May 24, 2024
A large scale bacterial attraction assay: a new quantitative bacterial migration assay suitable for genetic screens
Peer-reviewed method
- Thomas Quiroz Monnens1,
- Alice Boulanger1
- 1LIPME, Université de Toulouse, INRAE, CNRS, Université Paul Sabatier, 31320, Castanet-Tolosan, France
- PLOS ONE Lab Protocols
- Spotlight series
Collection Citation: Thomas Quiroz Monnens, Alice Boulanger 2024. A large scale bacterial attraction assay: a new quantitative bacterial migration assay suitable for genetic screens. protocols.io https://dx.doi.org/10.17504/protocols.io.n2bvjn65bgk5/v1
License: This is an open access collection distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this collection and it's working
Created: May 03, 2024
Last Modified: October 09, 2024
Collection Integer ID: 99639
Keywords: large scale bacterial attraction assay, new quantitative bacterial migration assay, bacterial attraction assay, genetic screens bacteria, attracted bacteria, large quantities of attracted bacteria, bacterial migration, motile bacterial cell, high numbers of attracted bacteria, rapid chemotactic responses to chemoeffector, chemotactic responses to chemoeffector, bacterial migration over time, rapid chemotactic response, chemotactic response, investigating chemoeffector kinetic, lasting chemoeffector gradient, chemoeffector kinetics in the setup, chemoeffector, microfluidic assay, chemoeffector gradient, chemotaxi, using chemoattractant, pathogenic bacterium xanthomonas campestris pv, genetic screen, suitable for genetic screen, forward genetic screen, assay, chemical gradient, various motility mechanism, environmental bacterium shewanella oneidensi, impaired strain
Funders Acknowledgements:
ANR JCJC
Grant ID: ANR-19-CE20-JCJC-0014-01
Disclaimer
Spotlight Video
The video below is a supplement with extra context and tips, as part of the protocols.io Spotlight series, featuring conversations with protocol authors.
This protocol was peer reviewed and accepted for publication in Plos One protocol
Abstract
Bacteria use various motility mechanisms to explore their environments. Chemotaxis is the ability of a motile bacterial cell to direct its movement in response to chemical gradients. A number of methods have been developed and widely used to study chemotactic responses to chemoeffectors including capillary, agar plug, microscopic slide, and microfluidic assays. While valuable, these assays are primarily designed to monitor rapid chemotactic responses to chemoeffectors on a small scale, which poses challenges in collecting large quantities of attracted bacteria. Consequently, these setups are not ideal for experiments like forward genetic screens. To overcome this limitation, we developed the Large Scale Bacterial Attraction assay (LSBA), which relies on the use of a Nalgene™ Reusable Filter Unit and other materials commonly found in laboratories. We validate the LSBA by investigating chemoeffector kinetics in the setup and by using chemoattractants to quantify the chemotactic response of wild-type, and motility impaired strains of the plant pathogenic bacterium Xanthomonas campestris pv. campestris and the environmental bacterium Shewanella oneidensis. We show that the LSBA establishes a long lasting chemoeffector gradient, that the setup can be used to quantify bacterial migration over time and that the LSBA offers the possibility to collect high numbers of attracted bacteria, making it suitable for genetic screens.
Troubleshooting
Spotlight Video
The video below is a supplement with extra context and tips, as part of the protocols.io Spotlight series, featuring conversations with protocol authors.
This protocol was peer reviewed and accepted for publication in Plos One protocol
Protocol references
1. Blanvillain S, Meyer D, Boulanger A, Lautier M, Guynet C, Denancé N, et al. Plant carbohydrate scavenging through tonB-dependent receptors: a feature shared by phytopathogenic and aquatic bacteria. PloS One. 2007;2: e224. doi:10.1371/journal.pone.0000224
2. Boyeldieu A, Poli J-P, Ali Chaouche A, Fierobe H-P, Giudici-Orticoni M-T, Méjean V, et al. Multiple detection of both attractants and repellents by the dCache-chemoreceptor SO_1056 of Shewanella oneidensis. FEBS J. 2022;289: 6752–6766. doi:10.1111/febs.16548
3. Cerutti A, Jauneau A, Auriac M-C, Lauber E, Martinez Y, Chiarenza S, et al. Immunity at Cauliflower Hydathodes Controls Systemic Infection by Xanthomonas campestris pv campestris. Plant Physiol. 2017;174: 700–716. doi:10.1104/pp.16.01852
4. Luneau JS, Baudin M, Quiroz Monnens T, Carrère S, Bouchez O, Jardinaud M-F, et al. Genome-wide identification of fitness determinants in the Xanthomonas campestris bacterial pathogen during early stages of plant infection. New Phytol. 2022. doi:10.1111/nph.18313
5. Dugé de Bernonville T, Noël LD, SanCristobal M, Danoun S, Becker A, Soreau P, et al. Transcriptional reprogramming and phenotypical changes associated with growth of Xanthomonas campestris pv. campestris in cabbage xylem sap. FEMS Microbiol Ecol. 2014;89: 527–541. doi:10.1111/1574-6941.12345