Construction and evaluation of genetically engineered replication-defective porcine reproductive and respiratory syndrome virus vaccine candidates

Welch, Siao Kun W.; Jolie, Rika; Pearce, Douglas S.; Koertje, William D.; Fuog, Eric; Shields, Shelly; Yoo, Dongwan; Calvert, Jay G.

doi:10.1016/j.vetimm.2004.09.022

Cited by 33 publications

(21 citation statements)

References 23 publications

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“…These may be attributed to the highly conserved sequences of ORFs 6 and 7. There are five missense mutations in ORF2 that have been reported to be critical for virus replication [36]. Residue changes at positions L 10 F and N 23 S (of the putative signal peptide sequence) have also been detected in comparisons between VR-2332 and 16244B strains [14].…”

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confidence: 97%

Molecular mutations associated with the in vitro passage of virulent porcine reproductive and respiratory syndrome virus

Han

Deng

et al. 2009

Virus Genes

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Mutants of a highly pathogenic, porcine reproductive, and respiratory syndrome virus (PRRSV), JXA1 strain, were prepared by continuous in vitro passage. Genomic sequence comparisons were made between mutants obtained at different passages and the parental strain JXA1. The mutant strain obtained at passage 80 contained a 12 nucleotide insertion and 108 nucleotide mutations that resulted in 45 amino acid changes. Most of these changes (89%) occurred between passage 10 and 45 and were genetically stable for the next 35-70 passages. A comparison of the mutants, their parental strain, and several American PRRSV strains, identified 13 characteristic amino acid changes. These sites, as well as the distinct 12 nucleotide insertion, represent possible genetic markers for the evaluation of live vaccine applications, particularly for additional studies of the safety and potency of live PRRSV vaccines.

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confidence: 97%

Molecular mutations associated with the in vitro passage of virulent porcine reproductive and respiratory syndrome virus

Han

Deng

et al. 2009

Virus Genes

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“…Several studies point towards an important role for the GP 5 and M proteins in PRRSV assembly (38,39), but nothing is yet known about the function of the minor structural proteins. Recently, Welch and coauthors (40) demonstrated that the ORF2a/b or ORF4 products are essential for a North American strain of PRRSV; mutant genomes from which these ORFs had been deleted did not generate infectivity for reasons that were not investigated. In the case of EAV, each of the structural proteins was shown to be required for the production of infectious progeny virus (26).…”

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confidence: 99%

Envelope Protein Requirements for the Assembly of Infectious Virions of Porcine Reproductive and Respiratory Syndrome Virus

Wissink

Kroese

Wijk

et al. 2005

J Virol

176

188

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Virions of porcine reproductive and respiratory syndrome virus (PRRSV) contain six membrane proteins: the major proteins GP 5 and M and the minor proteins GP 2a , E, GP 3 , and GP 4 . Here, we studied the envelope protein requirements for PRRSV particle formation and infectivity using full-length cDNA clones in which the genes encoding the membrane proteins were disrupted by site-directed mutagenesis. By transfection of RNAs transcribed from these cDNAs into BHK-21 cells and analysis of the culture medium using ultracentrifugation, radioimmunoprecipitation, and real-time reverse transcription-PCR, we observed that the production of viral particles is dependent on both major envelope proteins; no particles were released when either the GP 5 or the M protein was absent. In contrast, particle production was not dependent on the minor envelope proteins. Remarkably, in the absence of any one of the latter proteins, the incorporation of all other minor envelope proteins was affected, indicating that these proteins interact with each other and are assembled into virions as a multimeric complex. Independent evidence for such complexes was obtained by coexpression of the minor envelope proteins in BHK-21 cells using a Semliki Forest virus expression system. By analyzing the maturation of their N-linked oligosaccharides, we found that the glycoproteins were each retained in the endoplasmic reticulum unless expressed together, in which case they were collectively transported through the Golgi complex to the plasma membrane and were even detected in the extracellular medium. As the PRRSV particles lacking the minor envelope proteins are not infectious, we hypothesize that the virion surface structures formed by these proteins function in viral entry by mediating receptor binding and/or virus-cell fusion.Porcine reproductive and respiratory syndrome virus (PRRSV) belongs to the family of Arteriviridae, which also comprises Equine arteritis virus (EAV), Lactate dehydrogenase-elevating virus (LDV), and Simian hemorrhagic fever virus (24,42). This family belongs to the order of Nidovirales, together with the Coronaviridae, Toroviridae, and Roniviridae (2, 3). Arteriviruses are enveloped RNA viruses that contain a positive-strand RNA genome and synthesize a 3Ј nested set of six or eight subgenomic RNAs (sgRNAs) that encode the structural proteins. Analyses of purified virions of PRRSV have indicated that they are composed of seven proteins, i.e., four envelope glycoproteins named GP 2a (encoded by open reading frame 2a [ORF2a]), GP 3 (ORF3), GP 4 (ORF4), and GP 5 (ORF5); a nonglycosylated membrane protein M (ORF6); the nucleocapsid protein N (ORF7); and a nonglycosylated envelope protein, E, that is expressed from a second ORF (ORF2b) entirely contained within ORF2 (22,25,35,48). The 29-to 30-kDa GP 2a and 31-to 35-kDa GP 4 proteins are both putative class I integral membrane proteins with an N-terminal signal sequence and a C-terminal membrane anchor, containing two and four predicted N-glycosylation sites, respectively (22). T...

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“…The PRRSV genome encodes 14 non-structural and 8 structural proteins, several of which are well conserved and/or highly immunogenic [37–40]. Up to now, efforts to delete an entire gene of PRRSV have not been successful [20–23]. Therefore, the most feasible approach to develop a live-attenuated DIVA vaccine against PRRSV would be selectively eliminating a small protein fragment or an epitope, instead of deleting an entire protein.…”

Section: Discussionmentioning

confidence: 99%

“…Classically, live-attenuated DIVA vaccines are generated through deletion of an entire gene encoding an immunogenic, non-essential protein [17]. While technically straightforward in the case of some double-stranded DNA viruses like Pseudorabies Virus (Suid Herpesvirus 1), it is very difficult to delete an entire gene of a smaller RNA genome virus like PRRSV, whose genes are all essential for the productive viral infection [20–23]. An alternative approach to develop live-attenuated DIVA vaccines for RNA viruses is to selectively eliminate a small protein fragment or an epitope, instead of deleting the whole protein [24–26].…”

Section: Introductionmentioning

confidence: 99%

Characterization of a serologic marker candidate for development of a live-attenuated DIVA vaccine against porcine reproductive and respiratory syndrome virus

Kwon

Lima

et al. 2013

Vaccine

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DIVA (differentiating infected from vaccinated animals) vaccines have proven extremely useful for control and eradication of infectious diseases in livestock. We describe here the characterization of a serologic marker epitope, so-called epitope-M201, which can be a potential target for development of a live-attenuated DIVA vaccine against porcine reproductive and respiratory syndrome virus (PRRSV). Epitope-M201 is located at the carboxyl terminus (residues 161-174) of the viral M protein. The epitope is highly immunodominant and well-conserved among type-II PRRSV isolates. Rabbit polyclonal antibodies prepared against this epitope are non-neutralizing; thus, the epitope does not seem to contribute to the protective immunity against PRRSV infection. Importantly, the immunogenicity of epitope-M201 can be disrupted through the introduction of a single amino acid mutation which does not adversely affect the viral replication. All together, our results provide an important starting point for the development of a live-attenuated DIVA vaccine against type-II PRRSV.

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Construction and evaluation of genetically engineered replication-defective porcine reproductive and respiratory syndrome virus vaccine candidates

Cited by 33 publications

References 23 publications

Molecular mutations associated with the in vitro passage of virulent porcine reproductive and respiratory syndrome virus

Molecular mutations associated with the in vitro passage of virulent porcine reproductive and respiratory syndrome virus

Envelope Protein Requirements for the Assembly of Infectious Virions of Porcine Reproductive and Respiratory Syndrome Virus

Characterization of a serologic marker candidate for development of a live-attenuated DIVA vaccine against porcine reproductive and respiratory syndrome virus

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