1. Staph/Strep Peter Coschigano, Ph.D. Department of Biomedical Sciences OUCOM

2. Outline Cell wall review Staphylococcal infections and virulence mechanisms Streptococcal infections and virulence mechanisms Other bacterial agents of pneumonia

3. Membrane Structure: http://www.ppws.vt.edu/~sforza/prokaryote/cellwalls_compare.jpg Gram-negativeGram-positive Inner Membrane Outer Membrane Inner Membrane Peptido- glycan Peptidoglycan Periplasm Lipopolysaccharide (LPS) Teichoic acid

4. Staphylococcus –aureus –epidermidis –saprophyticus

5. Stained Staphylococcus aureus in tissues. [clumped, grape-like clusters of cocci] 10 µm

6. Characteristics of Staphylococcus aureus Most common cause of human staphylococcal illness Ubiquitous in nature, normal carriage in 20-40% of people on skin, in anterior nares (nose), in vagina Resistant to heat & desiccation  -hemolytic Can be spread by contact with inanimate objects (fomites) Illnesses –Skin Infections –Lung (pneumonia), blood (septicemia), systemic infection –Toxin-mediated diseases: food poisoning, toxic shock syndrome, scalded skin syndrome

7. S. aureus -pathogenesis Host compromise needed for infection (break in skin, foreign body insertion, obstructed hair follicle)

8. S. aureus- skin infections Superficial abscesses (hair follicles, sweat glands, sebaceous glands) Subcutaneous abscess (furuncles and carbuncles = boils) Impetigo (localized, superficial, spreading, crusty lesion) Cellulitis (diffuse, spreading, acute inflammation within solid tissues without necrosis or suppuration).

9. S. aureus (other infections) Septicemia (blood infection) Pneumonia (lung infection) Nosocomial (hospital acquired) infections –wound infections [surgical for ex] –catheter derived infections.

10. S. aureus diseases caused by secreted exotoxins Toxic shock syndrome (high fever,rash, shock) due to staphylococcal toxic shock toxin Food poisoning (gastroenteritis from staphylococcal enterotoxin-contaminated food) Scalded Skin syndrome (superficial blisters and skin loss due to staphylococcal exfoliative toxin)

11. S. aureus -virulence factors Factors that inhibit host immunity –Polysaccharide Capsule: outer bacterial slime layer that inhibits phagocytosis –Protein A: bacterial cell wall protein inhibits host antibody function Adhesive factors –Fibronectin binding protein: promotes bacterial attachment to host tissues Toxins (exotoxins) –Cytolytic exotoxins (hemolysins) damage immune cells & host tissues. –toxic shock toxin, exfoliative toxin

12. Streptococcus –pyogenes –pneumonia –agalactiae

13. Characteristics of Streptococcus pyogenes ( Group A  -hemolytic Streptococci ) Most common cause of bacterial sore throat (pharyngitis or “strep throat”) and tonsillitis, –10-30% of sore throat cases Common cause of skin infections –Impetigo, erysipelas, cellulitis, necrotizing fasciitis (rapid tissue death characteristic of “flesh-eating” bacteria) and Streptococcal toxic shock syndrome Spread person-to-person via respiratory droplets or skin contact. Carriers exist.

14. Gram-positive (blue) cocci in chains (streptococci). Diameter of a coccus = ~ 1 micron

15. Complications of streptococcal pharyngitis “Scarlet fever” = skin rash on neck, trunk, extremities caused by streptococcal pyrogenic exotoxin. (also called erythrogenic toxin) Infection sequelae (post-infection) –Acute rheumatic fever (inflammatory heart injury) –Acute poststreptococcal glomerulonephritis (inflammatory kidney injury)

16. Multiple episodes of “strep throat” are possible Infection by Group A streptococci leads to long term immunity to re-infection by the same bacterial strain, but there are >80 serotypes (M proteins) of this bacterium, each of which can cause separate episodes of streptococcal sore throat.

17. Strept. pyogenes virulence factors Factors that inhibit host immunity –M proteins and Hyaluronic acid capsule Adhesive/invasive factors –lipoteichoic acid, protein F1, M protein, and capsules –Enzymes (DNAse, hyaluronidase) Toxins (exotoxins) –Cytolytic exotoxins (hemolysins), SLO= streptolysin O –pyrogenic exotoxins, (erythrogenic toxin)

18. Lab ID of Staph vs. Strep Staph. aureus –Gram(+) coccal morphology –Catalase(+), Coagulase(+) –  -hemolytic –Multiple strains (groups I-IV) Strept. pyogenes –Gram(+) coccal morphology –Catalase(-), coagulase(-) –  -hemolytic –Multiple serotypes (80 M protein types)

19. Intact red blood cells in agar [lysed rbc zone around colonies of bacterial growth]

20. Slide test for catalase activity to differentiate Staphylococci from Streptococci Mix a drop of 3% hydrogen peroxide with cells on a glass slide. Vigorous bubbles indicates catalase activity.

21. Treatment of Staph. And Strep Infections Staph. aureus –Antibiotics, but multiple antibiotic resistance (Penicillin resistance, methicillin- resistance-MRSA), –Incision & drainage of lesions –Hygienic control –No commercial vaccine available Strep. pyogenes –Penicillin –No commercial vaccine available

22. Streptococcus pneumoniae Gram positive cocci (pair) Encapulated, complex carbohydrate, ~90 serotypes. Most commom (23) polysaccharides used in polyvalent vaccine. Normal flora. Infections can come from self or other.  -hemolytic.

23.  -hemolysis

24. Hemolysis Pattern

25. Streptococcus pneumoniae virulence factors Capsule (variable) Surface protein adhesins Pneumolysin –Binds cholesterol in CM and creates pores –Damages ciliated epithelial cells. Teichoic acid (& other CW components) Hydrogen peroxide (reactive oxygen)

26. Other bacterial agents of pneumonia

27. Membrane Structure: http://www.ppws.vt.edu/~sforza/prokaryote/cellwalls_compare.jpg Gram-negativeGram-positive Inner Membrane Outer Membrane Inner Membrane Peptido- glycan Peptidoglycan Periplasm Lipopolysaccharide (LPS) Teichoic acid

28. Klebsiella pneumoniae Gram neg. rod. Prominent capsule. Community acquired primary lobar pneumonia. –Alcohol abuse/ compromised pulmonary function can increase risk. Necrotic destruction, cavity formation, blood tinged sputum (“currant jelly sputum”).

29. Gram-negative rods

30. Legionella pneumophila Pleomorphic Gram neg. rod. Facultative intracellular parasite Penetrate cells by endocytosis. Phagolysosomal fusion inhibited. Proliferate in intracellular vacuole. Produce NZ which kill host when vacuole lyses. No person-to-person spread. Legionnaire’s disease-multi organ (CNS, GI, liver, kidneys) pneumonia with multilobar consolidation, inflammation and microabscess.

31. Life Cycle of L. pneumophila in macrophages Bacterial Pathogenesis, 2nd ed., 2002, Salyers and Whitt, ASM Press

32. Mycoplasma pneumoniae Smallest free-living bacteria (asymmetrical). No peptidoglycan cell wall-resistant to antibiotics that interfere with CW biosynthesis. Sterol in membrane. Slow growing (1-6 hr generation time). Extracellular pathogen. Adhere to respiratory epithelium via cytadherence. Destroy cilia and ciliated epithelial cells. Spread by nasal secretion (close contact necessary). Primary atypical (“walking”) pneumonia (age 6-20).

33. Mechanisms to avoid host defenses Multiple serotypes Capsule Replicate inside host cells Atypical “cell wall” Other –Adhesion –Cause cell damage