Active tuberculosis (TB) often presents with advanced pulmonary disease, including irreversible lung damage and cavities. Cavitary pathology contributes to antibiotic failure, transmission, morbidity and mortality. Matrix metalloproteinases (MMPs), in particular MMP-1 are implicated in TB pathogenesis. We explored the mechanisms relating MMP/TIMP imbalance to cavity formation in a modified rabbit model of cavitary TB. Our model results in consistent progression of consolidation to human-like cavities (100% by day 28) with resultant bacillary burdens (>107 CFU/g) far greater than those found in matched granulomatous tissue (105 CFU/g). Using a novel, breath-hold computerized tomography scanning and image analysis protocol. We show that cavities develop rapidly from areas of densely consolidated tissue. Radiological change correlated with a decrease in functional lung tissue as estimated by changes in lung density during controlled pulmonary expansion (R2=0.6356, p<0.0001). We demonstrated that the expression of interstitial collagenase (MMP-1) is specifically greater in cavitary compared to granulomatous lesions (p<0.01), and that TIMP-3 significantly decreases at the cavity surface. Our findings demonstrate that an MMP-1/TIMP imbalance, is associated with the progression of consolidated regions to cavities containing very high bacterial burdens. Our model provided mechanistic insight, correlating with human disease at the pathological, microbiological and molecular levels,. It also provides a strategy to investigate therapeutics in the context of complex TB pathology. We used these findings to predict a MMP/TIMP balance in active TB; and confirmed this in human plasma, revealing the potential of MMP/TIMP levels as key components of a diagnostic matrix aimed at distinguishing active from latent TB (PPV=92.9%; 95%CI 66.1–99.8%, NPV=85.6%; 95%CI 77.0–91.9%).

The chromosome of Mycobacterium tuberculosis (Mtb) encodes forty seven toxin-antitoxin modules belonging to the VapBC family. The role of these modules in the physiology of Mtb and the function(s) served by their expansion are unknown. We investigated ten vapBC modules from Mtb and the single vapBC from M. smegmatis. Of the Mtb vapCs assessed, only Rv0549c, Rv0595c, Rv2549c and Rv2829c were toxic when expressed from a tetracycline-regulated promoter in M. smegmatis. The same genes displayed toxicity when conditionally expressed in Mtb. Toxicity of Rv2549c in M. smegmatis correlated with the level of protein expressed, suggesting that the VapC level must exceed a threshold for toxicity to be observed. In addition, the level of Rv2456 protein induced in M. smegmatis was markedly lower than Rv2549c, which may account for the lack of toxicity of this and other VapCs scored as ‘non-toxic’. The growth inhibitory effects of toxic VapCs were neutralized by expression of the cognate VapB as part of a vapBC operon or from a different chromosomal locus, while that of non-cognate antitoxins did not. These results demonstrated a specificity of interaction between VapCs and their cognate VapBs, a finding corroborated by yeast two-hybrid analyses. Deletion of selected vapC or vapBC genes did not affect mycobacterial growth in vitro, but rendered the organisms more susceptible to growth inhibition following toxic VapC expression. However, toxicity of ‘non-toxic’ VapCs was not unveiled in deletion mutant strains, even when the mutation eliminated the corresponding cognate VapB, presumably due to insufficient levels of VapC protein. Together with the ribonuclease (RNase) activity demonstrated for Rv0065 and Rv0617 – VapC proteins with similarity to Rv0549c and Rv3320c, respectively – these results suggest that the VapBC family potentially provides an abundant source of RNase activity in Mtb, which may profoundly impact the physiology of the organism.

Background Current tuberculosis treatments leave patients with clinically significant lung injury and increased allcause mortality post-cure. Adjunctive host-directed therapies could protect the lungs, improve long-term survival, and shorten treatment duration; however, few have been tested clinically. Therefore, we aimed to assess the safety and preliminary efficacy of four host-directed therapies for tuberculosis.Methods In this prospective, open-label, phase 2, randomised controlled trial, patients with pulmonary tuberculosis were recruited at three clinical sites in South Africa. Eligible patients were aged 18-65 years, HIV-1-negative, and had rifampicin-susceptible Mycobacterium tuberculosis, a sputum Xpert cycle threshold of less than 20, and moderately advanced or far advanced disease on chest radiography. By use of numbers generated in blocks of ten and stratification by site, eligible patients were randomly assigned (1:1:1:1:1) to receive one of the four oral host-directed treatments plus standard tuberculosis treatment or standard treatment alone (the control group). Host-directed treatments were: CC-11050 (200 mg twice daily, taken with food; day 1-112); everolimus (0•5 mg/day; day 1-112); auranofin (3 mg/day for seven doses, then 6 mg/day; day 1-112); and ergocalciferol (5 mg on day 1, then 2•5 mg on day 28 and day 56). All study participants received oral rifabutin-substituted standard tuberculosis treatment for 180 days. Patients and clinicians were not masked to treatment assignment. Spirometry and sputum culture with solid and liquid media were done at baseline and up to 180 days at specified intervals throughout treatment. The primary endpoint was safety and tolerability up to day 210. Secondary preliminary efficacy endpoints were treatment effects on sputum microbiology (culture status at day 56 and the hazard ratio for stable culture conversion up to day 180) and lung function (FEV 1 and forced vital capacity [FVC]) measured by spirometry at day 56, day 180, and day 540. Safety was analysed in the intention-to-treat population and preliminary efficacy primarily in the per-protocol population. The trial is registered at ClinicalTrials.gov, NCT02968927. Post-treatment follow-up was completed in 2020.

COVID-19, the disease caused by SARS-CoV-2 (1), was declared a pandemic by the World Health Organization (WHO) in March 2020 (2). While awaiting a vaccine, several antivirals are being used to manage the disease with limited success (3, 4). To expand this arsenal, we screened 4 compound libraries: a United States Food and Drug Administration (FDA) approved drug library, an angiotensin converting enzyme-2 (ACE2) targeted compound library, a flavonoid compound library as well as a natural product library. Of the 121 compounds identified with activity against SARS-CoV-2, 7 were shortlisted for validation. We show for the first time that the active form of Vitamin D, calcitriol, exhibits significant potent activity against SARS-CoV-2. This finding paves the way for consideration of host-directed therapies for ring prophylaxis of contacts of SARS-CoV-2 patients.

COVID-19 was declared a pandemic by the World Health Organization on March 11, 2020. This novel coronavirus disease, caused by the SARS-CoV-2 virus, has resulted in severe and unprecedented social and economic disruptions globally. Since the discovery of COVID-19 in December 2019, numerous antivirals have been tested for efficacy against SARS-CoV-2 in vitro and also clinically to treat this disease. This review article discusses the main antiviral strategies currently employed and summarizes reported in vitro and in vivo efficacies of key antiviral compounds in use.

Reliable methods to detect the presence of SARS‐CoV‐2 at venues where people gather are essential for epidemiological surveillance to guide public policy. Communal screening of air in a highly crowded space has the potential to provide early warning on the presence and potential transmission of SARS‐CoV‐2 as suggested by studies early in the epidemic. As hospitals and public facilities apply varying degrees of restrictions and regulations, it is important to provide multiple methodological options to enable environmental SARS‐CoV‐2 surveillance under different conditions. This study assessed the feasibility of using high‐flowrate air samplers combined with RNA extraction kit designed for environmental sample to perform airborne SARS‐CoV‐2 surveillance in hospital setting, tested by RT‐qPCR. The success rate of the air samples in detecting SARS‐CoV‐2 was then compared with surface swab samples collected in the same proximity. Additionally, positive RT‐qPCR samples underwent viral culture to assess the viability of the sampled SARS‐CoV‐2. The study was performed in inpatient ward environments of a quaternary care university teaching hospital in Singapore housing active COVID‐19 patients within the period of February to May 2020. Two types of wards were tested, naturally ventilated open‐cohort ward and mechanically ventilated isolation ward. Distances between the site of air sampling and the patient cluster in the investigated wards were also recorded. No successful detection of airborne SARS‐CoV‐2 was recorded when 50 L/min air samplers were used. Upon increasing the sampling flowrate to 150 L/min, our results showed a high success rate in detecting the presence of SARS‐CoV‐2 from the air samples (72%) compared to the surface swab samples (9.6%). The positive detection rate of the air samples along with the corresponding viral load could be associated with the distance between sampling site and patient. The furthest distance from patient with PCR‐positive air samples was 5.5 m. The airborne SARS‐CoV‐2 detection was comparable between the two types of wards with 60%–87.5% success rate. High prevalence of the virus was found in toilet areas, both on surfaces and in air. Finally, no successful culture attempt was recorded from the environmental air or surface samples.

cWith phosphodiesterase inhibitors (PDE-Is) showing significant promise in shortening tuberculosis treatment, we assessed the effect of roflumilast, an FDA-approved type 4 PDE-I, in both acute and chronic murine models of tuberculosis. Alone, roflumilast had no effect on lung bacillary burden and mortality. However, when roflumilast was used in combination with isoniazid, a reduction in lung bacillary burden was observed. These data suggest that roflumilast may be a good candidate for tuberculosis host-directed therapy (HDT). The treatment of tuberculosis (TB) still takes at least 6 months of multidrug therapy, and adjunctive host-directed therapies (HDTs) have the potential to shorten its duration and prevent drug resistance (1). We have previously shown that type 3 and 5 phosphodiesterase inhibitors (PDE-Is) (cilostazol and sildenafil, respectively) are beneficial in shortening TB first-line standard treatment (2). While type 4 PDE-I (PDE4-I) therapies have been hampered by adverse-effect profiles, two have been approved by the Food and Drug Administration (FDA) since 2011 and others are in development (3). PDE4-Is have recently been studied as adjunctive therapies for TB (1,4). Although the synergism of the PDE4-I CC-3052 with isoniazid (INH) has been reported (5), our group found that, when used as part of a complete tuberculosis multidrug regimen, the PDE4-I rolipram was detrimental to bacterial clearance (6). Since PDE4 has four subtypes (PDE4A to -D), with at least 25 splice variants (7), these contradictory observations may be due to action on different PDE isoforms and the different isoform tissue distributions (7). Additionally, off-target effects or effects on the pharmacokinetics of anti-TB drugs cannot be ruled out.At the start of this study, the only PDE4-I approved by the FDA was roflumilast (trade names, Daxas and Daliresp) (8). Roflumilast is indicated for the management of severe chronic obstructive pulmonary disease (COPD). This agent induces accumulation of host cyclic AMP (cAMP) in lung cells, blocks neutrophil recruitment, and reduces inflammatory cytokines such as tumor necrosis factor alpha (TNF-␣), all of which delay the progression of lung pathology in COPD. To determine whether these anti-inflammatory, tissue-protective effects of roflumilast might also be useful in the management of TB, we assessed the effect of roflumilast as an adjunctive, host-directed therapy on (i) the survival of Mycobacterium tuberculosis-infected mice, (ii) lung cytokine levels during infection, and (iii) bacillary burden during monotherapy and combination therapy with isoniazid.Six-week-old female BALB/c mice were aerosol infected with a high dose of M. tuberculosis H37Rv (4.06 Ϯ 0.20 log 10 CFU). Daily oral gavage (5 days per week) of each group of 20 mice was started the day following infection with roflumilast at the anti-inflammatory dose of 5 mg/kg of body weight/day (9) or isoniazid at 25 mg/kg/day (10) or sham treatment (water). As expected, isoniazid significantly prolonged mouse survival time (P Ͻ 0....

The global tuberculosis (TB) epidemic and the spread of multi- and extensively-drug resistant strains of Mycobacterium tuberculosis (M.tb) have been fueled by low adherence to following lengthy treatment protocols, and the rapid spread of HIV (Human Immunodeficiency Virus). Persistence of the infection in immunocompetent individuals follows from the ability of M.tb to subvert host immune responses in favor of survival within macrophages. Alternative host-directed strategies are therefore being currently sought to improve treatment efficacy and duration. In this study, we evaluated tofacitinib, a new oral Janus kinase (JAK) blocker with anti-inflammatory properties, in shortening tuberculosis treatment. BALB/c mice, which are immunocompetent, showed acceleration of M.tb clearance achieving apparent sterilization after 16 weeks of adjunctive tofacitinib therapy at average exposures higher than recommended in humans, while mice receiving standard treatment alone did not achieve clearance until 24 weeks. True sterilization with tofacitinib was not achieved until five months. C3HeB/FeJ mice, which show reduced pro-inflammatory cytokines during M.tb infection, did not show improved clearance with adjunctive tofacitinib therapy, indicating that the nature of granulomatous lesions and host immunity may influence responsiveness to tofacitinib. Our findings suggest that the JAK pathway could be explored further for host-directed therapy in immunocompetent individuals.