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Monoclonal antibodies (MAbs) to three overlapping antigenic sites (designated 12, 2, and 23) on the hemagglutinin-neuraminidase glycoprotein (HN) of Newcastle disease virus (NDV) were previously shown to inhibit neuraminidase activity (NA) on neuraminlactose (R. M. Iorio and M. A. Bratt, 1984a, J. Immunol. 133, 2215-2219; R. M. Iorio et al., 1989, Virus Res. 13, 245-262). However, a competitive inhibitor of NA blocks the binding of only MAbs to site 23, suggesting that the domain they recognize may be closely related to the NA site. Antigenic variants selected with site 23 MAbs have single amino acid substitutions at HN residues 192, 193, or 200. Virions of variants, which have a substitution at residue 193 or 200, have alterations in NA which are not attributable to a commensurate change in HN content. A revertant of a temperature-sensitive mutant, which has markedly diminished NA relative to the wild type, has an amino acid substitution at residue 175. A second step revertant having partially restored NA has an additional substitution at residue 192 identical to that in one of the site 23 variants, which, in turn, also makes the revertant resistant to neutralization by site 23 MAbs. Thus, an amino acid substitution at residue 175, 193, or 200 of the HN of NDV can have marked effects on the NA of the protein. The amino acids in the region around residue 175 are highly conserved between the HNs of NDV and other paramyxoviruses, suggesting that this domain is important to the integrity of the NA site in this group of viruses.
The relationship between phagocytic activity of peritoneal macrophages and serum interferon (IF) titers was evaluated in mice challenged with Newcastle disease virus (NDV). Time course studies indicated peak serum IF titers between 6 and 12 hr, whereas Fc receptor-mediated macrophage phagocytosis was maximal 18 hr after viral administration. Both responses decreased in parallel as the inoculated dose of the virus was reduced. Splenectomy, shown by others to decrease the NDV-induced serum IF titers, significantly decreased the stimulation of phagocytosis. The role of T cells in the response to the virus was studied with nude mice raised under germfree conditions. NDV-induced serum IF titers and macrophage phagocytosis were both diminished in BALB/c nudes compared with their heterozygous littermates. Both responses could be partially restored by transfer of thymocytes obtained from heterozygous mice. The results provide further evidence that in vivo macrophage stimulation by NDV is mediated by induced IF. The experiments with nude mice also indicate that the IF response to NDV is regulated by T lymphocytes.
The main function region of the fusion (F) protein gene of 124 strains of Newcastle disease virus isolated from 2002 to 2004 in China was amplified and sequenced for further phylogenetic and residue substitutive analysis. Most of the isolates were classified into genotype VIIc, VIId, VIf, and VIb, while others into genotype IX, III, or II. The genotype IX, a unique genotype which includes strain F48, the first Chinese virulent NDV strain isolated in 1948, were still found inducing sporadic infections in certain areas. Subgenotype VIIc, VIId, and VIIe viruses, which were distributed in clusters in the phylogenetic tree distinct from members of subgenotypes VIIa and VIIb, were responsible for most outbreaks in China and circulated predominantly in China in recent years. Strain NDV03-026, an isolate of the genotype II which was normally lentogenic, was found carrying (112)RRQKRF(117) motif at the cleavage site of F protein as the virulent strain.
A recombinant baculovirus expressing the nucleocapsid gene (NP) of Newcastle disease virus (NDV), a member of the genus Rubulavirus, has been generated and shown to express the native protein to high levels in insect cells. In contrast to the NP protein of the rubulavirus human parainfluenza virus 2, the NDV protein has been demonstrated by electron microscopy and caesium chloride gradient analysis to be capable of self-assembly in vivo to form nucleocapsid-like structures in the absence of other NDV proteins. These structures, which contained RNA that was resistant to micrococcal nuclease digestion, were also observed when the protein was expressed in E. coli, a phenomenon which was not inhibited by the presence of a 40 amino acid fusion region at the amino terminus of the protein. Further, the formation of these structures was inhibited by the co-expression of the phosphoprotein (P). Therefore, we conclude that the P protein acts as a chaperone, preventing uncontrolled encapsidation of non-viral RNA by NP protein.
Using primary chicken kidney (PCK) cells, a plaque assay was carried out with an avirulent strain of Newcastle disease virus (NDV) without adding trypsin to the agar overlay medium. The plaque assay was done in less than 4 days in PCK cells, by 3 days earlier than in primary chicken embryo (CE) cells maintained in the presence of trypsin. The curves of the progeny virus production began to rise 6 h after the infection and reached a plateau at 12 h. Equal amounts of virus were produced in PCK cells both in the presence and absence of trypsin in the culture medium. Viral peptide analysis revealed that a large portion of the HN and F precursor envelope glycoproteins of the NDV-Ulster strain remained uncleaved in PCK-grown virions. This suggests that a marginal proteolytic cleavage of these glycoprotein suffices for the full growth of the progeny virus in PCK cells. A higher shut-off in the host protein synthesis occurred in the virus-infected PCK cells than in the infected CE cells.
Twenty-nine strains of Newcastle disease virus (NDV) isolated from outbreaks in chicken and goose flocks in several regions of China during 1985-2001 were characterized pathotypically and genotypically. All except one of these strains were velogenic. For genotyping, phylogenetic tree analysis (nt 47-420), restriction site mapping (nt 334-1682) and residue substitution analysis (residues 4-124) of the F gene were performed using sequences of our isolates and sequences of selected NDV strains from GenBank. The results revealed that most of these newly characterized strains belonged to six novel genetic groups that were designated as VIf, VIg, VIIc, VIId, VIIe and IX. The genotype IX viruses, to which the China challenge strain F48E8 used for vaccine evaluation belonged, were found only in China and still induced sporadic infections in certain areas. Isolates belonging to group VIf and VIg were distinct from previously reported members of genotype VI, with genetic distance from 2.5 to 12.1%. Subgenotype VIIc, VIId and VIIe viruses, which were distributed in clusters in the phylogenetic tree distinct from members of subgenotypes VIIa and VIIb, were responsible for disease outbreaks in chicken and goose flocks and circulated predominantly in southern China in recent years. Finally, cross-protective testing showed that specific-pathogen free (SPF) chickens vaccinated with La Sota vaccines can be fully protected against challenge by strains from genetic groups VIb, VIg, VIId and IX, indicating that the antigenic differences between strains of various genotypes are insufficient to change the cross-protection conferred by the commonly used vaccine.
This study was designed to examine whether vaccine or virulent strains of Newcastle disease virus (NDV) would potentiate the disease caused by Pasteurella anatipestifer infection in turkeys. The studies were conducted in turkeys of two age groups. There were three experiments. In two experiments four-week-old turkeys were exposed either to vaccine or virulent strains of NDV after experimental P anatipestifer infection. In the third experiment 14-week-old turkeys were first exposed to virulent NDV and superimposed with P anatipestifer infection. In experiment 1, one bird died where P anatipestifer was given in combination with the vaccine strain of NDV. However, there was no difference in the clinical signs, gross lesions and histopathology compared with turkeys given P anatipestifer alone. In experiment 3 where turkeys received a virulent strain of NDV in combination with P anatipestifer, birds became dyspnoeic and showed signs of illness. There was a difference in the course of the disease, gross lesions and histopathology when compared with turkeys that received P anatipestifer only.
The feasibility of using fragments of chorioallantois attached to egg shell (C-S) as an alternative to embryonated eggs for isolating Newcastle disease virus (NDV) was assessed. In paired titrations of 6 strains of virus, embryonated eggs detected an average of 6-to-7-fold greater amounts of virus than did C-S fragments. The efficiency of isolation of viscerotropic velogenic NDV from 240 swab samples was 97% in embryonated eggs and 84% in C-S fragments. Sensitivity differences between the 2 host systems were revealed only in tests of samples expected to contain small amounts of virus. These results suggest that C-S fragments can be reliably used in some but not all instances, thereby relieving logistic constraints sometimes associated with the use of embryonated eggs.
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A pseudo-crystalline array of subunits has been observed on particles of the La Sota, in contrast to the Ulster , strain of Newcastle disease virus (NDV) grown in MDBK tissue culture without trypsin. This regular arrangement of subunits was associated with the semi-permissive nature of the tissue culture system, as it disappeared when trypsin, which allows infectious virus to be made, was added. The phenomenon described was considered to be related to the crystalline array of matrix protein which has been described inside the envelope of Sendai virus and NDV by others.
Surgical removal of the bursa of Fabricius from newly hatched chicks resulted in a depletion of immunoglobulin A (IgA) from serum and bile of 55 and 67% of the birds, respectively, up to 11 weeks of age. The occurrence of IgG and IgM in serum and IgG in bile was not affected by neonatal bursectomy (Bx). A slight compensatory increase in the occurrence of IgM in bile was noted in Bx birds. When exposed to a lentogenic strain of Newcastle disease virus by either intramuscular or intratracheal routes, the Bx group produced a level of serum hemagglutinating antibody and tracheal wash neutralizing antibody indistinguishable from that reached in the sham-Bx group. All of the vaccinated Bx birds challenged by intratracheal exposure to a velogenic strain of Newcastle disease virus remained healthy. These results suggest that IgA is not essential for the development of immunity in the chicken and that other locally produced immunoglobulins or transuded serum antibody may protect the tracheal mucosa in the absence of IgA.
Several immunological traits were compared in lines of Japanese quail selected for high and low secondary immune responses to anti-Newcastle disease virus (NDV). The high line of quail, selected for high ability to produce antibodies, was more responsive in both their primary response to NDV antigen used in the selection as well as to other antigens used in selection process including NDV (Ishi strain), tissue culture-derived Newcastle disease (TCND strain), NDV strain B1, influenza virus, sheep erythrocytes, and Salmonella pullorum than the low line, selected for low ability to produce antibody titers. Mitogenic response to phytohemagglutinin mitogen in the blood lymphocytes from high line quail was more reactive than those from low line quail. Significant line differences were also found in the number of spontaneous rosette cells produced by blood lymphocytes with both fowl and rabbit erythrocytes; the number in the high line exceeded those in the low line.
Cultures of L cells were synchronized with respect to deoxyribonucleic acid (DNA) synthesis with thymidine and 5-fluoro-2'-deoxyuridine (FUdR) and infected with Newcastle disease virus (NDV), mengovirus, or reovirus 3. Inhibition of incorporation of (3)H-cytidine into the DNA of synchronized cells is partially inhibited 2 hr after infection with NDV or mengovirus and nearly completely suppressed 4 hr after infection. With NDV and mengovirus, no evidence was obtained of differences in sensitivity of cells during early S phase as compared to later stages in DNA synthesis. When cells were infected with reovirus at the time of release from FUdR block, inhibition of cellular DNA synthesis was evident at 2 to 3 hr, and it was complete at 4 to 5 hr after infection. However, when cells were infected several hours prerelease, synthesis of DNA occurred in early S phase in spite of the fact that the cells had been infected for up to 6 hr. The results indicate that DNA synthesis in early S phase is relatively insensitive to the inhibitory function of reovirus. Colorimetric determinations (diphenylamine reaction) of the amounts of DNA produced in synchronized cells have substantiated the inhibition of DNA synthesis observed by isotope incorporation techniques.