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Questioning the fetal microbiome illustrates pitfalls of low-biomass microbial studies

Abstract

Whether the human fetus and the prenatal intrauterine environment (amniotic fluid and placenta) are stably colonized by microbial communities in a healthy pregnancy remains a subject of debate. Here we evaluate recent studies that characterized microbial populations in human fetuses from the perspectives of reproductive biology, microbial ecology, bioinformatics, immunology, clinical microbiology and gnotobiology, and assess possible mechanisms by which the fetus might interact with microorganisms. Our analysis indicates that the detected microbial signals are likely the result of contamination during the clinical procedures to obtain fetal samples or during DNA extraction and DNA sequencing. Furthermore, the existence of live and replicating microbial populations in healthy fetal tissues is not compatible with fundamental concepts of immunology, clinical microbiology and the derivation of germ-free mammals. These conclusions are important to our understanding of human immune development and illustrate common pitfalls in the microbial analyses of many other low-biomass environments. The pursuit of a fetal microbiome serves as a cautionary example of the challenges of sequence-based microbiome studies when biomass is low or absent, and emphasizes the need for a trans-disciplinary approach that goes beyond contamination controls by also incorporating biological, ecological and mechanistic concepts.

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Fig. 1: Relative abundance of bacterial taxa from three recent fetal studies.
Fig. 2: Reagent contamination in meconium samples from extremely premature infants.
Fig. 3: Relative abundance of bacterial taxa in samples from Rackaityte et al.

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Acknowledgements

T.B. receives funding from the Deutsche Forschungsgemeinschaft (German Research Foundation no. BR2925 10-1 & PL241 16-1). F.D.B. is funded by AI045008, AI120489, R33HL137063, CA219871, AI139240, and the PennCHOP Microbiome Program. J.D. acknowledges funding from the European Research Council (ERC) under the European Union’s Horizon 2020 research and innovation programme (grant agreement ERC-2017-AdG no. 788191—Homo.symbiosus). W.M.d.V. is supported by the Gravitation grant 024.002.002 of the Netherlands Organization for Scientific Research. W.M.d.V. and A.S. are supported by the Academy of Finland (grants 1308255 and 1325103). A.M.E. is funded in part with Federal funds from the National Institute of Allergy and Infectious Diseases (NIAID), National Institutes of Health, Department of Health and Human Services, under grant number U19AI110818 to the Broad Institute. M.A.E. is funded through grants R01HD102318, R01HD098867 and R01NR014784. S.C.G.-V. was funded through a Peter Hans Hofschneider Professorship provided by the Stiftung Molekulare Biomedizin. M.G.G. and D.M.S are funded by the Canada Research Chairs Program. L.J.H. is supported by Wellcome Trust Investigator Awards 100974/C/13/Z and 220876/Z/20/Z and by the Biotechnology and Biological Sciences Research Council (BBSRC) Institute Strategic Programme Gut Microbes and Health BB/R012490/1 and its constituent projects BBS/E/F/000PR10353 and BBS/E/F/000PR10356. M.W.H. has received funding from the ERC under the European Union’s Horizon 2020 research and innovation programme (grant agreement no. 101019157). S.L. has received funding from the ERC under the European Union’s Horizon 2020 research and innovation programme (grant agreement 852600 Lacto-Be). A.J.M. receives funding from ERCAd HHMM-Neonates and Swiss National Science Sinergia. O.K. is supported by the ERC under the European Union’s Horizon 2020 research and innovation programme (grant agreement ERC-2020-COG no. 101001355). Work in the laboratories of P.W.O., L.O. and J.W. is supported by Science Foundation Ireland (SFI) through a Centre award (APC/SFI/12/RC/2273_P2) to APC Microbiome Ireland. J.W. acknowledges support through an SFI Professorship (19/RP/6853) and thanks V. McMahon for coordination of this review and R. O’Callaghan for encouragement. J.R. acknowledges funding from the Interuniversity Special Research Fund (iBOF) Flanders (FLEXIGUT R-11423), the Rega Institute, VIB and KU Leuven. N.S. receives funding from the ERC (ERC-STG project MetaPG-716575 and ERC-CoG microTOUCH-101045015) and from the European H2020 program (ONCOBIOME-825410 project, MASTER-818368 project and IHMCSA-964590). F.S. is supported in part by Science Foundation Ireland. G.C.S.S. acknowledges funding from the Medical Research Council (UK; MR/K021133/1) and the National Institute for Health Research (NIHR) Cambridge Biomedical Research Centre (Women’s Health theme). D.M.S. is funded by the Canadian Institute for Health Research and the Canada Research Chairs Program. A.W.W. receives core funding support from the Scottish Government’s Rural and Environment Science and Analytical Services (RESAS). M.Y. is supported by the Azrieli Faculty Fellowship.

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N.S. and J.W. conceived the project. K.M.K. and M.C.d.G. performed analyses and generated figures. K.M.K., M.C.d.G., M.E.P.-M., F.D.B., M.A.E., S.C.G.-V., M.G.G., M.W.H., A.J.M., R.C.M., E.G.P., J.P., F.S., D.M.S., G.C.S.S., G.W.T., A.W.W., and J.W. wrote the draft. All authors provided feedback, participated in discussions and contributed to the final version of the manuscript.

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Kennedy, K.M., de Goffau, M.C., Perez-Muñoz, M.E. et al. Questioning the fetal microbiome illustrates pitfalls of low-biomass microbial studies. Nature 613, 639–649 (2023). https://doi.org/10.1038/s41586-022-05546-8

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