Gram-positive diplococci in Gram stain of sputum
Gram-positive diplococci in Gram stain of sputum as shown above picture and Streptococcus pneumoniae are lancet-shaped (ovoid) cocci in short chains, diplococci, and single cocci. Therefore, the possible pathogen is S. pneumonia,e and let’s know about this etiological agent.
Pathogenicity of Streptococcus pneumoniae
S. pneumoniae causes pneumonia, acute sinusitis, otitis media, meningitis, bacteremia, sepsis, osteomyelitis, septic arthritis, endocarditis, peritonitis, pericarditis, cellulitis, and brain abscess. It is the most common cause of bacterial meningitis in adults and children and is one of the top two isolates found an ear infection, otitis media. S. pneumoniae has a polysaccharide capsule that acts as a virulence factor for the organism; more than 90 different serotypes are known, and these types differ in virulence.
Laboratory Diagnosis of Streptococcus pneumoniae
Gram positive diplococci
S. pneumoniae, Streptococcus salivarius, Streptococcus viridans are referred to collectively as viridans streptococci, a name derived from viridis (Latin for “green”), referring to the green pigment formed by the partial hemolysis of blood agar.
- Optochin Susceptibility Test
Optochin Susceptibility test uses for presumptive identification of Streptococcus pneumoniae from viridans streptococci.
Procedure For Optochin Susceptibility Test
- Select and streak using a straight wire, three to four well-isolated colonies of the suspected organism over a blood agar.
- Place an Optochin disk was over it firmly.
- Incubate the plate at 35°C for 18-24 hours in a CO2 incubator (5-10% CO2).
- Observe the zone of inhibition around the disk. was ≥ 14 mm around the disk, the organism was considered as S. pneumoniae.
Result And Interpretation
zone of inhibition of optochin disk 14 mm or greater: Optochin sensitive: Positive
No Zone of inhibition of optochin disk: Optochin resistance: Negative
The bile solubility test distinguishes Streptococcus pneumoniae from all other alpha-hemolytic (viridans) streptococci. Streptococcus pneumoniae is bile soluble whereas all other alpha-hemolytic streptococci are bile resistant.
Procedure Of Bile Solubility Test
- Prepare a heavy suspension of the organism (equivalent to 1 McFarland standard) from growth on agar media in 0.5 ml of phosphate-buffered saline, pH 7.0.
- Then, add 5 drops of 10% sodium deoxycholate to one of the tubes (labeled ‘test’), and 5 drops of sterile normal saline were added to another tube (labeled’ control’).
- Agitate both the tubes gently and place in a water bath at 35°C for 3 hours, checking hourly for clearing.
Result And Interpretations Of Bile Solubility Test
Positive Bile solubility test: Clearance of turbidity
Negative bile solubility test: No clearance of turbidity
Further Readings
- Cowan & Steel’s Manual for identification of Medical Bacteria. Editors: G.I. Barron & R.K. Felthani, 3rd ed 1993, Publisher Cambridge University Press.
- Bailey & Scott’s Diagnostic Microbiology. Editors: Bettey A. Forbes, Daniel F. Sahm & Alice S. Weissfeld, 12th ed 2007, Publisher Elsevier.
- Clinical Microbiology Procedure Handbook Vol. I & II, Chief in editor H.D. Isenberg, Albert Einstein College of Medicine, New York, Publisher ASM (American Society for Microbiology), Washington DC.
- Colour Atlas and Textbook of Diagnostic Microbiology. Editors: Koneman E.W., Allen D.D., Dowell V.R. Jr, and Sommers H.M.
- Jawetz, Melnick and Adelberg’s Medical Microbiology. Editors: Geo. F. Brook, Janet S. Butel & Stephen A. Morse, 21st ed 1998, Publisher Appleton & Lance, Co Stamford Connecticut.
- Mackie and Mc Cartney Practical Medical Microbiology. Editors: J.G. Colle, A.G. Fraser, B.P. Marmion, A. Simmous, 4th ed, Publisher Churchill Living Stone, New York, Melborne, Sans Franscisco 1996.
- Textbook of Diagnostic Microbiology. Editors: Connie R. Mahon, Donald G. Lehman & George Manuselis, 3rd edition2007, Publisher Elsevier.