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

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...

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Published in:PloS one 2019-01, Vol.14 (1), p.e0210970-e0210970
Main Authors: Versluis, Dennis, de J Bello González, Teresita, Zoetendal, Erwin G, Passel, Mark W J van, Smidt, Hauke
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description 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.
doi_str_mv 10.1371/journal.pone.0210970
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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.</abstract><cop>United States</cop><pub>Public Library of Science</pub><pmid>30653573</pmid><doi>10.1371/journal.pone.0210970</doi><tpages>e0210970</tpages><orcidid>https://orcid.org/0000-0001-8430-4500</orcidid><oa>free_for_read</oa></addata></record>
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identifier ISSN: 1932-6203
ispartof PloS one, 2019-01, Vol.14 (1), p.e0210970-e0210970
issn 1932-6203
1932-6203
language eng
recordid cdi_plos_journals_2168161386
source Publicly Available Content Database; PubMed Central
subjects Aluminum
Aluminum Oxide
Amphotericin B
Anaerobiosis
Antibiotic Prophylaxis
Antibiotic resistance
Antibiotics
Antifungal agents
Bacteria
Bacterial infections
Bacteriological Techniques
Biology and Life Sciences
Care and treatment
Cefotaxime
Chips
Clostridiales - drug effects
Clostridiales - growth & development
Clostridiales - isolation & purification
Cultivation
Culture
Culture media
Decontamination
Decontamination - methods
Digestive system
Digestive tract
Disease Reservoirs - microbiology
Drug resistance
Drug Resistance, Bacterial - genetics
Enterobacteriaceae - drug effects
Enterobacteriaceae - genetics
Enterobacteriaceae - isolation & purification
Feces - microbiology
Gastrointestinal Microbiome - drug effects
Gastrointestinal Microbiome - genetics
Gastrointestinal tract
Gene sequencing
Genes
Health risks
High-Throughput Screening Assays
Humans
Imipenem
Intensive care
Intensive Care Units
Intestinal microflora
Laboratories
Laboratorium voor Microbiologie
Medical screening
Medicine and Health Sciences
Methods
Microbial activity
Microbial drug resistance
Microbial Sensitivity Tests
Microbiological Laboratory
Microbiologie
Microbiology
Microbiota
Microbiota (Symbiotic organisms)
Microorganisms
MolEco
Pathogens
Porosity
Public health
Pure culture
Relative abundance
RNA, Bacterial - genetics
RNA, Ribosomal, 16S - genetics
rRNA 16S
Ruminococcaceae
Tobramycin
VLAG
WIMEK
title High throughput cultivation-based screening on porous aluminum oxide chips allows targeted isolation of antibiotic resistant human gut bacteria
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