Mycobacterium tuberculosis, the cause of Tuberculosis (TB), infects one third of the world’s population and causes substantial mortality worldwide. In its shortest format, treatment of TB requires six months of multidrug therapy with a mixture of broad spectrum and mycobacterial specific antibiotics, and treatment of multidrug resistant TB is longer. The widespread use of this regimen makes this one of the largest exposures of humans to antimicrobials, yet the effects of TB treatment on intestinal microbiome composition and long-term stability are unknown. We compared the microbiome composition, assessed by both 16S rDNA and metagenomic DNA sequencing, of TB cases during antimycobacterial treatment and following cure by 6 months of antibiotics. TB treatment does not perturb overall diversity, but nonetheless dramatically depletes multiple immunologically significant commensal bacteria. The microbiomic perturbation of TB therapy can persist for at least 1.2 years, indicating that the effects of TB treatment are long lasting. These results demonstrate that TB treatment has dramatic effects on the intestinal microbiome and highlight unexpected durable consequences of treatment for the world’s most common infection on human ecology.

The ability of science and medicine to control the pathogen Mycobacterium tuberculosis (Mtb) requires an understanding of the complex host environment within which it resides. Pathological and biological evidence overwhelmingly demonstrate how the mammalian steroid cholesterol is present throughout the course of infection. Better understanding Mtb requires a more complete understanding of how it utilizes molecules like cholesterol in this environment to sustain the infection of the host. Cholesterol uptake, catabolism, and broader utilization are important for maintenance of the pathogen in the host and it has been experimentally validated to contribute to virulence and pathogenesis. Cholesterol is catabolized by at least three distinct sub-pathways, two for the ring system and one for the side chain, yielding dozens of steroid intermediates with varying biochemical properties. Our ability to control this worldwide infectious agent requires a greater knowledge of how Mtb uses cholesterol to its advantage throughout the course of infection. Herein, the current state of knowledge of cholesterol metabolism by Mtb is reviewed from a biochemical perspective with a focus on the metabolic genes and pathways responsible for cholesterol steroid catabolism.

BackgroundEffective treatment of Mycobacterium tuberculosis (Mtb) infection requires at least 6 months of daily therapy with multiple orally administered antibiotics. Although this drug regimen is administered annually to millions worldwide, the impact of such intensive antimicrobial treatment on the host microbiome has never been formally investigated. Here, we characterized the longitudinal outcome of conventional isoniazid-rifampin-pyrazinamide (HRZ) TB drug administration on the diversity and composition of the intestinal microbiota in Mtb-infected mice by means of 16S rRNA sequencing. We also investigated the effects of each of the individual antibiotics alone and in different combinations.ResultsWhile inducing only a transient decrease in microbial diversity, HRZ treatment triggered a marked, immediate and reproducible alteration in community structure that persisted for the entire course of therapy and for at least 3 months following its cessation. Members of order Clostridiales were among the taxa that decreased in relative frequencies during treatment and family Porphyromonadaceae significantly increased post treatment. Experiments comparing monotherapy and different combination therapies identified rifampin as the major driver of the observed alterations induced by the HRZ cocktail but also revealed unexpected effects of isoniazid and pyrazinamide in certain drug pairings.ConclusionsThis report provides the first detailed analysis of the longitudinal changes in the intestinal microbiota due to anti-tuberculosis therapy. Importantly, many of the affected taxa have been previously shown in other systems to be associated with modifications in immunologic function. Together, our findings reveal that the antibiotics used in conventional TB treatment induce a distinct and long lasting dysbiosis. In addition, they establish a murine model for studying the potential impact of this dysbiosis on host resistance and physiology.Electronic supplementary materialThe online version of this article (doi:10.1186/s40168-017-0286-2) contains supplementary material, which is available to authorized users.

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The ability of the pathogen Mycobacterium tuberculosis to metabolize steroids like cholesterol and the roles that these compounds play in the virulence and pathogenesis of this organism are increasingly evident. Here, we demonstrate through experiments and bioinformatic analysis the existence of an architecturally distinct subfamily of acyl coenzyme A (acyl-CoA) dehydrogenase (ACAD) enzymes that are ␣ 2 ␤ 2 heterotetramers with two active sites. These enzymes are encoded by two adjacent ACAD (fadE) genes that are regulated by cholesterol.

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The metabolism of host cholesterol by Mycobacterium tuberculosis (Mtb) is an important factor for both its virulence and pathogenesis, although how and why cholesterol metabolism is required is not fully understood. Mtb uses a unique set of catabolic enzymes that are homologous to those required for classical β-oxidation of fatty acids but are specific for steroid-derived substrates. Here, we identify and assign the substrate specificities of two of these enzymes, ChsE4-ChsE5 (Rv3504-Rv3505) and ChsE3 (Rv3573c), that carry out cholesterol side chain oxidation in Mtb. Steady-state assays demonstrate that ChsE4-ChsE5 preferentially catalyzes the oxidation of 3-oxo-cholest-4-en-26-oyl CoA in the first cycle of cholesterol side chain β-oxidation that ultimately yields propionyl-CoA, whereas ChsE3 specifically catalyzes the oxidation of 3-oxo-chol-4-en-24-oyl CoA in the second cycle of β-oxidation that generates acetyl-CoA. However, ChsE4-ChsE5 can catalyze the oxidation of 3-oxo-chol-4-en-24-oyl CoA as well as 3-oxo-4-pregnene-20-carboxyl-CoA. The functional redundancy of ChsE4-ChsE5 explains the in vivo phenotype of the igr knockout strain of Mycobacterium tuberculosis; the loss of ChsE1-ChsE2 can be compensated for by ChsE4-ChsE5 during the chronic phase of infection. The X-ray crystallographic structure of ChsE4-ChsE5 was determined to a resolution of 2.0 Å and represents the first high-resolution structure of a heterotetrameric acyl-CoA dehydrogenase (ACAD). Unlike typical homotetrameric ACADs that bind four flavin adenine dinucleotide (FAD) cofactors, ChsE4-ChsE5 binds one FAD at each dimer interface, resulting in only two substrate-binding sites rather than the classical four active sites. A comparison of the ChsE4-ChsE5 substrate-binding site to those of known mammalian ACADs reveals an enlarged binding cavity that accommodates steroid substrates and highlights novel prospects for designing inhibitors against the committed β-oxidation step in the first cycle of cholesterol side chain degradation by Mtb.

Tuberculosis (TB) is an ancient infectious disease of humans that has been extensively studied both clinically and experimentally. Although susceptibility to Mycobacterium tuberculosis infection is clearly influenced by factors such as nutrition, immune status, and both mycobacterial and host genetics, the variable pathogenesis of TB in infected individuals remains poorly understood.