The infant gut resistome associates with E. coli, environmental exposures, gut microbiome maturity, and asthma-associated bacterial composition Graphical abstract Highlights d Distribution of infant gut resistome is bimodal, mainly driven by E. coli d The infant gut resistome is significantly affected by environmental factors d Low maturity of microbiome associates with high ARG load d A similar asthma-associated gut bacterial composition associates with high ARG load

Rationale: A major priority in tuberculosis (TB) is to reduce effective treatment times and emergence of resistance. Recent studies in macrophages and zebrafish show that inhibition of mycobacterial efflux pumps with verapamil reduces the bacterial drug tolerance and may enhance drug efficacy. Objectives: Using mice, a mammalian model known to predict human treatment responses, and selecting conservative human bioequivalent doses, we tested verapamil as an adjunctive drug together with standard TB chemotherapy. As verapamil is a substrate for CYP3A4, which is induced by rifampin, we evaluated the pharmacokinetic/ pharmacodynamic relationships of verapamil and rifampin coadministration in mice. Methods: Using doses that achieve human bioequivalent levels matched to those of standard verapamil, but lower than those of extended release verapamil, we evaluated the activity of verapamil added to standard chemotherapy in both C3HeB/FeJ (which produce necrotic granulomas) and the wild-type background C3H/HeJ mouse strains. Relapse rates were assessed after 16, 20, and 24 weeks of treatment in mice. Measurements and Main Results: We determined that a dose adjustment of verapamil by 1.5-fold is required to compensate for concurrent use of rifampin during TB treatment. We found that standard TB chemotherapy plus verapamil accelerates bacterial clearance in C3HeB/FeJ mice with near sterilization, and significantly lowers relapse rates in just 4 months of treatment when compared with mice receiving standard therapy alone.Conclusions: These data demonstrate treatment shortening by verapamil adjunctive therapy in mice, and strongly support further study of verapamil and other efflux pump inhibitors in human TB.Keywords: human equivalent doses; verapamil; efflux; Mycobacterium tuberculosis Tuberculosis (TB), a disease affecting millions of people worldwide, requires treatment of patients with multiple drugs for several months. The complexity and length of therapy has led to the emergence of extensively drug-resistant Mycobacterium tuberculosis, which poses a global threat (1). The limited number of new antimicrobials in the TB drug development pipeline further emphasizes the need for fresh approaches in TB therapy. Repurposing existing approved drugs that may serve as treatmentshortening adjuvants is a promising and relatively efficient approach (2). Efflux pump inhibitors (EPIs) are potential agents in this category, and there have been initial promising reports for licensed agents, such as verapamil, reserpine, and piperine (3).Recently, Adams and colleagues (4) showed that the bacterial efflux pump-encoding gene, Rv1258c promotes intracellular bacterial survival, and may mediate drug tolerance. Addition of verapamil reduced tolerance to rifampin in both M. tuberculosis and Mycobacterium marinum, and incubation of M. marinuminfected macrophages with verapamil reduced intracellular bacterial growth. When verapamil was added to isoniazidand rifampin-treated M. marinum-infected macrophages, it restored antibiotic killing by red...

Mycobacterium tuberculosis is an important human pathogen, and yet diagnosis remains challenging. Little research has focused on the impact of M. tuberculosis on the gut microbiota, despite the significant immunological and homeostatic functions of the gastrointestinal tract. To determine the effect of M. tuberculosis infection on the gut microbiota, we followed mice from M. tuberculosis aerosol infection until death, using 16S rRNA sequencing. We saw a rapid change in the gut microbiota in response to infection, with all mice showing a loss and then recovery of microbial community diversity, and found that pre-infection samples clustered separately from post-infection samples, using ecological beta-diversity measures. The effect on the fecal microbiota was observed as rapidly as six days following lung infection. Analysis of additional mice infected by a different M. tuberculosis strain corroborated these results, together demonstrating that the mouse gut microbiota significantly changes with M. tuberculosis infection.