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Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages.

ABSTRACT: Tuberculosis, caused by the intracellular pathogen Mycobacterium tuberculosis, remains the world's deadliest infectious disease. Sterilizing chemotherapy requires at least 6 months of multidrug therapy. Difficulty visualizing the subcellular localization of antibiotics in infected host cells means that it is unclear whether antibiotics penetrate all mycobacteria-containing compartments in the cell. Here, we combined correlated light, electron, and ion microscopy to image the distribution of bedaquiline in infected human macrophages at submicrometer resolution. Bedaquiline accumulated primarily in host cell lipid droplets, but heterogeneously in mycobacteria within a variety of intracellular compartments. Furthermore, lipid droplets did not sequester antibiotic but constituted a transferable reservoir that enhanced antibacterial efficacy. Thus, strong lipid binding facilitated drug trafficking by host organelles to an intracellular target during antimicrobial treatment.

SUBMITTER: Greenwood DJ

PROVIDER: S-EPMC7012645 | biostudies-literature | 2019 Jun

REPOSITORIES: biostudies-literature

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Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages.

Greenwood Daniel J DJ Dos Santos Mariana Silva MS Huang Song S Russell Matthew R G MRG Collinson Lucy M LM MacRae James I JI West Andy A Jiang Haibo H Gutierrez Maximiliano G MG

Science (New York, N.Y.) 20190627 6447

Tuberculosis, caused by the intracellular pathogen <i>Mycobacterium tuberculosis</i>, remains the world's deadliest infectious disease. Sterilizing chemotherapy requires at least 6 months of multidrug therapy. Difficulty visualizing the subcellular localization of antibiotics in infected host cells means that it is unclear whether antibiotics penetrate all mycobacteria-containing compartments in the cell. Here, we combined correlated light, electron, and ion microscopy to image the distribution ...[more]

PMID: 31249058

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Project description:A human immunodeficiency virus (HIV) infection cure requires an understanding of the cellular and anatomical sites harboring virus that contribute to viral rebound upon treatment interruption. Despite antiretroviral therapy (ART), HIV-associated neurocognitive disorders (HAND) are reported in HIV-infected individuals on ART. Biomarkers for macrophage activation and neuronal damage in cerebrospinal fluid (CSF) of HIV-infected individuals demonstrate continued effects of HIV in brain and suggest that the central nervous system (CNS) may serve as a viral reservoir. Using a simian immunodeficiency virus (SIV)/macaque model for HIV encephalitis and AIDS, we evaluated whether infected cells persist in brain despite ART. Eight SIV-infected pig-tailed macaques were virally suppressed with ART, and plasma and CSF viremia levels were analyzed longitudinally. To assess whether virus persisted in brain macrophages (BrMΦ) in these macaques, we used a macrophage quantitative viral outgrowth assay (MΦ-QVOA), PCR, and in situ hybridization (ISH) to measure the frequency of infected cells and the levels of viral RNA and DNA in brain. Viral RNA in brain tissue of suppressed macaques was undetectable, although viral DNA was detected in all animals. The MΦ-QVOA demonstrated that the majority of suppressed animals contained latently infected BrMΦ. We also showed that virus produced in the MΦ-QVOAs was replication competent, suggesting that latently infected BrMΦ are capable of reestablishing productive infection upon treatment interruption. This report provides the first confirmation of the presence of replication-competent SIV in BrMΦ of ART-suppressed macaques and suggests that the highly debated issue of viral latency in macrophages, at least in brain, has been addressed in SIV-infected macaques treated with ART.IMPORTANCE Resting CD4+ T cells are currently the only cells that fit the definition of a latent reservoir. However, recent evidence suggests that HIV/SIV-infected macrophages persist despite ART. Markers of macrophage activation and neuronal damage are observed in the CSF of HIV-infected individuals and of SIV-infected macaques on suppressive ART regimens, suggesting that the CNS has continued virus infection and latent infection. A controversy exists as to whether brain macrophages represent a latent source of replication-competent virus capable of reestablishing infection upon treatment interruption. In this study, we demonstrated the presence of the latent macrophage reservoir in brains of SIV-infected ART-treated macaques and analyzed the reservoir using our established outgrowth assay to quantitate macrophages harboring replication-competent SIV genomes. Our results support the idea of the existence of other latent reservoirs in addition to resting CD4+ T cells and underscore the importance of macrophages in developing strategies to eradicate HIV.

Project description:BackgroundYamuna, a major tributary of Ganga, which flows through the national capital region of Delhi, is among the major polluted rivers in India. The accumulation of various effluents, toxic chemicals, heavy metals, and increased organic load in the Yamuna directly affects the organisms that thrive inside or around this river. It also makes it an ideal site for studying the impact of pollution on the river microflora, which are sentinels of the water quality.ResultsIn this study, the microbial community structure and functional diversity of the Yamuna river water was assessed from the New Delhi region. The community structure of Yamuna during pre-monsoon (June) was found to be significantly different from the post-monsoon (November) time, with Acinetobacter being the most abundant genus during June, and Aeromonas during November. The functional characterization revealed the higher abundance of Methyl-accepting chemotaxis protein in the river water, which could be important for the microbial chemosensory adaptation in the environment. A higher abundance of genes related to nitrogen and sulfur metabolism, metal tolerance, and xenobiotic degradation, and complete degradation pathways of aromatic compounds such as toluene, xylene, benzene and phenol were identified. Further, the results showed the presence of a pool of antibiotic resistance genes in the bacterial microbiome in the Yamuna alongside a large number of broad-spectrum antibiotics, such as carbapenemases and metallo-β-lactamases. Efflux mechanism of resistance was found to dominate among these microbes conferring multi-drug resistance. The Principal Coordinate Analysis of the taxonomic composition of the Yamuna River water with publicly available freshwater and sewage datasets revealed significant differences in the two Yamuna samples and a greater resemblance of pre-monsoon Yamuna sample to sewage sample owing to the higher pollution levels in Yamuna in the pre-monsoon time.ConclusionThe metagenomic study of the Yamuna river provides the first insights on the bacterial microbiome composition of this large polluted river, and also helps to understand the dynamics in the community structure and functions due to seasonal variations. The presence of antibiotic resistance genes and functional insights on the metabolic potential of a polluted river microbiome are likely to have several applications in health, biotechnology and bioremediation.

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Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages.

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Subcellular antibiotic visualization reveals a dynamic drug reservoir in infected macrophages.

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