Vaccine 29 (2011) 867–872 Contents lists available at ScienceDirect Vaccine journal homepage: www.elsevier.com/locate/vaccine Short communication Swine adipose stromal cells loaded with recombinant bovine herpesvirus 4 virions expressing a foreign antigen induce potent humoral immune responses in pigs Gaetano Donofrio a,∗ , Simone Taddei a , Valentina Franceschi a , Antonio Capocefalo a , Sandro Cavirani a , Nicola Martinelli b , Simone Ottonello c , Maura Ferrari b a Dipartimento di Salute Animale, Università di Parma, Sezione di Malattie Infettive degli Animali, Via del Taglio 10, 43126 Parma, Italy b Istituto Zooprofilattico Sperimentale della Lombardia e dell’Emilia-Romagna, Via A. Bianchi 7, 25124 Brescia, Italy c Dipartimento di Biochimica e Biologia Molecolare, Università di Parma, V.le G.P. Usberti, 23/A, Campus Universitario, 43124 Parma, Italy article info Article history: Received 29 September 2010 Received in revised form 3 November 2010 Accepted 14 November 2010 Available online 27 November 2010 Keywords: Adipose derived stromal cells Bovine herpesvirus vector Pig Serum neutralizing antibodies abstract Increasingly effective vaccination strategies are needed to counteract the high incidence of contagious diseases associated with intensive swine breeding. Recombinant viral vaccines are a promising new avenue in this direction. Key features of viral vectors suitable for immunoprophylaxis are safety, ease of manipulation and the ability to replicate in a variety of hosts. Most of the above requirements are met by bovine herpesvirus 4 (BoHV-4), a non-pathogenic dsDNA virus capable of infecting a broad range of cell types in vitro. Here we report the results of an exploratory study using an engineered BoHV-4 virus (eBoHV-4) expressing two unrelated glycoprotein antigens from bovine viral diarrhea virus (BVDV) and bovine herpesvirus 1 (BoHV-1), to assess the potential of recombinant BoHV-4 as a self-adjuvanted immunogen in pigs. Free eBoHV-4 virions and virions preloaded into homologous swine adipose-derived stromal cells (SADSC) were tested. Neither virus formulation elicited neutralizing anti-BoHV-4 antibodies, nor any disease symptom, yet both induced specific immune responses against the heterologous antigens. However, a much earlier (18 vs 28 days post-infection) and more robust neutralizing response against BVDV and BoHV-1 viruses was elicited by eBoHV-4-preinfected SADSCs compared to free virions. The data validate BoHV-4 as a safe and effective heterologous antigen carrier/producer and identify SADSCs as helpful tools for the formulation of increasingly efficacious recombinant immunogens for pig vaccination. © 2010 Elsevier Ltd. All rights reserved. 1. Introduction Bovine herpesvirus 4 (BoHV-4) is a dsDNA virus belonging to the Herpesviridae family,  -herpesviridae subfamily [1]. Although replication of most -herpesviruses is restricted to their natural host species, BoHV-4 is highly promiscuous. It has been recovered from a variety of ruminants besides cattle, plus sporadic isolates from species as diverse as lions, cats and owl monkeys [2,3]. As to live animals, BoHV-4 has been reported to infect goats [4], guinea pigs, and rabbits [5]. Despite compelling evidence regarding its involvement in post- partum metritis [6–8], BoHV-4 has been proposed, and is currently being tested, as a gene delivery vector for immunoprophylaxis and gene therapy due to its extremely favourable biological proper- ties [9,10]: (i) the lack of oncogenic potential; (ii) a large genome (144 ± 4 kb) capable of accommodating large fragments of foreign ∗ Corresponding author. Tel.: +39 0521902677; fax: +39 0521902672. E-mail address: gaetano.donofrio@unipr.it (G. Donofrio). DNA; and (iii) the ability to replicate in a variety of host species, including some human cell lines [9,11,12]. The construction of infectious, BoHV-4-derived Bacterial Arti- ficial Chromosome (BAC) clones in Escherichia coli, followed by progeny virion reconstitution via transfection into permissive eukaryotic cells, has been successfully pursued as a means to rapidly modify the viral genome and adapt it to specific needs [10,13,14]. Animals such as rabbits, sheep, cows and chickens have been successfully immunized with recombinant BoHV-4-BAC derivatives expressing heterologous antigens [10,14–17]. Despite the strong economical impact of the pig industry and the fact that swine cells have been shown to be susceptible to BoHV-4 infection [9], there are as yet no data regarding the ability of for- eign antigens expressed from recombinant BoHV-4 to produce an immune response in this important livestock species. We thus wished to assess the potential of BoHV-4-vectorialized foreign antigens as prototype recombinant vaccines in pigs. Being concerned with the possible occurrence of anti-vector side- responses, which may lead to a rapid clearance of the virus, a formulation consisting of homologous cells preinfected with 0264-410X/$ – see front matter © 2010 Elsevier Ltd. All rights reserved. doi:10.1016/j.vaccine.2010.11.048