The effect of 4-(2-hydroxyethyl piperazine-1-ethanesulfonic acid) on antigen titre of live attenuated Bovine Pleuro-pneumonia (CBPP) T1-44 vaccine during propagation, formulation and storage

Abstract

Contagious bovine pleuro-pneumonia is a highly contagious and infectious disease of cattle and water buffaloes caused by Mycoplasma mycoides subsp. mycoides small colony that affect the respiratory tract of animals. Vaccination with CBPP T1-44 live attenuated strain is the most effective method of controlling disease, though the African manufacturers are unable to produce the CBPP vaccine of the recommended titre per dose of 10⁸. Including a 4-(2-Hydroxyethyl piperazine-1-Ethanesulfonic Acid) (HEPES) buffer in growth media helps maintain pH and obtain optimal antigen and vaccine titre. This study aimed to evaluate the potential effects of HEPES buffer in improving CBPP T1-44 strain vaccine titre. Vaccines were produced by culturing in the PPLO media buffered with HEPES at different concentrations of 0.05M, 0.075M, and 0.1M, with Na₂HPO₄ as a control unit. Harvest pH was fixed at 6.5±0.3. The titre of the vaccine was obtained by titration whereby a total viable number of Mycoplasma was determined by colour change unit 50, and the titre was calculated according to Spearman and Karber's formula. Mean titre per dose were log₁₀ 1x10^9.3, log₁₀ 1x10^9.23, log₁₀ 1x10^9.133 and log₁₀ 1x10^6.982 for HEPES buffered vaccine at 0.05M, 0.075M, 0.1M and Na₂HPO₄ respectively. The post hoc test revealed the statistically significant difference between all concentrations of HEPES tested and Na₂HPO₄. Adding relatively inexpensive HEPES to most current media formulations is essential for producing a quality, efficacious, stable vaccine at a minimal cost with optimal recommended titre.