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Tuberculosis is the deadliest infection of our time. In contrast, about 11,000 people died of Ebola between 2014 and 2016. Despite this manifest difference in mortality, there is now a vaccine licensed in the United States and by the European Medicines Agency, with up to 100% efficacy against Ebola. The developments that led to the trialing of the Ebola vaccine were historic and unprecedented. The single licensed TB vaccine (BCG) has limited efficacy. There is a dire need for a more efficacious TB vaccine. To deploy such vaccines, trials are needed in sites that combine high disease incidence and research infrastructure. We describe our twelve-year experience building a TB vaccine trial site in contrast to the process in the recent Ebola outbreak. There are additional differences. Relative to the Ebola pipeline, TB vaccines have fewer trials and a paucity of government and industry led trials. While pathogens have varying levels of difficulty in the development of new vaccine candidates, there yet appears to be greater interest in funding and coordinating Ebola interventions. TB is a global threat that requires similar concerted effort for elimination.

Tuberculosis is the deadliest infection of our time, with about 1.3 million deaths in 2017. In contrast, about 11,000 people died of Ebola between 2014 and 2016. Despite this manifest difference in mortality, there is now a licensed vaccine with up to 100% efficacy against Ebola. The developments that led to the trialing and licensure of the Ebola vaccine were historic and unprecedented. The single licensed TB vaccine (BCG) has limited efficacy. There is a dire need for a new, more efficacious TB vaccine. In order to license and deploy such vaccines, trials are needed in a limited number of sites that combine high disease incidence, research infrastructure and expertise. We first chronologically describe our twelve-year experience in building a TB vaccine trial site, an arduous and incremental process in contrast to the recent Ebola outbreak, which illustrates that sites can be set up under great adversity. Due to the striking differences in the vaccine development of the two diseases, we also compare the number of respective human trials between the two diseases, using publicly available data. Relative to the Ebola pipeline, TB vaccines have fewer trials and a paucity of government and industry led trials. While acknowledging pathogens have varying levels of difficulty in the development of new vaccine candidates, there yet appears to be greater interest in funding and coordinating Ebola interventions. We are convinced that TB is a global threat that requires similar concerted effort for elimination. Funding Statement: AERAS, EDCTP and GSK funded the studies included in the manuscript. None had a role in interpreting results in the article or the decision to publish. Declaration of Interests: The authors declare no competing interests. Ethics Approval Statement: The authors stated that ethics approval was obtained for all the studies included, but was not applicable to this article.

A large proportion of people with latent tuberculosis live in malaria-endemic areas, so co-infection with these two organisms is likely to be common. To determine whether there might be a biologic interaction between these two pathogens in vivo, we infected mice with Mycobacterium tuberculosis and then with a non-lethal strain of Plasmo- dium yoelii eight weeks later. Mice chronically infected with M. tuberculosis simulate the equilibrium between pathogen and host thought to exist in human latent infection. Co-infected mice were less able to contain growth of M. tuberculosis in lung, spleen, and liver (mean ± SEM log10 colony-forming units 5.50 ± 0.11 versus 5.12 ± 0.08, 4.58 ± 0.07 versus 4.13 ± 0.10, and 2.86 ± 0.10 versus 2.49 ± 0.10, respectively) and had increased mortality. In populations where both diseases are endemic, there may be implications for increased incidence of clinically detectable tuberculosis. More than one-third of the global population harbors the latent form of tuberculosis infection. During latency, Myco- bacterium tuberculosis is postulated to exist in a dormant state where the host can effectively contain the pathogen. 1 Some aspects of this equilibrium have been modeled using a strain of mouse C57BL/6 (B6), which is relatively resistant to infec- tion with M. tuberculosis. One day after low-dose aerosol infection, 50�100 bacilli can be cultured from the lung of infected mice. The bacilli grow logarithmically in the lung reaching a plateau at 10 5 -10 6 colony-forming units (CFU) within four weeks. Cell-mediated immune responses pro- moted by the release of Th1 cytokines interferon- (IFN-) and interleukin-12 (IL-12) effect containment of the infec- tion, allowing B6 mice to survive for almost a year. 2 Although there is a plateau in the bacterial growth, the murine host fails to eliminate the pathogen. 3 B6 mice are highly susceptible to infection with a murine malaria parasite Plasmodium yoelii. 4 Intraperitoneal infec- tion using parasitized erythrocytes results in a steady increase in parasitemia, reaching up to 60% of the red blood cells. With a lethal P. yoelii strain, mice die within eight days of infection. With the non-lethal P. yoelii 17X strain, however, blood stage infection completely resolves and death is rarely seen. 5 Infection with this and other strains of rodent malaria parasites (P. chabaudi chabaudi, P. vinckei vinckei, and P. berghei) is accompanied by initial Th1 type immune re

Techniques to improve the sensitivity of smear microscopy would facilitate early tuberculosis (TB) diagnosis and disease control, especially in low-income countries where the positive predictive value is high. C 18 -carboxypropylbetaine (CB-18) is a zwitterionic detergent that helps to compensate for the innate buoyancy of mycobacteria, potentially enhancing recovery by centrifugation. Previous data suggest that CB-18 may increase the sensitivity of smear, culture, and molecular amplification diagnostic testing. The goal of the present study was to evaluate if the sensitivity of the smear technique using light microscopy could be improved by treating respiratory samples with CB-18. In the first phase, respiratory specimens were collected consecutively from patients with suspected pulmonary tuberculosis in a tertiary-care hospital in Rio de Janeiro, Brazil (236 specimens were analyzed). After protocol modifications, another 120 respiratory specimens were evaluated. The standard technique was N -acetyl- l -cysteine with sodium hydroxide (NALC-NaOH) treatment, smear concentration with centrifugation, and Ziehl-Neelsen staining. Culture on Löwenstein-Jensen slants was performed on all specimens for use as the “gold standard.” No specimens from patients undergoing active TB treatment were included. The initial protocol for CB-18 processing resulted in a sensitivity of 59.6% and specificity of 96.8% compared to standard processing with a sensitivity of 66.0% and specificity of 96.8%. Using the modified protocol, the sensitivity of CB-18 increased to 71.4% with a specificity of 97.0% versus standard processing with a sensitivity of 61.9% and a specificity of 99.0%. The diagnostic yield of acid-fast bacillus smear with CB-18 in the absence of fluorescence microscopy and PCR compared to standard processing with NALC-NaOH was not significantly different, although the power to detect a difference by the modified assay was low.

The pathogenesis of tuberculosis involves multiple phases and is believed to involve both a carefully deployed series of adaptive bacterial virulence factors and inappropriate host immune responses that lead to tissue damage. A definedMycobacterium tuberculosismutant strain lacking thesigH-encoded transcription factor showed a distinctive infection phenotype. In resistant C57BL/6 mice, the mutant achieved high bacterial counts in lung and spleen that persisted in tissues in a pattern identical to those of wild-type bacteria. Despite a high bacterial burden, the mutant produced a blunted, delayed pulmonary inflammatory response, and recruited fewer CD4+and CD8+T cells to the lung in the early stages of infection. In susceptible C3H mice, the mutant again showed diminished immunopathology and was nonlethal at over 170 days after intravenous infection, in contrast to isogenic wild-type bacilli, which killed with a median time to death of 52 days. Complete genomic microarray analysis revealed thatM. tuberculosis sigHmay mediate the transcription of at least 31 genes directly and that it modulates the expression of about 150 others; the SigH regulon governs thioredoxin recycling and may be involved in the maintenance of intrabacterial reducing capacity. These data show that theM. tuberculosis sigHgene is dispensable for bacterial growth and survival within the host, but is required for the production of immunopathology and lethality. This phenotype demonstrates that beyond an ability to grow and persist within the host,M. tuberculosishas distinct virulence mechanisms that elicit deleterious host responses and progressive pulmonary disease.

Mycobacterium microti is phylogenetically closely related to Mycobacterium tuberculosis and is a member of that complex of organisms. It is a curved, acid-fast bacillus that is naturally attenuated with a narrow host range for Microtus species only. In this study, we confirm the unique susceptibility of voles to infection with M. microti and the relative resistance of mice with a significantly lower organism burden after 8 weeks of infection. In addition, histopathologic examination of lungs reveals a lack of cellular, granulomatous aggregates characteristically seen in murine M. tuberculosis infection. In the past, M. microti has been used successfully in humans as a vaccine against tuberculosis but was associated with cutaneous reactions. In an attempt to circumvent this adverse effect, we report the efficacy of aerosol and oral vaccination with M. microti. High-dose orogastric vaccination with M. microti resulted in a statistically significant improvement in protection against aerosol challenge with virulent M. tuberculosis in the murine model compared with subcutaneous M. bovis BCG Pasteur vaccination.

Fluoroquinolone resistance in tuberculosis may rapidly emerge. Mice infected with high titers of aerosolized Mycobacterium tuberculosis and treated for 8 weeks with four concentrations of moxifloxacin (0.125, 0.25, 0.50, and 1.0%) mixed into the diet had drug concentrations of 2.4, 4.1, 5.3, and 17.9 μg/ml, respectively, in blood. Selection of fluoroquinolone-resistant mutants occurred in all surviving mice.

A large proportion of people with latent tuberculosis live in malaria-endemic areas, so co-infection with these two organisms is likely to be common. To determine whether there might be a biologic interaction between these two pathogens in vivo, we infected mice with Mycobacterium tuberculosis and then with a non-lethal strain of Plasmodium yoelii eight weeks later. Mice chronically infected with M. tuberculosis simulate the equilibrium between pathogen and host thought to exist in human latent infection. Co-infected mice were less able to contain growth of M. tuberculosis in lung, spleen, and liver (mean +/- SEM log10 colony-forming units = 5.50 +/- 0.11 versus 5.12 +/- 0.08, 4.58 +/- 0.07 versus 4.13 +/- 0.10, and 2.86 +/- 0.10 versus 2.49 +/- 0.10, respectively) and had increased mortality. In populations where both diseases are endemic, there may be implications for increased incidence of clinically detectable tuberculosis.

Techniques to improve the sensitivity of smear microscopy would facilitate early tuberculosis (TB) diagnosis and disease control, especially in low-income countries where the positive predictive value is high. C(18)-carboxypropylbetaine (CB-18) is a zwitterionic detergent that helps to compensate for the innate buoyancy of mycobacteria, potentially enhancing recovery by centrifugation. Previous data suggest that CB-18 may increase the sensitivity of smear, culture, and molecular amplification diagnostic testing. The goal of the present study was to evaluate if the sensitivity of the smear technique using light microscopy could be improved by treating respiratory samples with CB-18. In the first phase, respiratory specimens were collected consecutively from patients with suspected pulmonary tuberculosis in a tertiary-care hospital in Rio de Janeiro, Brazil (236 specimens were analyzed). After protocol modifications, another 120 respiratory specimens were evaluated. The standard technique was N-acetyl-L-cysteine with sodium hydroxide (NALC-NaOH) treatment, smear concentration with centrifugation, and Ziehl-Neelsen staining. Culture on Löwenstein-Jensen slants was performed on all specimens for use as the "gold standard." No specimens from patients undergoing active TB treatment were included. The initial protocol for CB-18 processing resulted in a sensitivity of 59.6% and specificity of 96.8% compared to standard processing with a sensitivity of 66.0% and specificity of 96.8%. Using the modified protocol, the sensitivity of CB-18 increased to 71.4% with a specificity of 97.0% versus standard processing with a sensitivity of 61.9% and a specificity of 99.0%. The diagnostic yield of acid-fast bacillus smear with CB-18 in the absence of fluorescence microscopy and PCR compared to standard processing with NALC-NaOH was not significantly different, although the power to detect a difference by the modified assay was low.