Chlamydia pneumoniae: Culture Methods

  • C.-C. Kuo


After T’ang et al. in Peking first successfully cultivated trachoma organisms in chick embryo in 1957, egg culture became a standard method of isolation and growth of Chlamydia trachomatis [1]. The researchers’ success was owed to the use of streptomycin, but not penicillin, for control of contamination. Since egg culture is not only cumbersome for cultivation but also for purification of organisms from yolk sacs, finding a sensitive cell culture method was vigorously pursued. In 1965, Gordon and Quan described a method for isolation of trachoma agents in McCoy cell culture using a flat-bottomed culture vial [2]. To enhance the sensitivity, McCoy cells were irradiated with γ-radiation and centrifugation was applied during the absorption period after specimens were inoculated to culture vials. The procedure was simplified by Ripa and Mårdh, who replaced γ-radiation with the use of cycloheximide [3]. In their method non-irradiated McCoy cells were used. To enhance the sensitivity of McCoy cells to C. trachomatis growth, cycloheximide was added to culture medium for cultivating infected cells. Cycloheximide inhibits host cell protein synthesis but does not affect chlamydial metabolism; thus, it reduces the competition for nutrients between host cells and parasites and enhances the growth of chlamydial organisms. This cell culture method was quickly adapted, and replaced egg culture for isolation and growth of C. trachomatis. Other modified methods have appeared and been used, such as the use of DEAE-dextrantreated HeLa 229 cells [4]. However, the crucial factors are centrifugation and cycloheximide. The use of 96-well microtiter plates for isolation of C. trachomatis was first introduced by McComb and Puzniak in 1974 [5] and later popularized by Yoder et al. in 1981 for handling large volumes of clinical specimens [6].


Respiratory Syncytial Virus Chlamydia Trachomatis McCoy Cell Chlamydial Organism Chlamydial Culture 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. 1.
    T’ang F, Chang H, Huang Y, Wang K (1951) Studies on the etiology of trachoma with special reference to isolation of the virus in chick embryo. Chin Med J 75: 429–447Google Scholar
  2. 2.
    Gordon FB, Quan AL (1965) Isolation of the trachoma agent in cell culture. Proc Soc Exp Biol Med 118: 354–359PubMedGoogle Scholar
  3. 3.
    Ripa KT, Mårdh PA (1977) Cultivation of Chlamydia trachomatis in cycloheximidetreated McCoy cells. J Clin Microbiol 6: 328–331PubMedGoogle Scholar
  4. 4.
    Kuo C-C, Wang S-P, Wentworth B, Grayston JT (1972) Primary isolation of TRIC organisms in HeLa 229 cells treated with DEAE-dextran. J Infect Dis 125: 665–668PubMedCrossRefGoogle Scholar
  5. 5.
    McComb DE, Puzniak CI (1974) Micro cell culture method for isolation of Chlamydia trachomatis. Appl Microbiol 28: 727–729PubMedGoogle Scholar
  6. 6.
    Yoder BL, Stamm WE, Koester CM, Alexander ER (1981) Microtest procedure for isolation of Chlamydia trachomatis. J Clin Microbiol 13: 1036–1039PubMedGoogle Scholar
  7. 7.
    Kuo C-C, Chen H-H, Wang S-P, Grayston JT (1986) Identification of a new group of Chlamydia psittaci strains called TWAR. J Clin Microbiol 24: 1034–1037PubMedGoogle Scholar
  8. 8.
    Cles LD, Stamm WE (1990) Use of HL cells for improved isolation and passage of Chlamydia pneumoniae. J Clin Microbiol 28: 938–940PubMedGoogle Scholar
  9. 9.
    Kuo C-C, Grayston JT (1990) A sensitive cell line, HL cells, for isolation and propagation of Chlamydia pneumoniae strain TWAR. J Infect Dis 162: 755–758PubMedCrossRefGoogle Scholar
  10. 10.
    Wong KH, Skelton SK, Chan YK (1992) Efficient culture of Chlamydia pneumoniaewith cell lines derived from the human respiratory tract. J Clin Microbiol 30: 1625–1630PubMedGoogle Scholar
  11. 11.
    Roblin PM, Dumornay W, Hammerschlag MR (1992) Use of Hep-2 cells for improved isolation and passage of Chlamydia pneumoniae. J Clin Microbiol 30: 1968–1971PubMedGoogle Scholar
  12. 12.
    Kuo C-C, Wang S-P, Grayston JT (1977) Growth of trachoma organisms in HeLa 229 cell culture. In: Hobson D, Holmes KK (eds) Nongonococcal urethritis and related infections. American Society for Microbiology, Washington DC, pp 328–336Google Scholar
  13. 13.
    Cavallaro JJ, Monto AS (1972) HL cells, a sensitive line for the isolation and propagation of respiratory syncytial virus. Proc Soc Exp Biol Med 140: 507–510PubMedGoogle Scholar
  14. 14.
    Moulder JW, Hatch TP, Kuo C-C, Schachter J, Storz J (1984) Chlamydia Jones, Rake and Sterns. In: Krieg NR (ed) Bergey’s manual of systematic bacteriology, vol 1. Williams & Wilkins, Baltimore, pp 729–735Google Scholar
  15. 15.
    Wang S-P, Grayston JT (1991) Chlamydia pneumoniae elementary body antigenic reactivity with fluorescent antibody is destroyed by methanol. J Clin Microbiol 29: 1539–1541PubMedGoogle Scholar
  16. 16.
    Maass M, Bartels C, Engel PM, Mamat U, Sievers HH (1998) Endovascular presence of viable Chlamydia pneumoniae is a common phenomenon in coronary artery disease. J Am Coll Cardiol 31: 827–832PubMedCrossRefGoogle Scholar
  17. 17.
    Grayston JT, Kuo C-C, Wang S-P, Altman J (1986) A new Chlamydia psittaci strain TWAR, isolated in acute respiratory tract infection. N Engl J Med 315: 161–168PubMedCrossRefGoogle Scholar
  18. 18.
    Ekman M-R, Grayston JT, Visakorpi R, Kleemola M, Kuo C-C, Saikku P (1993) An epidemic of infections due to Chlamydia pneumoniae in military conscripts. Clin Infect Dis 17: 420–425PubMedCrossRefGoogle Scholar
  19. 19.
    Boman J, Allard A, Persson K, Lundborg M, Juto P, Wadell G (1997) Rapid diagnosis of respiratory Chlamydia pneumoniae infection by nested touchdown polymerase chain reaction compared with culture and antigen detection by EIA. J Infect Dis 175: 1523–1526PubMedCrossRefGoogle Scholar
  20. 20.
    Kuo C-C, Grayston JT (1988) Factors affecting viability and growth in HeLa 229 cells of Chlamydia sp. strain TWAR. J Clin Microbiol 26: 812–815PubMedGoogle Scholar
  21. 21.
    Maass M, Dalhoff K (1995) Transport and storage conditions for cultural recovery of Chlamydia pneumoniae. J Clin Microbiol 33: 1793–1796PubMedGoogle Scholar

Copyright information

© Springer-Verlag Italia, Milano 1999

Authors and Affiliations

  • C.-C. Kuo

There are no affiliations available

Personalised recommendations