1. 24/02/2008Dr Ekta, Microbiology, GMCA CHLAMYDIA II MBBS

2. 24/02/2008Dr Ekta, Microbiology, GMCA KEY WORDS Intracellular Energy parasites Elementary body Reticulate body Inclusion bodies – HP, LCL Biotypes/ serotypes Tissue culture C. trachomatis Trachoma Inclusion conjunctivitis Lymphogranuloma venereum Frei test Genital chlamydiasis C. psittaci Psittacosis Ornithosis C. pneumoniae

3. 24/02/2008Dr Ekta, Microbiology, GMCA INTRODUCTION Obligate intracellular parasites of humans, animals & birds Resemble bacteria except it cannot multiply outside living cells/ tissues (like viruses) Cannot synthesize ATP – depends on host cell for energy & nutrient sources. Hence, called Energy Parasites.

4. 24/02/2008Dr Ekta, Microbiology, GMCA CHLAMYDIA 4 species in the Genus Chlamydia – C. trachomatis, C. pneumoniae, affects humans C. psittaci and C. pecorum affects ruminants All are non-motile, gram negative; share antigens, have both DNA and RNA.

5. 24/02/2008Dr Ekta, Microbiology, GMCA Classification C.trachomatis : eye & genital infections, infant pneumonia, and LGV (Lymphogranuloma Venereum) in adults C.pneumoniae : different types of respiratory infections. C.psittaci : psittacosis in man, ornithosis in birds

6. 24/02/2008Dr Ekta, Microbiology, GMCA Life cycle Chlamydiae occur in 2 forms : 1.Elementary body – extracellular, infective form 2.Reticulate body – intracellular, growing & replicative form Chlamydial microcolony within the host cell is called Inclusion body. Mature inclusion body contains 100 - 500 elementary bodies

7. 24/02/2008Dr Ekta, Microbiology, GMCA

8. 24/02/2008Dr Ekta, Microbiology, GMCA Life cycle C. psittaci – host cell is severely damaged, EBs are released within 48 hrs by cell lysis C.trachomatis – mature inclusion body appears to be exocytosed in 72- 96 hrs.

9. 24/02/2008Dr Ekta, Microbiology, GMCA Antigenic Properties Three major Ags 1.Genus specific Ag – heat stable, common to all chlamydiae, a lipopolysaccharide resembling LPS of GNB. Present in all stages. 2.Species specific protein Ags – present at the envelope surface, help in classifying chlamydia into species 3.Ag for Intraspecies typing – found only in some members of a species, located on major OMP (MOMP), demonstrated by micro- IF. Classifies species into serovars/ serotypes

10. 24/02/2008Dr Ekta, Microbiology, GMCA Variants of Chlamydia C. trachomatis – 2 biovars : TRIC & LGV 1.TRIC – Trachoma, Inclusion conjunctivitis - divided into 12 serovars 2.LGV – Lymphogranuloma venereum – 3 serovars

11. 24/02/2008Dr Ekta, Microbiology, GMCA Human diseases SpeciesSerotypeDisease C. trachomatis A, B, Ba, CEndemic blinding trachoma C. trachomatis D to KInclusion conjunctivitis. Genital chlamydiasis C. trachomatis L1, L2, L3Lymphogranuloma venereum C. psittaci Many serotypesPsittacosis C. pneumoniae Acute resp. disease

12. 24/02/2008Dr Ekta, Microbiology, GMCA Laboratory Diagnosis 4 approaches available: 1.Microscopic demonstration of inclusion or elementary bodies 2.Isolation of chlamydia 3.Demonstration of chlamydial Ag 4.Demonstration of Abs or hypersensitivity

13. 24/02/2008Dr Ekta, Microbiology, GMCA Microscopy Gram negative but stained better by Giemsa, Castaneda or Machiavello stains. Giemsa Stain: Elementary body & the Reticulate body stains blue in cytoplasm Lugol’s iodine: rapid & simple screening method for ocular infections, stains glycogen matrix of C. trachomatis Immunoflurescence staining: more sensitive & specific, by using monoclonal Abs. Identifies inclusion bodies as well as extracellular elementary bodies. Used for ocular, cervical or urethral specimens.

14. 24/02/2008Dr Ekta, Microbiology, GMCA IF staining

15. 24/02/2008Dr Ekta, Microbiology, GMCA Culture Yolk sac of 6 - 8 days old chick embryo. Tissue culture – McCoy, HeLa cell lines * C. psittaci carry the risk of laboratory infection.

16. 24/02/2008Dr Ekta, Microbiology, GMCA Microscopic appearance A monolayer of tissue culture cells has been exposed to cells of chlamydia trachomatis. Infected cells within the cell sheet have a cytoplasm with a granular appearance.

17. 24/02/2008Dr Ekta, Microbiology, GMCA Demonstration of antigens Micro – IF : infected ocular or genital samples are stained with fluorescent conjugated Ab ELISA – best for screening large number of specimens, detects LPS Ag Molecular methods - PCR

18. 24/02/2008Dr Ekta, Microbiology, GMCA Chlamydia trachomatis Leading cause of ocular & genital infections Pathogenicities : 1.Trachoma 2.Inclusion conjunctivitis 3.Infant pneumonia 4.Genital infections – genital chlamydiasis, LGV

19. 24/02/2008Dr Ekta, Microbiology, GMCA Trachoma Greek word trakkus – rough (roughness of conjunctiva) Caused by C. trachomatis types A, B & C. Chronic keratoconjunctivitis Transmitted by fingers, fomites, flies or dust Established trachoma passes through 4 stages (I – IV). Infectivity is maximum in early cases, stage IV is non infectious.

20. 24/02/2008Dr Ekta, Microbiology, GMCA Laboratory diagnosis of Trachoma Demonstration of characteristic inclusion bodies (Halberstaedter Prowazek or HP bodies) in conjunctival scrapings by Giemsa. Culture – yolk sac, cell lines

21. 24/02/2008Dr Ekta, Microbiology, GMCA Treatment & Control Local application of antibiotics Oral administration - Tetracycline or Doxycycline for several weeks Single dose Azithromycin Control – mass education & chemotherapy

22. 24/02/2008Dr Ekta, Microbiology, GMCA Inclusion Conjunctivitis Caused by C. trachomatis types D to K Naturally present in the genital tract Neonatal form - “Inclusion Blenorrhoea ”, develops when the infant is in birth canal, appears 5-12 days after birth, prevented by local application of antibiotics Adult form – “Swimming Pool Conjunctivitis ” – associated with bathing in community swimming pools contaminated with chlamydia from genital secretions.

23. 24/02/2008Dr Ekta, Microbiology, GMCA Infant Pneumonia Usually occurs around 4-16 wks of age. Cough & wheezing. Rarely fever Conjunctivitis often precedes pneumonia

24. 24/02/2008Dr Ekta, Microbiology, GMCA Genital Infections Two types : 1.Genital chlamydiasis 2.Lymphogranuloma venereum

25. 24/02/2008Dr Ekta, Microbiology, GMCA Genital chlamydiasis Most common STD, clinical spectrum similar to gonococcal infections. Men – urethritis (NGU), epididymitis, proctitis & Reiter’s syndrome Women – acute urethral syndrome, mucopurulent cervicitis, endometritis, salpingitis, PID, infertility, ectopic pregnancy, premature delivery, postpartum fever. Diagnosis – gram stained smears of urogenital exudates showing –more than 4 neutrophils / OIF in urethritis, – >30 / OIF in cervicitis. Confirmatory tests – culture, micro-IF, ELISA, PCR

26. 24/02/2008Dr Ekta, Microbiology, GMCA Lymphogranuloma venereum Most commonly caused by L2 type Site – regional lymph nodes Incubation period – 3 days to 5 wks 1° lesion – small painless papulovesicular lesion on external genitalia 2° stage – after 2 wks, lymphatic spread to draining LNs (men – inguinal, women – intrapelvic & pararectal)

27. 24/02/2008Dr Ekta, Microbiology, GMCA Lymphogranuloma venereum Nodes enlarge, suppurate, become adherent to the skin & break down to form discharging sinuses. Metastatic complications – may occur, involves joints, eyes & meninges 3° stage – chronic, lasts for several years; scarring & lymphatic blockage Late sequelae more distressing in women – rectal strictures, elephantiasis of vulva (esthiomene)

28. 24/02/2008Dr Ekta, Microbiology, GMCA Laboratory Diagnosis of LGV Demonstration of elementary bodies in materials aspirated from bubos (inguinal) Isolation – cell cultures Serology – detection of Abs 1.CFT 1 : 64 or more 2.Micro- IF 1 : 512 or more Frei Test – ID test using crude chlamydial Ag, not done now.

29. 24/02/2008Dr Ekta, Microbiology, GMCA Chlamydia psittaci Causes Psittacosis – disease of parrots Shed in the droppings or nasal discharges and aerosols are liberated. Humans – occupational disease as in poultry workers, pigeon farmers, petshop owners, veterinarians; laboratory acquired infection Infection by inhalation. Mild influenza like syndrome to fatal pneumonia

30. 24/02/2008Dr Ekta, Microbiology, GMCA Laboratory Diagnosis Specimen – blood (early stages), later sputum Demonstration of LCL (Levinthal-Cole-Lillie) Inclusion bodies in alveolar macrophages, mouse brain, yolk sac, cell cultures LCL bodies - more diffuse & irrregular, not stained by iodine Serology – CFT, micro-IF

31. 24/02/2008Dr Ekta, Microbiology, GMCA Chlamydia pneumoniae Isolated in 1986 from acute respiratory diseases in adults in Taiwan – C. psittaci strain TWAR Later classified as a separate species. Common cause of respiratory disease in older children & adults. C/F – pharyngitis, sinusitis, bronchitis & pneumonia (atypical pneumonia) Outbreaks in closed communities. Reinfections are common.

32. 24/02/2008Dr Ekta, Microbiology, GMCA Chlamydia pneumoniae Diagnosis – Ag detection by EIA, direct IF, PCR, as isolation is very difficult. Treatment – clarithromycin or azithromycin Newer findings – suspected to be associated with atherosclerosis and its clinical effects like coronary, carotid & cerebral arterial disease. Evidences – detection of chlamydial Ags in plaques, isolation of chlamydia from coronary artery plaques, experimental induction of atheroma in rabbits infected with chlamydia

33. 24/02/2008Dr Ekta, Microbiology, GMCA So…What’s Chlamydia anyways? How is it detected, treated and prevented? If it happens to me, what do I do? What are the possible complications? Chlamidia