I'm working with some pathogenic E. coli.
I want to grow them in a way that they will express their virulence and antibiotic resistances as much as possible
Any suggestions?
- Media?
- Growth conditions?
- Supplements?
NOTE: Preferably in liquid culture
By the way, they grow perfectly in NB and such rich media.
Hello Amir Nakar
If you need to express anything in the host, you bring the requirement for that expression. The antibiotic resistance will be expressed only in the presence of an antibiotic. Doesnt matter what medium or supplements you use. If you are looking for histidine competence, then you should add a minimal media without histidine. like wise you have act.
To know the working concentration of the antibiotic, You might want to look into the plasmid and identify what is the dosage it can tolerate. if you cant find it out, then i would suggest you to go with readily available strips that has gradient concentrations of antibiotics. It will show you the perfect concentration at which your host works perfectly.
All the best !!
Please do contact me if your doubt is not cleared
You must cultivate the bacteria under test in medium containing ihe factor that you need to express i.e the antibiotic .
Hi Amir,
Assuming those were separate questions, to express the virulence genes of pathogenic E. coli one would have to expose the bacteria to conditions that mimicked the host environment. Different virulence genes will be expressed at different stages of the infection so there isn't a single answer but please see below for some examples:
Here is a review: Article Environment Controls LEE Regulation in Enteropathogenic Esch...
Oxygen:
Article Enterotoxigenic E. Coli virulence gene regulation in human infections
The most common method for upregulating the T3SS for EPEC, EHEC and Citrobacter is LB switch to DMEM eg.
EPEC E69 strains grown for 16 h ON in LB shaking and primed by sub-culturing 1:100 in DMEM low glucose with no additives supplemented with
appropriate antibiotics for 3 h, with no agitation in an atmosphere of 5% CO2.
EHEC grown for 8 h in LB shaking and primed by sub-culturing 1:500 into DMEM low glucose with no additives supplemented with appropriate antibiotics ON.
Conditions will vary depending on what your working on and which virulence genes you want to induce. Hope this helps. Good luck.
You must expose E.coli to severe condition and seperate the survival then you can test the virulence factors morphologically then using pcr to investigate the genes responsible of these factors.
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