Sylvatic circulation of African swine fever virus (ASFV) in warthogs and Ornithodoros ticks that live in warthog burrows historically occurred in northern South Africa. Outbreaks of the disease in domestic pigs originated in this region. A controlled area was declared in the north in 1935 and regulations were implemented to prevent transfer of potentially infected suids or products to the rest of the country. However, over the past six decades, warthogs have been widely translocated to the south where the extralimital animals have flourished to become an invasive species. Since 2016, there have been outbreaks of ASF in pigs outside the controlled area that cannot be linked to transfer of infected animals or products from the north. An investigation in 2008–2012 revealed that the presence of Ornithodoros ticks and ASFV in warthog burrows extended marginally across the boundary of the controlled area. We found serological evidence of ASFV circulation in extralimital warthogs further south in the central part of the country.

Reagents were prepared for use in ELISAs to determine the concentration of the antigenic components of Clostridium botulinum type C and D. The results obtained were compared with the L+dose assay and a good correlation was found between the two assays for measurement of the C and D neurotoxin concentration. These ELISAs were also used to determine the concentration of the neurotoxins in toxoid form. The relationship between the C neurotoxin dose, in toxoid form, and the immune response in guinea pigs could be deduced from the data obtained. The relationship for the D neurotoxin was not that clear, as the same concentration of the antigen resulted in variable potency values. However, these ELISAs can be used to formulate the concentration of the C and D components in the final bivalent vaccine. Replacement of the preliminary potency assay on the monovalent components after production with the in vitro assays will shorten the total production time of the vaccine by about 60 days. The economical and ethical implications are the reduction in the use of animals to evaluate the vaccine. ß

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