Glycoconjugate vaccine using a genetically modified O antigen induces protective antibodies toFrancisella tularensis

Abstract

Francisella tularensisis the causative agent of tularemia, a category A bioterrorism agent. The lipopolysaccharide (LPS) O antigen (OAg) ofF. tularensishas been considered for use in a glycoconjugate vaccine, but conjugate vaccines tested so far have failed to confer protection necessary against aerosolized pulmonary bacterial challenge. WhenF. tularensisOAg was purified under standard conditions, the antigen had a small molecular size [25 kDa, low molecular weight (LMW)]. Using milder extraction conditions, we found the native OAg had a larger molecular size [80 kDa, high molecular weight (HMW)], and in a mouse model of tularemia, a glycoconjugate vaccine made with the HMW polysaccharide coupled to tetanus toxoid (HMW-TT) conferred better protection against intranasal challenge than a conjugate made with the LMW polysaccharide (LMW-TT). To further investigate the role of OAg size in protection, we created anF. tularensislive vaccine strain (LVS) mutant with a significantly increased OAg size [220 kDa, very high molecular weight (VHMW)] by expressingin F. tularensisa heterologous chain-length regulator gene (wzz) from the related speciesFrancisella novicida. Immunization with VHMW-TT provided markedly increased protection over that obtained with TT glycoconjugates made using smaller OAgs. We found that protective antibodies recognize a length-dependent epitope better expressed on HMW and VHMW antigens, which bind with higher affinity to the organism.