Medium for selective isolation of Fusobacterium nucleatum from human periodontal pockets

Abstract

A selective medium, CVE agar, was developed for the isolation of Fusobacterium nucleatum from subgingival plaque of periodontally diseased patients. The medium contained 1.0% Trypticase (BBL Microbiology Systems), 0.5% yeast extract, 0.5% NaCl, 0.2% glucose, 0.02% l-tryptophan, 1.5% agar, and 5% defibrinated whole sheep blood. Erythromycin and crystal violet were added as the selective inhibitory agents at concentrations of 4 and 5 μg/ml, respectively. The medium permitted almost total recovery of F. nucleatum when compared with a nonselective medium and suppressed the recovery of most remaining species by 6 to 8 orders of magnitude. Microorganisms suppressed to a lesser degree included Selenomonas sputigena, Actinobacillus actinomycetemcomitans, Eikenella corrodens, and some strains of Peptostreptococcus. The distinct colonial morphology of F. nucleatum on CVE agar made differentiation relatively easy when contaminants were present. With this medium, F. nucleatum was enumerated from 278 subgingival plaque samples and accounted for <1.0 to >25% of the cultivatable microbiota.