Evaluation in Mice of a Conjugate Vaccine for Cholera Made from Vibrio cholerae O1 (Ogawa) O-Specific Polysaccharide

Background: Protective immunity against cholera is serogroup specific. Serogroup specificity in Vibrio cholerae is determined by the O-specific polysaccharide (OSP) of lipopolysaccharide (LPS). Generally, polysaccharides are poorly immunogenic, especially in young children. Methodology Here we report the evaluation in mice of a conjugate vaccine for cholera (OSP:TThc) made from V. cholerae O1 Ogawa O-Specific Polysaccharide–core (OSP) and recombinant tetanus toxoid heavy chain fragment (TThc). We immunized mice intramuscularly on days 0, 21, and 42 with OSP:TThc or OSP only, with or without dmLT, a non-toxigenic immunoadjuvant derived from heat labile toxin of Escherichia coli. Principal Findings We detected significant serum IgG antibody responses targeting OSP following a single immunization in mice receiving OSP:TThc with or without adjuvant. Anti-LPS IgG responses were detected following a second immunization in these cohorts. No anti-OSP or anti-LPS IgG responses were detected at any time in animals receiving un-conjugated OSP with or without immunoadjuvant, and in animals receiving immunoadjuvant alone. Responses were highest following immunization with adjuvant. Serum anti-OSP IgM responses were detected in mice receiving OSP:TThc with or without immunoadjuvant, and in mice receiving unconjugated OSP. Serum anti-LPS IgM and vibriocidal responses were detected in all vaccine cohorts except in mice receiving immunoadjuvant alone. No significant IgA anti-OSP or anti-LPS responses developed in any group. Administration of OSP:TThc and adjuvant also induced memory B cell responses targeting OSP and resulted in 95% protective efficacy in a mouse lethality cholera challenge model. Conclusion: We describe a protectively immunogenic cholera conjugate in mice. Development of a cholera conjugate vaccine could assist in inducing long-term protective immunity, especially in young children who respond poorly to polysaccharide antigens

Survival likelihood of neonatal CD-1 mice following oral challenge with wild-type O1 Ogawa <i>V. cholerae</i> O395.

<p>Three- to five-day-old pups (cohort size 20) were orally gavaged with 50 µl of a preparation containing 2.3×10⁹ CFU of wild type <i>V. cholerae</i> O395 mixed with a 1∶250 dilution of pooled day 56 serum from mice intramuscularly immunized with conjugate vaccine (OSP:TThc) and immunoadjuvantative dmLT, or dmLT alone. Survival curves were compared by log rank testing.</p

Vibriocidal responses in mice intramuscularly immunized with OSP:TThc (with dmLT), OSP (with dmLT), OSP:TThc (no dmLT), OSP (no dmLT), or dmLT alone.

<p>The columns indicate mean reciprocal end titers, and error bars represent the standard errors of the mean. An asterisk denotes a statistically significant difference (<i>P</i><0.05) from baseline (day 0) titer. Responder frequencies are also listed. #, statistically significant difference among the vaccine cohorts (<i>P</i><0.05).</p

Serum anti-OSP-BSA and anti-LPS IgG antibody responses in mice intramuscularly immunized with OSP:TThc (with dmLT), OSP (with dmLT), OSP:TThc (no dmLT), OSP (no dmLT), or dmLT alone.

<p>Mean and standard error of the mean are reported for each group. An asterisk denotes a statistically significant difference (<i>P</i><0.05) from baseline (day 0) titer. Responder frequencies are also listed. #, statistically significant difference among the compared cohorts (<i>P</i><0.05).</p

Structure of Ogawa OSP:TThc conjugate.

<p>6.4 moles OSP per mole conjugate. TThc = recombinant tetanus toxoid heavy chain fragment.</p

OSP-BSA specific IgG memory B cell responses in mice immunized with different vaccine antigens.

*<p>Data are expressed as responder frequencies (see text).</p