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Comprehensive insights into transcriptional adaptation of intracellular mycobacteria by microbe-enriched dual RNA sequencing

  • Rienksma, Rienk A1
  • Suarez-Diez, Maria1
  • Mollenkopf, Hans-Joachim2
  • Dolganov, Gregory M3
  • Dorhoi, Anca4
  • Schoolnik, Gary K3
  • Martins dos Santos, Vitor AP1, 5
  • Kaufmann, Stefan HE4
  • Schaap, Peter J1
  • Gengenbacher, Martin4, 6
  • 1 Laboratory of Systems and Synthetic Biology, Wageningen University and Research Centre, Dreijenplein 10, Wageningen, HB, 6703, the Netherlands , Wageningen (Netherlands)
  • 2 Core Facility Microarray/Genomics, Max Planck Institute for Infection Biology, Charitéplatz 1, Berlin, 10117, Germany , Berlin (Germany)
  • 3 Stanford University School of Medicine, Department of Microbiology and Immunology, 300 Pasteur Drive, Stanford, CA, 94305-5124, USA , Stanford (United States)
  • 4 Max Planck Institute for Infection Biology, Department of Immunology, Charitéplatz 1, Berlin, 10117, Germany , Berlin (Germany)
  • 5 LifeGlimmer GmbH, Markelstrasse 38, Berlin, 12163, Germany , Berlin (Germany)
  • 6 Yong Loo Lin School of Medicine, National University of Singapore, Present address: Department of Microbiology, 5 Science Drive 2, Singapore, 117545, Singapore , Singapore (Singapore)
Published Article
BMC Genomics
Springer (Biomed Central Ltd.)
Publication Date
Feb 05, 2015
DOI: 10.1186/s12864-014-1197-2
Springer Nature


BackgroundThe human pathogen Mycobacterium tuberculosis has the capacity to escape eradication by professional phagocytes. During infection, M. tuberculosis resists the harsh environment of phagosomes and actively manipulates macrophages and dendritic cells to ensure prolonged intracellular survival. In contrast to other intracellular pathogens, it has remained difficult to capture the transcriptome of mycobacteria during infection due to an unfavorable host-to-pathogen ratio.ResultsWe infected the human macrophage-like cell line THP-1 with the attenuated M. tuberculosis surrogate M. bovis Bacillus Calmette–Guérin (M. bovis BCG). Mycobacterial RNA was up to 1000-fold underrepresented in total RNA preparations of infected host cells. We employed microbial enrichment combined with specific ribosomal RNA depletion to simultaneously analyze the transcriptional responses of host and pathogen during infection by dual RNA sequencing. Our results confirm that mycobacterial pathways for cholesterol degradation and iron acquisition are upregulated during infection. In addition, genes involved in the methylcitrate cycle, aspartate metabolism and recycling of mycolic acids were induced. In response to M. bovis BCG infection, host cells upregulated de novo cholesterol biosynthesis presumably to compensate for the loss of this metabolite by bacterial catabolism.ConclusionsDual RNA sequencing allows simultaneous capture of the global transcriptome of host and pathogen, during infection. However, mycobacteria remained problematic due to their relatively low number per host cell resulting in an unfavorable bacterium-to-host RNA ratio. Here, we use a strategy that combines enrichment for bacterial transcripts and dual RNA sequencing to provide the most comprehensive transcriptome of intracellular mycobacteria to date. The knowledge acquired into the pathogen and host pathways regulated during infection may contribute to a solid basis for the deployment of novel intervention strategies to tackle infection.

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