The absolute concentration method determines if 1%

or more of an inoculum will grow after being cultured on

media containing critical concentrations of a drug on the

plate. It requires growth of the patient strain on drug free

medium to demonstrate the viability, but does not compare

the colony numbers on drug free and drug containing

media so that the inoculum must be carefully standardized.

The resistance ratio is similar to the absolute

concentration method except that the patient strain is

compared with the growth of a standard laboratory strain.

Results are reported as the ratio of the MIC of the patient

strain to that of the laboratory strain. A patient strain with

a ratio of 8:1 is considered resistant, while 4:1 is suggestive

of resistance. This method is more tolerant to variation in

concentration of drugs within different batches of media.

The proportion method compares the growth of a patient

strain in the presence and absence of a drug. If 1% or more

of the inoculum produces colonies on media that contains an

agent at the critical concentration compared with controls,

the isolate is considered to be resistant. This method is the

most popular and is relatively simple to perform and interpret.

Susceptibility Testing of Mycobacteria

Eleven drugs are used in the treatment of tuberculosis.

Five are considered “primary” and include streptomycin,

isoniazid, rifampin, pyrazinamide and ethambutol, while

the remaining six, Ethionamide, ciprofloxacin, kanamycin,

D-cycloserine, para-aminosalicylic acid and amikacin are

considered “secondary” and used only when resistance

develops to the primary drugs.

Although drugs have been incorporated in inspissated

egg-based media for conducting susceptibility tests,

many laboratories internationally now prefer using

Middlebrook 7H11 or 7H10 as a base medium, adding

the drugs after cooling the agar to 45°C. Adding the drugs

to the agar medium after autoclaving decreases the loss

of activity that can occur in egg-based medium such LJ

during inspisstion. An additional loss of drug activity may

occur in egg-based media with binding of some agents to

egg albumin and other proteins.

Drug Concentrations for Proportion Method

Susceptibility Testing using Various Culture Media*

Drug concentration (µg/mL)

Drug 7H10 7H11 Lowenstein-Jensen

Isoniazid 0.2,1.0 0.2,1.0 0.2,1.0

p-Aminosalicylic acid 2.0 8.0 0.5

Streptomycin 2.0 2.0 4.0

Rifampin 1.0 1.0 40.0

Ethambutol 2.0 7.5 2.0

Ethionamide 5.0 10.0 0.0

Kanamycin 5.0 6.0 20.0

Capreomycin 10.0 10.0 20.0

D-Cycloserine 20.0 30.0 30.0

Pyrazinamide 50.0 — 100.0

Microbiology and Bacteriology 861