Selective medium with DNase test agar and a modified toluidine blue O technique for primary isolation of Branhamella catarrhalis in sputum

Abstract

A selective medium with DNase test agar and incorporating vancomycin (10 micrograms/ml), trimethoprim (8 micrograms/ml), and amphotericin B (2 micrograms/ml) supported the growth of 305 Branhamella catarrhalis isolates. A modified toluidine blue O technique was used after 48 h of incubation in CO2 to overlay suspected B. catarrhalis colonies. A metachromatic color change was observed in 15 min, indicating DNase production. In 200 unselected sputum samples of hospitalized patients, this method was compared with routine microbiologic procedures; 31 B. catarrhalis isolates were recovered with the method, compared with 22 isolated from the clinical laboratory. This medium will be particularly useful for culture of sputum, which shows inflammatory cells and gram-negative diplococci on Gram-stained smears.