High throughput cultivation-based screening on porous aluminum oxide chips allows targeted isolation of antibiotic resistant human gut bacteria

Dennis Versluis*, Teresita de J. Bello González, Erwin G. Zoetendal, Mark W.J. van Passel, Hauke Smidt

*Corresponding author for this work

Research output: Contribution to journalArticleAcademicpeer-review

1 Citation (Scopus)

Abstract

The emergence of bacterial pathogens that are resistant to clinical antibiotics poses an increasing risk to human health. An important reservoir from which bacterial pathogens can acquire resistance is the human gut microbiota. However, thus far, a substantial fraction of the gut microbiota remains uncultivated and has been little-studied with respect to its resistance reservoir-function. Here, we aimed to isolate yet uncultivated resistant gut bacteria by a targeted approach. Therefore, faecal samples from 20 intensive care patients who had received the prophylactic antibiotic treatment selective digestive decontamination (SDD), i.e. tobramycin, polymyxin E, amphotericin B and cefotaxime, were inoculated anaerobically on porous aluminium oxide chips placed on top of poor and rich agar media, including media supplemented with the SDD antibiotics. Biomass growing on the chips was analysed by 16S rRNA gene amplicon sequencing, showing large inter-individual differences in bacterial cultivability, and enrichment of a range of taxonomically diverse operational taxonomic units (OTUs). Furthermore, growth of Ruminococcaceae (2 OTUs), Enterobacteriaceae (6 OTUs) and Lachnospiraceae (4 OTUs) was significantly inhibited by the SDD antibiotics. Strains belonging to 16 OTUs were candidates for cultivation to pure culture as they shared 95% sequence identity with the closest type strain and had a relative abundance of 2%. Six of these OTUs were detected on media containing SDD antibiotics, and as such were prime candidates to be studied regarding antibiotic resistance. One of these six OTUs was obtained in pure culture using targeted isolation. This novel strain was resistant to the antibiotics metrodinazole and imipenem. It was initially classified as member of the Ruminococcaceae, though later it was found to share 99% nucleotide identity with the recently published Sellimonas intestinalis BR72T. In conclusion, we show that high-throughput cultivation-based screening of microbial communities can guide targeted isolation of bacteria that serve as reservoirs of antibiotic resistance.

Original languageEnglish
Article numbere0210970
JournalPLoS ONE
Volume14
Issue number1
DOIs
Publication statusPublished - 17 Jan 2019

Fingerprint

aluminum oxide
Aluminum Oxide
Bacteria
Screening
Decontamination
digestive system
antibiotics
decontamination
Throughput
Anti-Bacterial Agents
screening
bacteria
Microbial Drug Resistance
intestinal microorganisms
antibiotic resistance
Pathogens
polymyxins
tobramycin
imipenem
Colistin

Cite this

@article{240f16ca5cd241939d82eb3e0640bb34,
title = "High throughput cultivation-based screening on porous aluminum oxide chips allows targeted isolation of antibiotic resistant human gut bacteria",
abstract = "The emergence of bacterial pathogens that are resistant to clinical antibiotics poses an increasing risk to human health. An important reservoir from which bacterial pathogens can acquire resistance is the human gut microbiota. However, thus far, a substantial fraction of the gut microbiota remains uncultivated and has been little-studied with respect to its resistance reservoir-function. Here, we aimed to isolate yet uncultivated resistant gut bacteria by a targeted approach. Therefore, faecal samples from 20 intensive care patients who had received the prophylactic antibiotic treatment selective digestive decontamination (SDD), i.e. tobramycin, polymyxin E, amphotericin B and cefotaxime, were inoculated anaerobically on porous aluminium oxide chips placed on top of poor and rich agar media, including media supplemented with the SDD antibiotics. Biomass growing on the chips was analysed by 16S rRNA gene amplicon sequencing, showing large inter-individual differences in bacterial cultivability, and enrichment of a range of taxonomically diverse operational taxonomic units (OTUs). Furthermore, growth of Ruminococcaceae (2 OTUs), Enterobacteriaceae (6 OTUs) and Lachnospiraceae (4 OTUs) was significantly inhibited by the SDD antibiotics. Strains belonging to 16 OTUs were candidates for cultivation to pure culture as they shared 95{\%} sequence identity with the closest type strain and had a relative abundance of 2{\%}. Six of these OTUs were detected on media containing SDD antibiotics, and as such were prime candidates to be studied regarding antibiotic resistance. One of these six OTUs was obtained in pure culture using targeted isolation. This novel strain was resistant to the antibiotics metrodinazole and imipenem. It was initially classified as member of the Ruminococcaceae, though later it was found to share 99{\%} nucleotide identity with the recently published Sellimonas intestinalis BR72T. In conclusion, we show that high-throughput cultivation-based screening of microbial communities can guide targeted isolation of bacteria that serve as reservoirs of antibiotic resistance.",
author = "Dennis Versluis and {de J. Bello Gonz{\'a}lez}, Teresita and Zoetendal, {Erwin G.} and {van Passel}, {Mark W.J.} and Hauke Smidt",
year = "2019",
month = "1",
day = "17",
doi = "10.1371/journal.pone.0210970",
language = "English",
volume = "14",
journal = "PLoS ONE",
issn = "1932-6203",
publisher = "Public Library of Science",
number = "1",

}

High throughput cultivation-based screening on porous aluminum oxide chips allows targeted isolation of antibiotic resistant human gut bacteria. / Versluis, Dennis; de J. Bello González, Teresita; Zoetendal, Erwin G.; van Passel, Mark W.J.; Smidt, Hauke.

In: PLoS ONE, Vol. 14, No. 1, e0210970, 17.01.2019.

Research output: Contribution to journalArticleAcademicpeer-review

TY - JOUR

T1 - High throughput cultivation-based screening on porous aluminum oxide chips allows targeted isolation of antibiotic resistant human gut bacteria

AU - Versluis, Dennis

AU - de J. Bello González, Teresita

AU - Zoetendal, Erwin G.

AU - van Passel, Mark W.J.

AU - Smidt, Hauke

PY - 2019/1/17

Y1 - 2019/1/17

N2 - The emergence of bacterial pathogens that are resistant to clinical antibiotics poses an increasing risk to human health. An important reservoir from which bacterial pathogens can acquire resistance is the human gut microbiota. However, thus far, a substantial fraction of the gut microbiota remains uncultivated and has been little-studied with respect to its resistance reservoir-function. Here, we aimed to isolate yet uncultivated resistant gut bacteria by a targeted approach. Therefore, faecal samples from 20 intensive care patients who had received the prophylactic antibiotic treatment selective digestive decontamination (SDD), i.e. tobramycin, polymyxin E, amphotericin B and cefotaxime, were inoculated anaerobically on porous aluminium oxide chips placed on top of poor and rich agar media, including media supplemented with the SDD antibiotics. Biomass growing on the chips was analysed by 16S rRNA gene amplicon sequencing, showing large inter-individual differences in bacterial cultivability, and enrichment of a range of taxonomically diverse operational taxonomic units (OTUs). Furthermore, growth of Ruminococcaceae (2 OTUs), Enterobacteriaceae (6 OTUs) and Lachnospiraceae (4 OTUs) was significantly inhibited by the SDD antibiotics. Strains belonging to 16 OTUs were candidates for cultivation to pure culture as they shared 95% sequence identity with the closest type strain and had a relative abundance of 2%. Six of these OTUs were detected on media containing SDD antibiotics, and as such were prime candidates to be studied regarding antibiotic resistance. One of these six OTUs was obtained in pure culture using targeted isolation. This novel strain was resistant to the antibiotics metrodinazole and imipenem. It was initially classified as member of the Ruminococcaceae, though later it was found to share 99% nucleotide identity with the recently published Sellimonas intestinalis BR72T. In conclusion, we show that high-throughput cultivation-based screening of microbial communities can guide targeted isolation of bacteria that serve as reservoirs of antibiotic resistance.

AB - The emergence of bacterial pathogens that are resistant to clinical antibiotics poses an increasing risk to human health. An important reservoir from which bacterial pathogens can acquire resistance is the human gut microbiota. However, thus far, a substantial fraction of the gut microbiota remains uncultivated and has been little-studied with respect to its resistance reservoir-function. Here, we aimed to isolate yet uncultivated resistant gut bacteria by a targeted approach. Therefore, faecal samples from 20 intensive care patients who had received the prophylactic antibiotic treatment selective digestive decontamination (SDD), i.e. tobramycin, polymyxin E, amphotericin B and cefotaxime, were inoculated anaerobically on porous aluminium oxide chips placed on top of poor and rich agar media, including media supplemented with the SDD antibiotics. Biomass growing on the chips was analysed by 16S rRNA gene amplicon sequencing, showing large inter-individual differences in bacterial cultivability, and enrichment of a range of taxonomically diverse operational taxonomic units (OTUs). Furthermore, growth of Ruminococcaceae (2 OTUs), Enterobacteriaceae (6 OTUs) and Lachnospiraceae (4 OTUs) was significantly inhibited by the SDD antibiotics. Strains belonging to 16 OTUs were candidates for cultivation to pure culture as they shared 95% sequence identity with the closest type strain and had a relative abundance of 2%. Six of these OTUs were detected on media containing SDD antibiotics, and as such were prime candidates to be studied regarding antibiotic resistance. One of these six OTUs was obtained in pure culture using targeted isolation. This novel strain was resistant to the antibiotics metrodinazole and imipenem. It was initially classified as member of the Ruminococcaceae, though later it was found to share 99% nucleotide identity with the recently published Sellimonas intestinalis BR72T. In conclusion, we show that high-throughput cultivation-based screening of microbial communities can guide targeted isolation of bacteria that serve as reservoirs of antibiotic resistance.

U2 - 10.1371/journal.pone.0210970

DO - 10.1371/journal.pone.0210970

M3 - Article

VL - 14

JO - PLoS ONE

JF - PLoS ONE

SN - 1932-6203

IS - 1

M1 - e0210970

ER -