African horse sickness (AHS) is a debilitating and often fatal viral disease affecting horses in much
of Africa, caused by the dsRNA orbivirus African horse sickness virus (AHSV). Vaccination remains
the single most effective weapon in combatting AHS, as there is no treatment for the disease
apart from good animal husbandry. However, the only commercially available vaccine is a liveattenuated
version of the virus (LAV). The threat of outbreaks of the disease outside its endemic
region and the fact that the LAV is not licensed for use elsewhere in the world, have spurred
attempts to develop an alternative safer, yet cost-effective recombinant vaccine. Here, we report
the plant-based production of a virus-like particle (VLP) AHSV serotype five candidate vaccine by
Agrobacterium tumefaciens-mediated transient expression of all four capsid proteins in Nicotiana
benthamiana using the cowpea mosaic virus-based HyperTrans (CPMV-HT) and associated pEAQ
plant expression vector system. The production process is fast and simple, scalable, economically
viable, and most importantly, guinea pig antiserum raised against the vaccine was shown to
neutralize live virus in cell-based assays. To our knowledge, this is the first report of AHSV VLPs
produced in plants, which has important implications for the containment of, and fight against
the spread of, this deadly disease.
Figure S1 Optimization of plant-based expression of recombinant
AHSV-5 structural proteins.
Figure S2 Increased formation of AHSV-5 VLPs incorporating a
mutated version of VP7.
Figure S3 Purification of AHSV-5 VLPs.
Figure S4 Mass spectrometry analysis of the 4 protein bands
recovered from SDS-PAGE separation of density gradient fractions
from leaves co-infiltrated with Agrobacterium AGL1 pEAQ
recombinants for co-expression of AHSV capsid proteins VP2,
VP3, VP5 and VP7mu.
Figure S5 Quantification of AHSV-5 VLPs by gel densitometry.
Table S1 Virus neutralizing antibody titres of serum samples from
vaccinated and control guinea pigs V1 and C1.