Dissecting serotype-specific contributions to live oral cholera vaccine efficacy.

The O1 serogroup of Vibrio cholerae causes pandemic cholera and is divided into the Ogawa and Inaba serotypes. The O-antigen is V. cholerae's immunodominant antigen, and the two serotypes, which differ by the presence or absence of a terminally methylated O-antigen, likely influence development of immunity to cholera and oral cholera vaccines (OCVs). However, there is no consensus regarding the relative immunological potency of each serotype, in part because previous studies relied on genetically heterogeneous strains. Here, we engineered matched serotype variants of a live OCV candidate, HaitiV, and used a germfree mouse model to evaluate the immunogenicity and protective efficacy of each vaccine serotype. By combining vibriocidal antibody quantification with single- and mixed-strain infection assays, we found that all three HaitiV variants-InabaV, OgawaV, and HikoV (bivalent Inaba/Ogawa)-were immunogenic and protective. None of the vaccine serotypes were superior across both of these vaccine metrics, suggesting that the impact of O1-serotype variation in OCV design, although detectable, is subtle. However, all three live vaccines significantly outperformed formalin-killed HikoV, supporting the idea that live OCV usage will bolster current cholera control practices. The potency of OCVs was found to be challenge strain-dependent, emphasizing the importance of appropriate strain selection for cholera challenge studies. Our findings and experimental approaches will be valuable for guiding the development of live OCVs and oral vaccines for additional pathogens.

Correlates of Protection for Cholera.

A correlate of protection (CoP) is a measured adaptive immune response to vaccination or infection that is associated with protection against disease. However, the degree to which a CoP can serve as a surrogate end point for vaccine efficacy should depend on the robustness of this association. While cholera toxin is a dominant target of the human antibody response to Vibrio cholerae infection, antitoxin responses are not associated with long-term immunity, and are not effective CoPs for cholera. Instead, protection appears to be mediated by functional antibodies that target the O-polysaccharide coated V. cholerae outer membrane. Vibriocidal antibodies, which are complement-dependent bactericidal antibodies, remain the most accepted CoP for cholera and are used as surrogate end points in some vaccine studies. However, the association between vibriocidal antibody titers and immunity is not absolute, and they are unlikely to reflect a mechanistic correlate of protection against cholera.

Gut Microbiota and Development of Vibrio cholerae-Specific Long-Term Memory B Cells in Adults after Whole-Cell Killed Oral Cholera Vaccine.

Cholera is a diarrheal disease caused by Vibrio cholerae that continues to be a major public health concern in populations without access to safe water. IgG- and IgA-secreting memory B cells (MBC) targeting the V. cholerae O-specific polysaccharide (OSP) correlate with protection from infection in persons exposed to V. cholerae and may be a major determinant of long-term protection against cholera. Shanchol, a widely used oral cholera vaccine (OCV), stimulates OSP MBC responses in only some people after vaccination, and the gut microbiota is a possible determinant of variable immune responses observed after OCV. Using 16S rRNA sequencing of feces from the time of vaccination, we compared the gut microbiota among adults with and without MBC responses to OCV. Gut microbial diversity measures were not associated with MBC isotype or OSP-specific responses, but individuals with a higher abundance of Clostridiales and lower abundance of Enterobacterales were more likely to develop an MBC response. We applied protein-normalized fecal supernatants of high and low MBC responders to THP-1-derived human macrophages to investigate the effect of microbial factors at the time of vaccination. Feces from individuals with higher MBC responses induced significantly different IL-1ß and IL-6 levels than individuals with lower responses, indicating that the gut microbiota at the time of vaccination may "prime" the mucosal immune response to vaccine antigens. Our results suggest the gut microbiota could impact immune responses to OCVs, and further study of microbial metabolites as potential vaccine adjuvants is warranted.

Impaired mucociliary motility enhances antigen-specific nasal IgA immune responses to a cholera toxin-based nasal vaccine.

Nasal mucosal tissues are equipped with physical barriers, mucus and cilia, on their surface. The mucus layer captures inhaled materials, and the cilia remove the inhaled materials from the epithelial layer by asymmetrical beating. The effect of nasal physical barriers on the vaccine efficacy remains to be investigated. Tubulin tyrosine ligase-like family, member 1 (Ttll1) is an essential enzyme for appropriate movement of the cilia on respiratory epithelium, and its deficiency (Ttll1-KO) leads to mucus accumulation in the nasal cavity. Here, when mice were intra-nasally immunized with pneumococcal surface protein A (PspA, as vaccine antigen) together with cholera toxin (CT, as mucosal adjuvant), Ttll1-KO mice showed higher levels of PspA-specific IgA in the nasal wash and increased numbers of PspA-specific IgA-producing plasma cells in the nasal passages when compared with Ttll1 hetero (He) mice. Mucus removal by N-acetylcysteine did not affect the enhanced immune responses in Ttll1-KO mice versus Ttll1-He mice. Immunohistological and flow cytometry analyses revealed that retention time of PspA in the nasal cavity in Ttll1-KO mice was longer than that in Ttll1-He mice. Consistently, uptake of PspA by dendritic cells was higher in the nasopharynx-associated lymphoid tissue (NALT) of Ttll1-KO mice than that of Ttll1-He mice. These results indicate that the ciliary function of removing vaccine antigen from the NALT epithelial layer is a critical determinant of the efficacy of nasal vaccine.

Transient Intestinal Colonization by a Live-Attenuated Oral Cholera Vaccine Induces Protective Immune Responses in Streptomycin-Treated Mice.

Current mouse models for evaluating the efficacy of live oral cholera vaccines (OCVs) have important limitations. Conventionally raised adult mice are resistant to intestinal colonization by Vibrio cholerae, but germfree mice can be colonized and have been used to study OCV immunogenicity. However, germfree animals have impaired immune systems and intestinal physiology; also, live OCVs colonize germfree mice for many months, which does not mimic the clearance kinetics of live OCVs in humans. In this study, we leveraged antibiotic-treated, conventionally raised adult mice to study the effects of transient intestinal colonization by a live OCV V. cholerae strain. In a single-dose vaccination regimen, we found that HaitiV, a live-attenuated OCV candidate, was cleared by streptomycin-treated adult mice within 2 weeks after oral inoculation. This transient colonization elicited far stronger adaptive immune correlates of protection against cholera than did inactivated whole-cell HaitiV. Infant mice from HaitiV-vaccinated dams were also significantly more protected from choleric disease than pups from inactivated-HaitiV-vaccinated dams. Our findings establish the benefits of antibiotic-treated mice for live-OCV studies as well as their limitations and underscore the immunogenicity of HaitiV.IMPORTANCE Oral cholera vaccines (OCVs) are being deployed to combat cholera, but current killed OCVs require multiple doses and show little efficacy in young children. Live OCVs have the potential to overcome these limitations, but small-animal models for testing OCVs have shortcomings. We used an antibiotic treatment protocol for conventional adult mice to study the effects of short-term colonization by a single dose of HaitiV, a live-OCV candidate. Vaccinated mice developed vibriocidal antibodies against V. cholerae and delivered pups that were resistant to cholera, whereas mice vaccinated with inactivated HaitiV did not. These findings demonstrate HaitiV's immunogenicity and suggest that this antibiotic treatment protocol will be useful for evaluating the efficacy of live OCVs.

Immunogenicity and protective efficacy of a live, oral cholera vaccine formulation stored outside-the-cold-chain for 140 days.

BACKGROUND: Cholera, an acute watery diarrhoeal disease caused by Vibrio cholerae serogroup O1 and O139 across the continents. Replacing the existing WHO licensed killed multiple-dose oral cholera vaccines that demand 'cold chain supply' at 2-8 °C with a live, single-dose and cold chain-free vaccine would relieve the significant bottlenecks and cost determinants in cholera vaccination campaigns. In this direction, a prototype cold chain-free live attenuated cholera vaccine formulation (LACV) was developed against the toxigenic wild-type (WT) V. cholerae O139 serogroup. LACV was found stable and retained its viability (5 × 106 CFU/mL), purity and potency at room temperature (25 °C ± 2 °C, and 60% ± 5% relative humidity) for 140 days in contrast to all the existing WHO licensed cold-chain supply (2-8 °C) dependent killed oral cholera vaccines. RESULTS: The LACV was evaluated for its colonization potential, reactogenicity, immunogenicity and protective efficacy in animal models after its storage at room temperature for 140 days. In suckling mice colonization assay, the LACV recorded the highest recovery of (7.2 × 107 CFU/mL) compared to those of unformulated VCUSM14P (5.6 × 107 CFU/mL) and the WT O139 strain (3.5 × 107 CFU/mL). The LACV showed no reactogenicity even at an inoculation dose of 104-106 CFU/mL in a rabbit ileal loop model. The rabbits vaccinated with the LACV or unformulated VCUSM14P survived a challenge with WT O139 and showed no signs of diarrhoea or death in the reversible intestinal tie adult rabbit diarrhoea (RITARD) model. Vaccinated rabbits recorded a 275-fold increase in anti-CT IgG and a 15-fold increase in anti-CT IgA antibodies compared to those of rabbits vaccinated with unformulated VCUSM14P. Vibriocidal antibodies were increased by 31-fold with the LACV and 14-fold with unformulated VCUSM14P. CONCLUSION: The vaccine formulation mimics a natural infection, is non-reactogenic and highly immunogenic in vivo and protects animals from lethal wild-type V. cholerae O139 challenge. The single dose LACV formulation was found to be stable at room temperature (25 ± 2 °C) for 140 days and it would result in significant cost savings during mass cholera vaccination campaigns.

Immunohistochemical, histological and ultrastructural evaluation of protection provided by cholera vaccine against V. cholerae O139.

In our previous study, complete protection was observed in rabbit immunized with 1 × 1010 CFU of live attenuated VCUSM21P vaccine against challenge with 1 × 109 CFU Vibrio cholerae O139. In the present study, we investigated whether the vaccines can effectively protect immunized animals from any pathologic changes using histological, immunohistochemical and ultrastructural techniques. Severe pathology is evident in wild type injected ileum in non-immunized, showing extensive villous destruction, edema, necrosis and inflammation with infiltration of large numbers of inflammatory cells, extensive damage to the villi and microvilli with pore formation. Histology of ileum injected with wild type in immunized rabbit shows no significant pathological changes except for a few inflammatory cells in lamina propria with mild edema in mucosa and submucosa. immunohistochemical staining revealed O139 antigens of wild type are seen in the lamina propria of edematous villi, muscularis mucosa and submucosa with weak presence in the muscle coat in non-immunized rabbit after challenged with wild type in non-immunized rabbits, but in immunized rabbit localisation of the O139 LPS antigen is seen at the tips of the intact villi, within lamina propria and muscularis mucosa only. These observations suggest that the vaccine can effectively protect animals from any pathologic changes and eliminate V. cholerae O139 from the immunized animals.

Obtaining and Characteristic of Antibodies to Vibrio cholerae Protective Antigens Conjugated with Gold Nanoparticles.

Gold nanoparticle conjugates with Vibrio cholerae antigens were synthesized. The animals were immunized with the obtained conjugates. Rabbit polyclonal antibodies to the antigens were obtained, which showed high specific activity. On the model of white laboratory mice, the protective activity of conjugates of cholera antigens with nanoparticles during infection of vaccinated animals was evaluated using a commercial vaccine as a control. It was shown that in terms of immunogenicity, the created prototypes of cholera vaccine using gold nanoparticles as a carrier and adjuvant complied with the production regulations for the Russian national cholera chemical vaccine.

Efficacy of a Single-Dose, Inactivated Oral Cholera Vaccine in Bangladesh.

BACKGROUND: A single-dose regimen of the current killed oral cholera vaccines that have been prequalified by the World Health Organization would make them more attractive for use against endemic and epidemic cholera. We conducted an efficacy trial of a single dose of the killed oral cholera vaccine Shanchol, which is currently given in a two-dose schedule, in an urban area in which cholera is highly endemic. METHODS: Nonpregnant residents of Dhaka, Bangladesh, who were 1 year of age or older were randomly assigned to receive a single dose of oral cholera vaccine or oral placebo. The primary outcome was vaccine protective efficacy against culture-confirmed cholera occurring 7 to 180 days after dosing. Prespecified secondary outcomes included protective efficacy against severely dehydrating culture-confirmed cholera during the same interval, against cholera and severe cholera occurring 7 to 90 versus 91 to 180 days after dosing, and against cholera and severe cholera according to age at baseline. RESULTS: A total of 101 episodes of cholera, 37 associated with severe dehydration, were detected among the 204,700 persons who received one dose of vaccine or placebo. The vaccine protective efficacy was 40% (95% confidence interval [CI], 11 to 60%; 0.37 cases per 1000 vaccine recipients vs. 0.62 cases per 1000 placebo recipients) against all cholera episodes, 63% (95% CI, 24 to 82%; 0.10 vs. 0.26 cases per 1000 recipients) against severely dehydrating cholera episodes, and 63% (95% CI, -39 to 90%), 56% (95% CI, 16 to 77%), and 16% (95% CI, -49% to 53%) against all cholera episodes among persons vaccinated at the age of 5 to 14 years, 15 or more years, and 1 to 4 years, respectively, although the differences according to age were not significant (P=0.25). Adverse events occurred at similar frequencies in the two groups. CONCLUSIONS: A single dose of the oral cholera vaccine was efficacious in older children (≥5 years of age) and in adults in a setting with a high level of cholera endemicity. (Funded by the Bill and Melinda Gates Foundation and others; ClinicalTrials.gov number, NCT02027207.).

"It depends how one understands it:" a qualitative study on differential uptake of oral cholera vaccine in three compounds in Lusaka, Zambia.

BACKGROUND: The Zambian Ministry of Health implemented a reactive one-dose Oral Cholera Vaccine (OCV) campaign in April 2016 in three Lusaka compounds, followed by a pre-emptive second-round in December. Understanding uptake of this first-ever two-dose OCV campaign is critical to design effective OCV campaigns and for delivery of oral vaccines in the country and the region. METHODS: We conducted 12 Focus Group Discussions (FGDs) with men and women who self-reported taking no OCV doses and six with those self-reporting taking both doses. Simple descriptive analysis was conducted on socio-demographic and cholera-related data collected using a short questionnaire. We analyzed transcribed FGDs using the framework of dose, gender and geographic location. RESULTS: No differences were found by gender and location. All participants thought cholera to be severe and the reactive OCV campaign as relevant if efficacious. Most reported not receiving information on OCV side-effects and duration of protection. Those who took both doses listed more risk factors (including 'wind') and felt personally susceptible to cholera and protected by OCV. Some described OCV side-effects, mostly diarrhoea, vomiting and dizziness, as the expulsion of causative agents. Those who did not take OCV felt protected by their good personal hygiene practices or, thought of themselves and OCV as powerless against the multiple causes of cholera including poor living conditions, water, wind, and curse. Most of those who did not take OCV feared side-effects reported by others. Some interpreted side-effects as 'western' malevolence. Though > 80% discussants reported not knowing duration of protection, some who did not vaccinate, suggested that rather than rely on OCV which could lose potency, collective action should be taken to change the physical and economic environment to prevent cholera. CONCLUSIONS: Due to incomplete information, individual decision-making was complex, rooted in theories of disease causation, perceived susceptibility, circulating narratives, colonial past, and observable outcomes of vaccination. To increase coverage, future OCV campaigns may benefit from better communication on eligibility and susceptibility, expected side effects, mechanism of action, and duration of protection. Governmental improvements in the physical and economic environment may increase confidence in OCV and other public health interventions among residents in Lusaka compounds.

Safety of a bivalent, killed, whole-cell oral cholera vaccine in pregnant women in Bangladesh: evidence from a randomized placebo-controlled trial.

BACKGROUND: Cholera increases the risk of harmful effects on foetuses. We prospectively followed pregnant women unaware of their pregnancy status who received a study agent in a clinical trial evaluating the association between exposure to an oral cholera vaccine (OCV) and foetal survival. METHODS: Study participants were selected from a randomized placebo-controlled trial conducted in Dhaka, Bangladesh. The vaccination campaign was conducted between January 10 and February 4, 2014. We enrolled women who were exposed to an OCV or placebo during pregnancy (Cohort 1) and women who were pregnant after the vaccination was completed (Cohort 2). Our primary endpoint was pregnancy loss (spontaneous miscarriage or stillbirth), and the secondary endpoints were preterm delivery and low birth weight. We employed a log-binomial regression to calculate the relative risk of having adverse outcomes among OCV recipients compared to that among placebo recipients. RESULT: There were 231 OCV and 234 placebo recipients in Cohort 1 and 277 OCV and 299 placebo recipients in Cohort 2. In Cohort 1, the incidence of pregnancy loss was 113/1000 and 115/1000 among OCV and placebo recipients, respectively. The adjusted relative risk for pregnancy loss was 0.97 (95% CI: 0.58-1.61; p = 0.91) in Cohort 1. We did not observe any variation in the risk of pregnancy loss between the two cohorts. The risks for preterm delivery and low birth weight were not significantly different between the groups in both cohorts. CONCLUSIONS: Our study provides additional evidence that exposure to an OCV during pregnancy does not increase the risk of pregnancy loss, preterm delivery, or low birth weight, suggesting that pregnant women in cholera-affected regions should not be excluded in a mass vaccination campaign. TRIAL REGISTRATION: The study is registered at ( http://clinicaltrials.gov ). Identifier: NCT02027207 .

Progress and Challenges in Using Oral Cholera Vaccines to Control Outbreaks: The Médecins Sans Frontières Experience.

The use of oral cholera vaccine (OCV) has increased since 2011, when Shanchol, the first OCV suitable for large-scale use, became available. Médecins Sans Frontières considers OCVs an essential cholera outbreak control tool and has contributed to generating new evidence on OCV use in outbreaks. We showed that large-scale mass campaigns are feasible during outbreaks, documented high short-term effectiveness and showed that vaccines are likely safe in pregnancy. We found that a single-dose regimen has high short-term effectiveness, making rapid delivery of vaccine during outbreaks easier, especially given the on-going global vaccine shortage. Despite progress, OCV has still not been used widely in some of the largest recent outbreaks and thousands of cholera deaths are reported every year. While working towards improving our tools to protect those most at-risk of cholera, we must strive to use all available effective interventions in efficient ways, including OCV, to prevent avoidable deaths today.

Cholera: Immunity and Prospects in Vaccine Development.

Vibrio cholerae is a prototypical noninvasive mucosal pathogen, yet infection generates long-lasting protection against subsequent disease. Vibriocidal antibody responses are an imperfect but established correlate of protection against cholera following both infection and vaccination. However, vibriocidal antibody responses are likely a surrogate marker for longer-lasting functional immune responses that target the O-polysaccharide antigen at the mucosal surface. While the current bivalent inactivated oral whole cell vaccine is being increasingly used to prevent cholera in areas where the disease is a threat, the most significant limitation of this vaccine is it offers relatively limited direct protection in young children. Future strategies for cholera vaccination include the development of cholera conjugate vaccines and the further development of live attenuated vaccines. Ultimately, the goal of a multivalent vaccine for cholera and other childhood enteric infections that can be incorporated into a standard immunization schedule should be realized.

A Self-Assembling Whole-Cell Vaccine Antigen Presentation Platform.

Diarrhea is the most common infection in children under the age of 5 years worldwide. In spite of this, only a few vaccines to treat infectious diarrhea exist, and many of the available vaccines are sparingly and sporadically administered. Major obstacles to the development and widespread implementation of vaccination include the ease and cost of production, distribution, and delivery. Here we present a novel, customizable, and self-assembling vaccine platform that exploits the Vibrio cholerae bacterial biofilm matrix for antigen presentation. We use this technology to create a proof-of-concept, live-attenuated whole-cell vaccine that is boosted by spontaneous association of a secreted protein antigen with the cell surface. Sublingual administration of this live-attenuated vaccine to mice confers protection against V. cholerae challenge and elicits the production of antigen-specific IgA in stool. The platform presented here enables the development of antigen-boosted vaccines that are simple to produce and deliver, addressing many of the obstacles to vaccination against diarrheal diseases. This may also serve as a paradigm for the development of broadly protective biofilm-based vaccines against other mucosal infections.IMPORTANCE Diarrheal disease is the most common infection afflicting children worldwide. In resource-poor settings, these infections are correlated with cognitive delay, stunted growth, and premature death. With the development of efficacious, affordable, and easily administered vaccines, such infections could be prevented. While a major focus of research on biofilms has been their elimination, here we harness the bacterial biofilm to create a customizable platform for cost-effective, whole-cell mucosal vaccines that self-incorporate secreted protein antigens. We use this platform to develop a sublingually administered live-attenuated prototype vaccine based on Vibrio cholerae This serves not only as a proof of concept for a multivalent vaccine against common bacterial enteric pathogens but also as a paradigm for vaccines utilizing other bacterial biofilms to target mucosal infections.

Immunogenicity and Protection From a Single Dose of Internationally Available Killed Oral Cholera Vaccine: A Systematic Review and Metaanalysis.

In addition to improved water supply and sanitation, the 2-dose killed oral cholera vaccine (OCV) is an important tool for the prevention and control of cholera. We aimed to document the immunogenicity and protection (efficacy and effectiveness) conferred by a single OCV dose against cholera. The metaanalysis showed that an estimated 73% and 77% of individuals seroconverted to the Ogawa and Inaba serotypes, respectively, after an OCV first dose. The estimates of single-dose vaccine protection from available studies are 87% at 2 months decreasing to 33% at 2 years. Current immunologic and clinical data suggest that protection conferred by a single dose of killed OCV may be sufficient to reduce short-term risk in outbreaks or other high-risk settings, which may be especially useful when vaccine supply is limited. However, until more data suggest otherwise, a second dose should be given as soon as circumstances allow to ensure robust protection.

Cholera: recent updates.

PURPOSE OF REVIEW: In this review, we will examine updates in cholera epidemiology, advances in our understanding of pathogenesis and protective immunity, and changes to prevention strategies. RECENT FINDINGS: New modeling techniques and molecular epidemiology have led to advancements in our understanding of how Vibrio cholerae has persisted and re-emerged in new areas during the seventh pandemic. Use of next-generation sequencing has shed new light on immune responses to disease and vaccination, and the role of the gut microbiome in cholera. Increased efficacy and availability of vaccines have made long-term goals of global control of cholera more achievable. SUMMARY: Advancements in our understanding of immunity and susceptibility to V. cholerae, in addition to an increased global commitment to disease prevention, have led to optimism for the future of cholera prevention.

Efficacy of a bivalent killed whole-cell cholera vaccine over five years: a re-analysis of a cluster-randomized trial.

BACKGROUND: Oral cholera vaccine (OCV) is a feasible tool to prevent or mitigate cholera outbreaks. A better understanding of the vaccine's efficacy among different age groups and how rapidly its protection wanes could help guide vaccination policy. METHODS: To estimate the level and duration of OCV efficacy, we re-analyzed data from a previously published cluster-randomized, double-blind, placebo controlled trial with five years of follow-up. We used a Cox proportional hazards model and modeled the potentially time-dependent effect of age categories on both vaccine efficacy and risk of infection in the placebo group. In addition, we investigated the impact of an outbreak period on model estimation. RESULTS: Vaccine efficacy was 38% (95% CI: -2%,62%) for those vaccinated from ages 1 to under 5 years old, 85% (95% CI: 67%,93%) for those 5 to under 15 years, and 69% (95% CI: 49%,81%) for those vaccinated at ages 15 years and older. Among adult vaccinees, efficacy did not appear to wane during the trial, but there was insufficient data to assess the waning of efficacy among child vaccinees. CONCLUSIONS: Through this re-analysis we were able to detect a statistically significant difference in OCV efficacy when the vaccine was administered to children under 5 years old vs. children 5 years and older. The estimated efficacies are more similar to the previously published analysis based on the first two years of follow-up than the analysis based on all five years. TRIAL REGISTRATION: ClinicalTrials.gov identifier NCT00289224.

Immunity Provided by an Outer Membrane Vesicle Cholera Vaccine Is Due to O-Antigen-Specific Antibodies Inhibiting Bacterial Motility.

An outer membrane vesicle (OMV)-based cholera vaccine is highly efficacious in preventing intestinal colonization in the suckling mouse model. Immunity from OMVs comes from immunoglobulin (Ig), particularly IgG, in the milk of mucosally immunized dams. Anti-OMV IgG renders Vibrio cholerae organisms immotile, thus they pass through the small intestine without colonizing. However, the importance of motility inhibition for protection and the mechanism by which motility is inhibited remain unclear. By using both in vitro and in vivo experiments, we found that IgG inhibits motility by specifically binding to the O-antigen of V. cholerae We demonstrate that the bivalent structure of IgG, although not required for binding to the O-antigen, is required for motility inhibition. Finally, we show using competition assays in suckling mice that inhibition of motility appears to be responsible for most, if not all, of the protection engendered by OMV vaccination, thus providing insight into the mechanism of immune protection.

Conjugate Vaccines from Bacterial Antigens by Squaric Acid Chemistry: A Closer Look.

By using O-SP-core (O-SPcNH2 ) polysaccharide, isolated from Vibrio cholera O1 lipopolysaccharide (LPS) and related synthetic substances, a detailed study of factors that affect conjugation of bacterial polysaccharides to protein carriers through squaric acid chemistry to form conjugate vaccines has been carried out. Several previously unrecognized processes that take place during the squarate labeling of the O-SPcNH2 and subsequent conjugation of the formed squarate (O-SPcNH-SqOMe) have been identified. The efficiency of conjugation at pH 8.5, 9.0, and 9.5 to bovine serum albumin (BSA) and to the recombinant tetanus toxin fragment C (rTT-Hc) has been determined. The study led to a protocol for more efficient labeling of O-SPcNH2 antigen with the methyl squarate group, to yield a higher-quality, more potent squarate conjugation reagent. Its use resulted in about twofold increases in conjugation efficiency (from 23-26 % on BSA to 51 % on BSA and 55 % on rTT-Hc). The spent conjugation reagent could be recovered and regenerated by treatment with MeI in the absence of additional base. The immunological properties of the experimental vaccine made from the regenerated conjugation reagent were comparable with those of the immunogen made from the parent O-SPcNH-SqOMe.

Delivery cost analysis of a reactive mass cholera vaccination campaign: a case study of Shanchol™ vaccine use in Lake Chilwa, Malawi.

BACKGROUND: Cholera is a diarrheal disease that produces rapid dehydration. The infection is a significant cause of mortality and morbidity. Oral cholera vaccine (OCV) has been propagated for the prevention of cholera. Evidence on OCV delivery cost is insufficient in the African context. This study aims to analyze Shanchol vaccine delivery costs, focusing on the vaccination campaign in response of a cholera outbreak in Lake Chilwa, Malawi. METHODS: The vaccination campaign was implemented in two rounds in February and March 2016. Structured questionnaires were used to collect costs incurred for each vaccination related activity, including vaccine procurement and shipment, training, microplanning, sensitization, social mobilization and vaccination rounds. Costs collected, including financial and economic costs were analyzed using Choltool, a standardized cholera cost calculator. RESULTS: In total, 67,240 persons received two complete doses of the vaccine. Vaccine coverage was higher in the first round than in the second. The two-dose coverage measured with the immunization card was estimated at 58%. The total financial cost incurred in implementing the campaign was US$480275 while the economic cost was US$588637. The total financial and economic costs per fully vaccinated person were US$7.14 and US$8.75, respectively, with delivery costs amounting to US$1.94 and US$3.55, respectively. Vaccine procurement and shipment accounted respectively for 73% and 59% of total financial and economic costs of the total vaccination campaign costs while the incurred personnel cost accounted for 13% and 29% of total financial and economic costs. Cost for delivering a single dose of Shanchol was estimated at US$0.97. CONCLUSION: This study provides new evidence on economic and financial costs of a reactive campaign implemented by international partners in collaboration with MoH. It shows that involvement of international partners' personnel may represent a substantial share of campaign's costs, affecting unit and vaccine delivery costs.