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Monitoring Plasmid Replication in Live Mammalian Cells over Multiple Generations by Fluorescence Microscopy

Monitoring Plasmid Replication in Live Mammalian Cells over Multiple Generations by Fluorescence Microscopy

Published on: December 13, 2012

Real-time plasmid transmission detection pipeline.

Natalie Scherff^1,2, Jörg Rothgänger², Thomas Weniger²

¹1Institute of Hygiene, University Hospital Münster, Münster, Germany.

Microbiology Spectrum

|October 29, 2024

Summary

This study introduces a new pipeline for real-time detection of plasmid transmission in clinical microbiology. The developed system effectively identifies antimicrobial resistance spread, aiding in infection control.

Keywords:

antimicrobial resistance horizontal gene transfer long-read sequencing plasmid transmission whole-genome sequencing

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Plasmid Stability Analysis with Open-Source Droplet Microfluidics

Plasmid Stability Analysis with Open-Source Droplet Microfluidics

Published on: December 27, 2024

Area of Science:

Microbiology
Bioinformatics
Genomics

Background:

Antimicrobial resistance spread via plasmids is a significant clinical challenge.
Current methods often miss plasmid-mediated transmissions, focusing mainly on clonal spread.
Advances in long-read sequencing allow plasmid reconstruction but lack user-friendly tools.

Purpose of the Study:

To evaluate a novel real-time plasmid transmission detection pipeline integrated into SeqSphere+ software.
To assess the pipeline's utility for early warning alerts (EWAs) of plasmid spread.
To determine the pipeline's suitability for clinical microbiology settings.

Main Methods:

Utilized a local Mash plasmid database for early warning alerts (EWAs) with a distance threshold of 0.001.
Employed core-genome multi-locus sequence typing for clonal transmission detection.
Integrated tools like MOB-suite, AMRFinderPlus, and MUMmer for plasmid characterization and comparison.

Main Results:

The SeqSphere+ pipeline successfully identified clonal and carbapenemase-carrying plasmid clusters, matching published data.
Early warning alerts (EWAs) were generated in the correct chronological order.
Optimal performance was achieved using a sketch size of 1,000 with plasmid size correction enabled.

Conclusions:

The developed pipeline is suitable for clinical microbiology due to automated analyses and an alert system.
Its integration into the GUI-based SeqSphere+ platform requires limited bioinformatics knowledge.
The pipeline enables (near) real-time plasmid transmission detection in routine diagnostics using long-read sequencing.