Immunogenicity and Protective Efficacy of a Multi-Antigen Subunit Vaccine in Mice.

There is an urgent need for an effective TB vaccine capable of controlling both acute and chronic infection in populations with diverse genetic backgrounds. In this study, we characterised the immunogenicity and protective efficacy of a novel protein-in-adjuvant subunit vaccine. The protein component is a fusion protein of three different antigens, which we termed CysVac5: CysD, a major component of the sulfate activation pathway that is highly expressed during the chronic stage of infection, is fused with two major secreted mycobacterial antigens, Ag85B and MPT83.

Characterization of the Protective Immune Responses Conferred by Recombinant BCG Overexpressing Components of Sec Protein Export System.

Bacillus Calmette-Guérin (BCG) is the only approved vaccine against tuberculosis (TB). However, its efficacy in preventing pulmonary TB in adults is limited. Despite its variable efficacy, BCG offers a number of unique and beneficial characteristics, which make it suitable as a vaccine vehicle to express recombinant molecules.

Advax adjuvant formulations promote protective immunity against aerosol Mycobacterium tuberculosis in the absence of deleterious inflammation and reactogenicity.

The development of safe and effective adjuvants is a critical goal of vaccine development programs. In this report, we defined the immunostimulatory profile and protective effect against aerosol Mycobacterium tuberculosis infection of vaccine formulations incorporating the semi-crystalline adjuvant δ-inulin (Advax). Advax formulated with CpG oligonucleotide and the QS-21 saponin (Advax) was the most effective combination, demonstrated by the capacity of CysVac2/Advax to significantly reduce the bacterial burden in the lungs of M.

Paper Versus Digital Data Collection Methods for Road Safety Observations: Comparative Efficiency Analysis of Cost, Timeliness, Reliability, and Results.

Background: Roadside observational studies play a fundamental role in designing evidence-informed strategies to address the pressing global health problem of road traffic injuries. Paper-based data collection has been the standard method for such studies, although digital methods are gaining popularity in all types of primary data collection.

Objective: This study aims to understand the reliability, productivity, and efficiency of paper vs digital data collection based on three different road user behaviors: helmet use, seatbelt use, and speeding.

Protective efficacy of recombinant BCG over-expressing protective, stage-specific antigens of Mycobacterium tuberculosis.

Tuberculosis (TB) remains a major cause of mortality and morbidity worldwide, yet current control strategies, including the existing BCG vaccine, have had little impact on disease control. CysVac2, a fusion protein comprising stage-specific Mycobacterium tuberculosis antigens, provided superior protective efficacy against chronic M. tuberculosis infection in mice, compared to BCG.

Delta inulin-based adjuvants promote the generation of polyfunctional CD4 T cell responses and protection against Mycobacterium tuberculosis infection.

There is an urgent need for the rational design of safe and effective vaccines to protect against chronic bacterial pathogens such as Mycobacterium tuberculosis. Advax™ is a novel adjuvant based on delta inulin microparticles that enhances immunity with a minimal inflammatory profile and has entered human trials to protect against viral pathogens. In this report we determined if Advax displays broad applicability against important human pathogens by assessing protective immunity against infection with M.

components expressed during chronic infection of the lung contribute to long-term control of pulmonary tuberculosis in mice.

Tuberculosis (TB) remains a major cause of mortality and morbidity worldwide, yet current control strategies, including the existing BCG vaccine, have had little impact on disease control. The tubercle bacillus modifies protein expression to adapt to chronic infection of the host, and this can potentially be exploited to develop novel therapeutics. We identified the gene encoding the first step of the sulphur assimilation pathway, as highly induced during chronic infection in the mouse lung, suggesting therapies based on CysD could be used to target infection.

Influence of phthiocerol dimycocerosate on CD4(+) T cell priming and persistence during Mycobacterium tuberculosis infection.

The characterisation of mycobacterial factors that influence or modulate the host immune response may aid the development of more efficacious TB vaccines. We have previously reported that Mycobacterium tuberculosis deficient in export of Phthiocerol Dimycocerosates (DIM) (MT103(ΔdrrC)) is more attenuated than wild type M. tuberculosis and provides sustained protective immunity compared to the existing BCG vaccine.

Homogentisate 1-2-Dioxygenase Downregulation in the Chronic Persistence of Pseudomonas aeruginosa Australian Epidemic Strain-1 in the CF Lung.

Some Pseudomonas aeruginosa strains including Australian Epidemic Strain-1 (AES-1 or AUS-01) cause persistent chronic infection in cystic fibrosis (CF) patients, with greater morbidity and mortality. Factors conferring persistence are largely unknown. Previously we analysed the transcriptomes of AES-1 grown in Luria broth, nematode growth medium for Caenorhabditis elegans assay (both aerobic) and artificial sputum medium (mainly hypoxic).

Host cell-induced components of the sulfate assimilation pathway are major protective antigens of Mycobacterium tuberculosis.

New therapies to control tuberculosis are urgently required because of the inability of the only available vaccine, BCG, to adequately protect against tuberculosis. Here we demonstrate that proteins of the Mycobacterium tuberculosis sulfate-assimilation pathway (SAP) represent major immunogenic targets of the bacillus, as defined by strong T-cell recognition by both mice and humans infected with M. tuberculosis.

The secreted lipoprotein, MPT83, of Mycobacterium tuberculosis is recognized during human tuberculosis and stimulates protective immunity in mice.

The long-term control of tuberculosis (TB) will require the development of more effective anti-TB vaccines, as the only licensed vaccine, Mycobacterium bovis bacille Calmette-Guérin (BCG), has limited protective efficacy against infectious pulmonary TB. Subunit vaccines have an improved safety profile over live, attenuated vaccines, such as BCG, and may be used in immuno-compromised individuals. MPT83 (Rv2873) is a secreted mycobacterial lipoprotein expressed on the surface of Mycobacterium tuberculosis.

Protective immunity afforded by attenuated, PhoP-deficient Mycobacterium tuberculosis is associated with sustained generation of CD4+ T-cell memory.

Definition of protective immunity induced by effective vaccines is important for the design of new pathogen control strategies. Inactivation of the PhoP response-regulator in Mycobacterium tuberculosis results in a highly attenuated strain that demonstrates impressive protective efficacy in pre-clinical models of tuberculosis. In this report we demonstrate that the protection afforded by the M.

Cutaneous immunosurveillance by self-renewing dermal gammadelta T cells.

The presence of γδ T cell receptor (TCR)-expressing cells in the epidermis of mice, termed dendritic epidermal T cells (DETCs), is well established. Because of their strict epidermal localization, it is likely that DETCs primarily respond to epithelial stress, such as infections or the presence of transformed cells, whereas they may not participate directly in dermal immune responses. In this study, we describe a prominent population of resident dermal γδ T cells, which differ from DETCs in TCR usage, phenotype, and migratory behavior.

Cutinase-like protein-6 of Mycobacterium tuberculosis is recognised in tuberculosis patients and protects mice against pulmonary infection as a single and fusion protein vaccine.

Infection with Mycobacterium tuberculosis continues to be a leading cause of death in many regions of the world, and control of this disease is hampered by the lack of a safe and effective vaccine. Secreted proteins of M. tuberculosis are an important group of antigens for subunit vaccines which target this infection.

Sulfite reduction in mycobacteria.

Mycobacterium tuberculosis places an enormous burden on the welfare of humanity. Its ability to grow and its pathogenicity are linked to sulfur metabolism, which is considered a fertile area for the development of antibiotics, particularly because many of the sulfur acquisition steps in the bacterium are not found in the host. Sulfite reduction is one such mycobacterium-specific step and is the central focus of this paper.

Contribution of L-alanine dehydrogenase to in vivo persistence and protective efficacy of the BCG vaccine.

The tuberculosis (TB) vaccine strain Mycobacterium bovis BCG is unable to utilise alanine and this deficiency is thought to inhibit the growth of the vaccine in vivo and limit vaccine efficacy. In this report we demonstrate that L-alanine catabolism can be conferred on BCG by introduction of the gene encoding L-alanine dehydrogenase (Ald) of Mycobacterium tuberculosis. Restoration of Ald activity did not change the in vivo growth of BCG in macrophages or mice, and protection against aerosol M.

The trifunctional sulfate-activating complex (SAC) of Mycobacterium tuberculosis.

The sulfate activation pathway is essential for the assimilation of sulfate and, in many bacteria, is comprised of three reactions: the synthesis of adenosine 5'-phosphosulfate (APS), the hydrolysis of GTP, and the 3'-phosphorylation of APS to produce 3'-phosphoadenosine 5'-phosphosulfate (PAPS), whose sulfuryl group is reduced or transferred to other metabolites. The entire sulfate activation pathway is organized into a single complex in Mycobacterium tuberculosis. Although present in many bacteria, these tripartite complexes have not been studied in detail.

The Mycobacterium tuberculosis cysD and cysNC genes form a stress-induced operon that encodes a tri-functional sulfate-activating complex.

Sulfur metabolism has been implicated in the virulence, antibiotic resistance and anti-oxidant defence of Mycobacterium tuberculosis. Despite its human disease relevance, sulfur metabolism in mycobacteria has not yet been fully characterized. ATP sulfurylase catalyses the synthesis of activated sulfate (adenosine 5'-phosphosulfate, APS), the first step in the reductive assimilation of sulfate.

Mycobacterium tuberculosis defective in phthiocerol dimycocerosate translocation provides greater protective immunity against tuberculosis than the existing bacille Calmette-Guérin vaccine.

We demonstrate that Mycobacterium tuberculosis that is unable to export the complex lipid phthiocerol dimycocerosate has a decreased capacity to replicate in mice and affords sustained protective immunity against M. tuberculosis infection Protection was significantly better than that provided by the existing vaccine, Mycobacterium bovis bacille Calmette-Guérin (BCG), and this improved protective efficacy was maintained for at least 24 weeks after vaccination. Protection afforded by this attenuated strain coincided with a number of factors that were not associated with BCG vaccination: long-term persistence of the strain within the host, sustained and potent induction of antimycobacterial interferon-gamma-secreting cells equal to that induced by virulent M.

Destabilized green fluorescent protein for monitoring transient changes in mycobacterial gene expression.

The green fluorescent protein (GFP) is a useful reporter for the study of gene expression and protein localisation within living cells. The stability of GFP permits its intracellular accumulation and detection, but renders it less useful for assessing transient changes in gene expression. We have developed a destabilized form of GFP for monitoring gene expression in mycobacteria.

Regulation of mutY and nature of mutator mutations in Escherichia coli populations under nutrient limitation.

Previous analysis of aerobic, glucose-limited continuous cultures of Escherichia coli revealed that G:C-to-T:A (G:C-->T:A) transversions were the most commonly occurring type of spontaneous mutation. One possible explanation for the preponderance of these mutations was that nutrient limitation repressed MutY-dependent DNA repair, resulting in increased proportions of G:C-->T:A transversions. The regulation of the mutY-dependent DNA repair system was therefore studied with a transcriptional mutY-lacZ fusion recombined into the chromosome.