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Efficacy of Bifidobacterium animalis subsp. lactis (BB-12), B. infantis and Lactobacillus acidophilus (La-5) probiotics to prevent gut dysbiosis in preterm infants of 28+0–32+6 weeks of gestation: a randomised, placebo-controlled, double-blind, multicentre trial: the PRIMAL Clinical Study Protocol
  1. Janina Marißen1,
  2. Annette Haiß1,
  3. Claudius Meyer2,
  4. Thea Van Rossum3,
  5. Lisa Marie Bünte1,
  6. David Frommhold4,
  7. Christian Gille5,
  8. Sybelle Goedicke-Fritz6,
  9. Wolfgang Göpel1,
  10. Hannes Hudalla7,
  11. Julia Pagel1,
  12. Sabine Pirr8,
  13. Bastian Siller1,
  14. Dorothee Viemann8,
  15. Maren Vens9,
  16. Inke König9,
  17. Egbert Herting1,
  18. Michael Zemlin6,
  19. Stephan Gehring2,
  20. Peer Bork3,
  21. Philipp Henneke10,
  22. Christoph Härtel1
  23. for the PRIMAL consortium
  1. 1Department of Paediatrics, University of Lübeck, Lübeck, Germany
  2. 2Department of Paediatrics, University Medical Centre, Mainz, Germany
  3. 3Structural and Computational Biology Unit, European Molecular Biology Laboratory, Heidelberg, Germany
  4. 4Children's Hospital Memmingen, Memmingen, Germany
  5. 5Department of Neonatology, University of Tübingen, Tübingen, Germany
  6. 6Department of General Paediatrics and Neonatology, Saarland University, Homburg, Germany
  7. 7Department of Neonatology, University of Heidelberg, Heidelberg, Germany
  8. 8Department of Paediatric Pneumology, Allergology and Neonatology, Hannover Medical School, Hannover, Germany
  9. 9Institute of Medical Biometry and Statistics, University of Lübeck, Lübeck, Germany
  10. 10Insitute for Immunodeficiency (CCI) and Centre for Paediatrics and Adolescent Medicine, Medical Centre and Faculty of Medicine, University of Freiburg, Freiburg, Germany
  1. Correspondence to Professor Christoph Härtel; christoph.haertel{at}


Introduction The healthy ‘eubiosis’ microbiome in infancy is regarded as the microbiome derived from term, vaginally delivered, antibiotic free, breastfed infants at 4–6 months. Dysbiosis is regarded as a deviation from a healthy state with reduced microbial diversity and deficient capacity to control drug-resistant organisms. Preterm infants are highly sensitive to early gut dysbiosis. Latter has been associated with sepsis and necrotising enterocolitis, but may also contribute to long-term health problems. Probiotics hold promise to reduce the risk for adverse short-term outcomes but the evidence from clinical trials remains inconclusive and none has directly assessed the effects of probiotics on the microbiome at high resolution.

Methods and analysis A randomised, double blind, placebo-controlled study has been designed to assess the safety and efficacy of the probiotic mix of Bifidobacterium animalis subsp. lactis, B. infantis and Lactobacillus acidophilus in the prevention of gut dysbiosis in preterm infants between 28+0 and 32+6 weeks of gestation. The study is conducted in 18 German neonatal intensive care units. Between April 2018 and March 2020, 654 preterm infants of 28+0–32+6 weeks of gestation will be randomised in the first 48 hours of life to 28 days of once daily treatment with either probiotics or placebo. The efficacy endpoint is the prevention of gut dysbiosis at day 30 of life. A compound definition of gut dysbosis is used: (1) colonisation with multidrug-resistant organisms or gram-negative bacteria with high epidemic potential or (2) a significant deviation of the gut microbiota composition as compared with healthy term infants. Dysbiosis is determined by (1) conventional microbiological culture and (2) phylogenetic microbiome analysis by high-throughput 16S rRNA and metagenome sequencing. Persistence of dysbiosis will be assessed at 12-month follow-up visits. Side effects and adverse events related to the intervention will be recorded. Key secondary endpoint(s) are putative consequences of dysbiosis. A subgroup of infants will be thoroughly phenotyped for immune parameters using chipcytometry.

Ethics and dissemination Ethics approval was obtained in all participating sites. Results of the trial will be published in peer-review journals, at scientific meetings, on the website ( and via social media of parent organisations.

Trial registration number DRKS00013197; Pre-results.

  • probiotics
  • preterm infants
  • immuno-phenotyping
  • microbiome
  • dysbiosis
  • metagenome sequencing

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  • JM and AH contributed equally.

  • Contributors CH, JM, PH, MZ, SG, PB and EH conceptualised the study, obtained funding, obtained endorsement by PRIMAL consortium and wrote the protocol. IK and MV provided statistical expertise. AH, JM and CH wrote the first draft of the manuscript. CM, TVR, LMB, DF, CG, SG-F, WG, HH, JP, SP, BS, DV and EH provided critical review of the protocol and built enrolment capacity at their sites. All authors read and approved the final manuscript and agree to be accountable for all aspects of the work.

  • Funding This study is supported by the German Federal Ministry of Education and Research (Bundesministerium für Bildung und Forschung, BMBF) grant (01GL1746B).

  • Competing interests The verum was kindly provided by the company Metagenics (Aliso Viejo, California, USA).

  • Patient consent for publication Not required.

  • Ethics approval Institutional review board approvals have been obtained at all participating sites, specifically Ethical Board of the Medical Chamber of the Hansestadt Bremen, Ethical Board of the University of Bonn, Ethical Board of the Medical Chamber of the federal state of Mecklenburg-Vorpommern, Ethical Board of the University of Tübingen, Ethical Board of the Medical School Hannover, Ethical Board of the University of Cologne, Ethical Board of the University of Essen, Ethical Board of the Medical Chamber of the Westphalia-Lippe region, Ethical Board of the Medical Chamber of Hessen, Ethical Board of the Medical Chamber of Hamburg, Ethical Board of the University of Bochum, Ethical Board of the University of Freiburg, Ethical Board of the Medical Chamber of the federal state of Bavaria, Ethical Board of the University of Heidelberg, Ethical Board of the EMBL Heidelberg, Ethical Board of the University of Lübeck, Ethical Board of the University of Jena, Ethical Board of the Saar University Homburg.

  • Provenance and peer review Not commissioned; externally peer reviewed.

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