L.J. Slopes for BCG Vaccines

Principle And Interpretation

Solid media used for isolation and cultivation of Mycobacteria are either egg-based or agar-based. Egg-based media contain whole eggs or egg yolk, potato flour, salts and glycerol and are solidified by inspissation. Of the egg-based media, Lowenstein Jensen Medium is most commonly used (1). L.J. Medium was originally formulated by Lowenstein, containing congo red and malachite green dyes (2). Jensen (3) modified Lowensteins medium by altering the citrate and phosphate contents, eliminating the congo red dye and by increasing the malachite green concentration. Gruft (4,5) further modified L. J. Medium with the addition of two antimicrobics to increase selectivity. his medium supports the growth of a wide variety of Mycobacteria and can also be used for niacin testing (6). Penicillin and alidi ic acid along with malachite green prevents growth of the ma ority of contaminants surviving decontamination of the specimen while encouraging earliest possible growth of Mycobacteria. RA acts as stimulant and help to increase the isolation rate of Mycobacteria. Do not add glycerol to the medium if bovine or other glycerophobic strains are to be cultured (7). Malachite green serves as an inhibitor and also as pH indicator. Formation of blue one indicates a decrease in pH by gram-positive contaminants (e.g. Streptococci) and yellow ones of dye destruction by gram-negative bacilli. Proteolytic contaminants cause locali ed or complete digestion of medium. Hardy et al (8) recommended each specimen to be inoculated and incubated in triplicate,

1. o identify saprophytes at room temperature (25 C).
2. o identify presence or absence of pigmentation by photochromogenes and scotochromogenes at 35 C alternately in light and dark as per the type of organism.

Routinely cultivation is carried out aerobically at 35 C.