Bovis Bacille Calmette-Guérin Vaccination Research Articles

Differences in responses to the intracellular macrophage environment between Mycobacterium bovis BCG vaccine strains Moreau and Pasteur.

The Bacille Calmette-Guérin (BCG) vaccine comprises a family of strains with variable protective efficacy against pulmonary tuberculosis (TB) and leprosy, partly due to genetic differences between strains. Previous data highlighting differences between the genomes and proteomic profiles of BCG strains Moreau and Pasteur led us to evaluate their behaviour in the macrophage microenvironment, capable of stimulating molecular responses that can impact the protective effect of the vaccine. Strain infectivity, viability, co-localisation with acidified vesicles, macrophage secretion of IL-1 and MCP-1 and lipid droplet biogenesis were evaluated after infection. We found that BCG Moreau is internalised more efficiently, with significantly better intracellular survival up to 96 h p.i., whereas more BCG Pasteur bacilli were found co-localised in acidified vesicles up to 6 h p.i. IL-1β and MCP-1 secretion and lipid droplet biogenesis by infected macrophages were more prominent in response to BCG Pasteur. Overall, our results show that, compared to Pasteur, BCG Moreau has increased fitness and better endurance in the harsh intracellular environment, also regulating anti-microbial responses (lower IL-1b and MCP-1). These findings contribute to the understanding of the physiology of BCG Moreau and Pasteur in response to the intraphagosomal environment in a THP-1 macrophage model.

Histopathologic differences in granulomas of Mycobacterium bovis bacille Calmette Guérin (BCG) vaccinated and non-vaccinated cattle with bovine tuberculosis.

Mycobacterium bovis (M. bovis) is the zoonotic bacterium responsible for bovine tuberculosis. An attenuated form of M. bovis, Bacillus Calmette-Guerin (BCG), is a modified live vaccine known to provide variable protection in cattle and other species. Protection for this vaccine is defined as a reduction in disease severity rather than prevention of infection and is determined by evaluation of the characteristic lesion of tuberculosis: the granuloma. Despite its recognized ability to decrease disease severity, the mechanism by which BCG imparts protection remains poorly understood. Understanding the histopathologic differences between granulomas which form in BCG vaccinates compared to non-vaccinates may help identify how BCG imparts protection and lead to an improved vaccine. Utilizing special stains and image analysis software, we examined 88 lymph nodes obtained from BGC-vaccinated and non-vaccinated animals experimentally infected with M. bovis. We evaluated the number of granulomas, their size, severity (grade), density of multinucleated giant cells (MNGC), and the amounts of necrosis, mineralization, and fibrosis. BCG vaccinates had fewer granulomas overall and smaller high-grade granulomas with less necrosis than non-vaccinates. The relative numbers of high- and low- grade lesions were similar as were the amounts of mineralization and the density of MNGC. The amount of fibrosis was higher in low-grade granulomas from vaccinates compared to non-vaccinates. Collectively, these findings suggest that BCG vaccination reduces bacterial establishment, resulting in the formation of fewer granulomas. In granulomas that form, BCG has a protective effect by containing their size, reducing the relative amount of necrosis, and increasing fibrosis in low-grade lesions. Vaccination did not affect the amount of mineralization or density of MNGC.

Progressive Host-Directed Strategies to Potentiate BCG Vaccination Against Tuberculosis.

The pursuit to improve the TB control program comprising one approved vaccine, M. bovis Bacille Calmette-Guerin (BCG) has directed researchers to explore progressive approaches to halt the eternal TB pandemic. Mycobacterium tuberculosis (M.tb) was first identified as the causative agent of TB in 1882 by Dr. Robert Koch. However, TB has plagued living beings since ancient times and continues to endure as an eternal scourge ravaging even with existing chemoprophylaxis and preventive therapy. We have scientifically come a long way since then, but despite accessibility to the standard antimycobacterial antibiotics and prophylactic vaccine, almost one-fourth of humankind is infected latently with M.tb. Existing therapeutics fail to control TB, due to the upsurge of drug-resistant strains and increasing incidents of co-infections in immune-compromised individuals. Unresponsiveness to established antibiotics leaves patients with no therapeutic possibilities. Hence the search for an efficacious TB immunization strategy is a global health priority. Researchers are paving the course for efficient vaccination strategies with the radically advanced operation of core principles of protective immune responses against M.tb. In this review; we have reassessed the progression of the TB vaccination program comprising BCG immunization in children and potential stratagems to reinforce BCG-induced protection in adults.

A next generation BCG vaccine moves forward

The Mycobacterium Bovis bacille Calmette–Guerin (BCG) vaccine is now over 100 years old. During this century, we have learnt a lot about what the BCG vaccination does and does not do, including that it does not provide consistent or optimal protection against tuberculosis (TB), and particularly, the most common pulmonary form of the disease, that is responsible for Mycobacterium tuberculosis transmission.1,2 Many new candidate TB vaccines are in the development pipeline, including individual antigens delivered by viral vectors or with adjuvant and live mycobacterial vaccines such as genetically modified BCG or M tuberculosis itself.

Protective immunity against tuberculosis in a free-living badger population vaccinated orally with Mycobacterium bovis Bacille Calmette-Guérin.

Vaccination of badgers with Mycobacterium bovis Bacille Calmette-Guérin (BCG) has been shown to protect badgers against tuberculosis in experimental trials. During the 3-year County Kilkenny BCG vaccine field study, badgers were treated orally with placebo (100% in Zone A), BCG (100% in Zone C) or randomly assigned 50%: 50% treatment with BCG or placebo (Zone B). At the end of the study, 275 badgers were removed from the trial area and subjected to detailed post-mortem examination followed by histology and culture for M. bovis. Among these badgers, 83 (30.2%) were captured for the first time across the three zones, representing a non-treated proportion of the population. Analysis of the data based on the infection status of treated animals showed a prevalence of 52% (95% CI: 40%-63%) infection in Zone A (placebo), 39% (95% CI: 17%-64%) in Zone B (placebo) and 44% (95% CI: 20%-70%) in Zone B (BCG vaccinated) and 24% (95% CI: 14%-36%) in Zone C (BCG vaccinated). There were no statistically significant differences in the proportion of animals with infection involving the lung and thoracic lymph nodes, extra-thoracic infection or in the distribution and severity scores of histological lesions. Among the 83 non-treated badgers removed at the end of the study, the infection prevalence of animals in Zone A (prevalence=46%, 95% CI: 32%-61%) and Zone B (prevalence=44%, 95% CI: 23%-67%) was similar to the treated animals in these zones. However, in Zone C, no evidence of infection was found in any of the untreated badgers (prevalence=0%, 95% CI: 0%-14%). This is consistent with an indirect protective effect in the non-vaccinated badgers leading to a high level of population immunity. The results suggest that BCG vaccination of badgers could be a highly effective means of reducing the incidence of tuberculosis in badger populations.

Exploiting Pre-Existing CD4+ T Cell Help from Bacille Calmette-Guérin Vaccination to Improve Antiviral Antibody Responses.

The continuing emergence of viral pathogens and their rapid spread into heavily populated areas around the world underscore the urgency for development of highly effective vaccines to generate protective antiviral Ab responses. Many established and newly emerging viral pathogens, including HIV and Ebola viruses, are most prevalent in regions of the world in which Mycobacterium tuberculosis infection remains endemic and vaccination at birth with M. bovis bacille Calmette-Guérin (BCG) is widely used. We have investigated the potential for using CD4+ T cells arising in response to BCG as a source of help for driving Ab responses against viral vaccines. To test this approach, we designed vaccines comprised of protein immunogens fused to an immunodominant CD4+ T cell epitope of the secreted Ag 85B protein of BCG. Proof-of-concept experiments showed that the presence of BCG-specific Th cells in previously BCG-vaccinated mice had a dose-sparing effect for subsequent vaccination with fusion proteins containing the Ag 85B epitope and consistently induced isotype switching to the IgG2c subclass. Studies using an Ebola virus glycoprotein fused to the Ag 85B epitope showed that prior BCG vaccination promoted high-affinity IgG1 responses that neutralized viral infection. The design of fusion protein vaccines with the ability to recruit BCG-specific CD4+ Th cells may be a useful and broadly applicable approach to generating improved vaccines against a range of established and newly emergent viral pathogens.

Phenotypic and functional characterization of lung resident lymphocytes of BCG vaccinated mice

Abstract Mycobacterium tuberculosis (Mtb) remains to be a leading cause of morbidity and mortality, causing an estimated 1.3 million deaths annually. It is known that intranasal Mycobacterium bovis Bacille Calmette-Guérin (BCG) vaccination in mice has proven to provide superior protection against pulmonary TB, as compared to parenterally administered BCG. In the current study, we determined the phenotype and function of lung-resident lymphocytes in intranasally BCG vaccinated mice. C57BL/6 mice were vaccinated intranasally with live-attenuated BCG and lung lymphocytes were isolated and stained for immunophenotyping via flow cytometry at 24, 48, and 72 hours post-vaccination. At 48 hours post-vaccination, we have observed an expansion of CD69+, CD103+, and CD69+CD103+ lymphocytes within the lung of vaccinated mice. Studies are underway to determine the phenotype and function of lung-resident lymphocyte subsets and their expansion capacity at three months after BCG vaccination and Mtb infection.

Vaccination of white-tailed deer (Odocoileus virginianus) with Mycobacterium bovis bacille Calmette-Guérin (BCG) results in positive tuberculin skin test results in a dose-dependent fashion

Mycobacterium bovis is the cause of tuberculosis in most mammalian species, most notably cattle and other members of the family Bovidae; however, many species of the family Cervidae are also susceptible. In North America, tuberculosis has been identified in both captive and free-ranging cervids. Captive cervids are tested for tuberculosis following many of the same guidelines applied to cattle, including intradermal tuberculin testing using M. bovis purified protein derivative (PPD). Both captive and free-ranging deer and elk have been implicated as the source of infection for many cattle herds. Vaccination with the human vaccine M. bovis BCG has been considered as one possible tool to aid in eradication of tuberculosis from cattle and both captive and free-ranging cervids. Studies in cattle have demonstrated that BCG vaccination can induce false positive intradermal tuberculin test reactions in some cattle. Similar findings have been reported for red deer. We orally vaccinated white-tailed deer with BCG and showed that vaccination can induce false positive skin test reactions in some deer and that the rate of false positive reactions is greater with a higher vaccine dose.

SapM mutation to improve the BCG vaccine: Genomic, transcriptomic and preclinical safety characterization

The Mycobacterium bovis Bacille Calmette Guérin (BCG) vaccine shows variable efficacy in protection against adult tuberculosis (TB). Earlier, we have described a BCG mutant vaccine with a transposon insertion in the gene coding for the secreted acid phosphatase SapM, which led to enhanced long-term survival of vaccinated mice challenged with TB infection. To facilitate development of this mutation as part of a future improved live attenuated TB vaccine, we have now characterized the genome and transcriptome of this sapM::Tn mutant versus parental BCG Pasteur. Furthermore, we show that the sapM::Tn mutant had an equal low pathogenicity as WT BCG upon intravenous administration to immunocompromised SCID mice, passing this important safety test. Subsequently, we investigated the clearance of this improved vaccine strain following vaccination and found a more effective innate immune control over the sapM::Tn vaccine bacteria as compared to WT BCG. This leads to a fast contraction of IFNγ producing Th1 and Tc1 cells after sapM::Tn BCG vaccination. These findings corroborate that a live attenuated vaccine that affords improved long-term survival upon TB infection can be obtained by a mutation that further attenuates BCG. These findings suggest that an analysis of the effectiveness of innate immune control of the vaccine bacteria could be instructive also for other live attenuated TB vaccines that are currently under development, and encourage further studies of SapM mutation as a strategy in developing a more protective live attenuated TB vaccine.

Characterizing the BCG induced T cell independent mechanisms for defense against Mycobacterium tuberculosis

Abstract Mycobacterium bovis Bacille Calmette Guérin (BCG) is a potent immune stimulator that activates innate and adaptive immunity. In the C57BL/6 mouse model of tuberculosis, BCG stimulated immunity causes a significant reduction of M. tuberculosis burden after pulmonary infection. The majority of work has focused on BCG induced T cell immunity as CD4+ T cells play a significant role in protection against tuberculosis. We hypothesized that BCG also induced a T cell independent mechanism that was sufficient to significantly restrict mycobacterial growth. BCG robustly activates innate immunity and our preliminary experiments showed that BCG induced a significant reduction in colony forming units (CFU) in the lungs of mice vaccinated 7 days before pulmonary infection. Our studies indicated that although BCG was administered through subcutaneous inoculation, increased numbers of Ly6C+ monocytes were observed in the lungs within 7 days. Selective depletion also determined that neutrophils played a role in priming innate immunity. Others have identified a role for innate immunity during BCG vaccination termed trained innate immunity. However, our results suggested that at day 7 post BCG vaccination trained innate immunity did not cause the CFU reduction, as the NOD2 receptor or NK cell activation were not absolute requirements, despite being linked to trained immunity. Furthermore, in vivo killing of mycobacterium was dependent on BCG being viable and was monocyte derived, highlighting a novel mechanism. Taken together our data suggest that an early innate mechanism induced by viable BCG is responsible for reducing the mycobacterial burden and precedes trained innate immunity.

Mycobacterium bovis Osteitis Following Immunization with Bacille Calmette-Guérin (BCG) in Korea.

BackgroundMycobacterium bovis Bacille Calmette-Guérin (BCG) osteitis, a rare complication of BCG vaccination, has not been well investigated in Korea. This study aimed to evaluate the clinical characteristics of BCG osteitis during the recent 10 years in Korea.MethodsChildren diagnosed with BCG osteitis at the Seoul National University Children's Hospital from January 2007 to March 2018 were included. M. bovis BCG was confirmed by multiplex polymerase chain reaction (PCR) in the affected bone. BCG immunization status and clinical information were reviewed retrospectively.ResultsTwenty-one patients were diagnosed with BCG osteitis and their median symptom onset from BCG vaccination was 13.8 months (range, 6.0–32.5). Sixteen children (76.2%) received Tokyo-172 vaccine by percutaneous multiple puncture method, while four (19.0%) and one (4.8%) received intradermal Tokyo-172 and Danish strain, respectively. Common presenting symptoms were swelling (76.2%), limited movement of the affected site (63.2%), and pain (61.9%) while fever was only accompanied in 19.0%. Femur (33.3%) and the tarsal bones (23.8%) were the most frequently involved sites; and demarcated osteolytic lesions (63.1%) and cortical breakages (42.1%) were observed on plain radiographs. Surgical drainage was performed in 90.5%, and 33.3% of them required repeated surgical interventions due to persistent symptoms. Antituberculosis medications were administered for a median duration of 12 months (range, 12–31). Most patients recovered without evident sequelae.ConclusionHighly suspecting BCG osteitis based on clinical manifestations is important for prompt management. A comprehensive national surveillance system is needed to understand the exact incidence of serious adverse reactions following BCG vaccination and establish safe vaccination policy in Korea.

BCG vaccination for bovine tuberculosis; conclusions from the Jerusalem One Health workshop.

The global burden of bovine tuberculosis (bTB) remains poorly characterized, with spill-over impacts on multiple species. The "One Health" concept is especially relevant given the bidirectional risk of cattle infecting humans with Mycobacterium bovis and humans infecting cattle with Mycobacterium tuberculosis. "Test and cull" is the traditional bTB control method, but the strategy may not be economically feasible or culturally acceptable where cattle are highly prized or their killing is a religious taboo; it is also less effective when there are wildlife reservoirs of infection. Vaccination with M.bovis bacille Calmette-Guerin (BCG) provides protection against bTB, but its use in animals has been limited. The Jerusalem One Health workshop considered key bTB knowledge gaps and innovative solutions. Knowledge gaps identified included (a) the poorly quantified prevalence of M.bovis infection and disease in cattle, domestic camelids and human populations in developing countries, (b) the absence of alternatives to a "test and cull" strategy in settings where the killing of infected animals is culturally or economically unacceptable, or where affected species are protected and (c) an understanding of the induction of mucosal immunity against bTB. We summarize discussions on the use of BCG vaccination in domestic animals and wildlife and list potential projects to address the knowledge gaps identified.

Efficacy and Safety of BCG Vaccine for Control of Tuberculosis in Domestic Livestock and Wildlife.

Bovine tuberculosis (TB) continues to be an intractable problem in many countries, particularly where “test and slaughter” policies cannot be implemented or where wildlife reservoirs of Mycobacterium bovis infection serve as a recurrent source of infection for domestic livestock. Alternative control measures are urgently required and vaccination is a promising option. Although the M. bovis bacille Calmette-Guérin (BCG) vaccine has been used in humans for nearly a century, its use in animals has been limited, principally as protection against TB has been incomplete and vaccination may result in animals reacting in the tuberculin skin test. Valuable insights have been gained over the past 25 years to optimise protection induced by BCG vaccine in animals and in the development of tests to differentiate infected from vaccinated animals (DIVA). This review examines factors affecting the efficacy of BCG vaccine in cattle, recent field trials, use of DIVA tests and the effectiveness of BCG vaccine in other domestic livestock as well as in wildlife. Oral delivery of BCG vaccine to wildlife reservoirs of infection such as European badgers, brushtail possums, wild boar, and deer has been shown to induce protection against TB and could prove to be a practical means to vaccinate these species at scale. Testing of BCG vaccine in a wide range of animal species has indicated that it is safe and vaccination has the potential to be a valuable tool to assist in the control of TB in both domestic livestock and wildlife.

Novel vaccination approaches to prevent tuberculosis in children.

Pediatric tuberculosis (TB) is an underappreciated problem and accounts for 10 % of all TB deaths worldwide. Children are highly susceptible to infection with Mycobacterium tuberculosis and interrupting TB spread would require the development of effective strategies to control TB transmission in pediatric populations. The current vaccine for TB, M. bovis Bacille Calmette-Guérin (BCG), can afford some level of protection against TB meningitis and severe forms of disseminated TB in children; however, its efficacy against pulmonary TB is variable and the vaccine does not afford life-long protective immunity. For these reasons there is considerable interest in the development of new vaccines to control TB in children. Multiple vaccine strategies are being assessed and include recombinant forms of the existing BCG vaccine, protein or viral candidates designed to boost BCG-induced immunity, or live attenuated forms of M. tuberculosis. A number of these candidates have entered clinical trials; however, no vaccine has shown improved protective efficacy compared to BCG in humans. The current challenge is to identify the most suitable candidates to progress from early to late stage clinical trials, in order to deliver a vaccine that can control and hopefully eliminate the global threat of TB.

Effect of Experimental Parameters on Alginate/Chitosan Microparticles for BCG Encapsulation.

The aim of the present study was to develop novel Mycobacterium bovis bacille Calmette-Guérin (BCG)-loaded polymeric microparticles with optimized particle surface characteristics and biocompatibility, so that whole live attenuated bacteria could be further used for pre-exposure vaccination against Mycobacterium tuberculosis by the intranasal route. BCG was encapsulated in chitosan and alginate microparticles through three different polyionic complexation methods by high speed stirring. For comparison purposes, similar formulations were prepared with high shear homogenization and sonication. Additional optimization studies were conducted with polymers of different quality specifications in a wide range of pH values, and with three different cryoprotectors. Particle morphology, size distribution, encapsulation efficiency, surface charge, physicochemical properties and biocompatibility were assessed. Particles exhibited a micrometer size and a spherical morphology. Chitosan addition to BCG shifted the bacilli surface charge from negative zeta potential values to strongly positive ones. Chitosan of low molecular weight produced particle suspensions of lower size distribution and higher stability, allowing efficient BCG encapsulation and biocompatibility. Particle formulation consistency was improved when the availability of functional groups from alginate and chitosan was close to stoichiometric proportion. Thus, the herein described microparticulate system constitutes a promising strategy to deliver BCG vaccine by the intranasal route.

Optimization and scale-up of cell culture and purification processes for production of an adenovirus-vectored tuberculosis vaccine candidate

Tuberculosis (TB) is the second leading cause of death by infectious disease worldwide. The only available TB vaccine is the Bacille Calmette–Guerin (BCG). However, parenterally administered Mycobacterium bovis BCG vaccine confers only limited immune protection from pulmonary tuberculosis in humans. There is a need for developing effective boosting vaccination strategies. AdAg85A, an adenoviral vector expressing the mycobacterial protein Ag85A, is a new tuberculosis vaccine candidate, and has shown promising results in pre-clinical studies and phase I trial. This adenovirus vectored vaccine is produced using HEK 293 cell culture.Here we report on the optimization of cell culture conditions, scale-up of production and purification of the AdAg85A at different scales. Four commercial serum-free media were evaluated under various conditions for supporting the growth of HEK293 cell and production of AdAg85A. A culturing strategy was employed to take advantages of two culture media with respective strengths in supporting the cell growth and virus production, which enabled to maintain virus productivity at higher cell densities and resulted in more than two folds of increases in culture titer. The production of AdAg85A was successfully scaled up and validated at 60L bioreactor under the optimal conditions.The AdAg85A generated from the 3L and 60L bioreactor runs was purified through several purification steps. More than 98% of total cellular proteins was removed, over 60% of viral particles was recovered after the purification process, and purity of AdAg85A was similar to that of the ATCC VR-1516 Ad5 standard. Vaccination of mice with the purified AdAg85A demonstrated a very good level of Ag85A-specific antibody responses. The optimized production and purification conditions were transferred to a GMP facility for manufacturing of AdAg85A for generation of clinical grade material to support clinical trials.

A Birth Cohort Study of Maternal and Infant Serum PCB-153 and DDE Concentrations and Responses to Infant Tuberculosis Vaccination.

Background:Reasons for the highly variable and often poor protection conferred by the Mycobacterium bovis bacille Calmette–Guérin (BCG) vaccine are multifaceted and poorly understood.Objectives:We aimed to determine whether early-life exposure to PCBs (polychlorinated biphenyls) and DDE [1,1-dichloro-2,2-bis(p-chlorophenyl)ethylene] reduces 6-month infant BCG vaccine response.Methods:Data came from families participating in a prospective birth cohort in eastern Slovakia. At birth, maternal and cord blood were collected for chemical analyses, and infants were immunized with BCG. Blood was collected from infants for chemical analyses and to determine 6-month BCG-specific immunoglobulin (Ig) G and IgA levels. Multivariable linear regression models were fit to examine chemical–BCG associations among approximately 500 mother–infant pairs, with adjustment for confounders.Results:The median 6-month infant concentration of the prevalent congener PCB-153 was 113 ng/g lipid [interquartile range (IQR): 37–248], and 388 ng/g lipid (IQR: 115–847) for DDE. Higher 6-month infant concentrations of PCB-153 and DDE were strongly associated with lower 6-month BCG-specific antibody levels. For instance, BCG-specific IgG levels were 37% lower for infants with PCB-153 concentrations at the 75th percentile compared to the 25th percentile (95% CI: –42, –32; p < 0.001). Results were similar in magnitude and precision for DDE. There was also evidence of PCB–DDE additivity, where exposure to both compounds reduced anti-BCG levels more than exposure to either compound alone.Conclusions:The associations observed in this study indicate that environmental exposures may be overlooked contributors to poorer responses to BCG vaccine. The overall association between these exposures and tuberculosis incidence is unknown.Citation:Jusko TA, De Roos AJ, Lee SY, Thevenet-Morrison K, Schwartz SM, Verner MA, Palkovicova Murinova L, Drobná B, Kočan A, Fabišiková A, Čonka K, Trnovec T, Hertz-Picciotto I, Lawrence BP. 2016. A birth cohort study of maternal and infant serum PCB-153 and DDE concentrations and responses to infant tuberculosis vaccination. Environ Health Perspect 124:813–821; http://dx.doi.org/10.1289/ehp.1510101

Display of Antigens on Polyester Inclusions Lowers the Antigen Concentration Required for a Bovine Tuberculosis Skin Test.

The tuberculin skin test is the primary screening test for the diagnosis of bovine tuberculosis (TB), and use of this test has been very valuable in the control of this disease in many countries. However, the test lacks specificity when cattle have been exposed to environmental mycobacteria or vaccinated with Mycobacterium bovis bacille Calmette-Guérin (BCG). Recent studies showed that the use of three or four recombinant mycobacterial proteins, including 6-kDa early secretory antigenic target (ESAT6), 10-kDa culture filtrate protein (CFP10), Rv3615c, and Rv3020c, or a peptide cocktail derived from those proteins, in the skin test greatly enhanced test specificity, with minimal loss of test sensitivity. The proteins are present in members of the pathogenic Mycobacterium tuberculosis complex but are absent in or not expressed by the majority of environmental mycobacteria and the BCG vaccine strain. To produce a low-cost skin test reagent, the proteins were displayed at high density on polyester beads through translational fusion to a polyhydroxyalkanoate synthase that mediates the formation of antigen-displaying inclusions in recombinant Escherichia coli. Display of the proteins on the polyester beads greatly increased their immunogenicity, allowing for the use of very low concentrations of proteins (0.1 to 3 μg of mycobacterial protein/inoculum) in the skin test. Polyester beads simultaneously displaying all four proteins were produced in a single fermentation process. The polyester beads displaying three or four mycobacterial proteins were shown to have high sensitivity for detection of M. bovis-infected cattle and induced minimal responses in animals exposed to environmental mycobacteria or vaccinated with BCG.

Childhood BCG vaccination does not influence control of Mycobacterium tuberculosis growth by human bronchoalveolar lavage cells

Childhood vaccination with Mycobacterium bovis Bacille Calmette-Guérin (BCG) reduces the risk of infection with Mycobacterium tuberculosis and the risk of severe forms of tuberculosis in children. The protection of adults from pulmonary tuberculosis is doubtful. This study evaluated the effect of the vaccination on the growth of M.tuberculosis human bronchoalveolar mononuclear cells (BALMC). Healthy, adult healthcare workers who were regularly exposed to M.tuberculosis, household tuberculosis contacts, and cured tuberculosis patients were recruited in a multicentre study conducted in Germany. BALMC were co-cultured with different strains of M.tuberculosis in growth inhibition assays (MGIAs). MGIAs on BALMC were conducted in 90 contact persons (known vaccination status, n=75) and 62 former tuberculosis patients (known status, n=22). Growth rates for M.tuberculosis H37Rv in BALMC were independent of the vaccination status, both in healthy contacts and in cured tuberculosis patients. This finding was validated in growth inhibition assays using two different Haarlem M.tuberculosis outbreak strains. Subgroup analyses based on the Interferon-gamma release assay status found no impact of the vaccination on mycobacterial growth. This study suggests that M.bovis BCG vaccination does not alter the anti-mycobacterial capacity of BALMC as assessed in ex-vivo growth inhibition assays.

Attrition of T-cell functions and simultaneous upregulation of inhibitory markers correspond with the waning of BCG-induced protection against tuberculosis in mice.

Mycobacterium bovis bacille Calmette-Guérin (BCG) is the most widely used live attenuated vaccine. However, the correlates of protection and waning of its immunity against tuberculosis is poorly understood. In this study, we correlated the longitudinal changes in the magnitude and functional quality of CD4+ and CD8+ T-cell response over a period of two years after mucosal or parenteral BCG vaccination with the strength of protection against Mycobacterium tuberculosis in mice. The BCG vaccination-induced CD4+ and CD8+ T cells exhibited comparable response kinetics but distinct functional attributes in-terms of IFN-γ, IL-2 and TNF-α co-production and CD62L memory marker expression. Despite a near life-long BCG persistence and the induction of enduring CD4+ T-cell responses characterized by IFN-γ and/or TNF-α production with comparable protection, the protective efficacy waned regardless of the route of vaccination. The progressive decline in the multifactorial functional abilities of CD4+ and CD8+ T cells in-terms of type-1 cytokine production, proliferation and cytolytic potential corresponded with the waning of protection against M. tuberculosis infection. In addition, simultaneous increase in the dysfunctional and terminally-differentiated T cells expressing CTLA-4, KLRG-1 and IL-10 during the contraction phase of BCG-induced response coincided with the loss of protection. Our results question the empirical development of BCG-booster vaccines and emphasize the pursuit of strategies that maintain superior T-cell functional capacity. Furthermore, our results underscore the importance of understanding the comprehensive functional dynamics of antigen-specific T-cell responses in addition to cytokine polyfunctionality in BCG-vaccinated hosts while optimizing novel vaccination strategies against tuberculosis.