IMPROVED NEWCASTLE DISEASE VIRUS VACCINES AND VECTORS
Manoharan, Vinoth Kumar
Samal, Siba K
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Newcastle disease (ND) is an economically important disease of poultry worldwide. The use of vaccines to control ND is necessary because of frequent outbreaks of the disease in enzootic countries. The fusion (F) and hemagglutinin-neuraminidase (HN) proteins of Newcastle disease virus (NDV) are multifunctional proteins that play critical roles during infection. The F protein of NDV is a type I membrane glycoprotein that mediates the fusion of viral envelope to the host cell membrane. The F protein activation initiates a series of conformational changes in the F protein leading to virus-cell membrane fusion, which occurs at the cell surface at neutral pH thus modulating NDV entry and spread. In the present study, we investigated the role of tyrosine to alanine mutation at amino acid position 524 and 527 in the F protein cytoplasmic tail (CT) of NDV strain LaSota by using reverse genetic techniques. Our results suggest that tyrosine residues at 524 and 527 position of F protein CT domain play a major role in fusogenicity and in replication thus modulating NDV infectivity. The F protein is synthesized as an inactive precursor, F0, which is functionally activated after cleavage by host cell proteases into F1 and F2 polypeptides, linked by disulfide bonds. The amino acid sequence surrounding the F protein cleavage site determines the virulence of NDV. We also studied the role of other avian paramyxovirus fusion protein cleavage site sequences in F protein cleavage of NDV strain Banjarmasin. This study has helped us to understand the requirement of F protein cleavage site in proteolytic processing, plaque formation and virus infectivity. Further, the role of these F cleavage site mutant viruses as genotype-matched vaccines for virulent NDV infection has been explored. Reverse genetics has also been used to develop NDV strains as a potential vaccine vectors for various human and animal pathogens, such as highly pathogenic avian influenza (H5N1), human immunodeficiency virus, severe acute respiratory syndrome coronavirus, ebola virus, respiratory syncytial virus and human parainfluenza virus type 3. NDV has several characteristics that makes it a suitable candidate for vaccine vector development. It is safe in humans and animals due to natural host range restriction, expresses foreign protein abundantly, infects via intranasal route, produces both humoral and mucosal immune responses, is antigenically distinct from human and animal pathogens, and lack of preexisting immunity to NDV in humans and animals. In one vaccine trial with non-human primates, the mesogenic NDV strain Beaudette C (BC) replicated to a high titer and induced a substantially higher antibody response compared to the lentogenic strain LaSota, and thus appeared to be more effective. However, NDV strains that have a polybasic cleavage site in the F protein and an intracerebral pathogenicity index (ICPI) >0.7 have been classified as Select Agents. Most mesogenic strains, including strain BC, fall into this category and therefore cannot be handled in BSL-2 conditions. In this study, we constructed a series of recombinant (rNDV) vectors containing the cleavage site sequence of avirulent strain LaSota and other avian paramyxoviruses, together with various regions of the F protein exchanged between NDV strains AKO-18 and BC. We used these modified rNDV vectors to express SIV gp160 envelope protein and immunized guinea pigs. Our results showed that rNDV/SIV vaccines were immunogenic and effectively neutralized SIV mac251 strain in vitro. These results support the idea of the use of NDV as a vaccine vector for expression of SIV immunogens capable of inducing neutralizing antibodies against diverse SIV strains, thus providing an improved vaccine vector platform for ultimately testing the NDV vectored vaccines in non-human primates and humans.