Enhanced Production of Shigella flexneri Polysaccharide from a Newly Devised Silicate Method as a Potential Vaccine Conjugate.

Abstract

Conventional glycoconjugates are prepared with polysaccharides (PS) isolated from bacterial sources by fermentation technique. This approach has some major challenges like lower yield of PS, impurities and usage of hazardous chemicals. Reports on efficient and enhanced production of PS from Shigella flexneri is meager in literature. Hence, in the current study, three different types of media namely Yeast extract medium, Shigella sonnei-defined medium and synthetic medium were utilized for the culture of S. flexneri. Among the selected media it was recognized that the culture of S. flexneri harvested in synthetic media produced significant quantity of PS in less time when compared to the other two media. Different purification techniques such as phenol chloroform extraction, acid precipitation, detergent method, chromatographic purification and a novel silicate method were carried out to refine the harvested PS from impurities. It was observed that large impurities such as bacterial protein, debris and media components were eliminated significantly by using chromatographic and silicate methods. The final yield of purified PS was approximately 20-35% higher in silicate method which is reported for the first time in this study for purification of PS. Further, the characterization of the purified PS was done using high-performance liquid chromatography and high-performance anion-exchange chromatography with pulsed amperometric detection. Hence, the robust process developed in the present study using synthetic media and chromatographic filtration technique along with the novel silicate treatment produced significant quantities of PS from S. flexneri in reduced cost and time, which could be further conjugated to a suitable carrier to generate a potential Shigella conjugate antigen.