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Laboratory Identification: Streptococcus pneumoniae

Bile solubility is a rapid, confirmatory test often used to differentiate S. pneumoniae from viridans streptococci, in which a drop of 10% deoxycholate is placed on an area of growth. The bacterial cells of S. pneumoniae are bile soluble, as shown by the disappearance (lysing) of the colonies in the area where a drop of 10% deoxycholate had been added to the surface of the agar (indicated by yellow arrows in top image). The colonies of viridans streptococci would remain intact and clearly visible.
Optochin or "P" disc susceptibility is a presumptive test using a filter paper disc impregnated with 6 mg of ethylhydrocupreine hydrochloride (optochin). In 2004, the discovery of the organism Streptococcus pseudopneumoniae made it important to perform optochin testing under increased CO2 conditions to differentiate it from S. pneumoniae. When testing is performed in an ambient atmosphere (35º C for 18-24 hours), BOTH S. pneumoniae AND S. pseudopneumniae are susceptible to optochin (zone >14 mm). If testing is performed in 5% CO2, S. pseudopneumoniae is resistant to optochin (zone ≤ 14 mm). Therefore, testing should be confirmed with bile solubility tests or serological identification. The chart summarizes this testing:

OrganismOptochin (O2-ambient) ResultsOptochin (5% CO2) ResultsBile Solubility Results
S. pneumoniaeSusceptible (zone >14 mm)Susceptible (zone >14 mm)Positive (soluble)
S. pseudopneumoniaeSusceptible (zone >14 mm)Resistant (zone ≤ 14 mm)Negative (insoluble)

Gram stain: Gram-positive cocci (GPC) in pairs, often demonstrating a lancet-shape. The organisms can be inside or outside of the polymorphonuclear (PMN) cells.
Appearance on blood agar plate (BAP): Alpha-hemolytic and often have a mucoid appearance (shown in lower image), suggesting the presence of a capsule around the cells.