains may not be representative of those of pathological measles circulating at that or any time in human populations. Taking into account these issues as well as the fact that so many studies have been based upon Ed-related strains, we have sequenced the entire genome of a recently isolated pathogenic strain, 9301B. Between this recent isolate and the classical Ed strain, there were 465 nucleotide differences (2.93%) and 114 amino acid differences (2.19%). Computation of nonsynonymous and synonymous substitutions in open reading frames as well as direct comparisons of noncoding regions of each gene and extracistronic regulatory regions clearly revealed the regions where changes have been permissible and nonpermissible. Notably, considerable nonsynonymous substitutions appeared to be permissible for the P frame to maintain a high degree of sequence conservation for the overlapping C frame. However, the cause and the effect were largely unclear for any substitution, indicating that there is a considerable gap between the two strains that cannot be filled. The sequence reported here would be useful as a reference of contemporary wild-type MeV.