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Fluorescent in situ hybridization of Mycobacterium tuberculosis rRNA species for analysis of ribosomal synthesis as a marker of bacterial growth of individual bacilli in distinct lesional microenvironments of lung granulomas from C3HeB/FeJ mice

Citation

Vasquez, Joshua; Maidji, Ekaterina; Karthikeyan, Dhuvarakesh; Eshun-Wilson, Franceen (2021), Fluorescent in situ hybridization of Mycobacterium tuberculosis rRNA species for analysis of ribosomal synthesis as a marker of bacterial growth of individual bacilli in distinct lesional microenvironments of lung granulomas from C3HeB/FeJ mice, Dryad, Dataset, https://doi.org/10.7272/Q6Z036DR

Abstract

There is urgent need for new drug regimens that more rapidly cure tuberculosis (TB). Existing TB drugs and regimens vary in treatment-shortening activity, but the molecular basis of these differences is unclear, and no existing assay directly quantifies the ability of a drug or regimen to shorten treatment. Drugs historically classified as sterilizing and non-sterilizing have distinct impacts on a fundamental aspect of Mycobacterium tuberculosis physiology: ribosomal RNA (rRNA) synthesis. In culture, in mice, and in human studies, measurement of precursor rRNA reveals that sterilizing drugs and highly effective drug regimens profoundly suppress M. tuberculosis rRNA synthesis, whereas non-sterilizing drugs and weaker regimens do not. The rRNA synthesis ratio provides a readout of drug effect that is orthogonal to traditional measures of bacterial burden. Here, we show that M. tuberculosis have markedly different levels of ongoing rRNA synthesis in varying key histologic microenvironments. 

Usage Notes

Raw images and datasets of single-bacillary quantitative image analysis of multiplexed in situ hybridization (ISH) of rRNA species in Mycobacterium tuberculosis (Mtb) in lung granulomas from C3HeB/FeJ mice. Granulomas were stained by fluorescent ISH for Mtb 23S rRNA and pre-rRNA, and counterstained with DAPI. Datasets are available as *csv and original images used for analysis are available as *tif or *czi. Observations from analysis of these data have been submitted for review to Nature Communications as figures 2b, 2e and extended figure 2, 3b and 3c from the manuscript entitled "Mycobacterium tuberculosis precursor rRNA indicates treatment-shortening activity of drugs and regimens".