An experimental study of the safety of a chemical monovalent tableted cholera vaccine in enteral administration]

The safety of experimental chemical cholera monovalent vaccine in tablets, produced by the institute "Microbe" (Saratov, USSR), has been studied. The study has shown that the vaccine, administered to adult rabbits and germ-free suckling rabbits by the enteral route, retains residual toxicity, mainly due to the presence of O-antigen. One or two administrations of 1-2 human doses of this preparation to adult rabbits induce minimal structural changes admissible from the viewpoint of safety. After immunization made in two administrations immunobiological transformation develops more rapidly and is more pronounced than after immunization in a single administration.     

Systemic immunity of experimental animals immunized with cholera vaccines

The levels of antitoxic and vibriocidal antibodies in the sera of suckling rabbits after their parenteral immunization with cholera vaccine, cholera toxoid and a combination of cholera vaccine and toxoid were examined. Cholera vaccine induces intensive production of vibriocidal antibodies, and cholera toxoid, of antitoxic antibodies. The parenteral administration of the serum of rabbits immunized with cholera toxoid neutralized the action of cholera toxin in the small intestine of suckling rabbits. The complex preparation combines the properties of the corpuscular vaccine and the toxoid, inducing the production of both vibriocidal and antitoxic antibodies.     

Experimental research on cholera vaccines. Local immunity

Parenteral immunization of rabbits with cholera vaccine decreased the number of Vibrio cholerae adhering to the mucous membrane of the small intestine. Cholera toxoid and the complex preparation ensure protection from the local action of cholera toxin on the ligated loop of the rabbit intestine, while cholera vaccine produces no effect under the same conditions. The use of three preparations under study leads to the decrease of exudative reaction to the introduction of live V. cholerae, the effectiveness of these vaccines growing in the following order: cholera vaccine, cholera toxoid, the complex preparation.     

A Cholera Conjugate Vaccine Containing O-specific Polysaccharide (OSP) of V. cholerae O1 Inaba and Recombinant Fragment of Tetanus Toxin Heavy Chain (OSP:rTTHc) Induces Serum,Memory and Lamina Proprial Responses against OSP and Is Protective in Mice

Background

Vibrio cholerae is the cause of cholera, a severe watery diarrhea. Protection against cholera is serogroup specific. Serogroup specificity is defined by the O-specific polysaccharide (OSP) component of lipopolysaccharide (LPS).

Methodology

Here we describe a conjugate vaccine for cholera prepared via squaric acid chemistry from the OSP of V. cholerae O1 Inaba strain PIC018 and a recombinant heavy chain fragment of tetanus toxin (OSP:rTTHc). We assessed a range of vaccine doses based on the OSP content of the vaccine (10-50 μg), vaccine compositions varying by molar loading ratio of OSP to rTTHc (3:1, 5:1, 10:1), effect of an adjuvant, and route of immunization.

Principle Findings

Immunized mice developed prominent anti-OSP and anti-TT serum IgG responses, as well as vibriocidal antibody and memory B cell responses following intramuscular or intradermal vaccination. Mice did not develop anti-squarate responses. Intestinal lamina proprial IgA responses targeting OSP occurred following intradermal vaccination. In general, we found comparable immune responses in mice immunized with these variations, although memory B cell and vibriocidal responses were blunted in mice receiving the highest dose of vaccine (50 μg). We found no appreciable change in immune responses when the conjugate vaccine was administered in the presence or absence of immunoadjuvant alum. Administration of OSP:rTTHc resulted in 55% protective efficacy in a mouse survival cholera challenge model.

Conclusion

We report development of an Inaba OSP:rTTHc conjugate vaccine that induces memory responses and protection against cholera in mice. Development of an effective cholera conjugate vaccine that induces high level and long-term immune responses against OSP would be beneficial, especially in young children who respond poorly to polysaccharide antigens.     

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.     

Progress towards development of a cholera subunit vaccine

Cholera, an enteric disease that can reach pandemic proportions, remains a world-wide problem that is positioned to increase in incidence as changes in global climate or armed conflict spawn the conditions that enhance transmission to humans and, thus, precipitate epidemic cholera. An effective subunit cholera vaccine that can provide protective immunity with one parenteral immunization would be a major advantage over the existing oral vaccines that can require two doses for optimal protection. The existing vaccines are clearly effective in some settings, but are less so in others, especially with respect to specific groups such as young (2-5 years) children. In our efforts to develop a cholera subunit vaccine, we focused on two Vibrio cholerae antigens, LPS (lipopolysaccharide) and TCP (toxin co-regulated pilus), that are known to induce protective antibodies in animal models and, in the case of anti-LPS antibodies, to be associated with clinical protection of V. cholerae exposed or vaccinated individuals. This review discusses the current cholera vaccines and compares the advantages of a cholera subunit vaccine to that of the whole cell vaccines. We discuss the possible subunit antigens and prospective targeted use of a subunit cholera vaccine.     

Immunoglobulins and antibodies of various classes in children immunized with live oral Shigella sonnei vaccine from the spontaneous mutant]

The authors determined the immunoglobulin and specific antibodies level in the blood serum of 17 children aged from 7 to 10 years during the immunization with live dysentery Some vaccine. Mancini's test demonstrated the absence of any differences in the amount of IgA and IgG in children of the given age group and in adults before the immunization; in comparison with adults, IgM was increased in children. 14 to 20 days after the immunization in children there was a significant elevation of the IgG only, whereas in adults the immunoglobulin level of all the 3 classes increased significantly. The titres of specific antibodies of the IgA-, IgG-classes and of hemagglutinins before the immunization detected by Coombs' test failed to differ in children from the titres of antibodies of these classes in adults; the level of IgM antibodies was much greater in children than in adults. The changes and accumulation of antibodies of various classes in children and adults during the enteral immunization with live dysentery vaccine differed significantly: in children the vaccine stimulated the IgA- and the IgM-antibody synthesis, whereas adults responded to the immunization by increased production of all the 3 antibody classes. On the basis of the noted immunological shifts a conclusions was drawn on a marked local immunization activity of the live enteral Sonne dysentery vaccine from the spontaneous mutant in children.     

吸附霍乱菌苗免疫后人群血清杀弧菌抗体观察

本文介绍了用霍乱杀弧菌抗体微量法,检测经Ｏ１型吸附霍乱菌苗免疫前后人群的Ｏ１型霍乱和Ｏ１３９型霍乱的血清杀弧菌抗体。结果表明,免前人群的Ｉｎａｂａ、Ｏｇａｗａ、Ｏ１３９型三种自然杀弧菌抗体很低（ＧＭＴ在５．４９以下）,免后Ｉｎａｂａ和Ｏｇａｗａ型杀弧菌抗体大幅度增加,ＧＭＴ分别达到７８２和５９２、Ｏ１３９型霍乱杀弧菌抗体免疫前后无变化     

Use of oral cholera vaccines in an outbreak in Vietnam: a case control study

Background

Killed oral cholera vaccines (OCVs) are available but not used routinely for cholera control except in Vietnam, which produces its own vaccine. In 2007–2008, unprecedented cholera outbreaks occurred in the capital, Hanoi, prompting immunization in two districts. In an outbreak investigation, we assessed the effectiveness of killed OCV use after a cholera outbreak began.

Methodology/Principal Findings

From 16 to 28 January 2008, vaccination campaigns with the Vietnamese killed OCV were held in two districts of Hanoi. No cholera cases were detected from 5 February to 4 March 2008, after which cases were again identified. Beginning 8 April 2008, residents of four districts of Hanoi admitted to one of five hospitals for acute diarrhea with onset after 5 March 2008 were recruited for a matched, hospital-based, case-control outbreak investigation. Cases were matched by hospital, admission date, district, gender, and age to controls admitted for non-diarrheal conditions. Subjects from the two vaccinated districts were evaluated to determine vaccine effectiveness. 54 case-control pairs from the vaccinated districts were included in the analysis. There were 8 (15%) and 16 (30%) vaccine recipients among cases and controls, respectively. The vaccine was 76% protective against cholera in this setting (95% CI 5% to 94%, P = 0.042) after adjusting for intake of dog meat or raw vegetables and not drinking boiled or bottled water most of the time.

Conclusions/Significance

This is the first study to explore the effectiveness of the reactive use of killed OCVs during a cholera outbreak. Our findings suggest that killed OCVs may have a role in controlling cholera outbreaks.     

Thermal Stability of an Oral Killed-Cholera-Whole-Cell Vaccine Containing Recombinant B-Subunit of Cholera Toxin

An oral killed cholera vaccine containing 1×10¹¹ cells of Vibrio cholerae O1 (heat- or formalin-killed) representing the Ogawa and Inaba biotypes and containing 1 mg of B-subunit of cholera toxin (CTB) produced by recombinant DNA technology (the WC/rCTB vaccine) was subjected to temperatures of 4 C, 30 C or 42 C for up to 6 months time. Lipopolysaccharide antigen (LPS) and CTB content of the vaccine samples determined at various times remained unchanged during the study except for the CTB component which decreased by about 50% after 6 months of storage at 42 C. Immunogenicity determined by immunization of rabbits with the vaccine in Freund's complete adjuvant and measuring anti-LPS and anti-CTB antibody titers in the serum by an ELISA was also found to be unaltered. Lyophilization of the vaccine and storage at room temperature for 7 days also did not have any adverse effect on antigen content or immunogenicity as tested above. There was up to one log reduction in serum antibody titers after immunization without using any adjuvant or using Freund's incomplete adjuvant, and up to two logs following oral immunization. Immunization by oral feeding of the vaccine followed by RITARD challenge with a virulent V. cholerae O1 strain showed evidence of protection against severe or lethal diarrhea. The results suggest that the vaccine retains its antigen content and ability to induce antibodies unchanged when maintained at elevated temperatures for relatively long periods of time.     

O1或O139霍乱疫苗对异群霍乱缺乏交叉保护

O1和O139霍乱弧菌是引起急性腹泻的病原微生物,用这两群菌的灭活全菌体与重组霍乱毒素B亚单位构建的亚单位／菌体型疫苗免疫队CA小鼠,对本群菌的攻击可提供良好免疫保护,而对异群霍乱菌则缺乏交叉保护作用。     

Serotype-independent protection against pneumococcal infections elicited by intranasal immunization with ethanol-killed pneumococcal strain,SPY1

The 23-valent polysaccharide vaccine and the 7-valent pneumococcal conjugate vaccine are licensed vaccines that protect against pneumococcal infections worldwide. However, the incidence of pneumococcal diseases remains high in low-income countries. Whole-cell vaccines with high safety and strong immunogenicity may be a favorable choice. We previously obtained a capsule-deficient Streptococcus pneumoniae mutant named SPY1 derived from strain D39. As an attenuated live pneumococcal vaccine, intranasal immunization with SPY1 elicits broad serotype-independent protection against pneumococcal infection. In this study, for safety consideration, we inactivated SPY1 with 70% ethanol and intranasally immunized BALB/c mice with killed SPY1 plus cholera toxin adjuvant for four times. Results showed that intranasal immunization with inactivated SPY1 induced strong humoral and cellular immune responses. Intranasal immunization with inactivated SPY1 plus cholera toxin adjuvant elicited effective serotype-independent protection against the colonization of pneumococcal strains 19F and 4 as well as lethal infection of pneumococcal serotypes 2, 3, 14, and 6B. The protection rates provided by inactivated SPY1 against lethal pneumococcal infection were comparable to those of currently used polysaccharide vaccines. In addition, vaccine-specific B-cell and T-cell immune responses mediated the protection elicited by SPY1. In conclusion, the 70% ethanol-inactivated pneumococcal whole-cell vaccine SPY1 is a potentially safe and less complex vaccine strategy that offers broad protection against S. pneumoniae.     

DNA and adenovirus tumor vaccine expressing truncated survivin generates specific immune responses and anti-tumor effects in a murine melanoma model

Survivin is overexpressed in major types of cancer and is considered an ideal "universal" tumor-associated antigen that can be targeted by immunotherapeutic vaccines. However, its anti-apoptosis function raises certain safety concerns. Here, a new truncated human survivin, devoid of the anti-apoptosis function, was generated as a candidate tumor vaccine. Interleukin 2 (IL-2) has been widely used as an adjuvant for vaccination against various diseases. Meanwhile, the DNA prime and recombinant adenovirus (rAd) boost heterologous immunization strategy has been proven to be highly effective in enhancing immune responses. Therefore, the efficacy of a new cancer vaccine based on a truncated form of survivin, combined with IL-2, DNA prime, and rAd boost, was tested. As prophylaxis, immunization with the DNA vaccine alone resulted in a weak immune response and modest anti-tumor effect, whereas the tumor inhibition ratio with the DNA vaccine administered with IL-2 increased to 89?% and was further increased to nearly 100?% by rAd boosting. Moreover, complete tumor rejection was observed in 5 of 15 mice. Efficacy of the vaccine administered therapeutically was enhanced by nearly 300?% when combined with carboplatin. These results indicated that vaccination with a truncated survivin vaccine using DNA prime-rAd boost combined with IL-2 adjuvant and carboplatin represents an attractive strategy to overcoming immune tolerance to tumors and has potential therapeutic benefits in melanoma cancer.     

Mucosal model of genital immunization in male rhesus macaques with a recombinant simian immunodeficiency virus p27 antigen.

Human immunodeficiency virus (HIV) can be transmitted through infected seminal fluid or vaginal or rectal secretions during heterosexual or homosexual intercourse. To prevent mucosal transmission and spread to the regional lymph nodes, an effective vaccine may need to stimulate immune responses at the genitourinary mucosa. In this study, we have developed a mucosal model of genital immunization in male rhesus macaques, by topical urethral immunization with recombinant simian immunodeficiency virus p27gag, expressed as a hybrid Ty virus-like particle (Ty-VLP) and covalently linked to cholera toxin B subunit. This treatment was augmented by oral immunization with the same vaccine but with added killed cholera vibrios. Polymeric secretory immunoglobulin A (sIgA) and IgG antibodies to p27 were induced in urethral secretions, urine, and seminal fluid. This raises the possibility that the antibodies may function as a primary mucosal defense barrier against SIV (HIV) infection. The regional lymph nodes which constitute the genital-associated lymphoid tissue contained p27-specific CD4+ proliferative and helper T cells for antibody synthesis by B cells, which may function as a secondary immune barrier to infection. Blood and splenic lymphocytes also showed p27-sensitized CD4+ T cells and B cells in addition to serum IgG and IgA p27-specific antibodies; this constitutes a third level of immunity against dissemination of the virus. A comparison of genito-oral with recto-oral and intramuscular routes of immunization suggests that only genito-oral immunization elicits specific sIgA and IgG antibodies in the urine, urethra, and seminal fluid. Both genito-oral and recto-oral immunizations induced T-cell and B-cell immune responses in regional lymph nodes, with preferential IgA antibody synthesis. The mucosal route of immunization may prevent not only virus transmission through the genital mucosa but also dissemination and latency of the virus in the draining lymph nodes.     

Evaluation of TcpF-A2-CTB Chimera and Evidence of Additive Protective Efficacy of Immunizing with TcpF and CTB in the Suckling Mouse Model of Cholera

The secreted colonization factor, TcpF, which is produced by Vibrio cholerae 01 and 0139, has generated interest as a potential protective antigen in the development of a subunit vaccine against cholera. This study evaluated immunogenicity/protective efficacy of a TcpF holotoxin-like chimera (TcpF-A2-CTB) following intraperitoneal immunization compared to TcpF alone, a TcpF+CTB mixture, or CTB alone. Immunization with the TcpF-A2-CTB chimera elicited significantly greater amounts of anti-TcpF IgG than immunization with the other antigens (P<0.05). Protective efficacy was measured using 6-day-old pups reared from immunized dams and orogastrically challenged with a lethal dose of El Tor V. cholerae 01 Inaba strain N16961. Protection from death, and weight loss analysis at 24 and 48 hours post-infection demonstrated that immunization with TcpF alone was poorly protective. However, immunization with TcpF+CTB was highly protective and showed a trend toward greater protection than immunization with CTB alone (82% vs 64% survival). Immunization with the TcpF-A2-CTB chimera demonstrated less protection (50% survival) than immunization with the TcpF+CTB mixture. The TcpF-A2-CTB chimera used for this study contained the heterologous classical CTB variant whereas the El Tor CTB variant (expressed by the challenge strain) was used in the other immunization groups. For all immunization groups that received CTB, quantitative ELISA data demonstrated that the amounts of serum IgG directed against the homologous immunizing CTB antigen was statistically greater than the amount to the heterologous CTB antigen (P≤0.003). This finding provides a likely explanation for the poorer protection observed following immunization with the TcpF-A2-CTB chimera and the relatively high level of protection seen after immunization with homologous CTB alone. Though immunization with TcpF alone provided no protection, the additive protective effect when TcpF was combined with CTB demonstrates its possible value as a component of a multivalent subunit vaccine against Vibrio cholerae 01 and 0139.     

Determination of Shigella antigens in the feces of people, immunized with enteral vaccine prepared from S. flexneri antigen

The dynamic determination of the presence of the specific antigen and its activity in the excreta of humans subjected to enteral immunization with vaccine prepared from S. flexneri antigen was made in the agglutination test and neutralization test with the use of, respectively, antibody and antigenic erythrocyte diagnosticums. In the feces and urine of the vaccinees antibodies occurred less commonly and, as a rule, they were less active than those detected in dysentery patients at the corresponding time from the beginning of the disease. The occurrence of Shigella antigens in the feces of the vaccinees was greater than in their urine at the corresponding time. Similarities and differences in the dynamics of the isolation of Shigella antigen from dysentery patients and from the vaccinees receiving enteral vaccine prepared from S. flexneri antigens were established.     

冻干口服霍乱rBS—WC菌苗安全及保护效果评价的小鼠模型探讨

本文仿照人口服免疫程序用小鼠为模型对冻干口服霍乱ｒＢＳ－ＷＣ菌苗制品进行了安全及保护率评价。给ＫＭ小鼠口服三种不同剂量的ｒＢＳ－ＷＣ菌苗后,无发病,无体重减轻,无死亡现象,表明该菌苗安全,无毒副作用。免疫组与安慰剂组小鼠用肠结扎攻毒试验评价保护效果,两组小鼠用吴江－２及滨－４３活菌攻击后,免疫组显示良好保护作用（Ｐ＜０．０５）。此法可以用于口服型ｒＢＳ－ＷＣ菌苗的评价,但也存在需要动物数较多,个体间差异较大,手术较麻烦的问题。     

The case for reactive mass oral cholera vaccinations

Introduction

The outbreak of cholera in Zimbabwe intensified interest in the control and prevention of cholera. While there is agreement that safe water, sanitation, and personal hygiene are ideal for the long term control of cholera, there is controversy about the role of newer approaches such as oral cholera vaccines (OCVs). In October 2009 the Strategic Advisory Group of Experts advised the World Health Organization to consider reactive vaccination campaigns in response to large cholera outbreaks. To evaluate the potential benefit of this pivotal change in WHO policy, we used existing data from cholera outbreaks to simulate the number of cholera cases preventable by reactive mass vaccination.

Methods

Datasets of cholera outbreaks from three sites with varying cholera endemicity—Zimbabwe, Kolkata (India), and Zanzibar (Tanzania)—were analysed to estimate the number of cholera cases preventable under differing response times, vaccine coverage, and vaccine doses.

Findings

The large cholera outbreak in Zimbabwe started in mid August 2008 and by July 2009, 98,591 cholera cases had been reported with 4,288 deaths attributed to cholera. If a rapid response had taken place and half of the population had been vaccinated once the first 400 cases had occurred, as many as 34,900 (40%) cholera cases and 1,695 deaths (40%) could have been prevented. In the sites with endemic cholera, Kolkata and Zanzibar, a significant number of cases could have been prevented but the impact would have been less dramatic. A brisk response is required for outbreaks with the majority of cases occurring during the early weeks. Even a delayed response can save a substantial number of cases and deaths in long, drawn-out outbreaks. If circumstances prevent a rapid response there are good reasons to roll out cholera mass vaccination campaigns well into the outbreak. Once a substantial proportion of a population is vaccinated, outbreaks in subsequent years may be reduced if not prevented. A single dose vaccine would be of advantage in short, small outbreaks.

Conclusions

We show that reactive vaccine use can prevent cholera cases and is a rational response to cholera outbreaks in endemic and non-endemic settings. In large and long outbreaks a reactive vaccination with a two-dose vaccine can prevent a substantial proportion of cases. To make mass vaccination campaigns successful, it would be essential to agree when to implement reactive vaccination campaigns and to have a dynamic and determined response team that is familiar with the logistic challenges on standby. Most importantly, the decision makers in donor and recipient countries have to be convinced of the benefit of reactive cholera vaccinations.     

Apudocytes in experimental cholera vaccination

The morphofunctional state of apudocytes in the gastrointestinal tract and immunocompetent organs (spleen, mesenteric lymph nodes) of mice immunized with chemical bivalent cholera vaccine was studied. The study revealed that the APUD system of the intestine and the argyrophil elements of the immunocompetent organs of white mice gave a response to the oral administration of commercial cholera vaccine. The reaction of the APUD system of the gastrointestinal tract was manifested by a significant increase in the number of apudocytes and their greater synthesizing activity in the immunized animals during the period of maximum immunological transformation of the macroorganism. The immunization of mice with Vibrio cholerae facilitated the maintenance of homeostasis in the macroorganism and prevented appearance of morphological disturbances in its organs and system after subsequent challenge with V. cholerae.     

Safety of the Recombinant Cholera Toxin B Subunit, Killed Whole-Cell (rBS-WC) Oral Cholera Vaccine in Pregnancy

Introduction

Mass vaccinations are a main strategy in the deployment of oral cholera vaccines. Campaigns avoid giving vaccine to pregnant women because of the absence of safety data of the killed whole-cell oral cholera (rBS-WC) vaccine. Balancing this concern is the known higher risk of cholera and of complications of pregnancy should cholera occur in these women, as well as the lack of expected adverse events from a killed oral bacterial vaccine.

Methodology/Principal Findings

From January to February 2009, a mass rBS-WC vaccination campaign of persons over two years of age was conducted in an urban and a rural area (population 51,151) in Zanzibar. Pregnant women were advised not to participate in the campaign. More than nine months after the last dose of the vaccine was administered, we visited all women between 15 and 50 years of age living in the study area. The outcome of pregnancies that were inadvertently exposed to at least one oral cholera vaccine dose and those that were not exposed was evaluated. 13,736 (94%) of the target women in the study site were interviewed. 1,151 (79%) of the 1,453 deliveries in 2009 occurred during the period when foetal exposure to the vaccine could have occurred. 955 (83%) out of these 1,151 mothers had not been vaccinated; the remaining 196 (17%) mothers had received at least one dose of the oral cholera vaccine. There were no statistically significant differences in the odds ratios for birth outcomes among the exposed and unexposed pregnancies.

Conclusions/Significance

We found no statistically significant evidence of a harmful effect of gestational exposure to the rBS-WC vaccine. These findings, along with the absence of a rational basis for expecting a risk from this killed oral bacterial vaccine, are reassuring but the study had insufficient power to detect infrequent events.

Trial Registration

ClinicalTrials.gov {"type":"clinical-trial","attrs":{"text":"NCT00709410","term_id":"NCT00709410"}}NCT00709410