In Situ Monitoring and Quantitative Determination of R27 Plasmid Conjugation

Horizontal gene transfer (HGT) by plasmid conjugation is a major driving force in the spread of antibiotic resistance among Enterobacteriaceae. Most of the conjugation studies are based on calculation of conjugation ratios (number of transconjugants/number of donors) after viable counting of transco...

Full description

Saved in:
Bibliographic Details
Published in:Life (Basel, Switzerland) Switzerland), 2022-08, Vol.12 (8), p.1212
Main Authors: Gibert, Marta, Jiménez, Carlos J., Comas, Jaume, Zechner, Ellen L., Madrid, Cristina, Balsalobre, Carlos
Format: Article
Language:English
Subjects:
Antibiotic resistance
Antibiotics
Confocal microscopy
Conjugation
E coli
Flow cytometry
Fluorescence
fluorophores
Gene transfer
Genes
Horizontal transfer
in situ monitoring
IncHI
Labeling
Lasers
Microscopy
Monitoring
Phenotypes
Physiology
plasmid conjugation
Plasmids
Proteins
R27
Spatial distribution
Yeast
Citations: Items that this one cites
Online Access:Get full text
Tags: Add Tag
No Tags, Be the first to tag this record!
Description
Summary:Horizontal gene transfer (HGT) by plasmid conjugation is a major driving force in the spread of antibiotic resistance among Enterobacteriaceae. Most of the conjugation studies are based on calculation of conjugation ratios (number of transconjugants/number of donors) after viable counting of transconjugant and donor cells. The development of robust, fast and reliable techniques for in situ monitoring and quantification of conjugation ratios might accelerate progress in understanding the impact of this cellular process in the HGT. The IncHI1 plasmids, involved in multiresistance phenotypes of relevant pathogens such as Salmonella and E. coli, are distinguished by the thermosensitivity of their conjugative transfer. Conjugation mediated by IncHI1 plasmids is more efficient at temperatures lower than 30 °C, suggesting that the transfer process takes place during the environmental transit of the bacteria. In this report, we described a methodology to monitor in situ the conjugation process during agar surface matings of the IncHI1 plasmid R27 and its derepressed derivative drR27 at different temperatures. A three-color-labeling strategy was used to visualize the spatial distribution of transconjugants within the heterogeneous environment by epifluorescence and confocal microscopy. Moreover, the fluorescent labelling was also used to quantify conjugation frequencies in liquid media by flow cytometry.
ISSN:2075-1729
2075-1729
DOI:10.3390/life12081212