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Developmental dynamics of the preterm infant gut microbiota and antibiotic resistome

Nature Microbiology volume 1, Article number: 16024 (2016) Cite this article

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Abstract

Development of the preterm infant gut microbiota is emerging as a critical research priority1. Since preterm infants almost universally receive early and often extended antibiotic therapy2, it is important to understand how these interventions alter gut microbiota development3–6. Analysis of 401 stools from 84 longitudinally sampled preterm infants demonstrates that meropenem, cefotaxime and ticarcillin–clavulanate are associated with significantly reduced species richness. In contrast, vancomycin and gentamicin, the antibiotics most commonly administered to preterm infants, have non-uniform effects on species richness, but these can be predicted with 85% accuracy based on the relative abundance of only two bacterial species and two antibiotic resistance (AR) genes at treatment initiation. To investigate resistome development, we functionally selected resistance to 16 antibiotics from 21 faecal metagenomic expression libraries. Of the 794 AR genes identified, 79% had not previously been classified as AR genes. Combined with deep shotgun sequencing of all stools, we find that multidrug-resistant members of the genera Escherichia, Klebsiella and Enterobacter, genera commonly associated with nosocomial infections, dominate the preterm infant gut microbiota. AR genes that are enriched following specific antibiotic treatments are generally unique to the specific treatment and are highly correlated with the abundance of a single species. The most notable exceptions include ticarcillin–clavulanate and ampicillin, both of which enrich for a large number of overlapping AR genes, and are correlated with Klebsiella pneumoniae. We find that all antibiotic treatments are associated with widespread collateral microbiome impact by enrichment of AR genes that have no known activity against the specific antibiotic driver.

Using a combination of metagenomic shotgun sequencing and functional selections, we deeply interrogated the gut microbiota and resistome of 401 stools from 84 preterm infants sampled longitudinally and stratified by antibiotic use during their hospitalization (Table 1, Supplementary Figs 2–3 and Supplementary Table 1a–c). All infants included were born prematurely (<33 weeks gestational age; median (interquartile range (IQR)) 27 weeks (25–29 weeks)) and of low birthweights (median (IQR) of 865 g (718–1,141 g)). In contrast to term infants7–10, we found the preterm infant gut microbiota is colonized by approximately tenfold fewer bacterial species, compositionally distinct, and established independent of delivery mode (Supplementary Fig. 1). While all but two infants amongst the 84 in our cohort received antibiotic therapy within 24 h of birth, 39% received no additional antibiotics after their first week of life (Table 1) and were used as controls to study the effect of specific antibiotic treatments administered between 1 and 10 weeks of life. Infants who were treated with antibiotics after the first week of life received a median (IQR) of 21 (16–32) total days of additional therapy during their hospitalization. Samples were analysed over time, ranging from 6.1 to 157.7 days of life, with a median (IQR) of 3 (2–6) samples analysed per individual. An extensive set of clinical covariates were considered for their impact on the developing gut microbiota (Supplementary Table 1a–b).

Table 1 Details of preterm infant cohort analysed in this study.
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Figure 1: Development of the preterm infant gut microbiota and antibiotic use.
Figure 2: Functional metagenomic selections identify novel antibiotic resistance gene clusters in Proteobacterial species in the preterm infant gut.
Figure 3: Species and antibiotic resistance gene enrichment following specific antibiotic treatments.

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Acknowledgements

We thank A. Moore for initial study conception and clinical insight, C. Hall-Moore for assistance in access to cohort samples, M. Wallace for cultured strain characterization, and members of the Dantas lab for discussions of the results and analyses. We also thank families and clinical staff in the Saint Louis Children's Hospital Neonatal Intensive Care Unit for their cooperation with study, and Laura Linneman and Julie Hoffmann for their efforts in enrolment and data accrual. This work is supported in part by awards to G.D. through the Children's Discovery Institute (MD-II-2011-117 and 127), the March of Dimes Foundation (6-FY12-394), and the National Institute of General Medical Sciences of the National Institutes of Health (R01-GM099538). This work is also supported in part by an award to Washington University School of Medicine through a Clinical and Translational Science Award (CTSA) Grant (UL1 TR000448). This collection was also supported by the US National Institutes of Health Grants UH3AI083265 and P30DK052574 (Biobank Core), along with funding from the Eunice Kennedy Shriver National Institute of Child Health and Human Development, and Foundation for the National Institutes of Health (made possible by support from the Gerber Foundation). M.K.G. is a Mr. and Mrs. Spencer T. Olin Fellow at Washington University and a National Science Foundation (NSF) graduate research fellow (DGE-1143954). The content is solely the responsibility of the authors and does not necessarily represent the official views of the funding agencies.

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Authors and Affiliations

  1. Center for Genome Sciences and Systems Biology, Washington University School of Medicine, St Louis, Missouri, USA

    Molly K. Gibson, Bin Wang, Sara Ahmadi & Gautam Dantas

  2. Department of Pathology and Immunology, Washington University School of Medicine, St Louis, Missouri, USA

    Bin Wang, Sara Ahmadi, Carey-Ann D. Burnham & Gautam Dantas

  3. Department of Pediatrics, Washington University School of Medicine, St Louis, Missouri, USA

    Carey-Ann D. Burnham, Phillip I. Tarr & Barbara B. Warner

  4. Department of Molecular Microbiology, Washington University School of Medicine, St Louis, Missouri, USA

    Phillip I. Tarr & Gautam Dantas

  5. Department of Biomedical Engineering, Washington University, St Louis, Missouri, USA

    Gautam Dantas

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Contributions

P.I.T. and B.B.W. assembled the cohort and developed protocols and infrastructure to obtain the biologics and clinical metadata from the cohort. M.K.G. and G.D. designed and conceived the study. M.K.G. performed 16S rRNA gene sequencing for comparison to term infants. B.W. prepared shotgun metagenomic sequencing libraries. S.A. created metagenomic libraries, performed functional selections, and prepared sequencing libraries. M.K.G. performed functional metagenomic data analysis, shotgun metagenomic data analysis, statistical modelling. M.K.G. wrote the manuscript with contributions from C.A.B., P.I.T., B.B.W., and G.D.

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Correspondence to Gautam Dantas.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Figures 1-9 (PDF 646 kb)

Supplementary Table 1

Metadata for 401 stool samples and 84 individuals. (XLSX 192 kb)

Supplementary Table 2

Functional metagenomic selections of 21 fecal metagenomic expression libraries to 16 antibiotics. (XLSX 39 kb)

Supplementary Table 3

VelvetOptimiser assembly stats for 312 assembled preterm infant meatgenomes. (XLSX 61 kb)

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Gibson, M., Wang, B., Ahmadi, S. et al. Developmental dynamics of the preterm infant gut microbiota and antibiotic resistome. Nat Microbiol 1, 16024 (2016). https://doi.org/10.1038/nmicrobiol.2016.24

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