title: Generation of porcine reproductive and respiratory syndrome (PRRS) virus-like-particles (VLPs) with different protein composition.
abstract: The causative agent of Porcine Reproductive and Respiratory Syndrome (PRRS) is an enveloped ssRNA (+) virus belonging to the Arteriviridae family. Gp5 and M proteins form disulfide-linked heterodimers that constitute the major components of PRRSV envelope. Gp2, Gp3, Gp4 and E are the minor structural proteins, being the first three incorporated as multimeric complexes in the virus surface. The disease has become one of the most important causes of economic losses in the swine industry. Despite efforts to design an effective vaccine, the available ones allow only partial protection. In the last years, VLPs have become good vaccine alternatives because of safety issues and their potential to activate both branches of the immunological response. The characteristics of recombinant baculoviruses as heterologous expression system have been exploited for the production of VLPs of a wide variety of viruses. In this work, two multiple baculovirus expression vectors (BEVs) with PRRS virus envelope proteins were engineered in order to generate PRRS VLPs: on the one hand, Gp5 and M cDNAs were cloned to generate the pBAC-Gp5M vector; on the other hand, Gp2, Gp3, Gp4 and E cDNAs have been cloned to generate the pBAC-Gp234E vector. The corresponding recombinant baculoviruses BAC-Gp5M and BAC-Gp234E were employed to produce two types of VLPs: basic Gp5M VLPs, by the simultaneous expression of Gp5 and M proteins; and complete VLPs, by the co-expression of the six PRRS proteins after co-infection. The characterization of VLPs by Western blot confirmed the presence of the recombinant proteins using the available specific antibodies (Abs). The analysis by Electron microscopy showed that the two types of VLPs were indistinguishable between them, being similar in shape and size to the native PRRS virus. This system represents a potential alternative for vaccine development and a useful tool to study the implication of specific PRRS proteins in the response against the virus.
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain. Electronic address: mgarcia@ingenasa.com.
authors.forename: Marga
authors.initials: M
authors.lastname: García Durán
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain.
authors.forename: Sofia
authors.initials: S
authors.lastname: Costa
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain.
authors.forename: Javier
authors.initials: J
authors.lastname: Sarraseca
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain.
authors.forename: Nuria
authors.initials: N
authors.lastname: de la Roja
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain.
authors.forename: Julia
authors.initials: J
authors.lastname: García
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain.
authors.forename: Isabel
authors.initials: I
authors.lastname: García
authors.affiliation: Inmunología y Genética Aplicada, S.A. (INGENASA), Hnos. Garcia Noblejas 41, 28037 Madrid, Spain.
authors.forename: Maria José
authors.initials: MJ
authors.lastname: Rodríguez
mesh_terms: D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D015153:Blotting, Western; D015870:Gene Expression; D005822:Genetic Vectors; D046529:Microscopy, Electron, Transmission; D019316:Porcine respiratory and reproductive syndrome virus / Q000235:genetics*; D011994:Recombinant Proteins / Q000032:analysis* / Q000235:genetics; D061987:Sf9 Cells; D018411:Spodoptera; D014764:Viral Proteins / Q000032:analysis* / Q000235:genetics; D022701:Virosomes / Q000737:chemistry* / Q000302:isolation & purification* / Q000648:ultrastructure
chemical_list: D011994:Recombinant Proteins; D014764:Viral Proteins; D022701:Virosomes