Hi,

There are different ways of antimicrobial susceptibility testing such as disk diffusion testing, MIC value determination by microdilution or macrodilution method. However, I would suggest international standards/references; for instance, EUCAST (http://www.eucast.org/ast_of_bacteria) and CLSI Guideline (https://clsi.org/standards/products/microbiology/) for both testing and interpretation (if conventional antibiotics are used).

Regards,

You should use VITEK 2 Compact.

I definitely recommend EUCAST and its website (eucast.org). EUCAST describes disc diffusion in detail and has a large Q/A section to work with. Disc diffusion is one of a few methods used in routine laboratories to do susceptibility testing. Another alternative is broth microdilution, which is available as automated system (e.g. Vitek, Phoenix, …) or for manual handling (different suppliers).

The method you describe in your question is an agardilution method (though not in its most effective way), which is usually not done in routine diagnostic labs.

Agardilution is much more interesting, if you want to test a large number of strains against one antibiotic (routine: one strain against a large number of antibiotics). For classical agardilution, however, the bacteria are not added to the agar but are spotted on agar plates, which contain a geometric dilution of an antibiotic to test again. As with each and every method, standardization is required. You are right when worrying that too high temperature of the agar has effects on the bacteria and the test outcome.

Hi, there are standard guides as to the methods of preparation, inoculation incubation and interpretation of results.making sure no moisture on the inoculated plates, time of incubation concentration.of antibiotics on disks are std procedures

I think we need to be clear,

You are not going to pour 0.5 McFarland culture but streak it on the MH agar if you are interested in the Kirby-Bauer method. Then you place your antimicrobial discs on the agar surface using forceps or using disc dispenser. There are available and approved protocols check CLSI, EUCAST or already published articles that have reported the procedure.

Cheers

Well there are variations in some of microbiological procedures. Nonetheless, the standard remains spreading the standard pure culture on the MH agar. It should be noted that at temperature of 50 C some bacteria will be inhibited.

If it is disc diffusion method you intend to use you must spread the 0.5 McFarland culture using sterile swab stick on the MH agar. The antibiotic discs can then be placed at equidistant using forceps or disc dispenser on the agar surface. Standard procedures and protocols for antibiotic susceptibility testing recommended by CLSI, EUCAST, BSAC, DIN are available online.

Dear Duman:

Please, find in the attached file my book "Theoratical and practical aspects of antimicrobial chemotherapy"

Hope it will answered all questions in this field.

Best and kind regard

It is highly risky as one cannot determine the ideal temperature of the agar before mixing bacterial suspension safely. Even the fractional inhibition would result in change in bacterial suspension and thus final reporting. So why take risk if you can simply lawn culture on solid agar. It also easy, safe and recommended by most scholars.

I would suggest you make use of CLSI standard and for the process of inoculation the spread plate method would be better for if your susceptibility method is the disc diffusion technique

For determination of AST, researchers should follow the CLSI guidelines strictly.

The method you described above resemble "Agar dilution method" which recommended by CLSI (document CLSI- M02-A12).

In this method, serial dilutions of the drug are prepared in agar and poured into plates. The Advantage of this method over The traditional disk diffusion method that many strains can be inoculated on each plate containing an antibiotic dilution.

Regards