Cholera Vaccine Research Articles

Assessment of Vaccine Herd Protection: Lessons Learned From Cholera and Typhoid Vaccine Trials.

Vaccine herd protection is the extension of the defense conferred by immunization beyond the vaccinated to unvaccinated persons in a population, as well as the enhancement of the protection among the vaccinated, due to vaccination of the surrounding population. Vaccine herd protection has traditionally been inferred from observations of disease trends after inclusion of a vaccine in national immunization schedules. Rather than awaiting outcomes of widescale vaccine deployment, earlier-stage evaluation of vaccine herd protection during trials or mass vaccination projects could help inform policy decisions about potential vaccine introduction. We describe the components, influencing factors, and implications of vaccine herd protection and discuss various methods for assessing herd protection, using examples from cholera and typhoid vaccine studies.

Defining Polysaccharide-Specific Antibody Targets against Vibrio cholerae O139 in Humans following O139 Cholera and following Vaccination with a Commercial Bivalent Oral Cholera Vaccine, and Evaluation of Conjugate Vaccines Targeting O139.

ABSTRACTCholera caused by Vibrio cholerae O139 could reemerge, and proactive development of an effective O139 vaccine would be prudent. To define immunoreactive and potentially immunogenic carbohydrate targets of Vibrio cholerae O139, we assessed immunoreactivities of various O-specific polysaccharide (OSP)-related saccharides with plasma from humans hospitalized with cholera caused by O139, comparing responses to those induced in recipients of a commercial oral whole-cell killed bivalent (O1 and O139) cholera vaccine (WC-O1/O139). We also assessed conjugate vaccines containing selected subsets of these saccharides for their ability to induce protective immunity using a mouse model of cholera. We found that patients with wild-type O139 cholera develop IgM, IgA, and IgG immune responses against O139 OSP and many of its fragments, but we were able to detect only a moderate IgM response to purified O139 OSP-core, and none to its fragments, in immunologically naive recipients of WC-O1/O139. We found that immunoreactivity of O139-specific polysaccharides with antibodies elicited by wild-type infection markedly increase when saccharides contain colitose and phosphate residues, that a synthetic terminal tetrasaccharide fragment of OSP is more immunoreactive and protectively immunogenic than complete OSP, that native OSP-core is a better protective immunogen than the synthetic OSP lacking core, and that functional vibriocidal activity of antibodies predicts in vivo protection in our model but depends on capsule thickness. Our results suggest that O139 OSP-specific responses are not prominent following vaccination with a currently available oral cholera vaccine in immunologically naive humans and that vaccines targeting V. cholerae O139 should be based on native OSP-core or terminal tetrasaccharide.IMPORTANCE Cholera is a severe dehydrating illness of humans caused by Vibrio cholerae serogroup O1 or O139. Protection against cholera is serogroup specific, and serogroup specificity is defined by O-specific polysaccharide (OSP). Little is known about immunity to O139 OSP. In this study, we used synthetic fragments of the O139 OSP to define immune responses to OSP in humans recovering from cholera caused by V. cholerae O139, compared these responses to those induced by the available O139 vaccine, and evaluated O139 fragments in next-generation conjugate vaccines. We found that the terminal tetrasaccharide of O139 is a primary immune target but that the currently available bivalent cholera vaccine poorly induces an anti-O139 OSP response in immunologically naive individuals.

A phase I/II study to evaluate safety, tolerability and immunogenicity of Hillchol®, an inactivated single Hikojima strain based oral cholera vaccine, in a sequentially age descending population in Bangladesh

BackgroundThe World Health Organization (WHO) recommends the use of oral cholera vaccines (OCVs) as part of an integrated control program, both in highly endemic settings and during cholera epidemics. The available and internationally recommended WHO-prequalified OCVs (Dukoral, Shanchol, Euvichol) contain multiple heat and formalin-killed V. cholerae strains of Inaba and Ogawa serotypes. MSD Wellcome Trust Hilleman Laboratories Pvt. Ltd. in technical collaboration with University of Gothenburg, Sweden has developed a new single strain OCV, Hillchol. This vaccine consists of formaldehyde-inactivated whole cell El Tor V. cholerae O1 bacteria engineered into the Hikojima serotype for stable expression of both the Ogawa (AB) and Inaba (AC) LPS antigens on the bacterial surface. We evaluated the safety and immunogenicity of this novel and potentially much less expensive OCV in comparison with Shanchol. MethodsWe conducted a randomized, non-inferiority, age-descending clinical trial of OCV (Hillchol vs. Shanchol) in the Mirpur area of Dhaka city from July 2016 to May 2017. This study was carried out in three different age cohorts (1–<5, 5–17 and ≥18 years old). Two doses of vaccine were given at 14 days intervals to 560 healthy participants. FindingsNo serious adverse events were reported. There were no significant differences in the rates of adverse events between the test vaccine (Hillchol) and the comparator (Shanchol) group. Serum vibriocidal antibody responses in all age groups combined were comparable for all the O1 Ogawa (59% vs. 67%; 90% CI of difference: −14.55, −0.84) and Inaba (70% vs. 71%; 90% CI of difference: −7.24, 5.77) serotypes, showing that the Hillchol vaccine was non-inferior to Shanchol. This new vaccine was also non-inferior to Shanchol in the different age strata. ConclusionThe safety and immunogenicity profile of the new OCV Hillchol is comparable to Shanchol in persons residing in a cholera-endemic setting. ClinicalTrials.gov number: NCT02823899.

Serum vibriocidal responses when second doses of oral cholera vaccine are delayed 6 months in Zambia

Two-dose killed oral cholera vaccines (OCV) are currently being used widely to control cholera. The standard dose-interval for OCV is 2 weeks; however, during emergency use of the vaccine, it may be more appropriate to use the available doses to quickly give a single dose to more people and give a delayed second dose when more vaccine becomes available.This study is an open label, randomized, phase 2 clinical trial of the vibriocidal response induced by OCV, comparing the responses when the second dose was given either 2 weeks (standard dose interval) or 6 months (extended dose interval) after the first dose. Vaccine was administered to healthy participants > 1 year of age living in the Lukanga Swamps area of Zambia. Three age cohorts (<5 years, 5–14 years, and ≥ 15 years) were randomized to the either dose-interval. The primary outcome was the vibriocidal GMT 14 days after the second dose.156 of 172 subjects enrolled in the study were included in this analysis. The Inaba vibriocidal titers were not significantly different 14 days post dose two for a standard dose-interval GMT: 45.6 (32–64.9), as compared to the GMT 47.6 (32.6–69.3), for the extended dose-interval, (p = 0.87). However, the Ogawa vibriocidal GMTs were significantly higher 14 days post dose two for the extended-dose interval at 87.6 (58.9–130.4) compared to the standard dose-interval group at 49.7 (34.1–72.3), p = 0.04. Vibriocidal seroconversion rates (a > 4-fold rise in vibriocidal titer) were not significantly different between dose-interval groups.This study demonstrated that vibriocidal titers 14 days after a second dose when given at an extended\\ dose interval were similar to the standard dose-interval. The findings suggest that a flexible dosing schedule may be considered when epidemiologically appropriate.The trial was registered at Clinical Trials.gov (NCT03373669).

CVD 103-HgR live, attenuated cholera vaccine strain viability in drinking waters from the US and Europe

CVD 103-HgR live, attenuated oral cholera vaccine strain is indicated for single dose immunization against Vibrio cholerae, the causative agent for cholera. The vaccine packets containing buffer powder and lyophilized CVD 103-HgR are reconstituted in water and consumed. Studies were performed to explore the viability of CVD 103-HgR in drinking waters from common sources. CVD 103-HgR vaccine was reconstituted in bottled and tap waters from the United States and Europe, and viability was measured via colony forming units assay. Chemical analysis of select water samples was used to identify chemicals that have a negative effect on CVD 103-HgR viability. CVD 103-HgR titers were stable in all bottled waters tested, including purified bottled water, bottled spring water, and sparkling waters. However, tap water from certain cities in the US and Europe affected viability and are not compatible with vaccine. Water chemistry revealed that these tap waters contained copper, likely leached from copper plumbing. These studies give high confidence in the stability of CVD 103-HgR reconstituted in a variety of bottled waters. Waters containing copper, including tap water, should not be used to reconstitute CVD 103-HgR strain oral vaccine due to the common use of copper plumbing.

Effectiveness of a killed whole-cell oral cholera vaccine in Bangladesh: further follow-up of a cluster-randomised trial

Killed whole-cell oral cholera vaccines (OCVs) are widely used for prevention of cholera in developing countries. However, few studies have evaluated the protection conferred by internationally recommended OCVs for durations beyond 2 years of follow-up. In this study, we followed up the participants of a cluster-randomised controlled trial for 2 years after the end of the original trial. Originally, we had randomised 90 geographical clusters in Dhaka slums in Bangladesh in equal numbers (1:1:1) to a two-dose regimen of OCV alone (targeted to people aged 1 year or older), a two-dose regimen of OCV plus a water-sanitation-hygiene (WASH) intervention, or no intervention. There was no masking of group assignment. The WASH intervention conferred little additional protection to OCV and was discontinued at 2 years of follow-up. Surveillance for severe cholera was continued for 4 years. Because of the short duration and effect of the WASH intervention, we combined the two OCV intervention groups. The primary outcomes were OCV overall protection (protection of all members of the intervention clusters) and total protection (protection of individuals who got vaccinated in the intervention clusters) against severe cholera, which we assessed by multivariable survival models appropriate for cluster-randomised trials. This trial is registered on ClinicalTrials.gov, NCT01339845. The study was done between April 17, 2011, and Nov 1, 2015. 268 896 participants were present at the time of the first dose, with 188 206 in the intervention group and 80 690 in the control group. OCV coverage of the two groups receiving OCV was 66% (123 659 of 187 214 participants). During 4 years of follow-up, 441 first episodes of severe cholera were detected (243 episodes in the vaccinated groups and as 198 episodes in the unvaccinated group). Overall OCV protection was 36% (95% CI 19 to 49%) and total OCV protection was 46% (95% CI 32 to 58). Cumulative total vaccine protection was notably lower for people vaccinated before the age of 5 years (24%; -30 to 56) than for people vaccinated at age 5 years or older (49%; 35 to 60), although the differences in protection for the two age groups were not significant (p=0·3308). Total vaccine protection dropped notably (p=0·0115) after 3 years in children vaccinated at 1-4 years of age. These findings provide further evidence of long-term effectiveness of killed whole-cell OCV, and therefore further support for the use of killed whole-cell OCVs to control endemic cholera, but indicate that protection is shorter-lived in children vaccinated before the age of 5 years than in people vaccinated at the age of 5 years or older. Bill & Melinda Gates Foundation. For the Bengali translation of the abstract see Supplementary Materials section.

Chitosan and alginate salt as biomaterials are potential natural adjuvants for killed cholera vaccine.

Chitosan and alginate salts are natural biopolymers that have gained recent attention in the biomedical sectors. Their properties allow them to become potential candidates as safe, cheap, and effective vaccine adjuvants. The present study aimed to enhance the immunogenic response of a current injectable killed cholera vaccine (KCV) using chitosan and alginate salt as natural adjuvants against alum. We tested KCV adjuvanted with alum, chitosan, and sodium alginate in mice. Mice were immunized intraperitoneally with KCV adjuvanted with alum, chitosan, or alginate salt and compared with a control unadjuvanted immunized group. Humoral, cellular, and functional immune responses were evaluated in all groups. The addition of adjuvants, particularly natural adjuvants, to KCV significantly improved the immune response as demonstrated by specific antibody increase, strong proliferation effects, and high protection rate against different challenge doses of cholera strains. Our findings demonstrate that chitosan and alginate salt are superior adjuvants for boosting the KCV immune response and highlights the requirement for further vaccine development.

Type II NKT Cell Agonist, Sulfatide, Is an Effective Adjuvant for Oral Heat-Killed Cholera Vaccines.

Oral vaccination has the potential to offer a safer and more efficacious approach for protection against enteric pathogens than injection-based approaches, especially in developing countries. One key advantage is the potential to induce intestinal immune responses in addition to systemic immunity. In general, antigen delivery via the oral route triggers weak immune responses or immunological tolerance. The effectiveness of oral vaccination can be improved by co-administering adjuvants. However, a major challenge is the absence of potent and safe oral adjuvants for clinical application. Here, the Type II NKT cell activator sulfatide is shown for the first time to be an effective oral adjuvant for Vibrio cholerae vaccine antigens in a mouse model. Specifically, administration of sulfatide with the oral cholera vaccine Dukoral® resulted in enhancement of intestinal antigen-specific IgA in addition to Th1 and Th17 immune responses. In summary, sulfatide is a promising adjuvant for inclusion in an oral cholera vaccine and our data further support the potential of adjuvants targeting NKT cells in new vaccine strategies.

Oral Cholera Vaccination as a Model to Assess Dietary Modulation of Gut Inflammation and Immunity

ObjectivesThe aim was to develop a human challenge model in which modulation of immune and inflammatory response by food ingredients can be evaluated. We hypothesized that oral cholera vaccination, in addition to inducing a specific antibody response, induces a significant increase in gut inflammatory response. MethodsTwenty healthy men (age 30.6 ± 1.8 y; BMI 24.9 ± 0.6) were enrolled in the study. Fecal and blood samples were collected at baseline. After a 2-week run-in period (D0-D14), subjects were vaccinated with oral cholera vaccine Dukoral (D15). After a period of 2 weeks, a second vaccination was administered (D29). Fecal samples were collected the day before (D14; D28) and the days (D16/D17; D30/D31) after each vaccination, as well as on D42. Blood samples were collected before vaccination (D15; D29), and on D16 and D17/D31, on D43. Primary outcome was fecal calprotectin concentration, secondary outcomes were serum levels of cholera toxin (CTB)-specific IgA and IgG. Other markers of local and systemic inflammatory response included beta-defensins, IP-10, IL-1 ra and hsCRP. Changes over time were tested by means of a linear mixed model. Outliers were identified with the 1.5xIQR rule and excluded from analysis. ResultsFecal calprotectin did not increase after the first vaccination. After the second vaccination, a significant increase was observed: from 12.8 ± 2.5 μg/g feces (mean ± SEM) on day 29 to 18.0 ± 2.9 μg/g on day 31 (P = 0.017). Plasma CTB-specific IgA and IgG were strongly increased after the first vaccination, with a further increase after the second vaccination. Plasma CRP slightly decreased on D17, compared to D15 (P = 0.016). IL-1ra significantly decreased 2 days after the first vaccination (P = 0.011), whereas no change was observed after the second vaccination. Beta-defensin was significantly increased at D31 compared to D29 (from 42.7 ± 7.0 to 80.7 ± 16.1 (P = 0.014)). IP-10 did not show any response to vaccination. ConclusionsIn addition to the expected antibody response, oral cholera vaccination induces an increase in fecal calprotectin and beta defensin, pointing to vaccine induced intestinal inflammation. These readouts may be added to intervention studies with dietary compounds to evaluate the potential for modulating immune responsiveness. Funding SourcesBio&Medical Technol Developm Program of the Natl Res Foundation, Min Science & ICT of Rep Korea.

Repurposing infectious diseases vaccines against cancer.

e14564 Background: Vaccines used to prevent infection have long been known to stimulate immune responses to cancer as illustrated by the approval of the Bacillus Calmette–Guérin (BCG) vaccine to treat bladder cancer since the 1970s. The recent approval of immunotherapies has rejuvenated this research area with reports of anti-tumour responses with existing infectious diseases vaccines used as such, either alone or in combination with approved immune checkpoint inhibitors. This prompted us to review recent research activities using approved vaccines to treat cancer. Methods: PubMed and clinicaltrials.gov databases were queried for all 31 infectious diseases having at least one preventive vaccine approved, and all hits were reviewed for relevance. Only articles and trial registrations about infectious diseases vaccines used in an unmodified form as treatment for cancer, were included. The search was restricted in time, capturing articles from the year 2000 until November 2020. Results: A total of 1,754 PubMed abstracts and 549 trial registrations were screened. Data supporting a cancer therapeutic use was found for 16 of the 31 infectious diseases vaccines. For 10 (BCG, diphtheria, tetanus, human papillomavirus (HPV), influenza, measles, pneumococcus, smallpox, typhoid and varicella-zoster), clinical trials have been conducted or are ongoing. Within the remaining 6, preclinical evidence supports further evaluation of the rotavirus, yellow fever and pertussis vaccine in carefully designed clinical trials. The mechanistic evidence for the cholera vaccine, combined with the observational data in colorectal cancer, is also supportive of clinical translation. There is limited data for the hepatitis B vaccine and for the mumps vaccine alone. Four findings are worth highlighting: first, the superiority of intra-vesical typhoid vaccine instillations over BCG instillations in a preclinical bladder cancer model, which is now the subject of a phase I trial; second, the perioperative use of the influenza vaccine to limit and prevent the NK cell dysfunction induced by cancer surgery; third, objective responses following intra-tumoral injections of the measles vaccine in cutaneous T-cell lymphoma; fourth, objective responses induced by the HPV vaccine in skin squamous cell carcinoma. Conclusions: Sixteen infectious diseases vaccines have been or are currently being explored for repurposing in oncology. All vaccines are intended to induce or improve an anti-tumour (immune) response. Next to biological and immunological mechanisms, that are very different from one vaccine to another, also the mode of administration and combinations and sequence with other (immuno-)therapies deserve more attention in future research.

Repurposing Infectious Diseases Vaccines Against Cancer.

Vaccines used to prevent infections have long been known to stimulate immune responses to cancer as illustrated by the approval of the Bacillus Calmette–Guérin (BCG) vaccine to treat bladder cancer since the 1970s. The recent approval of immunotherapies has rejuvenated this research area with reports of anti-tumor responses with existing infectious diseases vaccines used as such, either alone or in combination with immune checkpoint inhibitors. Here, we have reviewed and summarized research activities using approved vaccines to treat cancer. Data supporting a cancer therapeutic use was found for 16 vaccines. For 10 (BCG, diphtheria, tetanus, human papillomavirus, influenza, measles, pneumococcus, smallpox, typhoid and varicella-zoster), clinical trials have been conducted or are ongoing. Within the remaining 6, preclinical evidence supports further evaluation of the rotavirus, yellow fever and pertussis vaccine in carefully designed clinical trials. The mechanistic evidence for the cholera vaccine, combined with the observational data in colorectal cancer, is also supportive of clinical translation. There is limited data for the hepatitis B and mumps vaccine (without measles vaccine). Four findings are worth highlighting: the superiority of intravesical typhoid vaccine instillations over BCG in a preclinical bladder cancer model, which is now the subject of a phase I trial; the perioperative use of the influenza vaccine to limit and prevent the natural killer cell dysfunction induced by cancer surgery; objective responses following intratumoral injections of measles vaccine in cutaneous T-cell lymphoma; objective responses induced by human papillomavirus vaccine in cutaneous squamous cell carcinoma. All vaccines are intended to induce or improve an anti-tumor (immune) response. In addition to the biological and immunological mechanisms that vary between vaccines, the mode of administration and sequence with other (immuno-)therapies warrant more attention in future research.

Cholera Hot-Spots and Contextual Factors in Burundi, Planning for Elimination.

The Republic of Burundi first reported cholera cases in 1978 and outbreaks have been occurring nearly every year since then. From 2008–2020, 6949 cases and 43 deaths were officially reported. To evaluate Burundi’s potential to eliminate cholera, we identified hotspots using cholera incidence and disease persistence as suggested by the Global Task Force for Cholera Control. The mean annual incidence for each district that reported cholera ranged from 0.29 to 563.14 cases per 100,000 population per year from 2014–2020. Ten of 12 Health Districts which recorded cholera cases reported a mean annual incidence ≥5 per 100,000 for this time period. Cholera cases occur during the second half of the year in the areas near Lake Tanganyika and along the Ruzizi River, with the highest risk district being Bujumbura Centre. Additional research is needed to understand the role of Lake Tanganyika; risks associated with fishing; migration patterns; and other factors that may explain cholera’s seasonality. Due to the consistent epidemiological pattern and the relatively small area affected by cholera, control and elimination are feasible with an integrated program of campaigns using oral cholera vaccine over the short term and community-based interventions including WASH activities for sustained control.

A dysbiotic gut microbiome suppresses antibody-mediated protection against Vibrio cholerae

Abstract Vibrio cholerae is the etiologic agent of cholera, a severe diarrheal disease that represents a significant burden on global health and productivity. Given the need for more effective prophylactics, more research is needed to ascertain the interplay of protective immune responses and the resident gut microbiome, which in cholera-endemic areas is strongly and repeatedly modulated by malnutrition, cholera, and non-cholera infectious diarrhea. Initially, we conducted fecal transplants from healthy human donors into germ-free mice in order to characterize differential immune response activity. As we observed distinctive responses in our initial sampling, we moved to a more tractable antibiotic-treated mouse model where we assemble representative model communities of either human gut microbes resembling those of healthy individuals or those of individuals recovering from diarrhea or malnutrition. We establish these communities in a murine infection model, and show that the dysbiotic gut microbiome, commonly present in areas where malnutrition and diarrhea are common, suppresses the immune response against Vibrio cholerae through the action of CD4+ cells. Our findings suggest that the composition of the gut microbiome at time of infection may be pivotal for providing robust mucosal immunity, suggesting a target for the improvement of responses to oral cholera vaccines.

Immune Responses Induced in Chickens by Capsular Extract of P. Multocida Isolated From Farm Rat

An experiment was conducted to investigate the immune response induced in chickens by capsular extract of Pasteurella multocida isolated from rats wandering in and around the poultry farms. The rat isolate of P. multocida was isolated and identified by cultural, morphological, and biochemical characteristics, followed by capsular extract preparation and experimental vaccine development. The isolated P. multocida was found Gram-negative, non-motile, non-spore forming rod occurring singly or pains and occasionally as chains or filaments in Gram’s-staining method. The isolates consistently produced acid from dextrose, sucrose and mannitol but not fermented maltose or lactose. The Capsular antigen was extracted and confirmed by acriflavine test. Finally, experimental fowl cholera vaccine was prepared. Primary vaccination was performed at the dose rate of 5.6×107 CFU/ml through intramuscular and subcutaneous routes in birds of group A (10 birds) and group B (10 birds) and group C (10 birds) were control birds. Secondary vaccination was similarly performed after 15 days of primary vaccination in groups A and B. The levels of pre-vaccination and post-vaccination sera were determined by passive haemagglutination test. The passive haemagglutination antibody titre was recorded on 15 and 35 days of post vaccination in groups A and B. It was demonstrated that experimental capsular extract fowl cholera vaccine conferred 100% protection (p&lt;0.01) against challenge infection and found to be safe. It could be suggested that after thorough field trial, the experimentally prepared capsular extract FC vaccine using rat isolate of P. multocida may be used side by side with conventional FC vaccine.&#x0D; Res. Agric., Livest. Fish.8(1): 117-124, April 2021

An Update on Cholera Immunity and Current and Future Cholera Vaccines

Individual resistance to cholera infection and disease depends on both innate host factors and adaptive immunity acquired by a previous infection or vaccination. Locally produced, intestinal-mucosal secretory IgA (SIgA) antibodies against bacterial surface lipopolysaccharide (LPS) O antigens and/or secreted cholera toxins are responsible for the protective adaptive immunity, in conjunction with an effective mucosal immunologic memory that can elicit a rapid anamnestic SIgA antibody response upon re-exposure to the antigen/pathogen even many years later. Oral cholera vaccines (OCVs), based on inactivated Vibrio cholerae whole-cell components, either together with the cholera toxin B subunit (Dukoral™) or administered alone (Shanchol™/Euvichol-Plus™) were shown to be consistently safe and effective in large field trials in all settings. These OCVs are recommended by the World Health Organisation (WHO) for the control of both endemic cholera and epidemic cholera outbreaks. OCVs are now a cornerstone in WHO’s global strategy found in “Ending Cholera: A Global Roadmap to 2030.” However, the forecasted global demands for OCV, estimated by the Global Alliance for Vaccines and Immunization (GAVI) to 1.5 billion doses for the period 2020–2029, markedly exceed the existing manufacturing capacity. This calls for an increased production capacity of existing OCVs, as well as the rapid introduction of additional and improved vaccines under development.

Impact of Immunoglobulin Isotype and Epitope on the Functional Properties of Vibrio cholerae O-Specific Polysaccharide-Specific Monoclonal Antibodies.

Vibrio cholerae causes the severe diarrheal disease cholera. Clinical disease and current oral cholera vaccines generate antibody responses associated with protection. Immunity is thought to be largely mediated by lipopolysaccharide (LPS)-specific antibodies, primarily targeting the O-antigen. However, the properties and protective mechanism of functionally relevant antibodies have not been well defined. We previously reported on the early B cell response to cholera in a cohort of Bangladeshi patients, from which we characterized a panel of human monoclonal antibodies (MAbs) isolated from acutely induced plasmablasts. All antibodies in that previous study were expressed in an IgG1 backbone irrespective of their original isotype. To clearly determine the impact of affinity, immunoglobulin isotype and subclass on the functional properties of these MAbs, we re-engineered a subset of low- and high-affinity antibodies in different isotype and subclass immunoglobulin backbones and characterized the impact of these changes on binding, vibriocidal, agglutination, and motility inhibition activity. While the high-affinity antibodies bound similarly to O-antigen, irrespective of isotype, the low-affinity antibodies displayed significant avidity differences. Interestingly, despite exhibiting lower binding properties, variants derived from the low-affinity MAbs had comparable agglutination and motility inhibition properties to the potently binding antibodies, suggesting that how the MAb binds to the O-antigen may be critical to function. In addition, not only pentameric IgM and dimeric IgA, but also monomeric IgA, was remarkably more potent than their IgG counterparts at inhibiting motility. Finally, analyzing highly purified F(ab) versions of these antibodies, we show that LPS cross-linking is essential for motility inhibition.IMPORTANCE Immunity to the severe diarrheal disease cholera is largely mediated by lipopolysaccharide (LPS)-specific antibodies. However, the properties and protective mechanisms of functionally relevant antibodies have not been well defined. Here, we have engineered low and high-affinity LPS-specific antibodies in different immunoglobulin backbones in order to assess the impact of affinity, immunoglobulin isotype, and subclass on binding, vibriocidal, agglutination, and motility inhibition functional properties. Importantly, we found that affinity did not directly dictate functional potency since variants derived from the low-affinity MAbs had comparable agglutination and motility inhibition properties to the potently binding antibodies. This suggests that how the antibody binds sterically may be critical to function. In addition, not only pentameric IgM and dimeric IgA, but also monomeric IgA, was remarkably more potent than their IgG counterparts at inhibiting motility. Finally, analyzing highly purified F(ab) versions of these antibodies, we show that LPS cross-linking is essential for motility inhibition.

Parenteral Vaccination with a Cholera Conjugate Vaccine Boosts Vibriocidal and Anti-OSP Responses in Mice Previously Immunized with an Oral Cholera Vaccine.

.Oral cholera vaccination protects against cholera; however, responses in young children are low and of short duration. The best current correlates of protection against cholera target Vibrio cholerae O-specific polysaccharide (anti-OSP), including vibriocidal responses. A cholera conjugate vaccine has been developed that induces anti-OSP immune responses, including memory B-cell responses. To address whether cholera conjugate vaccine would boost immune responses following oral cholera vaccination, we immunized mice with oral cholera vaccine Inaba CVD 103-HgR or buffer only (placebo) on day 0, followed by parenteral boosting immunizations on days 14, 42, and 70 with cholera conjugate vaccine Inaba OSP: recombinant tetanus toxoid heavy chain fragment or phosphate buffered saline (PBS)/placebo. Compared with responses in mice immunized with oral vaccine alone or intramuscular cholera conjugate vaccine alone, mice receiving combination vaccination developed significantly higher vibriocidal, IgM OSP-specific serum responses and OSP-specific IgM memory B-cell responses. A combined vaccination approach, which includes oral cholera vaccination followed by parenteral cholera conjugate vaccine boosting, results in increased immune responses that have been associated with protection against cholera. These results suggest that such an approach should be evaluated in humans.

Ways to Reduce the Level of Contamination at the Stages of Tableted Chemical Cholera Vaccine Production

Objective of the study was an assessment of the degree of contamination of cholera chemical vaccine at the stages of preparation and determination of the ways to reduce it.Materials and methods. Liquid and lyophilized components of the cholera chemical vaccine used in the study: cholerogen-anatoxin and O-antigens of Vibrio cholerae 569B and V. cholerae M-41 strains, as well as auxiliary substances (sucrose, talc, calcium stearate, starch). Granulation was carried out in a device that works on a fluidized bed principle, GPCG 2 (GLATT, Germany). Subsequent tabletizing of the mixture was performed using MiniTabT compression machine (LUXNER, Germany). Studies were conducted on the evaluation of “microbiological purity” at the stages of manufacturing of the cholera chemical vaccine, tablets coated with an enteric coating. Positive or negative growth of microorganisms on Petri dishes with nutrient media was determined on visual inspection.Results and conclusions. The dynamics of changes in microbial contamination at certain technological stages of vaccine production has been revealed. It is shown that the solutions of antigens in the process of separation are subject to microbial contamination which is associated with the use of ammonium sulfate during precipitation and non-sterile water at the stage of dialysis. Sterility of semi-finished products has been achieved through twophase filtration of choleragen-anatoxin and sterilization of O-antigens of V. cholerae 569B and V. cholerae M-41 strains with flowing steam at (100±1) °C for 30 minutes. In order to decrease microbial contamination at the stage of granulation additional fine filters were installed in the air-supply system. Further on comparative assessment of microbial purity of vaccine batches obtained using both, direct compression and preliminary granulation, was carried out. It has been experimentally demonstrated that granulation of the components of a tablet mixture of cholera vaccine leads to a decrease in the level of bacterial contamination and improves the microbiological purity of the finished dosage form.

Genetic Characteristics of Antibiotic Resistance Gene of Vibrio cholerae Isolated

Vibrio cholerae is the factor of cholera disease in human and it has been becoming a serious problem in human health. This study was conducted to identify the genetic characteristics and antibiotic resistance of V. cholerae isolated in Tra Vinh Province. From the results, twenty-five (25) Vibrio spp. were isolated, including 6 strains of Vibrio (V.) cholerae (24%), 8 strains of V. paraheamolyticus (32%), 4 strains of V. vulnificus (16%), 5 strains of V. fluvialis (20%), and 2 strains of V. alginolyticus (8%). The serogroup results showed that all the 6 V. cholerae belonged to serogroup O1, with 50% positive to serotype Inaba, and 50% positive to Ogawa. No strain belonged to serogroup O139. The results also showed that 50% of V. cholerae were resistant to streptomycin and 33% were resistant to tetracycline and trimethoprim-sulfamethoxazole. Two of the 6 strains had an antibiotic resistance gene (gene tetA) encoding for a tetracycline-resistant factor. Gene blaSHV, gene aac(3)-IV and gene dhfrI encoding for β-lactam, aminoglycoside, trimethoprim resistant factors were not detected. Additionally, the results showed that adhesion indexes of V. cholerae strains T1 and T3 in the intestinal mucosa of rabbits shared the same antigens with the bacteria used to produce cholera vaccine (mORCVAX) available in Vietnam. V. cholerae was prevalent in the environment and clams in Tra Vinh province. Information from this research may be useful for further studies.

Inverse probability weighted estimators of vaccine effects accommodating partial interference and censoring.

Estimating population-level effects of a vaccine is challenging because there may be interference, that is, the outcome of one individual may depend on the vaccination status of another individual. Partial interference occurs when individuals can be partitioned into groups such that interference occurs only within groups. In the absence of interference, inverse probability weighted (IPW) estimators are commonly used to draw inference about causal effects of an exposure or treatment. Tchetgen Tchetgen and VanderWeele proposed a modified IPW estimator for causal effects in the presence of partial interference. Motivated by a cholera vaccine study in Bangladesh, this paper considers an extension of the Tchetgen Tchetgen and VanderWeele IPW estimator to the setting where the outcome is subject to right censoring using inverse probability of censoring weights (IPCW). Censoring weights are estimated using proportional hazards frailty models. The large sample properties of the IPCW estimators are derived, and simulation studies are presented demonstrating the estimators' performance in finite samples. The methods are then used to analyze data from the cholera vaccine study.