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Fetal meconium does not have a detectable microbiota before birth


Microbial colonization of the human intestine impacts host metabolism and immunity; however, exactly when colonization occurs is unclear. Although many studies have reported bacterial DNA in first-pass meconium samples, these samples are typically collected hours to days after birth. Here, we investigated whether bacteria could be detected in meconium before birth. Fetal meconium (n = 20) was collected by rectal swab during elective breech caesarean deliveries without labour and before antibiotics and compared to technical and procedural controls (n = 5), first-pass meconium (neonatal meconium; n = 14) and infant stool (n = 25). Unlike first-pass meconium, no microbial signal distinct from negative controls was detected in fetal meconium by 16S ribosomal RNA gene sequencing. Additionally, positive aerobic (n = 10 of 20) and anaerobic (n = 12 of 20) clinical cultures of fetal meconium (13 of 20 samples positive in at least one culture) were identified as likely skin contaminants, most frequently Staphylococcus epidermidis, and not detected by sequencing in most samples (same genera detected by culture and sequencing in 2 of 13 samples with positive culture). We conclude that fetal gut colonization of healthy term infants does not occur before birth and that microbial profiles of neonatal meconium reflect populations acquired during and after birth.

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Fig. 1: Diagram of the collection method for fetal meconium samples.
Fig. 2: Fetal meconium alpha and beta diversity do not differ from those of sampling negative controls.
Fig. 3: Neighbour-joining phylogenetic tree of all genera and presence/absence in each sample.

Data availability

All sequencing data associated with this study have been made publicly available in the National Center for Biotechnology Information Sequence Read Archive under project ID PRJNA666699. Source data are provided with this paper.

Code availability

Custom scripts for the microbiome analyses are available from the GitHub repository at


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We thank all the participants that were recruited in this study. We thank H. Brinkmann, L. Pasura, L. Maschirow and A. Schwickert for assisting with patient recruitment, L. Ehrlich with sample preparation and K. von Weizsaecker and W. Henrich for their external review of the microbiology protocol and advice on protocol improvements. We thank M. Shah for performing the genomic DNA extractions. T.B. and M.M.H. are supported by the Deutsche Forschungsgemeinschaft (German Research Foundation). K.M.K. is supported by a Farncombe Digestive Health Research Institute Student Fellowship. M.G.S. and D.M.S. are supported by the Canada Research Chairs Program. The laboratory analyses including sequencing were supported by funds from the Canadian Institute for Health Research Team Grant no. MWB 141879.

Author information

Authors and Affiliations



K.M.K. analysed the sequencing data and wrote the manuscript. M.J.G. contributed to sample collection and wrote the ‘Study design and sample collection’ section of the Methods. T.A. supervised the culture-based analyses. M.M.H. assisted with study design. L.R. assisted with study design and performed the V3–V4 amplifications and processing of raw sequencing data. M.G.S. assisted with study design and analysis of the sequencing data. D.M.S. contributed to data analysis and manuscript development. T.B. designed the study and contributed to sample collection. All authors discussed the analyses and results and edited the manuscript.

Corresponding authors

Correspondence to Deborah M. Sloboda or Thorsten Braun.

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The authors declare no competing interests.

Additional information

Peer review information Nature Microbiology thanks the anonymous reviewers for their contribution to the peer review of this work. Peer reviewer reports are available.

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Extended data

Extended Data Fig. 1 Strobe flowchart.

The study flowchart in line with the STROBE (Strengthening the Reporting of Observational Studies in Epidemiology) statement ( Pilot cohort 1A and pilot cohort 1B were used to optimize sample collection. The principal study cohort was analysed by culture and sequencing.

Extended Data Fig. 2 Optimization of collection to reduce contamination.

We considered potential problems related to contamination (birth mode, sampling, sample preparation, general) and sensitivity of culture methods and present our solutions to these problems (see Methods).

Source data

Extended Data Fig. 3 Detection of genera across technical replicates.

The abundance (read count) is shown for each sample by sequencing run (run 1 and run 2) for 30 cycles of PCR amplification. Genera are only shown if they were also detected within the same sample’s sequencing data from 40 cycles of PCR amplification.

Supplementary information

Reporting Summary

Peer Review File

Supplementary Tables

Supplementary Table 1. Pilot cohort culture results. Supplementary Table 2. ASV table of negative controls, fetal meconium, neonatal meconium and infant stool samples (listed in ST2). Supplementary Table 3. Sample data for ASV table (ST2). Supplementary Table 4. Agreement of sequencing runs. Supplementary Table 5. Use of negative controls in previous studies.

Source data

Source Data Fig. 2

a, Alpha diversity values for each sample. b, Axes 1 and 2 values for each sample. c, Bray–Curtis dissimilarity values for each sample pair.

Source Data Extended Data Fig. 2

Genus abundance data for each sample and each sequencing run.

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Kennedy, K.M., Gerlach, M.J., Adam, T. et al. Fetal meconium does not have a detectable microbiota before birth. Nat Microbiol 6, 865–873 (2021).

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