Non-toxigenic Vibrio cholerae challenge strains for evaluating vaccine efficacy and inferring mechanisms of protection

Human challenge studies are instrumental for testing cholera vaccines, but these studies use outdated strains and require inpatient facilities. Here, we created next-generation isogenic Ogawa and Inaba V. cholerae challenge strains (ZChol strains) derived from a contemporary Zambian clinical isolate representative of current dominant pandemic V. cholerae. To minimize the risk of severe diarrhea these strains were rendered non-toxigenic, since antibody responses which limit V. cholerae colonization are the primary mechanism of immune protection. These strains did not cause diarrhea in infant mice and proved to accurately gauge reduction in intestinal colonization mediated by effective vaccination. They are also valuable as targets for measuring vibriocidal antibody responses. Using barcoded ZChol strains, we discovered that vaccination tightens the infection bottleneck without restricting pathogen expansion in vivo. ZChol strains have the potential to enhance the safety, relevance, and scope of future cholera vaccine challenge studies and be valuable reagents for studies of immunity to cholera.

Effect of HIV status and retinol on immunogenicity to oral cholera vaccine in adult population living in an endemic area of Lukanga Swamps, Zambia

Background We set out to assess the impact of human immunodeficiency virus (HIV) and micronutrient deficiency as indicated by serum retinol levels on the immune responses to Oral Cholera Vaccine (Shanchol™) in a cohort of participants in Lukanga Swamps, Zambia. Cholera remains endemic in Zambia with vaccines being the only effective preventive measures. However, the effect of these vaccines on populations living with HIV has not been widely documented. Methods HIV testing and confirmation was done using the Alere Determine™ HIV-1/2 and Uni-Gold™ kits while vibriocidal antibody assay was applied for vaccine immunogenicity. Serum retinol analysis was assessed by Shimadzu Prominence HCT-2010 High Performance Liquid Chromatography (HPLC). The primary outcome was log transformed geometric mean titre. Results From 47 participants screened for HIV, 51% (24) tested positive. There was a statistically significant reduction in Ogawa geometric mean ratio (GMR) by 67% (GMR = 0.33; 95% CI: -0.15, 0.76; p-value = 0.009) attributable to HIV positivity with a non-significant reduction in Inaba GMR by about 50% due to HIV positivity. When doubling of retinol levels modelled, GMR reduction against Ogawa were non-significant but that against Inaba resulted in a significant reduction in geometric mean titer (GMT) (GMT-0.33, C.I 0.16–0.66, p-value 0.002). At 1000copies/ml viral load cut off and 350 cells/μl CD4 counts, Ogawa GMT was two times higher 11.16 (95%CI: 8.20–15.19) versus 6.06 (95%CI: 4.04–9.10) in low viremia participants, and three times higher in above threshold CD4 count participants; 24.81 (95%CI: 18.94–32.50) versus 7.07 (95%CI: 5.22–9.58). Conclusion Our results show that while Shanchol™ is immunogenic in both HIV+/- individuals, HIV + participants responded poorly. Viral load and CD4 count affected vaccine immunogenicity. More research is required for detailed understanding of this in order to appropriately inform policy and practice.

Optimizing one-dose and two-dose cholera vaccine allocation in outbreak settings: A modeling study

Background: A global stockpile of oral cholera vaccine (OCV) was established in 2013 for use in outbreak response and are licensed as two-dose regimens. Vaccine availability, however, remains limited. Previous studies have found that a single dose of OCV may provide substantial protection against cholera. Methods: Using a mathematical model with two age groups paired with optimization algorithms, we determine the optimal vaccination strategy with one and two doses of vaccine to minimize cumulative overall infections, symptomatic infections, and deaths. We explore counterfactual vaccination scenarios in three distinct settings: Maela, the largest refugee camp in Thailand, with high in- and out-migration; N'Djamena, Chad, a densely populated region; and Haiti, where departments are connected by rivers and roads. Results: Over the short term under limited vaccine supply, the optimal strategies for all objectives prioritize one dose to the older age group (over five years old), irrespective of setting and level of vaccination coverage. As more vaccine becomes available, it is optimal to administer a second dose for long-term protection. With enough vaccine to cover the whole population with one dose, the optimal strategies can avert up to 30% to 90% of deaths and 36% to 92% of symptomatic infections across the three settings over one year. The one-dose optimal strategies can avert 1.2 to 1.8 times as many cases and deaths as a two-dose pro-rata strategy. Conclusions: In an outbreak setting, speedy vaccination campaigns with a single dose of OCV may avert more cases and deaths than a two-dose pro-rata campaign under a limited vaccine supply.

IMPROVEMENT OF SPECIFIC CHOLERA PREVENTION USING IMMUNOMODULATORS

It is known that the combined use of vaccines, cytokines and various immunomodulatory drugs contributes to the development of a full-fledged immune response. This approach makes it possible to enhance the immunogenicity of modern vaccines and to direct the development of immune responses according to the humoral or cellular type, depending on the properties of the pathogen of a particular disease. The improvement of preventive drugs due to their combined use with cytokines and immunomodulators increases the intensity of immunity, increases the level of production of specific immunoglobulins, the protectivity of antigens, and also reduces the manifestation of adverse reactions leading to post-vaccination complications.Immunomodulators are already successfully used in drugs intended for the treatment and prevention of chronic herpes infections and influenza vaccines. Numerous experimental and clinical data indicate a positive effect of the use of immunomodulatory drugs in the vaccination of various viral and bacterial infections, including particularly dangerous ones.Improving the specific prevention of cholera can be achieved through immunomodulatory agents that can stimulate the formation of a local and systemic immune response.We conducted a comparative assessment of the feasibility of the combined use of the cholera bivalent chemical vaccine (the Federal Government Health Institution Antiplague Research Institute “Microbe”) and immunomodulators in order to increase the effectiveness of cholera vaccination.Since the cholera vaccine causes the activation of the humoral immune response, the production of specific immunoglobulins in the body of vaccinated experimental animals and the effect of immunomodulators on this process at different times of the post-vaccination period were evaluated.The ability of immunomodulators to increase the protective activity of the cholera vaccine was studied by infecting animals with a virulent strain of cholera one month and seven months after vaccination.It was found that immunomodulators increase the immunogenicity and protectivity of antigens that are part of the anti-cholera vaccine. The use of all immunopreparations increases the production of specific immunoglobulins in the serum of vaccinated experimental animals. It was shown that in the first month after vaccination, polyoxidonium most effectively stimulated the formation of antibodies, but lycopide contributed to a longer retention of anti-cholera immunoglobulins in the serum of vaccinated rabbits. The combined use of the vaccine and lycopide prevented the development of cholera in all animals taken in the experiment, including those vaccinated with a reduced dose. In the long-term post-vaccination period, this immunomodulator increased the protectiveness of the anti-cholera vaccine by three times. Polyoxidonium and derinate also increased the protective effect of the cholera vaccine, but were slightly inferior to lycopide. The combined use of cholera vaccine and immunomodulators, especially lycopide, can be used to improve specific cholera prevention.

Ecological safety and prospects of development of low-waste technologies in the biotechnology industry

Environmental pollution with industrial waste is an urgent problem today. A special place in the list of pollutants belongs to waste from biotechnological enterprises and industries, whose activities are related to the production of various drugs. Russian Research Anti-Plague Institute «Microbe» is the only manufacturer of unique immunobiological drugs in the Russian Federation – bivalent chemical cholera vaccine and rabies immunoglobulin from horse blood serum (AIG). At present, the institute actively uses fibrin as a basis for nutrient media – a waste in the production of AIG; a technology for the regeneration of alcohol waste has been developed; biologically active substances were obtained from the production waste of specific components of the cholera vaccine. The aim of the work was to assess the prospects of using waste products from the production of specific components of cholera vaccine (cholerogen-toxoid – X-AT, and O-antigen – O-AG) – formalized detoxified microbial-free filtrate (FMF), as a nutrient medium for the cultivation of industrial strains of microorganisms. It has been shown that the best methods for reducing formalin concentration are autoclaving and chemical neutralization with aqueous ammonia. During low-volume cultivation of Vibrio cholerae 569B and V. cholerae M-41 strains on all variants of experimental media based on PBP, an increase in biomass was noted. The production of Vibrio cholerae antigens at a level comparable to that of growing on a control nutrient medium was recorded in a medium variant based on O-AG production waste. The use of FMF as a nutrient medium in the future will reduce the volume of waste generated and reduce the load on the treatment facilities of the Institute, which will increase the environmental safety of production.

Longitudinal analysis of human humoral responses after vaccination with a live attenuated V. cholerae vaccine

Vibrio cholerae is a bacterial pathogen which causes the severe acute diarrheal disease cholera. Given that a symptomatic incident of cholera can lead to long term protection, a thorough understanding of the immune response to this pathogen is needed to identify parameters critical to the generation and durability of immunity. To approach this, we utilized a live attenuated cholera vaccine to model the response to V. cholerae infection in 12 naïve subjects. We found that this live attenuated vaccine induced durable vibriocidal antibody titers that were maintained at least one year after vaccination. Similar to what we previously reported in infected patients from Bangladesh, we found that vaccination induced plasmablast responses were primarily specific to the two immunodominant antigens lipopolysaccharide (LPS) and cholera toxin (CT). Interestingly, the magnitude of the early plasmablast response at day 7 predicted the serological outcome of vaccination at day 30. However, this correlation was no longer present at later timepoints. The acute responses displayed preferential immunoglobulin isotype usage, with LPS specific cells being largely IgM or IgA producing, while cholera toxin responses were predominantly IgG. Finally, CCR9 was highly expressed on vaccine induced plasmablasts, especially on IgM and IgA producing cells, suggesting a role in migration to the gastrointestinal tract. Collectively, these findings demonstrate that the use of a live attenuated cholera vaccine is an effective tool to examine the primary and long-term immune response following V. cholerae exposure. Additionally, it provides insight into the phenotype and specificity of the cells which likely return to and mediate immunity at the intestinal mucosa. A thorough understanding of these properties both in peripheral blood and in the intestinal mucosae will inform future vaccine development against both cholera and other mucosal pathogens. Trial Registration: NCT03251495.

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

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).

The 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 from 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 and OSP-specific responses, but individuals with a higher abundance of Clostridiales and lower 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-1β 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 that the gut microbiota could impact immune responses to OCVs, and further study of microbial metabolites as potential vaccine adjuvants is warranted.