Newcastle Disease Virus (NDV) Marker Vaccine: an Immunodominant Epitope on the Nucleoprotein Gene of NDV Can Be Deleted or Replaced by a Foreign Epitope

Abstract

The nucleoprotein (NP) of Newcastle disease virus (NDV) functions primarily to encapsidate the virus genome for the purpose of RNA transcription, replication, and packaging. This conserved multifunctional protein is also efficient in inducing NDV-specific antibody in chickens. Here, we localized a conserved B-cell immunodominant epitope (IDE) spanning residues 447 to 455 and successfully generated a recombinant NDV lacking the IDE by reverse genetics. Despite deletion of NP residues 443 to 460 encompassing the NP-IDE, the mutant NDV propagated in embryonated specific-pathogen-free chicken eggs to a level comparable to that of the parent virus. In addition, a B-cell epitope of the S2 glycoprotein of murine hepatitis virus (MHV) was inserted in-frame to replace the NP-IDE. Recombinant viruses properly expressing the introduced MHV epitope were successfully generated, demonstrating that the NP-IDE not only is dispensable for virus replication but also can be replaced by foreign sequences. Chickens immunized with the hybrid recombinants produced specific antibodies against the S2 glycoprotein of MHV and completely lacked antibodies directed against the NP-IDE. These marked-NDV recombinants, in conjunction with a diagnostic test, enable serological differentiation of vaccinated animals from infected animals and may be useful tools in ND eradication programs. The identification of a mutation-permissive region on the NP gene allows a rational approach to the insertion of protective epitopes and may be relevant for the design of NDV-based cross-protective marker vaccines.The negative-strand RNA virus genome of Newcastle disease virus (NDV) contains six genes encoding six major structural proteins: nucleoprotein (NP), phosphoprotein (P), matrix protein (M), fusion protein (F), hemagglutinin-neuraminidase (HN), and RNA-dependent RNA polymerase (L). The RNA together with NP, P, and L proteins forms the ribonucleoprotein complex (RNP), which serves as a template for RNA synthesis (15). The NP together with the polymerase proteins, P and L, plays an eminent role in encapsidating the RNA. Moreover, NP regulates transcription and replication of the viral genome by interacting with P alone, with P and L, or with itself (NP-NP interaction). For Sendai paramyxovirus, it was shown that a conserved N-terminal region of NP was involved in NP-RNA and NP-NP interaction (5), whereas the carboxyterminal domain was shown to be required for template function (8). Most of the NP is thus absolutely essential for virus replication due to multifold engagement of NP in the assembly and biologic activity of the RNP. In addition, NPs of negativestrand RNA viruses are highly immunogenic in nature and have been used as antigens for diagnostic purposes, including

Characterization of a Recombinant Newcastle Disease Virus Vaccine Strain

“…Since the first isolation of NDV from cDNA in 1999, various recombinant NDV vaccine strains with low virulence have been developed (14,20,34,35,43,44,46). However, NDV vaccine strains with all the desirable characteristics including virulence low enough to allow for in ovo vaccination, antigenic contemporaneity, and genetic stability are still elusive.…”

“…The dibasic amino acids at the proteolytic cleavage site ( 113 R-X-K/R-R 116 ) in the F protein are relevant to systemic replication, and the HN protein determines the tropisms of NDV (10,19,39). The V protein which is expressed via RNA editing of the P gene and the structure of noncoding regions of the viral genome such as gene start, gene end, and intergenic region (IGR) also contribute to virulence and replication of Paramyxovirus (12,23,35,48,54).…”

Analyses of the results of different test systems in the 2005 global proficiency testing schemes for infectious bursal disease virus and Newcastle disease virus antibody detection in chicken serum

“…When international reference sera are included in these studies, the quantitative results of such sera would provide very useful information on the comparability of the test results of this study to those of other studies. This study shows that when a laboratory reports a correlation between an antibody titre for IBDV or NDV and protection (Thayer et al ., 1987;Van den Berg & Meulemans, 1991;Maas et al ., 2001Maas et al ., , 2003Rahman et al ., 2004), or vaccine efficacy (Maas et al ., 2001(Maas et al ., , 2003Mebatsion et al ., 2002;Claassen et al ., 2004), or breakthrough titres for vaccination, and so on, without including results for any international reference sera, these titres cannot be interpreted correctly by other laboratories or authorities. Without the use of reference sera it is not possible to know the variation between laboratories or the comparability with other laboratories.…”

Generation of a Candidate Live Marker Vaccine for Equine Arteritis Virus by Deletion of the Major Virus Neutralization Domain

“…These vaccines have become attractive or mandatory in programs aimed at controlling or eradicating virus infections in food as well as in companion animals (2)(3)(4)65). Herpesvirus (glycoprotein gEdeleted pseudorabies virus and bovine herpesvirus) marker vaccines were among the first to be developed and used in the field (66), followed later by various genetically modified RNA viruses such as classical swine fever virus (64,69) and Newcastle disease virus (47,51). Marker vaccines can be based on live and inactivated viruses as well as on viral subunits, but they also include DNA and vector vaccines.…”