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Trust is good, control is better: technical considerations in blood microbiome analysis
  1. Robert Schierwagen1,
  2. Camila Alvarez-Silva2,
  3. Florence Servant3,
  4. Jonel Trebicka1,
  5. Benjamin Lelouvier3,
  6. Manimozhiyan Arumugam2
  1. 1 Translational Hepatology, Department of Internal Medicine I, Universitätsklinikum/Goethe-Universität, Frankfurt, Germany
  2. 2 The Novo Nordisk Foundation Center for Basic Metabolic Research, University of Copenhagen, Copenhagen, Denmark
  3. 3 Vaiomer SAS, Labège, France
  1. Correspondence to Dr Jonel Trebicka, Translational Hepatology Department of Internal Medicine I, Universitätsklinikum/ Goethe-Universität, Frankfurt, Germany; Jonel.Trebicka{at}

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We agree with Hornung et al 1 that studying blood microbiome is a major technical challenge with potential artefacts. At least three important challenges must be tackled:

  1. Low amount of bacterial DNA in blood.2

  2. High amounts of PCR inhibitors.

  3. Bacterial DNA contaminants from environment, reagents and consumables.

Measuring, reducing and controlling bacterial contaminants are key elements of optimisations made on the molecular pipeline used in our study3 as well as eight published studies on blood microbiome.2 4–7 The studies from Salter et al 8 and Laurence et al 9 are useful to understand the burden of bacterial contaminants when working with low bacterial abundance samples. In former publications,2 10 we have described our procedure and the controls performed to address such contamination. One must be careful when using a fixed list of bacterial contaminants, as each experiment has its own contamination burden. Therefore, two different experiments done under different conditions, will not have the same contaminants. What is essential, as pointed out by Hornung et al, is to include and analyse negative controls in each experiment. Although not explicitly mentioned before, our study3 included the following negative controls:

  1. Extraction negative controls (water at DNA extraction step).

  2. PCR negative controls (water at first PCR step).

We …

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