MICR 420

1. MICR 420 Emerging and Re-Emerging Infectious Diseases Lecture 7: C. trachomatis Dr. Nancy McQueen & Dr. Edith Porter

2. C. trachomatis The genus Chlamydia Morphology Growth and metabolic characteristics Virulence factors Diseases Diagnosis Direct antigen test Culture PCR Immune response Therapy Threats: Apparently under control Overview

3. Chlamydia • Obligate gram-negative intracellular bacteria • No peptidoglycan layer • Major outer membrane protein accounts for 60% of all surface protein • Small genome (~1.0 Mb) • Depend on host molecules for replication • Limited or no ATP synthesis • Two forms exist • Small elementary bodies (EB) • Survives outside host cells • Transferred to new host • Similar in function to a spore • RNA:DNA = 1:1 • Larger reticulate bodies (RB) • Grow within cells • RNA:DNA = 3:1 • Do not survive outside host Reticulate Elementary

4. Chlamydia Developmental Cycle • EB attach to epithelial host cell • Endocytosis • Prevention of fusion with lysosome • Development into RB • RB proliferation, formation of large inclusion bodies • Visible in light microscope • Development into EB • Rupture of host cell and release of EB 48h – 72h Persistent Forms

5. Chlamydia Species • C. pneumoniae • World wide • ~ 10% of pneumonias and 5% of all bronchitis cases • C. psittaci • Psittacosis • Zoonotic pneumonia • Inhalation of droppings from infected birds • Category B biological weapon • C. trachomatis • Various serotypes • Eye infections • STI

6. Chlamydia Infection of the Eyes • Inclusion conjunctivitis • Transmitted to newborn's eyes during passage through the birth canal • Preventive erythromycin • Spread through swimming pool water • Treated with tetracycline • Trachoma • Greatest cause of blindness worldwide

7. Trachoma • World wide ~ 80 (150?) million people infected and ~ 6 million blind • Mostly in developing countries • 3% of cause of all blindness world wide • Transmitted eye-hand-eye, eye-fomite-eye, flies • Infection occurs usually during childhood • Chronic follicular conjunctivitis inversion of eyelashes irritation of cornea corneal ulcerations, scarring vision loss typically at age 30 – 40

8. Trachoma

9. Histopathology of Trachoma http://www.lenntech.com/images/Water%20Borne%20Diseases/Trachoma.jpg

10. Simplified WHO System to Assess Trachoma (from Matthew J. Burton, British Medical Bulletin, 2007)

11. Pathogenesis of C. trachomatis (A) Chlamydia elementary bodies (EB) translocate Tarp into an associated cell to orchestrate rearrangement of host cell actin. The C-terminal domain of Tarp directly nucleates small actin filaments followed by hostmediated signaling involving tyrosine phosphorylation (*P) cascades and ultimately Arp2/3 to mobilize actin assembly necessary for invasion. (B) Throughout development, chlamydial inclusions intercept host-derived vesicles via recruitment of Rab GTPases (Rabs 1, 4, 10, and 11) and SNARE (Vamp 3, 7, and 8) molecules through interactions with Inc proteins. (C) Established inclusions are able to interfere with NF-kB (p50 and RelA) activation through ChlaDub1-mediated prevention of IkBa ubiquitination (*Ub) or CT441-mediated degradation of RelA. This would be predicted to interfere with proinflammatory signals originating from ligand binding to TNF family (TNFR) or pattern recognition (PRR) receptors.

12. Virulence Factors of C. trachomatis (CT) • Contribute to intracellular survival and proliferation • In part secreted via Type III secretion apparatus • Tarp (Translocatedactin recruiting phosphoprotein): effects actin rearrangement and endocytic uptake of CT • CT inserts proteins (Inc and others) in inclusion (endosome) membrane that prevent fusion of host lysosomes • CT releases proteins (ChlaDub1, CT441) into the cytoplasma that interfere with NFkB signal transduction, a key pathway of the acute inflammatory and innate immune response.

13. Immune Response to C. trachomatis • Not well understood • Infection controlled by cell mediated immune response • However: contributes also to the scarring • Antibodies are formed • Tear IgG actually enhance uptake • Tear IgA appear to be protective (Matthew J. Burton, British Medical Bulletin, 2007)

14. Diagnosis of Trachoma • No “gold standard” test • Direct antigen test • PCR • ELISA to measure patient antibodies against outer membrane proteins, conserved heat shock proteins • Culture • inoculating specimens onto cell culture monolayers • Visualization of inclusion bodies after 48 – 72 h using fluorescent labeled monoclonal antibodies against CT LPS or MOMP

15. Therapy and Prophylaxis of Trachoma • Eyelid surgery • Antibiotics to treat the infection • Tetracycline • Azithromycin • Education about facial cleanliness and personal hygiene • Environmental improvements WHO’s Global Alliance for the Elimination of Blinding Trachoma by 2020

16. Take Home Messages • Chlamydia are obligate intracellular bacteria that have a unique life cycle alternating between infectious elementary bodies and proliferative reticulate bodies. • Pathogenic factors include rearrangement of cytoskeleton promoting uptake, inhibition of fusion of lysosomes with endosomes and inhibition of NfkB activation. • C. trachomatis serotypes A-C (-K) cause trachoma, a chronic conjunctivitis leading to blindness. • Trachoma is diagnosed by direct antigen and nucleic acid based test and treated with tetracyline and azithromycin.

17. Resources • http://www.who.int/topics/trachoma/en/ • http://www.cdc.gov • Textbooks • Microbiology: A clinical Approach (2010) Garland Science • Prescott’s Principles of Microbiology (2009) McGraw-Hill • Microbiology: An Evolving Science (2009) Norton • Primary literature • Betts et al., Current Opinion in Microbiology, 2009, 12:81–87 • Matthew J. Burton, British Medical Bulletin, 2007