BacterPlate™ Mueller Hinton Agar (MHA)

Ready-to-use medium on 90mm plates for testing the antibiotic sensitivity of easily cultured bacteria.
Code: 05037
Packaging: 10 plates/ box or as per customer request.
Storage Conditions: 2–8°C.
Transportation: Ambient temperature.
Shelf Life: 04 months from the date of manufacture.

PRINCIPLES

BacterPlate™ Mueller Hinton Agar (MHA) is recognized by all experts as being the reference medium for the study of the susceptibility of bacteria to antibiotics and sulfamides. It is also useful for the isolation of Neisseria and is an excellent base medium for the preparation of blood agars.
The typical composition responds to that defined in the standards NF U47-106 and NF U47-107.
The packaging with semi-permeable Cellophane film helps balance the humidity of the environment during storage.

RESULTS

The choice of ingredients is determined in order to obtain a very low quantity of thymine and thymidine (substances known to inhibit the antibacterial activity of trimethoprim), and a very low quantity of para-aminobenzoic acid (PABA) and its structural analogues (which antagonize the activity of sulfonamides). As a result of the influence of calcium and magnesium on the sensitivity of Pseudomonas strains to aminoglycosides, Reller et al. recommended that the ion concentrations be included within the following limits :
- Calcium: 50–100 mg/liter
- Magnesium: 20–35 mg/
The Kirby-Bauer method is based on the diffusion of antibiotics impregnated in previously dried paper disks, deposited on the surface of the agar. When applied to the surface of the agar, the disks absorb a sufficient quantity of water to dissolve the antibiotic, which then diffuses into the medium according to physical laws of diffusion of molecules through a gel. In this way, a concentration gradient of antibiotic forms around each disk. At the same time as the antibiotics diffuse, the bacteria inoculated on the surface of the agar multiply. During the logarithmic phase of growth, bacterial multiplication is more rapid than the diffusion of the antibiotic and uninhibited bacterial cells continue to multiply until growth can be visualized. No growth appears when the antibiotic is present at inhibiting concentrations. It now becomes possible to measure the diameter of the inhibition zone, which is indirectly proportional to the minimal inhibitory concentrations found by the dilution method. Tables exist for interpreting the results in order to determine if the bacteria are sensitive or resistant to the antibiotic tested.

Antibiotic Susceptibility Testing (Kirby-Bauer Standard Method)

Use a pure culture from BacterTube^TM Tryptone Soy Broth (TSB) with turbidity matching the standard opacity of a barium sulfate suspension (0,5 on the McFarland scale).
Collect the inoculum using a sterile swab.
Inoculate the agar by streaking the swab 2 to 3 times across the entire agar surface to achieve uniform inoculum distribution.
Allow the plates to dry for 10 minutes before placing antibiotic disks.
Place the antibiotic disks with gentle pressure to ensure proper adhesion to the agar surface. Disks should be placed at least 15 mm from the plate edge and spaced sufficiently apart to avoid overlapping inhibition zones.
Incubate at 37°C for 18 to 24 hours.

Note: The Kirby-Bauer method is recognized as the most reliable and reproducible approach. However, other methods may also be employed, provided that the inoculum and inoculation procedures are thoroughly studied and standardized beforehand.

RESULTS

Measure the inhibition zones.
Refer to the table for interpreting inhibition zones furnished by the suppliers of antibiotic disks in order to establish the correlation between the inhibition zone and the minimal inhibitory concentration (M.I.C.).

BacterLab ensures the quality of each product batch by testing with ATCC reference strains:

Reference strains	Incubation conditions	Antibiotic discs	Sterility zone limit (mm)
P. aeruginosa ATCC 9027	35 – 37^oC for 18 – 24 hours	Gentamicin 10 μg	16 – 21
S. aureus ATCC 25923		Erythromucin 15 μg	22 – 30
E.coli ATCC 35218		Ampicillin 10 μg	16 – 22