Mycoplasma_Ureaplasma_RESPI_5154_2023_BENSON_Final.pptx

Mycoplasma and Ureaplasma (and bacteria that cause atypical pneumonia) November 29th, 2023 11 AM - 12 Noon https://www.cdc.gov/pneumonia/atypical/ mycoplasma/hcp/disease-specifics.html Marc Benson, Ph.D. • • • • Office: 317 E-mail: [email protected] Phone: (575) 674-2317 Office Hours: By appointment or stop by office Reading Material Murray et al. Medical Microbiology, 9th ed. Chapter 29, 33, 35 Metwally, Mirihan & Yassin, Aymen & Essam, Tamer & Hamouda, Hayam & Amin, Magdy. (2014). Detection, Characterization, and Molecular Typing of Human Mycoplasma spp. fro Major Hospitals in Cairo, Egypt. TheScientificWorldJournal. 2014. 549858. 10.1155/2014/549858. 5155: Respiratory System I 1 Copyright Notice • All reproduction or use of copyrighted materials shall comply with provisions of applicable law. Individuals are responsible for maintaining copyright compliance in good faith and with each intended use. Please consult BCOM Policy B5041 and the copyright guidelines located on https://bcomnm.org/copyrightguidelines/ for detailed information • BCOM prohibits duplication, distribution, or use of copyrighted materials by students, faculty and staff unless a fair use or other exemption applies, or permission has been obtained from the work's rights holder (which may or may not be the author/creator) • Infringement of copyright law may be considered a violation of the College’s Code of Professional Conduct. Anyone found liable for civil and/or criminal copyright infringement may be responsible for any monetary damages suffered by the College due to such violation(s) of this policy or related law or regulation 2 Lecture Outline Mycoplasma pneumoniae Other Mycoplasmatales Chlamydia (Chlamydophila) pneumoniae Legionella pneumophila 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. Pre-Lecture Clicker Questions Objectives Pneumonia basics General Information Epidemiology Lifecycle Clinical Disease Laboratory Diagnostics Summary Post-Lecture Clicker Questions Learning Catalytics Questions 3 Pre-Lecture Clicker Questions Lecture Outline Mycoplasma pneumoniae Other Mycoplasmatales Chlamydia (Chlamydophila) pneumoniae Legionella pneumophila 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. Pre-Lecture Clicker Questions Objectives Pneumonia basics General Information Epidemiology Lifecycle Clinical Disease Laboratory Diagnostics Summary Post-Lecture Clicker Questions Learning Catalytics Questions 5 Objectives • Describe the unique features of the Mycoplasma cell structure • List the most common bacterial agents that cause typical community-acquired pneumonia and recognize the difference between the clinical features of typical and atypical pneumonia • Recognize Mycoplasma pneumoniae as a major cause of atypical pneumonia based on the most common population infected, children and young adults • Recognize that Chlamydophila pneumoniae and Legionella pneumophila are also causes of atypical pneumonia and describe the unique features of each organism • Describe the most important virulence factor of Mycoplasma pneumoniae ( P1 adhesion) and its role in adhesion to ciliated epithelium • Describe the difficulty in establishing a definitive diagnosis of Mycoplasma pneumoniae, and the laboratory methods most commonly used for identification • List the clinically important Mycoplasmataceae found in the genitourinary tract • Compare and contrast Mycoplasma pneumoniae and Ureaplasma urealyticum, Mycoplasma hominis and Mycoplasma genitalium in terms of culture requirements ( growth media additives), carbohydrate utilization and antibiotic resistance • Explain the mechanism of action of antibiotics effective against Mycoplasma pneumoniae (Erythromycin, Tetracyclins, and fluoroquinolones) 6 Lecture Outline Mycoplasma pneumoniae Other Mycoplasmatales Chlamydia (Chlamydophila) pneumoniae Legionella pneumophila 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. Pre-Lecture Clicker Questions Objectives Pneumonia basics General Information Epidemiology Lifecycle Clinical Disease Laboratory Diagnostics Summary Post-Lecture Clicker Questions Learning Catalytics Questions 7 Pneumonia Pneumonias • Fever • Respiratory symptoms (cough, chest pain, dyspnea, etc.) Typical pneumonia • Respiratory symptoms (dyspnea, productive • Evidence of parenchymal involvement cough, bloody and/or purulent sputum) • Symptoms abrupt Lobar pneumonia • Constitutional symptoms (fever, malaise) • Involvement of a single lobe • Consolidation on x-ray Bronchopneumonia • Involvement of multiple areas in multiple lobes (patchy) Interstitial pneumonia • Involvement of tissue surrounding alveoli Community-acquired pneumonia • Pneumonia acquired outside of hospital Atypical pneumonia • Caused by bacteria that may not be detected by Gram stain • Respiratory symptoms (non-productive cough) • Gradual onset • Symptoms – more chronic-like • Constitutional signs (malaise, fever) • Interstitial patchy infiltrate on x-ray Hospital-acquired pneumonia • Pneumonia acquired in or within 48 hours of hospital stay Ventilator-associated pneumonia • Pneumonia when intubated Aspiration pneumonia • Inhalation of stomach acid, saliva, other material (food), which carries pathogens to lungs • Anaerobes from oral cavity 8 X-Rays (FYI) • • Lobar pneumonia Involvement of a distinct region of the lung X-ray shows dense areas in the lung due to consolidation (influx of fluid and leukocytes) • • Atypical pneumonia Multiple lobes involved X-ray shows diffuse patchy infiltrates Mycoplasma pneumoniae pneumonia Diffuse bilateral interstitial infiltrates with M. pneumoniae infection. Courtesy of Dwight A Powell, MD. These images are to show you the visual difference between lobar and atypical pneumonia. Interpretation will be taught by the clinicians at a later time. Etiology of Bacterial Pneumonia by Age Age Bacterial Etiology Newborns • • • Group B streptococcus E. coli Listeria monocytogenes (rare) Children • • • • • Streptococcus pneumoniae Haemophilus influenzae Moraxella catarrhalis Staphylococcus aureus Streptococcus pyogenes • • • • • • • S. pneumoniae (and 2° infection) H. influenzae (and 2° infection) Klebsiella pneumoniae Enterobacteriaceae Pseudomonas (cystic fibrosis) Anaerobic (aspiration) Staphylococcus aureus (2° infection; IV drugs) Streptococcus pyogenes (2° infection) Adults • Viral Etiology • Respiratory syncytial virus • Parainfluenza virus types 1,2, 3 (mostly) • Influenza A virus & B virus Atypicals • Mycoplasma pneumoniae • Chlamydia pneumoniae • Chlamydia trachomatis (infants) Atypicals • M. pneumoniae • C. pneumoniae • Legionella pneumophila Those in orange represent the most common for that age group • • • • Influenza virus A and B Respiratory syncytial virus Human metapneumovirus Adenovirus 4 and 7 10 Lecture Outline 1. 2. 3. 4. 5. Mycoplasma pneumoniae 6. Other Mycoplasmatales Chlamydia (Chlamydophila) pneumoniae 7. Legionella pneumophila 8. 9. 10. 11. Pre-Lecture Clicker Questions Objectives Pneumonia basics General Information Epidemiology Lifecycle Clinical Disease Laboratory Diagnostics Summary Post-Lecture Clicker Questions Learning Catalytics Questions 11 Causative Agents of Pneumonia Community-Acquired Pneumonia Typical Bacteria • Streptococcus pneumoniae • Haemophilus influenzae • Moraxella catarrhalis • Staphylococcus aureus • Enterobacteriaceae (Klebsiella pneumoniae) Atypical Bacteria • Mycoplasma pneumoniae • Chlamydia (Chlamydophila) pneumoniae • Legionella pneumophila Viral Pneumonia • Respiratory syncytial virus (RSV) • Parainfluenza virus types 1,2, 3 (mostly) • Influenza A virus & B virus • Human metapneumovirus • Adenovirus Hospital-Acquired Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Streptococcus spp. • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Serratia marcescens, Escherichia coli ) Ventilator-Associated Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Escherichia coli ) • Acinetobacter baumannii • Stenotrophomonas maltophilia Aspiration Pneumonia • Anaerobic oral flora (Bacteroides, Prevotella, Fusobacterium, Peptostreptococcus), and mixed with aerobic bacteria Zoonotic Pneumonia Bacteria • Bacillus anthracis • Coxiella burnetii • Yersinia pestis • Chlamydia psittaci • Pasteurella multocida • Francisella tularensis Viral • Hantavirus • Influenza • Coronavirus (SARS, MERS) Chronic Pneumonia Bacteria • Nocardia spp. • Actinomyces spp. • Mycobacterium spp. Fungi (also acute depending on exposure load) • Aspergillus spp. • Blastomyces dermatitidis • Coccidioides immitis • Cryptococcus neoformans (HIV/AIDS) • Paracoccidioides brasiliensis • Talaromyces marneffei (HIV/AIDS) • Histoplasma capsulatum Necrotizing Pneumonia and Lung Abscess • Anaerobic bacteria with or without mixed aerobes • Staphylococcus aureus • Klebsiella pneumoniae • Streptococcus pyogenes Less Common Causes of Viral Pneumonia • Rhinovirus, Enteroviruses, Echovirus, Coxsackievirus, Herpesvirus (EBV, CMV, VZV, HSV), Measles 12 Causative Agents of Pneumonia Community-Acquired Pneumonia Typical Bacteria • Streptococcus pneumoniae • Haemophilus influenzae • Moraxella catarrhalis • Staphylococcus aureus • Enterobacteriaceae (Klebsiella pneumoniae) Atypical Bacteria • Mycoplasma pneumoniae • Chlamydia (Chlamydophila) pneumoniae • Legionella pneumophila Viral Pneumonia • Respiratory syncytial virus (RSV) • Parainfluenza virus types 1,2, 3 (mostly) • Influenza A virus & B virus • Human metapneumovirus • Adenovirus Hospital-Acquired Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Streptococcus spp. • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Serratia marcescens, Escherichia coli ) Ventilator-Associated Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Escherichia coli ) • Acinetobacter baumannii • Stenotrophomonas maltophilia Aspiration Pneumonia • Anaerobic oral flora (Bacteroides, Prevotella, Fusobacterium, Peptostreptococcus), and mixed with aerobic bacteria Zoonotic Pneumonia Bacteria • Bacillus anthracis • Coxiella burnetii • Yersinia pestis • Chlamydia psittaci • Pasteurella multocida • Francisella tularensis Viral • Hantavirus • Influenza • Coronavirus (SARS, MERS) Chronic Pneumonia Bacteria • Nocardia spp. • Actinomyces spp. • Mycobacterium spp. Fungi (also acute depending on exposure load) • Aspergillus spp. • Blastomyces dermatitidis • Coccidioides immitis • Cryptococcus neoformans (HIV/AIDS) • Paracoccidioides brasiliensis • Talaromyces marneffei (HIV/AIDS) • Histoplasma capsulatum Necrotizing Pneumonia and Lung Abscess • Anaerobic bacteria with or without mixed aerobes • Staphylococcus aureus • Klebsiella pneumoniae • Streptococcus pyogenes Less Common Causes of Viral Pneumonia • Rhinovirus, Enteroviruses, Echovirus, Coxsackievirus, Herpesvirus (EBV, CMV, VZV, HSV), Measles 13 Neisseria meningitides Flow Chart Gram Negative Bacteria + Borrelia spp Treponema spp Leptospira interrogans Spirochete Maltose Fermentation Neisseria gonorrhoeae Moraxella catarrhalis + Growth on modified ThayerMartin agar - Gram Stain Gram Negative N/A Not Stained Atypical Mycoplasma spp Vibrio cholerae Campylobacter jejuni Intracellular Chlamydia/Chlamydophila spp Rickettsia spp Curved Diplococcus Shape + Haemophilus influenzae Pasteurella multocida Brucella melitensis Bordetella pertussis Legionella pneumophila Francisella tularensis Mycobacterium spp. Nocardia spp. Acid fast + Coccobacilli Bacilli - Escherichia coli Klebsiella spp Enterobacter spp Citrobacter spp Lactose Fermentation - + Oxidase test + H2S Production and Motility Pseudomonas aeruginosa Urease test - Salmonella spp Proteus + spp Shigella spp ONPG + Yersinia entercolitica 14 Mycoplasma pneumoniae General Biology • Atypical bacteria • No peptidoglycan (no cell wall) • Pleomorphic (oddly shaped) • Very small compared to most bacteria • Membrane contains cholesterol obtained from the host • Causes atypical pneumoniae (walking pneumonia) Epidemiology • Strict human pathogen • Causes community-acquired pneumonia in children (5-15 years old) • Transmission • Respiratory droplets • Spreads slowly due to long incubation time • Risk factors: • Children (5-15 years old) • Close contacts (miliary barracks, daycares, nursing homes) https://www.cdc.gov/pneumonia/atypical/ mycoplasma/hcp/disease-specifics.html 15 Mycoplasma pneumoniae Pathogenesis and Virulence Factors • Adherence and colonization • Attaches to respiratory epithelium, but does not invade • Uses a tip-like organelle (P1 adhesin) to adhere to host epithelium (major virulence factor) • Attachment leads to cilia arrest and subsequent cell death • Interferes with ciliary clearance (persistent cough) • Immune cell infiltration enhances cytotoxicity • Community Acquired Respiratory Distress Syndrome (CARDS) toxin • ADP-ribosyltransferase activity • Vacuolating activity P1 P1 From M. Modrzakowski aslides: Mycoplasma and Ureaplasma Clinical Microbiology and Infectious Diseases.Published January 1, 2008. Pages 63-63. © 2008. P1 https://www.cdc.gov/pneumonia/atypical/mycoplasma/hcp/disease-specifics.html 16 Mycoplasma pneumoniae Clinical Disease • Primary presentation is tracheobronchitis • Slow onset • May develop low grade fever, malaise, headache • Dry, nonproductive cough, no exudate • Atypical Pneumonia (walking pneumonia) • Patients generally feel well • Dry, nonproductive cough, no exudate • A common pneumonia in young adults • X-ray: Diffuse interstitial infiltrates • Can cause meningoencephalitis, paralysis, myelitis, pericarditis, hemolytic anemia, arthritis, mucocutaneous lesions • May be involved with autoimmunity https://emedicine.medscape.com/article/363083-overview 17 Mycoplasma pneumoniae Laboratory Diagnostics • Microscopy is not useful – does not stain – why? fried egg colony morphology • Cold agglutinins (non-specific) • Cross reactive anti-erythrocyte antibodies • Precipitate at cold temperatures • Culture requires prolonged incubation (weeks) • Eaton’s agar • Cholesterol in medium • Colonies look like fried eggs • Less so for M. pneumoniae • Serology • High titer or four-fold rise in convalescence • Polymerase chain reaction (PCR) • Nasopharyngeal samples • Specific and sensitive Mycoplasmas and their host: emerging and re-emerging minimal pathogens, Citti, Christine, Trends in Microbiology, Volume 21, Issue 4, 196-203, Mycoplasma under the microscope: Mycoplasma agalactiae as an example. (a) Transmission electron microscopy of M. agalactiae illustrating the mycoplasma cell morphology and size. (b) Mycoplasma colonies displaying a typical fried egg shape on solid..., Copyright © 2013 Elsevier Ltd Metwally, Mirihan & Yassin, Aymen & Essam, Tamer & Hamouda, Hayam & Amin, Magdy. (2014). Detection, Characterization, and Molecular Typing of Human Mycoplasma spp. from Major Hospitals in Cairo, Egypt. TheScientificWorldJournal. 2014. 549858. 10.1155/2014/549858. 18 Mycoplasma pneumoniae Treatments • Macrolides • Bind to the 50S subunit of the bacterial ribosome, thus inhibiting translation of mRNA • Tetracyclines (not for use in children) • Inhibit protein synthesis by blocking the attachment of charged aminoacyl-tRNA to the A site on the ribosome. Tetracycline binds to the 30S subunit of microbial ribosomes • Fluoroquinolones (not for use in children) • Block DNA replication by inhibiting DNA gyrase or DNA topoisomerase Mycoplasma pneumoniae General • M. pneumoniae is a strict aerobe • Non-motile, pleomorphic • Cell membrane contains cholesterol • No peptidoglycan; no cell wall Summary Epidemiology • Strict human pathogen • Causes community-acquired pneumonia in children (5-15 years old) • Transmission - Respiratory droplets • Risk factors - Children (5-15 years old) Pathogenesis • Tip-like organelle (P1) for tight adherence, cilia arrest, and cell death • Purulent discharge Community Acquired Respiratory Distress Syndrome (CARDS) toxin Disease • Primary presentation is tracheobronchitis • Slow onset, may develop low grade fever, malaise, headache, dry, nonproductive cough, no exudate • Atypical Pneumonia (walking pneumonia) • Patients generally feel well, dry, nonproductive cough, no exudate, X-ray: Diffuse interstitial infiltrates • Can cause meningoencephalitis, paralysis, myelitis, pericarditis, hemolytic anemia, arthritis, mucocutaneous lesions • May be involved with autoimmunity Lab Diagnosis • • • • • Fried egg colonies on Eaton’s agar Cold-agglutinins - autoantibodies that lead to erythrocyte destruction Serology: high titer or 4-fold increase in convalescence PCR Microscopy and Gram stain not useful 20 Organism Site Disease Lab Diagnostics Respiratory tract Tracheobronchitis, pharyngitis, pneumonia, secondary complications (neurologic, pericarditis, hemolytic anemia, arthritis, mucocutaneous lesions) Mycoplasma genitalium Genitourinary tract Nongonococcal urethritis, pelvic inflammatory disease Mycoplasma hominis Respiratory tract, genitourinary tract Pyelonephritis, postpartum fever, systemic infections in immunocompromised patients Ureaplasma urealyticum Respiratory tract, genitourinary tract Nongonococcal urethritis, pyelonephritis, spontaneous abortion, premature birth Mycoplasma pneumoniae • • • • M. pneumoniae is a strict aerobe; all others are facultative anaerobes Non-motile, pleomorphic Cell membrane contains cholesterol No peptidoglycan; no cell wall Treatment • • • NAAT cold agglutination glucose catabolism positive • • • Macrolides Tetracyclines Fluoroquinolones • • NAAT glucose catabolism positive • • Macrolides Fluoroquinolones • • NAAT glucose catabolism negative • • • Tetracyclines Clindamycin Fluoroquinolones • • • NAAT urea required for growth hydrolyze urea • • • Macrolides Tetracyclines Fluoroquinolones • M. genitalium, M hominis, and U. urealyticum colonize infants, particularly females • Carriage does not persist • Genitourinary colonization increases after puberty and sexual activity • 15% men and women colonized with M. hominis • 45-75% colonized with Ureaplasma Murray, Patrick R., PhD, Medical Microbiology, Chapter 33, Copyright © 2022, Elsevier Inc. All rights reserved. Other Mycoplasma and Ureaplasma spp. Causative Agents of Pneumonia Community-Acquired Pneumonia Typical Bacteria • Streptococcus pneumoniae • Haemophilus influenzae • Moraxella catarrhalis • Staphylococcus aureus • Enterobacteriaceae (Klebsiella pneumoniae) Atypical Bacteria • Mycoplasma pneumoniae • Chlamydia (Chlamydophila) pneumoniae • Legionella pneumophila Viral Pneumonia • Respiratory syncytial virus (RSV) • Parainfluenza virus types 1,2, 3 (mostly) • Influenza A virus & B virus • Human metapneumovirus • Adenovirus Hospital-Acquired Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Streptococcus spp. • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Serratia marcescens, Escherichia coli ) Ventilator-Associated Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Escherichia coli ) • Acinetobacter baumannii • Stenotrophomonas maltophilia Aspiration Pneumonia • Anaerobic oral flora (Bacteroides, Prevotella, Fusobacterium, Peptostreptococcus), and mixed with aerobic bacteria Zoonotic Pneumonia Bacteria • Bacillus anthracis • Coxiella burnetii • Yersinia pestis • Chlamydia psittaci • Pasteurella multocida • Francisella tularensis Viral • Hantavirus • Influenza • Coronavirus (SARS, MERS) Chronic Pneumonia Bacteria • Nocardia spp. • Actinomyces spp. • Mycobacterium spp. Fungi (also acute depending on exposure load) • Aspergillus spp. • Blastomyces dermatitidis • Coccidioides immitis • Cryptococcus neoformans (HIV/AIDS) • Paracoccidioides brasiliensis • Talaromyces marneffei (HIV/AIDS) • Histoplasma capsulatum Necrotizing Pneumonia and Lung Abscess • Anaerobic bacteria with or without mixed aerobes • Staphylococcus aureus • Klebsiella pneumoniae • Streptococcus pyogenes Less Common Causes of Viral Pneumonia • Rhinovirus, Enteroviruses, Echovirus, Coxsackievirus, Herpesvirus (EBV, CMV, VZV, HSV), Measles 22 Chlamydia pneumoniae • General Information FYI – You will see Chlamydophila and Chlamydia used interchangeably for C. pneumoniae and C. psittaci (C. trachomatis is always Chlamydia) • Causes sinusitis, pharyngitis, bronchitis, and atypical pneumonia • Obligate intracellular parasite • Cannot make ATP • Considered Gram negative (previously thought no muramic acid present, which is a standard component of peptidoglycan) • Very, very thin cell wall Epidemiology • Most infections are asymptomatic or mild • Does not cause seasonal infection; ubiquitous and worldwide • In U.S.: • Estimated 300,000 cases per year • Accounts for ~15% of community-acquired pneumonias among adults • ~50% of individuals have evidence of past infection by age 20 Risk factors • Smokers • School-aged children • Transmission • Respiratory secretions • Outbreaks among individuals living in close proximity: schools, prisoners, military, long-term care 23 Chlamydia pneumoniae Pathogenesis and Virulence Factors • Two morphological forms: • Elementary body (EB) is infectious • Resistant to harsh environmental conditions • Small: 300-400 nm • Infectious, but cannot replicate • Prevents fusion of phagosome with lysosome • Reticulate body (RB) is metabolically active • Replicates (binary fission) inside vesicles - inclusion bodies • Larger: 800-1,000 nm • Only intracellular • Not infectious • Antibiotics are only effective against the reticulate bodies • Biphasic development cycle • Invades nonciliated columnar, cuboidal, and transitional epithelial cells • EB conversion to RBs and multiply in inclusion bodies • RBs reorganize to EBs (mixed population) • Host cell ruptures and releases infectious EBs Inclusion body EB RB RB EB Citation: Chapter 39 Chlamydia, Ryan KJ. Sherris & Ryan's Medical Microbiology, 8e; 2022. Available at: https://accessmedicine.mhmedical.com/content.aspx?bookid=3107&sectionid=260928805 Accessed: October 22, 2022 Copyright © 2022 McGraw-Hill Education. All rights reserved 24 Chlamydia pneumoniae Pathogenesis and Virulence Factors • Type-three secretion system (molecular needle; injectisome) • Direct secretion of effectors directly into the host cytosol or inclusion membrane • Induce pro-survival signaling • Actin remodeling to support inclusion body • Avoids lysosomal degradation • Host tissue damage is caused by: • The direct destruction of cells during replication • The proinflammatory cytokine response Host Cell Membrane Outer membrane Type III Apparatus Cytoplasmic membrane Type III Toxins 25 Bacterial Secretion Host Cell Membrane Type I Extracellular Type II Type III Type IV Type V Type VI Outer Membrane Type VII Mycolic acid Periplasmic space Plasma Membrane Intracellular Type I Type II Type III Type IV Type V Type VI Type VII Gram Type Negative Negative Negative Negative Negative Negative Positive Process One step: Cytoplasm to outside Two step: Cytoplasm to periplasm to outside One step: Cytoplasm to host cytoplasm Substrates Proteases, lipases, adhesins Cholera Toxin, Exotoxin A Notes ABC transporters Sec/Tat components Two step: cytoplasm One step: periplasm, then One step: Two step: Cytoplasm to periplasm Cytoplasm to host tothe protein itself Cytoplasm to host or and through or bacterial forms a channel in bacterial cytoplasm peptidoglycan/mycolic acid cytoplasm outer membrane Neisseria Yersinia, gonorrohoeae (gene Shigella, acquisition), Pseudomonas Helicobacter toxins effectors Injectisome (protein Injectisome and nucleotides) (unfolded related to protein) conjugation system IgA protease in Neisseria Hydrolase Autotransporter system/ Sec/Tat components Spear gun, Bacterial communication Secretion 26 Chlamydia pneumoniae Clinical Disease • Atypical pneumonia • Usually asymptomatic or flu-like symptoms • X-ray: nonspecific: unilateral alveolar opacities; patchy infiltration, lobar opacification • May be associated with neurological disease (Alzheimer’s), atherosclerosis, asthma, and arthritis • Immunity is short-lived, not lifelong 27 Chlamydia pneumoniae Laboratory Diagnostics • Nucleic acid amplification test (NAAT) • Culture in tissue culture cells, embryonated eggs • Most specific method of diagnosis • Microscopy • Cytoplasmic inclusions seen on Giemsa-, iodine- stains • Gram stain is not useful for identifying this organism Giemsa Arrows denote the cytoplasmic inclusions The Eye. Klatt, Edward C., MD, Robbins and Cotran Atlas of Pathology, Chapter 20, 547-560.e3. Copyright © 2015, 2010, 2006 by Saunders, an imprint of Elsevier Inc. CDC 20874 28 Chlamydia pneumoniae Summary General • • Obligate intracellular parasite (does not make ATP) Considered Gram negative; very thin cell wall Epidemiology • • • • Ubiquitous and worldwide Transmission: respiratory secretion Risk factors: Children, smokers Outbreaks in populations living in close proximity EB RB Disease • Atypical pneumonia • Usually asymptomatic or flu-like symptoms • X-ray: nonspecific: unilateral alveolar opacities; patchy infiltration, lobar opacification • Immunity is short-lived, not lifelong Pathogenesis • Two morphological forms: • Elementary body: resistant to harsh conditions, small, infectious, cannot replicate • Reticulate body: replicates by binary fission inside inclusion bodies, larger, metabolically active • Invades nonciliated epithelial cells •Uses type 3 secretion system to secrete factors into the host cell that induce pro-survival signaling and actin remodeling • Avoids lysosomal degradation Lab Diagnosis • • NAAT Culture in tissue cells, microscopy to visualize cytoplasmic inclusions 29 Causative Agents of Pneumonia Community-Acquired Pneumonia Typical Bacteria • Streptococcus pneumoniae • Haemophilus influenzae • Moraxella catarrhalis • Staphylococcus aureus • Enterobacteriaceae (Klebsiella pneumoniae) Atypical Bacteria • Mycoplasma pneumoniae • Chlamydia (Chlamydophila) pneumoniae • Legionella pneumophila Viral Pneumonia • Respiratory syncytial virus (RSV) • Parainfluenza virus types 1,2, 3 (mostly) • Influenza A virus & B virus • Human metapneumovirus • Adenovirus Hospital-Acquired Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Streptococcus spp. • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Serratia marcescens, Escherichia coli ) Ventilator-Associated Pneumonia • Staphylococcus aureus • Pseudomonas aeruginosa • Gram-negative rods, Enterobacteriaceae (Klebsiella spp., Escherichia coli ) • Acinetobacter baumannii • Stenotrophomonas maltophilia Aspiration Pneumonia • Anaerobic oral flora (Bacteroides, Prevotella, Fusobacterium, Peptostreptococcus), and mixed with aerobic bacteria Zoonotic Pneumonia Bacteria • Bacillus anthracis • Coxiella burnetii • Yersinia pestis • Chlamydia psittaci • Pasteurella multocida • Francisella tularensis Viral • Hantavirus • Influenza • Coronavirus (SARS, MERS) Chronic Pneumonia Bacteria • Nocardia spp. • Actinomyces spp. • Mycobacterium spp. Fungi (also acute depending on exposure load) • Aspergillus spp. • Blastomyces dermatitidis • Coccidioides immitis • Cryptococcus neoformans (HIV/AIDS) • Paracoccidioides brasiliensis • Talaromyces marneffei (HIV/AIDS) • Histoplasma capsulatum Necrotizing Pneumonia and Lung Abscess • Anaerobic bacteria with or without mixed aerobes • Staphylococcus aureus • Klebsiella pneumoniae • Streptococcus pyogenes Less Common Causes of Viral Pneumonia • Rhinovirus, Enteroviruses, Echovirus, Coxsackievirus, Herpesvirus (EBV, CMV, VZV, HSV), Measles 30 Legionella pneumophila General Biology • Aerobic, Gram negative coccobacillus, motile • Facultative intracellular organism • Pneumonia due to environmental exposure • American Legion Convention in Philadelphia, 1976 • 182 ill, 146 hospitalized and 29 deaths • Causes atypical pneumonia • Risk factors: • Smokers, alcoholism • Chronic pulmonary disease • Individuals >40 years old • Immunosuppressed Air conditioners • • • • Water is a reservoir • Survives in free-living amoebae Outbreaks occur in summer/autumn Survives low and high temperatures • 2 minutes at 140°F, dormant under 68°F Transmission • Inhalation of aerosolized contaminated water • 3 Conditions • Heat, stasis, aerosolization • Air conditioners, shower heads, hot tubs • No human-to-human transmission https://www.indiamart.com/proddetail/used-air-conditioner-17965283591.html 31 Legionella pneumophila Pathogenesis and Virulence Factors • Opportunistic pathogen • Flagellated bacteria penetrate the mucus layer (motile) • Attach to cell with pili – invades alveolar macrophage • OMP - binds with C3 which causes opsonization • DOT (direct organelle trafficking) is a T4SS that prevents phagolysosomal fusion and modifies compartment to LCV (Legionella Containing Vacuole) • Toxic enzymes (phospholipases, metalloproteases) damage host cells • CMI - primary host defense mechanism 32 Bacterial Secretion Host Cell Membrane Type I Extracellular Type II Type III Type IV Type V Type VI Outer Membrane Type VII Mycolic acid Periplasmic space Plasma Membrane Intracellular Type I Type II Type III Type IV Type V Type VI Type VII Gram Type Negative Negative Negative Negative Negative Negative Positive Process One step: Cytoplasm to outside Two step: Cytoplasm to periplasm to outside One step: Cytoplasm to host cytoplasm Substrates Proteases, lipases, adhesins Cholera Toxin, Exotoxin A Notes ABC transporters Sec/Tat components Two step: cytoplasm One step: periplasm, then One step: Two step: Cytoplasm to periplasm Cytoplasm to host tothe protein itself Cytoplasm to host or and through or bacterial forms a channel in bacterial cytoplasm peptidoglycan/mycolic acid cytoplasm outer membrane Neisseria Yersinia, gonorrohoeae (gene Shigella, acquisition), Pseudomonas Helicobacter toxins effectors Injectisome (protein Injectisome and nucleotides) (unfolded related to protein) conjugation system IgA protease in Neisseria Hydrolase Autotransporter system/ Sec/Tat components Spear gun, Bacterial communication Secretion 33 Legionella pneumophila Pathogenesis and Virulence Factors • Bacteria induce phagocytosis in alveolar MΦ (OMP binds with C3 which causes opsonization) • Avoid phagolysosome killing (DOT and T4SS) • Endosome does not fuse to the lysosome • LCV forms • Recruits mitochondria, then rough ER • Multiplication occurs • The LCV ruptures to release bacteria LCV http://www.mgc.ac.cn/cgi-bin/VFs/genus.cgi?Genus=Legionella 34 Legionella pneumophila Clinical Disease • Asymptomatic infections common • Pontiac fever • Incubation 1-2 days • Self-limiting febrile influenza-like illness • Believed to be a hypersensitivity to a Legionella toxin • Legionnaire’s disease (legionellosis) • Incubation is 2-10 days • 15% mortality in healthy and 75% mortality in immunocompromised • Acute illness symptoms abrupt • Fever, chills, headache, nonproductive dry cough, diarrhea hyponatremia • Pneumonia • Inflammation and microabscesses in tissue (histology) • X-ray: • Patchy infiltrates that progress to consolidation • Multi-organ involvement is common https://www.cdc.gov/legionella/about/signs-symptoms.html https://emedicine.medscape.com/article/363083-overview 35 Legionella pneumophila Laboratory Diagnostics Gram Stain UpToDate Graphic 61367 Gram Negative Shape BCYE medium • Aerobic Gram negative coccobacillus, motile • Facultative intracellular organism • Isolation by culture (fastidious) (gold standard) • Growth requires L-cysteine • Buffered charcoal yeast extract (BCYE) agar • Charcoal absorbs molecules that prevent growth Coccobacilli • Urine antigen detection CDC 15328 https://www.thermofisher.com/order/catalog/product/R24680 • Direct fluorescence antibody 36 Legionella pneumophila General • • • Summary Aerobic, Gram negative coccobacillus; motile Pathogenesis Opportunist pathogen • Invasion of alveolar macrophage Facultative intracellular organism • OMP - binds with C3 which causes opsonization Epidemiology • DOT (direct organelle trafficking) is a T4SS that prevents • Water is a reservoir phagolysosomal fusion and modifies compartment to LCV • Outbreaks in summer/autumn (Legionella Containing Vacuole) • Survives in low and high temperatures • Toxic enzymes (phospholipases, metalloproteases) damage Transmission host cells • Inhalation of aerosolized contaminated water Disease Treatment: Azithromycin • Heat, stasis, aerosolization • Asymptomatic infections common • Air conditioners, showers, hot tubs • Pontiac fever - incubation 1-2 days; Self-limiting febrile Risk factors influenza-like illness • Smokers, alcoholism, chronic pulmonary • Legionnaire’s disease (legionellosis) - incubation is 2-10 days disease, individuals >40 years old, • 15% mortality in healthy and 75% mortality in immunosuppressed immunocompromised • Acute illness symptoms abrupt - fever, chills, headache, Lab Diagnosis • Aerobic Gram negative coccobacillus, motile nonproductive dry cough, diarrhea, hyponatremia • Grows on BCYE (buffered charcoal yeast extract) • Pneumonia - inflammation and microabscesses in tissue X• L-cysteine required ray: Patchy infiltrates that progress to consolidation • Urine antigen detection • Multiorgan involvement is common • Gram stain requires long safranin stain time or carbol fuchsin as counter stain 37 Lecture Outline Mycoplasma pneumoniae Other Mycoplasmatales Chlamydia (Chlamydophila) pneumoniae Legionella pneumophila 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. Pre-Lecture Clicker Questions Objectives Pneumonia basics General Information Epidemiology Lifecycle Clinical Disease Laboratory Diagnostics Summary Post-Lecture Clicker Questions Learning Catalytics Questions 38 Post-Lecture Clicker Questions Lecture Outline Mycoplasma pneumoniae Other Mycoplasmatales Chlamydia (Chlamydophila) pneumoniae Legionella pneumophila 1. 2. 3. 4. 5. 6. 7. 8. 9. 10. 11. Pre-Lecture Clicker Questions Objectives Pneumonia basics General Information Epidemiology Lifecycle Clinical Disease Laboratory Diagnostics Summary Post-Lecture Clicker Questions Learning Catalytics Questions: 55298239 40 Thank you 41

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