A novel Stx2B-based immunogen is able to confer total protection against Stx2 challenge

MEJIAS M.P; GHERSI G., ; PANEK C.A; FERNÁNDEZ-BRANDO R.J; CABRERA, G; ZYLBERMAN V; GOLDBAUM F; PALERMO M.S

Amsterdam, The Netherlands

Simposio; 8th International Symposium on Shiga Toxin (Verocytotoxin) Producing Escgerichi coli Infections; 2012

VTEC

Introduction &Objectives: Infection with Shiga Toxin II (Stx2) producing Escherichia coli (STEC) causes hemorrhagic colitis and can progress to the life-threatening complication known as Hemolytic Uremic Syndrome (HUS). The aim of this work was to generate and evaluate a novel immunogen against Stx2. For this purpose, the B subunit of Stx2 (Stx2B, binding subunit) was linked to a bacterial decameric protein scaffold (the resulting protein was named Chimera). This carrier is a highly immunogenic protein with adjuvant properties. Material & Methods: The stx2b coding sequence was cloned upstream to the bacterial carrier coding sequence (previously cloned in pET11-a vector) using a 10 aminoacid linker. This construct was used for expression and purification of the recombinant protein (rChimera). The chimera coding sequence was also sub-cloned into pCI-neo (pChimera) for DNA vaccination. BALB/c mice were immunized 3 times with rChimera (20µg Stx2B/dose) formulated in: A) Freund´s Adjuvant (FA)(intraperitoneal, i.p.), B) Alum Hydroxide (subcutaneous) and C) without adjuvant (i.p.). In addition, a prime boost protocol (D) was evaluated, with 3 doses of pChimera (100 µg/dose, intramuscular) and 1 dose of rChimera in FA (i.p.). Mice were immunized with recombinant Stx2B in FA (E)(i.p.) as control. Results: The Stx2B-specific IgG titers were determined by ELISA 45 days after the last immunization (n, Mean±SEM): A) 6, 124890±24289*; B) 4, 7285±2400; C) 4, 688±150; D) 4, 6910±2966; E) 4, 924±676. (*p