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Fast Identification Method for Screening Bacteria from Faecal Samples Using Oxford Nanopore Technologies MinION

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This study introduces a rapid method for identifying bacteria like lactobacilli and Bacteroides in human samples. It simplifies bacterial identification by reducing lengthy purification steps, offering quick and accurate results for culturomics.

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Area of Science:

  • Microbiology
  • Genomics
  • Molecular Biology

Background:

  • Traditional bacterial identification methods are time-consuming and costly due to extensive culturing, purification, and DNA extraction.
  • Isolating specific bacteria from complex environments like human faeces presents significant challenges.
  • Accurate and efficient bacterial identification is crucial for microbiome research and diagnostics.

Purpose of the Study:

  • To develop a fast, robust, and accurate method for identifying key gut bacteria, including lactobacilli, bifidobacteria, and Bacteroides, in human faecal samples.
  • To streamline the identification process by minimizing extensive culturing and DNA purification steps.
  • To provide a high-throughput screening method suitable for culturomics studies.

Main Methods:

  • Anaerobic culturing of bacteria from human faecal samples on selective media.
  • Sonication-based DNA extraction followed by near-complete 16S rRNA gene polymerase chain reaction amplification.
  • MinION sequencing using the Flongle adapter, with sequence analysis via NanoCLUST and graphics in RStudio.

Main Results:

  • The method accurately identified bacteria from 110 out of 125 investigated colonies, with 100% of reads assigned to a single species.
  • Fifteen colonies comprised mixtures of up to three different bacterial species, demonstrating the method's ability to resolve complex samples.
  • High accuracy and speed were achieved, significantly reducing the time and effort compared to conventional methods.

Conclusions:

  • The developed method offers a significant advantage for identifying specific bacteria, especially when exclusive selective media are unavailable.
  • It bypasses lengthy colony and DNA purification, enabling rapid and precise bacterial identification.
  • This technique is highly suitable for direct screening of pure bacterial cultures and for bacterial identification within culturomics studies.