Attenuated Mycobacterium Bovis Research Articles

An intranasal nanoparticle vaccine elicits protective immunity against Mycobacterium tuberculosis

Mucosal vaccines have the potential to elicit protective immune responses at the point of entry of respiratory pathogens, thus preventing even the initial seed infection. Unlike licensed injectable vaccines, mucosal vaccines comprising protein subunits are only in development. One of the primary challenges associated with mucosal vaccines has been identifying and characterizing safe yet effective mucosal adjuvants that can effectively prime multi-factorial mucosal immunity. In this study, we tested NanoSTING, a liposomal formulation of the endogenous activator of the stimulator of interferon genes (STING) pathway, cyclic guanosine adenosine monophosphate (cGAMP), as a mucosal adjuvant. We formulated a vaccine based on the H1 antigen (fusion protein of Ag85b and ESAT-6) adjuvanted with NanoSTING. Intranasal immunization of NanoSTING-H1 elicited a strong T-cell response in the lung of vaccinated animals characterized by (a) CXCR3+ KLRG1− lung resident T cells that are known to be essential for controlling bacterial infection, (b) IFNγ-secreting CD4+ T cells which is necessary for intracellular bactericidal activity, and (c) IL17-secreting CD4+ T cells that can confer protective immunity against multiple clinically relevant strains of Mtb. Upon challenge with aerosolized Mycobacterium tuberculosis Erdman strain, intranasal NanoSTING-H1 provides protection comparable to subcutaneous administration of the live attenuated Mycobacterium bovis vaccine strain Bacille–Calmette–Guérin (BCG). Our results indicate that NanoSTING adjuvanted protein vaccines can elicit a multi-factorial immune response that protects from infection by M. tuberculosis.

BCG vaccination reduces bovine tuberculosis transmission, improving prospects for elimination.

Bacillus Calmette-Guérin (BCG) is a routinely used vaccine for protecting children against Mycobacterium tuberculosis that comprises attenuated Mycobacterium bovis. BCG can also be used to protect livestock against M. bovis; however, its effectiveness has not been quantified for this use. We performed a natural transmission experiment to directly estimate the rate of transmission to and from vaccinated and unvaccinated calves over a 1-year exposure period. The results show a higher indirect efficacy of BCG to reduce transmission from vaccinated animals that subsequently become infected [74%; 95% credible interval (CrI): 46 to 98%] compared with direct protection against infection (58%; 95% CrI: 34 to 73%) and an estimated total efficacy of 89% (95% CrI: 74 to 96%). A mechanistic transmission model of bovine tuberculosis (bTB) spread within the Ethiopian dairy sector was developed and showed how the prospects for elimination may be enabled by routine BCG vaccination of cattle.

Live attenuated Mycobacterium bovis strains combined with the encapsulated H65 antigen as a vaccine strategy against bovine tuberculosis in a mouse model

Mycobacterium bovis is an etiological agent of bovine tuberculosis (bTB) that also infects other mammals, including humans. The lack of an effective vaccine for the control of bTB highlights the need for developing new vaccines. In this study, we developed and evaluated an M. bovis strain deleted in the virulence genes phoP, esxA and esxB as a vaccine candidate against bTB in BALBc mice. The evaluated strains were the new live vaccine and BCG, alone or in combination with ncH65vD. The immunogen ncH65vD is a fusion protein H65, encapsulated together with vitamin D3, within the oily body of a nanocapsule composed of an antigen-loading polymeric shell. All vaccines conferred protection against the M. bovis challenge. However, no significant differences were detected among the vaccinated groups regarding bacterial loads in lungs and spleen. Mice vaccinated with the mutant strain plus ncH65vD showed negative Ziehl Neelsen staining of mycobacteria in their lungs, which suggests better control of bacteria replication according to this protection parameter. Consistently, this vaccination scheme showed the highest proportion of CD4 + T cells expressing the protection markers PD-1 and CXCR3 among the vaccinated groups. Correlation studies showed that PD-1 and CXCR3 expression levels in lung-resident CD4 T cells negatively correlated with the number of colony forming units of M. bovis in the lungs of mice. Therefore, the results suggest a link between the presence of PD-1 + and CXCR3 + cells at the site of the immune response against mycobacteria and the level of mycobacterial loads.

Intranasal nanoparticle vaccine elicits protective immunity against Mycobacterium tuberculosis

Abstract Mucosal vaccines have the potential to elicit protective immune responses at the point of entry of respiratory pathogens thus preventing even the initial seed infection. Unlike licensed injectable vaccines, mucosal vaccines comprising protein subunits are only in the development stage. One of the primary challenges associated with mucosal vaccines has been the identification and characterization of safe yet effective mucosal adjuvants that can effectively prime multi-factorial mucosal immunity. In this study, we tested NanoSTING, a liposomal formulation of the endogenous activator of the stimulator of interferon genes (STING) pathway, cyclic guanosine adenosine monophosphate (cGAMP), as a mucosal adjuvant. Intranasal administration of this vaccine provides protection comparable to subcutaneous administration of the live attenuated Mycobacterium bovis vaccine strain Bacille–Calmette–Guérin (BCG) upon challenge with aerosolized Mycobacterium tuberculosis Erdman strain. Protection against the bacterium is likely due to CXCR3+ lung resident T cells that are known to be important for the control of bacterial infection. Intranasal delivery also induces IFNγ secreting CD4+ T cells in lung which is necessary for intracellular bactericidal activity. Our results indicate that NanoSTING adjuvanted protein vaccines can elicit a multi-factorial immune response that provides durable protection from infection by M. tuberculosis.

A novel multi-component protein vaccine ECP001 containing a protein polypeptide antigen nPstS1 riching in T-cell epitopes showed good immunogenicity and protection in mice.

Tuberculosis (TB) is an infectious disease that seriously affects human health. Until now, the only anti-TB vaccine approved for use is the live attenuated Mycobacterium bovis (M. bovis) vaccine - BCG vaccine, but its protective efficacy is relatively low and does not provide satisfactory protection against TB in adults. Therefore, there is an urgent need for more effective vaccines to reduce the global TB epidemic. In this study, ESAT-6, CFP-10, two antigens full-length and the T-cell epitope polypeptide antigen of PstS1, named nPstS1, were selected to form one multi-component protein antigens, named ECP001, which include two types, one is a mixed protein antigen named ECP001m, the other is a fusion expression protein antigen named ECP001f, as candidates for protein subunit vaccines. were prepared by constructing one novel subunit vaccine by mixing or fusing the three proteins and combining them with aluminum hydroxide adjuvant, and the immunogenicity and protective properties of the vaccine was evaluated in mice. The results showed that ECP001 stimulated mice to produce high titre levels of IgG, IgG1 and IgG2a antibodies; meanwhile, high levels of IFN-γ and a broad range of specific cytokines were secreted by mouse splenocytes; in addition, ECP001 inhibited the proliferation of Mycobacterium tuberculosis in vitro with a capacity comparable to that of BCG. It can be concluded that ECP001 is a novel effective multicomponent subunit vaccine candidate with potential as BCG Initial Immunisation-ECP001 Booster Immunisation or therapeutic vaccine for M. tuberculosis infection.

Neo-Adjuvant immunotherapies: Bladder cancer as a platform for drug development targeting mucosal immunity

Bacillus Calmette-Guerin (BCG) is a live attenuated Mycobacterium bovis strain, originally developed as a vaccine against tuberculosis. It is also the only bacterial cancer therapy approved by the US Food & Drug Administration for clinical use. BCG is delivered in the bladder, shortly after tumour resection, for patients with high-risk non-muscle invasive bladder cancer (NMIBC). Modulating mucosal immunity by exposing the urothelium to intravesical BCG has been the main therapeutic strategy for high-risk NMIBC over the last three decades. Thus, BCG provides a benchmark for the clinical development of bacteria—or other live attenuated pathogens—as cancer therapy. Currently, a myriad of immuno-oncology compounds is under clinical evaluation in BCG-unresponsive and BCG-naïve patients as an alternative therapy in the context of worldwide BCG shortages. For patients with non-metastatic muscle-invasive bladder cancer (MIBC), studies investigating neoadjuvant immunotherapy with either anti–PD-1/PD-L1 monoclonal antibodies in monotherapy or in combination with anti–CTLA-4 monoclonal antibodies have shown overall efficacy and acceptable safety profiles prior to radical cystectomy. Emerging clinical investigations are testing synergistic approaches by combining intravesical delivery of drugs with systemic immune checkpoint blockades in the neoadjuvant setting for patients with MIBC. Such novel strategy aims to prime a local anti-tumour immunity and reduce distant metastatic relapses by enhancing a systemic adaptive anti-tumour immune response. Here, we present and discuss some of the most promising clinical trials developing such novel therapeutic approaches.

Single-cell analysis reveals changes in BCG vaccine-injected mice modeling tuberculous meningitis brain infection

Tuberculous meningitis (TBM) is the most severe and deadly manifestation of tuberculosis. Neurological complications are observed in up to 50% of patients affected. Here, attenuated Mycobacterium bovis are injected into the cerebellum of mice, and histopathological images and cultured colonies confirm successful brain infection. Then, whole-brain tissue is dissected for 10X Genomics single-cell sequencing, and we acquire 15 cell types. Transcriptional changes of inflammation processes are found in multiple cell types. Specifically, Stat1 and IRF1 are shown to mediate inflammation in macrophages and microglia. For neurons, decreased oxidative phosphorylation activity in neurons is observed, which corresponds to TBM clinical symptoms of neurodegeneration. Finally, ependymal cells present prominent transcriptional changes, and decreased FERM domain containing 4A (Frmd4a) may contribute to TBM clinical symptoms of hydrocephalus and neurodegeneration. This study shows a single-cell transcriptome of M.bovis infection in mice and improves the understanding of brain infection and neurological complications in TBM.

DUSP1 mediates BCG induced apoptosis and inflammatory response in THP-1 cells via MAPKs/NF-κB signaling pathway

Tuberculosis (TB) is a zoonotic infectious disease caused by Mycobacterium tuberculosis (Mtb). Apoptosis and necrosis caused by the interaction between the host and the pathogen, as well as the host’s inflammatory response, play an important role in the pathogenesis of TB. Dual-specificity phosphatase 1 (DUSP1) plays a vital role in regulating the host immune responses. However, the role of DUSP1 in the regulation of THP-1 macrophage apoptosis induced by attenuated Mycobacterium bovis Bacillus Calmette-Guérin (BCG) infection remains unclear. In the present study, we report that infection with BCG significantly induces macrophage apoptosis and induces the production of DUSP1, TNF-α and IL-1β. DUSP1 knockdown significantly inhibited BCG-induced macrophage apoptosis and activation of MAPKs/NF-κB signaling pathway. In addition, DUSP1 knockdown suppressed BCG-induced inflammation in vivo. Taken together, this study demonstrates that DUSP1, as a regulator of MAPKs/NF-κB signaling pathway, plays a novel role in BCG-induced macrophage apoptosis and inflammatory response.

Galleria mellonella as an infection model for the virulent Mycobacterium tuberculosis H37Rv

ABSTRACT Tuberculosis (TB), caused by Mycobacterium tuberculosis (MTB), is a leading cause of infectious disease mortality. Animal infection models have contributed substantially to our understanding of TB, yet their biological and non-biological limitations are a research bottleneck. There is a need for more ethically acceptable, economical, and reproducible TB infection models capable of mimicking key aspects of disease. Here, we demonstrate and present a basic description of how Galleria mellonella (the greater wax moth, Gm) larvae can be used as a low cost, rapid, and ethically more acceptable model for TB research. This is the first study to infect Gm with the fully virulent MTB H37Rv, the most widely used strain in research. Infection of Gm with MTB resulted in a symptomatic lethal infection, the virulence of which differed from both attenuated Mycobacterium bovis BCG and auxotrophic MTB strains. The Gm-MTB model can also be used for anti-TB drug screening, although CFU enumeration from Gm is necessary for confirmation of mycobacterial load reducing activity of the tested compound. Furthermore, comparative virulence of MTB isogenic mutants can be determined in Gm. However, comparison of mutant phenotypes in Gm against conventional models must consider the limitations of innate immunity. Our findings indicate that Gm will be a practical, valuable, and advantageous additional model to be used alongside existing models to advance tuberculosis research.

Recombinant BCG to Enhance Its Immunomodulatory Activities.

The bacillus Calmette–Guérin (BCG) is an attenuated Mycobacterium bovis derivative that has been widely used as a live vaccine against tuberculosis for a century. In addition to its use as a tuberculosis vaccine, BCG has also been found to have utility in the prevention or treatment of unrelated diseases, including cancer. However, the protective and therapeutic efficacy of BCG against tuberculosis and other diseases is not perfect. For three decades, it has been possible to genetically modify BCG in an attempt to improve its efficacy. Various immune-modulatory molecules have been produced in recombinant BCG strains and tested for protection against tuberculosis or treatment of several cancers or inflammatory diseases. These molecules include cytokines, bacterial toxins or toxin fragments, as well as other protein and non-protein immune-modulatory molecules. The deletion of genes responsible for the immune-suppressive properties of BCG has also been explored for their effect on BCG-induced innate and adaptive immune responses. Most studies limited their investigations to the description of T cell immune responses that were modified by the genetic modifications of BCG. Some studies also reported improved protection by recombinant BCG against tuberculosis or enhanced therapeutic efficacy against various cancer forms or allergies. However, so far, these investigations have been limited to mouse models, and the prophylactic or therapeutic potential of recombinant BCG strains has not yet been illustrated in other species, including humans, with the exception of a genetically modified BCG strain that is now in late-stage clinical development as a vaccine against tuberculosis. In this review, we provide an overview of the different molecular engineering strategies adopted over the last three decades in order to enhance the immune-modulatory potential of BCG.

BCG Infection in Patients With Inborn Errors of Immunity Receiving the Russian BCG Strain

Bacillus Calmette-Guierin (BCG) vaccination complications are common in inborn errors of immunity (IEI) due to the inability to clear live attenuated Mycobacterium bovis. Various BCG-vaccine strains are used worldwide, and the profile of the Russian BCG strain vaccine complications in IEI is poorly characterized. To evaluate risks of BCG infection in a large cohort of patients with IEI vaccinated with the Russian BCG strain. We evaluated 778 patients with IEI vaccinated with the Russian BCG strain. A total of 114 (15%) developed BCG infection, 41 (36%) with local, 19 (17%) with regional, and 54 with (47%) disseminated disease. BCG infection was seen in 58% of the patients with severe combined immunodeficiency (SCID), 82% with chronic granulomatous disease, 50% with innate immune defects, 5% with combined immunodeficiency, and 2% with other IEI. BCG infection presented at a median age of 4 to 5 months in SCID, chronic granulomatous disease, combined immunodeficiency, and other IEI groups versus 12 months in patients with innate immune defects (P < .005). We found no influence of specific genetic defects, CD3+ and natural killer cell numbers in SCID, or dihydrorhodamine test stimulation index values in chronic granulomatous disease on the BCG-infection risks. All patients with SCID received antimycobacterial therapy at SCID diagnosis even in the absence of active BCG infection. More antimycobacterial agents were required in disseminated relative to local or regional infection (P < .0001). Only 1 of 114 patients (with SCID) died of BCG-related complications (<1%). BCG infection is common in patients with IEI receiving BCG vaccination. Rational early antimycobacterial therapy, combined with anticytokine agents for posttransplant inflammatory syndrome prevention, and treatment in SCID may prevent BCG-related mortality.

Intravenous BCG protects SIV+ macaques from tuberculosis.

Abstract Tuberculosis is the most common cause of death due to infection in people living with HIV (PLHIV). BCG, a live attenuated Mycobacterium bovis strain given intradermally to infants, is the only licensed vaccine to prevent TB. However, intradermal BCG offers little protection from pulmonary TB in adults and safety concerns limit its use in PLHIV. Recently, intravenous (IV) BCG has been shown to provide striking protection from TB in rhesus macaques. Given this dramatic success, we tested whether IV BCG could protect macaques with a pre-existing SIV infection using our established model of SIV/Mtb coinfection in Mauritian cynomolgus macaques (MCM). MCM were intrarectally infected with SIVmac239 and 5 months later were vaccinated with BCG at 8 ×107 CFU delivered IV. To prevent disseminated BCG disease in immunocompromised animals, four weeks after IV vaccination animals were treated with an 8-week regimen of isoniazid/rifampin/ethambutol (HRE). Four weeks after HRE cessation, animals were challenged with low-dose Mtb Erdman. SIV+ MCM exhibited no signs of BCG disease before or after HRE treatment. Following BCG vaccination, a rapid and sustained increase in airway T cells was observed in both SIV+ and SIV-naïve animals. Increased antibody titers to Mtb lysate were observed in plasma and airways following IV BCG. PBMC responses to Mtb-specific stimuli by IFNγ ELISpot indicated rapid and early clearance of Mtb in vaccinated animals regardless of SIV infection. All SIV-naïve (n=7) and 7 out of 10 SIV+, vaccinated animals were free of TB and without culturable bacilli in their tissues at 12 weeks post challenge. Flow cytometric analysis of PBMCs and tissues obtained at necropsy are underway to further define immune correlates of protection.

H65 fusion protein fails to improve the protection of a rationally attenuated live vaccine candidate against bovine tuberculosis in a mouse model of tuberculosis.

The fusion protein H65, composed of Mycobacterium tuberculosis (TB) ESX-secreted antigens, has improved the bacillus Calmette-Guerin-induced immune protection in a mouse model of bovine TB when formulated in the liposomal adjuvant CAF01. In this study, we aimed to evaluate the protective efficacy of an attenuated Mycobacterium bovis strain - a mutant in mce2 and phoP genes - combined with H65+CAF01 immunization. We evaluated the protection of MbΔmce2-phoP alone or combined with H65+CAF01 against M. bovis challenge in mice. Groups of BALBc mice were inoculated with the vaccine candidates or phosphate buffered saline (PBS), and 6 weeks after the last immunization, the animals were aerogenically challenged with virulent M. bovis. Bacterial load in organs was counted after 45 days of the challenge. One-way analysis of variance and Bonferroni's posttest were used for statistical analysis. All vaccinated mice showed reduced bacterial loads in lungs compared to unvaccinated animals. However, the protection level was similar between vaccinated groups. The MbΔmce2-phoP strain combined with three doses of H65+CAF01 induced equivalent protection than the MbΔmce2-phoP strain alone. Thus, the use of combined vaccination strategies requires a careful analysis of the potential interactions of each of their components with the host's immune system.

Characterizing the BCG Induced Macrophage and Neutrophil Mechanisms for Defense Against Mycobacterium tuberculosis.

The live attenuated Mycobacterium bovis strain, Bacille Calmette Guérin (BCG) is a potent innate immune stimulator. In the C57BL/6 mouse model of tuberculosis, BCG vaccination leads to a significant reduction of Mycobacterium tuberculosis burden after aerogenic infection. Our studies indicated that BCG induced protection against pulmonary tuberculosis was independent of T cells and present as early as 7 days after vaccination. This protection showed longevity, as it did not wane when conventional T cell and TNF-α deficient mice were infected 30 days post-vaccination. As BCG induced mycobacterial killing after 7 days, this study investigated the contributions of the innate immune system after BCG vaccination to better understand mechanisms required for mycobacterial killing. Subcutaneous BCG inoculation resulted in significant CD11b+F4/80+ monocyte subset recruitment into the lungs within 7 days. Further studies revealed that killing of mycobacteria was dependent on the viability of BCG, because irradiated BCG did not have the same effect. Although others have identified BCG as a facilitator of trained innate immunity, we found that BCG reduced the mycobacterial burden in the absence of mechanisms required for trained innate immunity, highlighting a role for macrophages and neutrophils for vaccine induced killing of M. tuberculosis.

BCG as a Case Study for Precision Vaccine Development: Lessons From Vaccine Heterogeneity, Trained Immunity, and Immune Ontogeny.

Vaccines have been traditionally developed with the presumption that they exert identical immunogenicity regardless of target population and that they provide protection solely against their target pathogen. However, it is increasingly appreciated that vaccines can have off-target effects and that vaccine immunogenicity can vary substantially with demographic factors such as age and sex. Bacille Calmette-Guérin (BCG), the live attenuated Mycobacterium bovis vaccine against tuberculosis (TB), represents a key example of these concepts. BCG vaccines are manufactured under different conditions across the globe generating divergent formulations. Epidemiologic studies have linked early life immunization with certain BCG formulations to an unanticipated reduction (∼50%) in all-cause mortality, especially in low birthweight males, greatly exceeding that attributable to TB prevention. This mortality benefit has been related to prevention of sepsis and respiratory infections suggesting that BCG induces “heterologous” protection against unrelated pathogens. Proposed mechanisms for heterologous protection include vaccine-induced immunometabolic shifts, epigenetic reprogramming of innate cell populations, and modulation of hematopoietic stem cell progenitors resulting in altered responses to subsequent stimuli, a phenomenon termed “trained immunity.” In addition to genetic differences, licensed BCG formulations differ markedly in content of viable mycobacteria key for innate immune activation, potentially contributing to differences in the ability of these diverse formulations to induce TB-specific and heterologous protection. BCG immunomodulatory properties have also sparked interest in its potential use to prevent or alleviate autoimmune and inflammatory diseases, including type 1 diabetes mellitus and multiple sclerosis. BCG can also serve as a model: nanoparticle vaccine formulations incorporating Toll-like receptor 8 agonists can mimic some of BCG’s innate immune activation, suggesting that aspects of BCG’s effects can be induced with non-replicating stimuli. Overall, BCG represents a paradigm for precision vaccinology, lessons from which will help inform next generation vaccines.

Effect of disaccharide-polyol systems on the thermal stability of freeze-dried Mycobacterium bovis

Live attenuated Mycobacterium bovis (M. bovis), marketed as Bacille Calmette-Guérin is the only FDA-approved vaccine against tuberculosis. The prerequisite of cold chain storage between 2 and 8 °C hinders the global vaccination effort. The study aims to investigate the effect of trehalose, sucrose and glycerol combinations in enhancing the stability of M. bovis. The bacilli were formulated in various ratios of trehalose-glycerol, sucrose-glycerol, trehalose-sucrose-glycerol systems (test samples) and sodium glutamate (control), freeze-dried and stored for 28 days at 4 °C, 25 °C and 37 °C. Bacteria viability at pre-, post-freeze-drying and after storage were quantified by its density in colony-forming unit per milliliter (CFU/mL) as obtained through the pour plate method. Formulations were characterized using differential scanning calorimetry. Structural collapsed cakes were found on all freeze-dried formulations because of the low Tg’. Comparing between binary and ternary formulations, trehalose-sucrose-glycerol was found to be a superior lyoprotectant. Upon storage, the viability of bacteria in disaccharide-polyol formulations was highest when stored at 4 °C followed by 25 °C. The lowest viability was found after storage at 37 °C. While the ternary disaccharide-polyol system may be used as a thermoprotectant up to 25 °C, sodium glutamate has a superior thermoprotective effect at temperature above 25 °C.

Combined immunodeficiency in a patient with c-Rel deficiency

This study reports a homozygous mutation in REL abrogating c-Rel protein expression in a patient with combined immunodeficiency characterized by susceptibility to Mycobacterium tuberculosis, Salmonella, Cryptosporidium, and cytomegalovirus.

Can immunization with Bacillus Calmette-Guérin (BCG) protect against Alzheimer’s disease?

Alzheimer’s disease (AD) is a neurodegenerative disorder which is the most prevalent cause of dementia in the western world. Currently, it is the most expensive disease in America, costing more than heart diseases and cancer and as the world population is getting older it is expected to become the most expensive medical disorder in the world. AD is characterized by three core pathologies: accumulation of amyloid β (Aβ) plaques, neurofibrillary tangles (NFT) and sustained inflammation. It is now believed that inflammation provides the link between Aβ and NFT. The immune system is therefore, a major player in the pathogenesis of AD. Here we propose that Bacillus Calmette-Guérin (BCG) could affect the incidence of AD.Bacillus Calmette-Guérin (BCG) is a live attenuated Mycobacterium bovis preparation first developed as a vaccine against M. tuberculosis. It has been shown to be moderately effective in preventing tuberculosis, while noted to induce modifications in inflammatory response and to regulate the immune system. Intra-vesical administration of BCG is used successfully in the past four decades to prevent recurrence of non-muscle invasive bladder cancer.In this manuscript we investigate the hypothesis that exposure to BCG decreases the prevalence of AD in elderly population and that this occurs through modulation of the immune system. Our hypothesis is based on several lines of evidence: lower prevalence of AD in countries with high BCG coverage, ability of BCG to ameliorate several conditions involving the immune system like type 1 diabetes mellitus and multiple sclerosis, animal models of AD in which BCG shows therapeutic potential and a plausible molecular mechanism which may be the basis for this hypothesis. Namely, elevated systemic levels of IL-2 (as found when BCG is given intra-vesically) that amplify Treg cells that inhibit AD associated inflammation, decreased plaque formation and restore cognitive function. To test this hypothesis one may study cognition in the large available “natural adult population” exposed to high dose of BCG through the bladder. Bladder cancer survivors not given BCG can serve as control group. This population can be used without adding any medical intervention.

Long-Term Reduction in Hyperglycemia in Advanced Type 1 Diabetes—The Value of Induced Aerobic Glycolysis by BCG Vaccinations

Mycobacteria are among the oldest co-evolutionary partners of humans. Attenuated Mycobacterium bovis, known as the Bacillus Calmette Guérin (BCG) vaccine, has been administered globally for 100 years for TB prevention. BCG vaccination shows clinical promise in numerous autoimmune diseases, including multiple sclerosis (MS) and type 1 diabetes (T1D). In published Phase II clinical trials in MS, BCG shows benefit starting 2 years after administration with continuing effectiveness at year 5. Here we report on the long-term stabilizing effect of BCG vaccination on blood sugar control in advanced T1D with 8 years of follow-up data. The data examines T1D subjects with long-term disease (&amp;gt;10 years duration) who received 2 doses of BCG 4 weeks apart. Starting after year 3, only BCG vaccinated T1D subjects had lowered HbA1c near normal (BCG treated 6.18+/-.34, placebo 7.07+/-.41, reference diabetic 7.22+/-.17, p=0.02; year 5 data with n=46 total subjects). Continued follow-up of 6 subjects for 8 years total confirmed the ability of BCG vaccinations to maintain lowered HbA1c levels without hypoglycemia (BCG treated 6.65+/-.26 vs. 7.22+/-.38, p=0.0002). Glucagon challenge tests were performed once HbA1cs were lowered but did not confirm a therapeutic return of C-peptide levels for improved blood sugar control. Pre-clinical NOD mouse data had shown in mice that BCG induces massive islet regeneration for restore murine blood sugars. In humans, BCG’s impact on blood sugars appears to be driven by a novel mechanism—a systemic shift in glucose metabolism from oxidative phosphorylation to aerobic glycolysis, a state of high glucose utilization as measured by metabolomics, RNAseq and cellular glucose uptake, in subjects receiving a long-term benefit with BCG vaccinations. These clinical and novel mechanistic findings, buttressed with further murine testing, set the stage for a safe and cost effective way to possibly improve blood sugars in humans with diabetes. Disclosure W. Kuhtreiber: None. L. Tran: None. B. Nguyen: None. S.E. Janes: None. A.A. DeFusco: None. D.L. Faustman: None.

Immunomodulator of Ethanol Extracts of The Leaves Azadirachta indica Against Macrophage Peritoneal Cell in Mice Induced The Vaccine BCG

This research was conducted to determine the immunomodulatory effect of ethanol extract of neem leaves (Azadirachta indica) to the number of peritoneal macrophages in mice wich induced by BCG vaccine. Bacillus Calmette-Guerin (BCG) vaccine contained an attenuated Mycobacterium bovis. Mycobacterium bovis belongs to the Mycobacterium Tuberculosis Complex (MTC) group that has a similar phenotype characteristic with Mycobacterium tuberculosis and similar clinical manifestations of tuberculosis.The type of the research that used in this study is laboratory experimental research with Post Test Design Design Only Control Group Design. The research was conducted at the Faculty of Veterinary Medicine of Airlangga University in July 2017 using 25 male mice divided into five groups. The dosage of ethanol extract of the neem leaves given was 200 mg / Kg BW with variation for two days, four days, and six days are given.In the result of statistical data analysis using Kruskal-walis test, it is known that the significance value p = 0,03 (p &lt;0,05), that means immunomodulatory of ethanol extract of neem leaves (Azadirachta indica) give an effect to peritoneal macrophage cell number in mice wich induced by BCG vaccine, so that neem leaves ethanol extract can be applied to tuberculosis patients.