Laboratory diagnosis, Treatment and Prevention of Streptococcus pneumoniae

Laboratory diagnosis of Streptococcus pneumoniae

Specimen: Sputum, blood, endotracheal aspirate, bronchoalveolar lavage, cerebrospinal fluid (CSF), pleural fluid, joint fluid, abscess fluid, bones, and other biopsy material.

Microscopy

• Gram staining of sputum shows lancet shaped Gram-positive cocci in pairs.
• Fresh emulsified sputum mixed with antiserum causes capsule swelling (the quellung reaction) for identification of pneumococci.
• In acute pneumococcal otitis media, Gram stain of an aspirated fluid smear from middle ear is useful to demonstrate the bacteria.

Culture

• Sputum or blood is plated on blood agar and incubated at 37° C in the presence of 5–10% carbon dioxide.
• Gray colonies with alpha-hemolysis are observed after overnight incubation.
• Diagnosis of pneumococcal meningitis is confirmed by CSF culture.

Identification of bacteria

• Optochin sensitivity test
  • S. pneumoniae is identified by its sensitivity to optochin (ethylhydrocupreine dihydrochloride).
  • The isolate is streaked onto a blood agar plate and a disk saturated with optochin is placed in the middle of the inoculum.
  • A zone of inhibited bacterial growth is seen around the disk after overnight incubation.

• Bile solubility test
  • It detects an autolytic enzyme, amidase, present in pneumococci, which breaks the bond between alanine and muramic acid of the peptidoglycan of the pneumococcal cell wall.
  • Isolates of S. pneumoniae are lysed rapidly when the autolysins are activated after exposure to bile.
  • Thus the organism can be identified by placing a drop of bile on an isolated colony.

• Inulin fermentation test
  • It ferments inulin and hence differentiate it from other streptococci.

Animal inoculation

• S. pneumoniae can be isolated from clinical specimens containing few pneumococci by intraperitoneal inoculation in mice.
• Pneumococci are demonstrated in the peritoneal exudate and heart blood of the mice, which die 1–3 days after inoculation.

Antigen detection

• Pneumococcal C polysaccharide is excreted in urine and can be detected using a commercially prepared immunoassay.
• The CIEP is a useful test to detect pneumococcal capsular polysaccharide antigen in the CSF for diagnosis of meningitis, and in the blood or urine for diagnosis of bacteremia and pneumonia.
• Latex agglutination test using the latex particles coated with anti-CRP antibody is employed to detect C reactive protein.
• The CRP is used as a prognostic marker in acute cases of acute pneumococcal pneumonia, acute rheumatic fever, and other infectious diseases.

Antibody detection

• The indirect hemagglutination, indirect fluorescent antibody test, and ELISA are used to demonstrate specific pneumococcal antibodies in invasive pneumococcal diseases.

Nucleic Acid–Based Tests

• Nucleic acid probes and PCR assays are used for identification of S. pneumoniae isolates in culture.

Treatment of Streptococcus pneumoniae

• Most pneumococci are susceptible to penicillin.
• Other antibiotics, such as macrolides or selected fluoroquinolones with activity against pneumococci, are available for patients who are allergic to penicillin.
• Penicillin-resistant pneumococci are being increasingly documented.
• The mechanism of this resistance is an alteration of one or more of the bacterium’s penicillin-binding proteins (PBPs) rather than production of beta-lactamase.
• Most resistant strains remain sensitive to third generation cephalosporins (such as cefotaxime or ceftriaxone), and all are still sensitive to vancomycin.
• For serious pneumococcal infections, treatment with a combination of antibiotics is recommended. Vancomycin combined with ceftriazone is used commonly for empiric treatment, followed by monotherapy with an effective cephalosporin, fluoroquinolone or vancomycin.
• Ceftriaxone can be used for meningitis caused by ceftriaxone-susceptible pneumococci.
• Amoxicillin is the drug of choice for treatment of otitis media, sinusitis, and pneumonia caused by penicillin-resistant pneumococci with intermediate resistance.
• Ceftriaxone is the drug of choice for non-CNS invasive pneumococcal diseases caused by penicillin- and ceftriaxone resistant pneumococci.
• Vancomycin is used if the pneumococcus is resistant to ceftriaxone.

Prevention and control of Streptococcus pneumoniae

• Pneumococcal vaccines: 23 valent pneumococcal polysaccharide vaccine (PPSV23) and polyvalent pneumococcal conjugate vaccine (PCV13) play an important role in prevention of pneumococcal diseases.
• Personal hygiene is another important factor.