Abstract
Classical Swine Fever Virus (CSFV) causes classical swine fever, a highly contagious hemorrhagic fever affecting both feral and domesticated pigs. Outbreaks of CSF in Europe, Asia, Africa and South America had significant adverse impacts on animal health, food security and the pig industry. The disease is generally contained by prevention of exposure through import restrictions (e.g. banning import of live pigs and pork products), localized vaccination programmes and culling of infected or at‐risk animals, often at very high cost. Current CSFV‐modified live virus vaccines are protective, but do not allow differentiation of infected from vaccinated animals (DIVA), a critical aspect of disease surveillance programmes. Alternatively, first‐generation subunit vaccines using the viral protein E2 allow for use of DIVA diagnostic tests, but are slow to induce a protective response, provide limited prevention of vertical transmission and may fail to block viral shedding. CSFV E2 subunit vaccines from a baculovirus/insect cell system have been developed for several vaccination campaigns in Europe and Asia. However, this expression system is considered expensive for a veterinary vaccine and is not ideal for wide‐spread deployment. To address the issues of scalability, cost of production and immunogenicity, we have employed an Agrobacterium‐mediated transient expression platform in Nicotiana benthamiana and formulated the purified antigen in novel oil‐in‐water emulsion adjuvants. We report the manufacturing of adjuvanted, plant‐made CSFV E2 subunit vaccine. The vaccine provided complete protection in challenged pigs, even after single‐dose vaccination, which was accompanied by strong virus neutralization antibody responses.
Highlights
Classical Swine Fever (CSF) is a highly contagious hemorrhagic disease affecting pigs
Insect cell-produced E2 antigens from Porcilisâ Pesti (MSD Animal Health, Germany) and Kansas State University (European Medicine Agency, 2000; Madera et al, 2016) were produced as transmembrane domain-deleted antigens. Both Classical swine fever virus (CSFV) E2 antigens were expressed in plants by vacuum agroinfiltration and purified by affinity chromatography
Antigen identity was confirmed by western blot, LiquidChromatography Mass Spectrometry (LC-MS) analysis of intact proteins and tryptic fragmentation of in-gel isolated protein (Figure 1c)
Summary
Classical Swine Fever (CSF) is a highly contagious hemorrhagic disease affecting pigs. Antigen production using the baculovirus/insect cell expression system remains expensive at manufacturing scale and limits widespread deployment of this particular subunit vaccine With this in mind, we chose to explore an alternative expression platform (Agrobacterium-mediated transient protein production in Nicotiana benthamiana plants) that has the potential to significantly reduce the cost to manufacture a novel CSFV E2 subunit vaccine (Nandi et al, 2016). Transient protein expression in N. benthamiana has become one of the main platforms used to produce plant-made therapeutics, with manufacturing facilities available to support clinical development and commercial launch (Holtz et al, 2015) This platform is well-adapted to the manufacture of veterinary products because it: (i) is easy to implement, (ii) provides flexible scales of production, (iii) offers rapid development timelines and (iv) has been demonstrated to produce products at lower cost than traditional cell culture-based expression platforms (Holtz et al, 2015; Kolotilin et al, 2014; Nandi et al, 2016; Shahid and Daniell, 2016). We describe the design and production of a N. benthamiana-produced recombinant CSFV E2 antigen, and present data assessing immunogenicity, safety and efficacy of the antigen against wild-type CSFV challenge in pigs
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