An adenoviral-vectored vaccine confers seroprotection against capsular group B meningococcal disease.

Adenoviral-vectored vaccines are licensed for prevention of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and Ebola virus, but, for bacterial proteins, expression in a eukaryotic cell may affect the antigen's localization and conformation or lead to unwanted glycosylation. Here, we investigated the potential use of an adenoviral-vectored vaccine platform for capsular group B meningococcus (MenB). Vector-based candidate vaccines expressing MenB antigen factor H binding protein (fHbp) were generated, and immunogenicity was assessed in mouse models, including the functional antibody response by serum bactericidal assay (SBA) using human complement. All adenovirus-based vaccine candidates induced high antigen-specific antibody and T cell responses. A single dose induced functional serum bactericidal responses with titers superior or equal to those induced by two doses of protein-based comparators, as well as longer persistence and a similar breadth. The fHbp transgene was further optimized for human use by incorporating a mutation abrogating binding to the human complement inhibitor factor H. The resulting vaccine candidate induced high and persistent SBA responses in transgenic mice expressing human factor H. The optimized transgene was inserted into the clinically relevant ChAdOx1 backbone, and this vaccine has now progressed to clinical development. The results of this preclinical vaccine development study underline the potential of vaccines based on genetic material to induce functional antibody responses against bacterial outer membrane proteins.

MET-PREVENT: metformin to improve physical performance in older people with sarcopenia and physical prefrailty/frailty - protocol for a double-blind, randomised controlled proof-of-concept trial.

INTRODUCTION: Skeletal muscle dysfunction is central to both sarcopenia and physical frailty, which are associated with a wide range of adverse outcomes including falls and fractures, longer hospital stays, dependency and the need for care. Resistance training may prevent and treat sarcopenia and physical frailty, but not everyone can or wants to exercise. Finding alternatives is critical to alleviate the burden of adverse outcomes associated with sarcopenia and physical frailty. This trial will provide proof-of-concept evidence as to whether metformin can improve physical performance in older people with sarcopenia and physical prefrailty or frailty. METHODS AND ANALYSIS: MET-PREVENT is a parallel group, double-blind, placebo-controlled proof-of-concept trial. Trial participants can participate from their own homes, including completing informed consent and screening assessments. Eligible participants with low grip strength or prolonged sit-to-stand time together with slow walk speed will be randomised to either oral metformin hydrochloride 500 mg tablets or matched placebo, taken three times a day for 4 months. The recruitment target is 80 participants from two secondary care hospitals in Newcastle and Gateshead, UK. Local primary care practices will act as participant identification centres. Randomisation will be performed using a web-based minimisation system with a random element, balancing on sex and baseline walk speed. Participants will be followed up for 4 months post-randomisation, with outcomes collected at baseline and 4 months. The primary outcome measure is the four metre walk speed at the 4-month follow-up visit. ETHICS AND DISSEMINATION: The trial has been approved by the Liverpool NHS Research Ethics Committee (20/NW/0470), the Medicines and Healthcare Regulatory Authority (EudraCT 2020-004023-16) and the UK Health Research Authority (IRAS 275219). Results will be made available to participants, their families, patients with sarcopenia, the public, regional and national clinical teams, and the international scientific community. TRIAL REGISTRATION NUMBER: ISRCTN29932357.

Application of a Neisseria meningitidis antigen microarray to identify candidate vaccine proteins from a human Phase I clinical trial.

Meningococcal meningitis is a rare but serious condition affecting mainly children and young adults. Outer membrane vesicles (OMV) from Neisseria meningitidis have been used successfully as vaccines against the disease, although they only provide protection against a limited number of the many existing variants. There have been many attempts to identify suitable protein antigens for use in defined vaccines that provide broad protection against the disease, such as that leading to the development of the four component 4CMenB vaccine. We previously reported the use of a protein antigen microarray to screen for IgG antibodies in sera derived from human recipients of an OMV-based vaccine, as part of a Phase I clinical trial. Here, we show that computational methods can be used to cluster antigens that elicit similar responses in the same individuals. Fitting of IgG antibody binding data to 4,005 linear regressions identified pairs of antigens that exhibited significant correlations. Some were from the same antigens in different quaternary states, whilst others might be correlated for functional or immunological reasons. We also conducted statistical analyses to examine correlations between individual serum bactericidal antibody (SBA) titres and IgG reactivity against specific antigens. Both Kendall's tau and Spearman's rank correlation coefficient statistics identified specific antigens that correlated with log(SBA) titre in five different isolates. The principal antigens identified were PorA and PorB, RmpM, OpcA, and the type IV pilus assembly secretin, PilQ. Other minor antigens identified included a lipoprotein, two proteins from the BAM complex and the efflux channel MtrE. Our results suggest that consideration of the entire antigen composition, and allowance for potential interaction between antigens, could be valuable in designing future meningococcal vaccines. Such an approach has the advantages that it uses data derived from human, rather than animal, immunization and that it avoids the need to screen individual antigens.

Viral vectors expressing group B meningococcal outer membrane proteins induce strong antibody responses but fail to induce functional bactericidal activity.

OBJECTIVE: Adenoviral vectored vaccines, with the appropriate gene insert, induce cellular and antibody responses against viruses, parasites and intracellular pathogens such as Mycobacterium tuberculosis. Here we explored their capacity to induce functional antibody responses to meningococcal transmembrane outer membrane proteins. METHODS: Vectors expressing porin A and ferric enterobactin receptor A antigens were generated, and their immunogenicity assessed in mice using binding and bactericidal assays. RESULTS: The viral vectors expressed the bacterial proteins in an in vitro cell-infection assay and, after immunisation of mice, induced higher titres (>105 end-point titre) and longer lasting (>32 weeks) transgene-specific antibody responses in vivo than did outer membrane vesicles containing the same antigens. However, bactericidal antibodies, which are the primary surrogate of protection against meningococcus, were undetectable, despite different designs to support the presentation of the protective B-cell epitopes. CONCLUSION: These results demonstrate that, while the transmembrane bacterial proteins expressed by the viral vector induced strong and persistent antigen-specific antibodies, this platform failed to induce bactericidal antibodies. The results suggest that conformation or post-translational modifications of bacterial outer membrane antigens produced in eukaryote cells might not result in presentation of the necessary epitopes for induction of functional antibodies.

Human B Cell Responses to Dominant and Subdominant Antigens Induced by a Meningococcal Outer Membrane Vesicle Vaccine in a Phase I Trial.

Neisseria meningitidis outer membrane vesicle (OMV) vaccines are safe and provide strain-specific protection against invasive meningococcal disease (IMD) primarily by inducing serum bactericidal antibodies against the outer membrane proteins (OMP). To design broader coverage vaccines, knowledge of the immunogenicity of all the antigens contained in OMVs is needed. In a Phase I clinical trial, an investigational meningococcal OMV vaccine, MenPF1, made from a meningococcus genetically modified to constitutively express the iron-regulated FetA induced bactericidal responses to both the PorA and the FetA antigen present in the OMP. Using peripheral blood mononuclear cells collected from this trial, we analyzed the kinetics of and relationships between IgG, IgA, and IgM B cell responses against recombinant PorA and FetA, including (i) antibody-secreting cells, (ii) memory B cells, and (iii) functional antibody responses (opsonophagocytic and bactericidal activities). Following MenPF1vaccination, PorA-specific IgG secreting cell responses were detected in up to 77% of participants and FetA-specific responses in up to 36%. Memory B cell responses to the vaccine were low or absent and mainly detected in participants who had evidence of preexisting immunity (P = 0.0069). Similarly, FetA-specific antibody titers and bactericidal activity increased in participants with preexisting immunity and is consistent with the idea that immune responses are elicited to minor antigens during asymptomatic Neisseria carriage, which can be boosted by OMV vaccines. IMPORTANCE Neisseria meningitidis outer membrane vesicles (OMV) are a component of the capsular group B meningococcal vaccine 4CMenB (Bexsero) and have been shown to induce 30% efficacy against gonococcal infection. They are composed of multiple antigens and are considered an interesting delivery platform for vaccines against several bacterial diseases. However, the protective antibody response after two or three doses of OMV-based meningococcal vaccines appears short-lived. We explored the B cell response induced to a dominant and a subdominant antigen in a meningococcal OMV vaccine in a clinical trial and showed that immune responses are elicited to minor antigens. However, memory B cell responses to the OMV were low or absent and mainly detected in participants who had evidence of preexisting immunity against the antigens. Failure to induce a strong B cell response may be linked with the low persistence of protective responses.

Minimally invasive versus conventional sternotomy for Mitral valve repair: protocol for a multicentre randomised controlled trial (UK Mini Mitral).

INTRODUCTION: Numbers of patients undergoing mitral valve repair (MVr) surgery for severe mitral regurgitation have grown and will continue to rise. MVr is routinely performed via median sternotomy; however, there is a move towards less invasive surgical approaches.There is debate within the clinical and National Health Service (NHS) commissioning community about widespread adoption of minimally invasive MVr surgery in the absence of robust research evidence; implementation requires investment in staff and infrastructure.The UK Mini Mitral trial will provide definitive evidence comparing patient, NHS and clinical outcomes in adult patients undergoing MVr surgery. It will establish the best surgical approach for MVr, setting a standard against which emerging percutaneous techniques can be measured. Findings will inform optimisation of cost-effective practice. METHODS AND ANALYSIS: UK Mini Mitral is a multicentre, expertise based randomised controlled trial of minimally invasive thoracoscopically guided right minithoracotomy versus conventional sternotomy for MVr. The trial is taking place in NHS cardiothoracic centres in the UK with established minimally invasive mitral valve surgery programmes. In each centre, consenting and eligible patients are randomised to receive surgery performed by consultant surgeons who meet protocol-defined surgical expertise criteria. Patients are followed for 1 year, and consent to longer term follow-up.Primary outcome is physical functioning 12 weeks following surgery, measured by change in Short Form Health Survey (SF-36v2) physical functioning scale. Early and 1 year echo data will be reported by a core laboratory. Estimates of key clinical and health economic outcomes will be reported up to 5 years.The primary economic outcome is cost effectiveness, measured as incremental cost per quality-adjusted life year gained over 52 weeks following index surgery. ETHICS AND DISSEMINATION: A favourable opinion was given by Wales REC 6 (16/WA/0156). Trial findings will be disseminated to patients, clinicians, commissioning groups and through peer reviewed publication. TRIAL REGISTRATION NUMBER: ISRCTN13930454.

Telomerase Activation to Reverse Immunosenescence in Elderly Patients With Acute Coronary Syndrome: Protocol for a Randomized Pilot Trial.

BACKGROUND: Inflammation plays a key role in the pathophysiology of coronary heart disease (CHD) and its acute manifestation, acute coronary syndrome (ACS). Aging is associated with a decline of the immune system, a process known as immunosenescence. This is characterized by an increase in highly proinflammatory T cells that are involved in CHD progression, plaque destabilization, and myocardial ischemia-reperfusion injury. Telomere dysfunction has been implicated in immunosenescence of T lymphocytes. Telomerase is the enzyme responsible for maintaining telomeres during cell divisions. It has a protective effect on cells under oxidative stress and helps regulate flow-mediated dilation in microvasculature. OBJECTIVE: The TACTIC (Telomerase ACTivator to reverse Immunosenescence in Acute Coronary Syndrome) trial will investigate whether a telomerase activator, TA-65MD, can reduce the proportion of senescent T cells in patients with ACS with confirmed CHD. It will also assess the effect of TA-65MD on decreasing telomere shortening, reducing oxidative stress, and improving endothelial function. METHODS: The study was designed as a single-center, randomized, double-blind, parallel-group, placebo-controlled phase II trial. Recruitment started in January 2019. A total of 90 patients, aged 65 years or older, with treated ACS who have had CHD confirmed by angiography will be enrolled. They will be randomized to one of two groups: TA-65MD oral therapy (8 mg twice daily) or placebo taken for 12 months. The primary outcome is the effect on immunosenescence determined by a decrease in the proportion of CD8+ TEMRA (T effector memory cells re-expressing CD45RA [CD45 expressing exon A]) cells at 12 months. Secondary outcomes include leukocyte telomere length, endothelial function, cardiac function as measured by echocardiography and NT-proBNP (N-terminal fragment of the prohormone brain-type natriuretic peptide), systemic inflammation, oxidative stress, and telomerase activity. RESULTS: The study received National Health Service (NHS) ethics approval on August 9, 2018; Medicines and Healthcare products Regulatory Agency approval on October 19, 2018; and NHS Health Research Authority approval on October 22, 2018. The trial began recruiting participants in January 2019 and completed recruitment in March 2020; the trial is due to report results in 2021. CONCLUSIONS: This pilot trial in older patients with CHD will explore outcomes not previously investigated outside in vitro or preclinical models. The robust design ensures that bias has been minimized. Should the results indicate reduced frequency of immunosenescent CD8+ T cells as well as improvements in telomere length and endothelial function, we will plan a larger, multicenter trial in patients to determine if TA-65MD is beneficial in the treatment of CHD in elderly patients. TRIAL REGISTRATION: ISRCTN Registry ISRCTN16613292; http://www.isrctn.com/ISRCTN16613292 and European Union Drug Regulating Authorities Clinical Trials Database (EudraCT), European Union Clinical Trials Register 2017-002876-26; https://tinyurl.com/y4m2so8g. INTERNATIONAL REGISTERED REPORT IDENTIFIER (IRRID): DERR1-10.2196/19456.

Cytomegalovirus infection is a risk factor for tuberculosis disease in infants.

Immune activation is associated with increased risk of tuberculosis (TB) disease in infants. We performed a case-control analysis to identify drivers of immune activation and disease risk. Among 49 infants who developed TB disease over the first 2 years of life, and 129 healthy matched controls, we found the cytomegalovirus-stimulated (CMV-stimulated) IFN-γ response to be associated with CD8+ T cell activation (Spearman's rho, P = 6 × 10-8). A CMV-specific IFN-γ response was also associated with increased risk of developing TB disease (conditional logistic regression; P = 0.043; OR, 2.2; 95% CI, 1.02-4.83) and shorter time to TB diagnosis (Log Rank Mantel-Cox, P = 0.037). CMV+ infants who developed TB disease had lower expression of NK cell-associated gene signatures and a lower frequency of CD3-CD4-CD8- lymphocytes. We identified transcriptional signatures predictive of TB disease risk among CMV ELISpot-positive (area under the receiver operating characteristic [AUROC], 0.98, accuracy, 92.57%) and -negative (AUROC, 0.9; accuracy, 79.3%) infants; the CMV- signature was validated in an independent infant study (AUROC, 0.71; accuracy, 63.9%). A 16-gene signature that previously identified adolescents at risk of developing TB disease did not accurately classify case and control infants in this study. Understanding the microbial drivers of T cell activation, such as CMV, could guide new strategies for prevention of TB disease in infants.

Immunogenicity profiling of protein antigens from capsular group B Neisseria meningitidis.

Outer membrane vesicle (OMV)- based vaccines have been used to provide strain-specific protection against capsular group B Neisseria meningitidis infections, but the full breadth of the immune response against the components of the OMV has not been established. Sera from adults vaccinated with an OMV vaccine were used to screen 91 outer membrane proteins (OMPs) incorporated in an antigen microarray panel. Antigen-specific IgG levels were quantified pre-vaccination, and after 12 and 18 weeks. These results were compared with IgG levels from mice vaccinated with the same OMV vaccine. The repertoires of highly responding antigens in humans and mice overlapped, but were not identical. The highest responding antigens to human IgG comprised four integral OMPs (PorA, PorB, OpcA and PilQ), a protein which promotes the stability of PorA and PorB (RmpM) and two lipoproteins (BamC and GNA1162). These observations will assist in evaluating the role of minor antigen components within OMVs in providing protection against meningococcal infection. In addition, the relative dominance of responses to integral OMPs in humans emphasizes the importance of this subclass and points to the value of maintaining conformational epitopes from integral membrane proteins in vaccine formulations.

Alternative vaccine administration by powder injection: Needle-free dermal delivery of the glycoconjugate meningococcal group Y vaccine.

Powder-injectors use gas propulsion to deposit lyophilised drug or vaccine particles in the epidermal and sub epidermal layers of the skin. We prepared dry-powder (Tg = 45.2 ± 0.5°C) microparticles (58.1 µm) of a MenY-CRM197 glyconjugate vaccine (0.5% wt.) for intradermal needle-free powder injection (NFPI). SFD used ultrasound atomisation of the liquid vaccine-containing excipient feed, followed by lyophilisation above the glass transition temperature (Tg' = - 29.9 ± 0.3°C). This resulted in robust particles (density~ 0.53 ±0.09 g/cm3) with a narrow volume size distribution (mean diameter 58.1 µm, and span = 1.2), and an impact parameter (ρvr ~ 11.5 kg/m·s) sufficient to breach the Stratum corneum (sc). The trehalose, manitol, dextran (10 kDa), dextran (150 kDa) formulation, or TMDD (3:3:3:1), protected the MenY-CRM197 glyconjugate during SFD with minimal loss, no detectable chemical degradation or physical aggregation. In a capsular group Y Neisseria meningitidis serum bactericidal assay (SBA) with human serum complement, the needle free vaccine, which contained no alum adjuvant, induced functional protective antibody responses in vivo of similar magnitude to the conventional vaccine injected by hypodermic needle and syringe and containing alum adjuvant. These results demonstrate that needle-free vaccination is both technically and immunologically valid, and could be considered for vaccines in development.

Correction: Temporal Expression of Peripheral Blood Leukocyte Biomarkers in a Macaca fascicularis Infection Model of Tuberculosis; Comparison with Human Datasets and Analysis with Parametric/Non-parametric Tools for Improved Diagnostic Biomarker Identification.

[This corrects the article DOI: 10.1371/journal.pone.0154320.].

Temporal Expression of Peripheral Blood Leukocyte Biomarkers in a Macaca fascicularis Infection Model of Tuberculosis; Comparison with Human Datasets and Analysis with Parametric/Non-parametric Tools for Improved Diagnostic Biomarker Identification.

A temporal study of gene expression in peripheral blood leukocytes (PBLs) from a Mycobacterium tuberculosis primary, pulmonary challenge model Macaca fascicularis has been conducted. PBL samples were taken prior to challenge and at one, two, four and six weeks post-challenge and labelled, purified RNAs hybridised to Operon Human Genome AROS V4.0 slides. Data analyses revealed a large number of differentially regulated gene entities, which exhibited temporal profiles of expression across the time course study. Further data refinements identified groups of key markers showing group-specific expression patterns, with a substantial reprogramming event evident at the four to six week interval. Selected statistically-significant gene entities from this study and other immune and apoptotic markers were validated using qPCR, which confirmed many of the results obtained using microarray hybridisation. These showed evidence of a step-change in gene expression from an 'early' FOS-associated response, to a 'late' predominantly type I interferon-driven response, with coincident reduction of expression of other markers. Loss of T-cell-associate marker expression was observed in responsive animals, with concordant elevation of markers which may be associated with a myeloid suppressor cell phenotype e.g. CD163. The animals in the study were of different lineages and these Chinese and Mauritian cynomolgous macaque lines showed clear evidence of differing susceptibilities to Tuberculosis challenge. We determined a number of key differences in response profiles between the groups, particularly in expression of T-cell and apoptotic makers, amongst others. These have provided interesting insights into innate susceptibility related to different host `phenotypes. Using a combination of parametric and non-parametric artificial neural network analyses we have identified key genes and regulatory pathways which may be important in early and adaptive responses to TB. Using comparisons between data outputs of each analytical pipeline and comparisons with previously published Human TB datasets, we have delineated a subset of gene entities which may be of use for biomarker diagnostic test development.

T-cell activation is an immune correlate of risk in BCG vaccinated infants.

Vaccines to protect against tuberculosis (TB) are urgently needed. We performed a case-control analysis to identify immune correlates of TB disease risk in Bacille Calmette-Guerin (BCG) immunized infants from the MVA85A efficacy trial. Among 53 TB case infants and 205 matched controls, the frequency of activated HLA-DR(+) CD4(+) T cells associates with increased TB disease risk (OR=1.828, 95% CI=1.25-2.68, P=0.002, FDR=0.04, conditional logistic regression). In an independent study of Mycobacterium tuberculosis-infected adolescents, activated HLA-DR(+) CD4(+) T cells also associate with increased TB disease risk (OR=1.387, 95% CI=1.068-1.801, P=0.014, conditional logistic regression). In infants, BCG-specific T cells secreting IFN-γ associate with reduced risk of TB (OR=0.502, 95% CI=0.29-0.86, P=0.013, FDR=0.14). The causes and impact of T-cell activation on disease risk should be considered when designing and testing TB vaccine candidates for these populations.

Impact of Reducing Complement Inhibitor Binding on the Immunogenicity of Native Neisseria meningitidis Outer Membrane Vesicles.

Neisseria meningitidis recruits host human complement inhibitors to its surface to down-regulate complement activation and enhance survival in blood. We have investigated whether such complement inhibitor binding occurs after vaccination with native outer membrane vesicles (nOMVs), and limits immunogenicity of such vaccines. To this end, nOMVs reactogenic lipopolysaccharide was detoxified by deletion of the lpxl1 gene (nOMVlpxl1). nOMVs unable to bind human complement factor H (hfH) were generated by additional deletions of the genes encoding factor H binding protein (fHbp) and neisserial surface protein A (NspA) (nOMVdis). Antibody responses elicited in mice with nOMVdis were compared to those elicited with nOMVlpxl1 in the presence of hfH. Results demonstrate that the administration of human fH to mice immunized with fHbp containing OMVlpxl1 decreased immunogenicity against fHbp (but not against the OMV as a whole). The majority of the OMV-induced bactericidal immune response (OMVlpxl1 or OMVdis) was versus PorA. Despite a considerable reduction of hfH binding to nOMVdis, and the absence of the vaccine antigen fHbp, immunogenicity in mice was not different from nOMVlpxl1, in the absence or presence of hfH (serum bactericidal titers of 1:64 vs 1:128 after one dose in the nOMVdis and nOMVlpxl1-immunized groups respectively). Therefore, partial inhibition of fH binding did not enhance immunity in this model.

The capsular group B meningococcal vaccine, 4CMenB : clinical experience and potential efficacy.

INTRODUCTION: Capsular group B meningococcal disease is a leading cause of childhood meningitis and septicaemia. Up to 10% of sufferers die, and sequelae remain in > 30% of survivors. A vaccine, four component meningococcal group B ( 4CMenB ), designed with the aim to induce broad coverage against this highly variable bacterium, has been licensed in countries including in the European Union, Canada and Australia. AREAS COVERED: Immunogenicity and safety data, published in peer-reviewed literature between 2004 and 2014, are presented in the context of the recent recommendation for the use of the vaccine in infants in the UK. EXPERT OPINION: 4CMenB induces significant reactogenicity when administered with routine infant vaccines, in particular with respect to fever rates. Fevers can be somewhat reduced using paracetamol. The efficacy of the vaccine is unknown but has been extrapolated from effectiveness data obtained from use of one of its components in New Zealand, immunogenicity data from clinical trials and estimation of coverage from in vitro studies. These data suggest that the vaccine will prevent a proportion of invasive meningococcal disease cases in infants and young children. Implementation and well-planned post-marketing surveillance will address uncertainties over field effectiveness.

Non-tuberculous mycobacteria have diverse effects on BCG efficacy against Mycobacterium tuberculosis.

The efficacy of Bacillus Calmette-Guerin (BCG) vaccination in protection against pulmonary tuberculosis (TB) is highly variable between populations. One possible explanation for this variability is increased exposure of certain populations to non-tuberculous mycobacteria (NTM). This study used a murine model to determine the effect that exposure to NTM after BCG vaccination had on the efficacy of BCG against aerosol Mycobacterium tuberculosis challenge. The effects of administering live Mycobacterium avium (MA) by an oral route and killed MA by a systemic route on BCG-induced protection were evaluated. CD4+ and CD8+ T cell responses were profiled to define the immunological mechanisms underlying any effect on BCG efficacy. BCG efficacy was enhanced by exposure to killed MA administered by a systemic route; T helper 1 and T helper 17 responses were associated with increased protection. BCG efficacy was reduced by exposure to live MA administered by the oral route; T helper 2 cells were associated with reduced protection. These findings demonstrate that exposure to NTM can induce opposite effects on BCG efficacy depending on route of exposure and viability of NTM. A reproducible model of NTM exposure would be valuable in the evaluation of novel TB vaccine candidates.

Evaluation of a human BCG challenge model to assess antimycobacterial immunity induced by BCG and a candidate tuberculosis vaccine, MVA85A, alone and in combination.

BACKGROUND: A new vaccine is urgently needed to combat tuberculosis. However, without a correlate of protection, selection of the vaccines to take forward into large-scale efficacy trials is difficult. Use of bacille Calmette-Guérin (BCG) as a surrogate for human Mycobacterium tuberculosis challenge is a novel model that could aid selection. METHODS: Healthy adults were assigned to groups A and B (BCG-naive) or groups C and D (BCG-vaccinated). Groups B and D received candidate tuberculosis vaccine MVA85A. Participants were challenged with intradermal BCG 4 weeks after those who received MVA85A. Skin biopsies of the challenge site were taken 2 weeks post challenge and BCG load quantified by culture and quantitative polymerase chain reaction (qPCR). RESULTS: Volunteers with a history of BCG showed some degree of protective immunity to challenge, having lower BCG loads compared with volunteers without prior BCG, regardless of MVA85A status. There was a significant inverse correlation between antimycobacterial immunity at peak response after MVA85A and BCG load detected by qPCR. CONCLUSION: Our results support previous findings that this BCG challenge model is able to detect differences in antimycobacterial immunity induced by vaccination and could aid in the selection of candidate tuberculosis vaccines for field efficacy testing.

Mycobacterial growth inhibition in murine splenocytes as a surrogate for protection against Mycobacterium tuberculosis (M. tb).

Development of an improved vaccine against tuberculosis (TB) is hindered by the lack of a surrogate of protection. Efficacy of new TB vaccines in humans can only be evaluated by expensive and time consuming efficacy trials within TB endemic areas. It is critical that vaccines with the greatest potential to protect are selected for these trials. Mycobacterial growth inhibition assays (MGIAs) have been developed with the hope that these in-vitro functional assays will correlate with protection, which could aid in the selection of the best vaccine candidates. The present study describes the use of the BACTEC system to perform MGIAs in mice. We demonstrate reproducible mycobacterial growth inhibition in splenocytes from BCG immunised mice compared with unimmunised mice (P < 0.023), which corresponded with in-vivo efficacy against Mycobacterium tuberculosis (M. tb) challenge. Microarray data showed extensive differential gene expression in splenocyte responses to ex-vivo BCG stimulation between unimmunised and BCG-immunised mice. TH1 responses, including IFN-γ, nitric oxide synthase (NOS2) and Interleukin -17 (IL-17) expression were enhanced in BCG immunised mice, indicating a possible mechanism for mycobacterial growth inhibition. Further investigation into whether the BACTEC MGIA can be used as a surrogate of protection in humans and preclinical animal models is now warranted.