Microbiology I Step I Review

1. Microbiology IStep I Review Devin Gillaspie

2. First the Basics… • Gram Positive Organisms • Stain purple/blue • Cell wall + cell membrane • Contains techoic acid – induces TNF and IL-1 • Gram Negative Organisms • Stain red • Outer membrane + Periplasmic space + cytoplasmic membrane • LPS in outer membrane

4. Who doesn’t Gram stain? • Treponema – too thin; use dark field microscopy or fluorescent Ab staining • Rickettsia, Legionellaand Chlamydia – intracellular. (Silver stain for Legionella) • Mycobacteria – high lipid content, use acid fast stain • Mycoplasma – no cell wall

5. Stains • Giemsa Stain – Borrelia, Plasmodium, trypanosomes, Chlamydia • PAS – Stains glycogen, mucopolysaccharides; to diagnose Whipple’s disease • Ziehl-Neelsen – Acid Fast organisms • India Ink – Cryptococcus neoformans (thick polysaccharide capsule) • Silver stain – Fungi and Legionella

6. Special Culture Media • H. influenza – Chocolate agar with factor V and X • N. gonorrhoea – Thayer-Martin media • B. pertussis – Bordet-Gengou • C. diphtheriae – Telluritepalte, Loffler’s media • M. pneumoniae – Eaton’s agar • Lactose fermenters – MacConkey’s agar (pink colonies); E. coli turns black-blue on EMB agar • Legionella– Charcoal yeast extract agar with cystein • Fungi – Sabouraud’s agar

7. Obligate Aerobes • Use oxygen dependent system to make ATP • Examples: • Nocardia • Pseudamonasaeruginosa • Mycobacterium tuberculosis (reactivation TB usually in apices of lungs which have highest PO2) • Bacillus

8. Obligate anaerobes • Lack catalase susceptible to superoxide dismutase • Foul smelling, difficult to culture, produce gas in tissue • Examples: • Clostridium • Bacteroides (GI tract normal flora) • Actinomyces

9. Intracellular Organisms • Obligate intracellular • Chalmydia, Rickettsia • Facultative intracellular • Salmonella, Neisseria, Brucella, Mycobacterium, Listeria, Fracncisella, Legionella

10. Encapsulated Bacteria • Capsules serve as anti-phagocytic virulence factor • Positive Quellung reaction – capsule swells when specific anticapsularantisera is added • Examples: • Klebsiellapneumoniae • Salmonella • Strep. pneumoniae • H. influenza type B • Neisseriameningitidis

11. Others • Urease positive bugs • Proteus, Klebsiella, H.pylori, Ureaplasma • Pigment producing bacteria • Actinomycesisralii– yellow “sulfur” granules • S. aureus– yellow pigment • Pseudamonasaeruginosa– blue-green pigment • Serratiamarcescens– red pigment

12. Bacterial Virulence Factors • Protein A – Binds Fc region of Ig, preventing opsonization and phagocytosis (S. aureus) • IgA protease – cleaves IgA, to colonize respiratory mucosa (S. penumoniae, H. flu type B, and Neisseria) • M protein – Prevents phagocytosis

13. Spores • Certain gram positive rods • Form at end of stationary phase – no metabolic activity in spore form • Highly resistant to destruction by heat and chemicals – autoclave required to kill spores

14. EndotoxinsvsExotoxins

15. Exotoxins and you • Super antigens • S. aureus– TSST-1 causes toxic shock syndrome • S. pyogenes – Scarlet fever-erythrogenic toxin • A-B toxins • C. diphtheriae – inactivates EF2; causespseudomembrane in throat • Vibriocholerae – inactivates Gs protein, stimulating adenylylcyclase and increase pumping Cl into gut and decreased Na absorption causing rice water diarrhea • E. coli (ETEC) – Heat-labile toxin (stimulates adenylatecyclase); Heat-stable toxin (stimulates Guanylatecyclase)  watery diarrhea • B. pertussis – increases cAMP by inhibiting Gαi whooping cough; inhibits chemokine receptor lymphocytosis

17. Other exotoxins • Clostridium perfringens – α toxin  gas gangrene • C. tetani – blocks release of inhibitory neurotransmitters GABA and glycine lockjaw • C. botulinum – Blocks release of Ach, causes anticholninergic symptoms, CNS paralysis • Bacillus anthracis – Edema factor, and adenylatecyclase • Shigella – Shiga toxin cleaves host cell rRNA (inactivates 60S ribosome), and enhances cytokine release  HUS • S. pyogenes – Streptolysin O, a hemolysin.

19. Bacterial Growth Curve

20. Bacterial Genetics • Transformation • Take up DNA from environment • Especially encapsulated bacteria • Conjugation • F+ plasmid contains genes required for conjugation (bacteria w/o plasmid are F-) • Plasmid is replicated and transferred through pilus to F- bacteria (no chromosomal DNA) • If F+ plasmid is incorporated into chromosomal DNA = Hfr • Replication of plasmid DNA may include some flanking genes

21. Bacterial Genetics • Transduction • Generalized – LyticBacteriophage infects bacteria  cleavage of host DNA  production of viral proteins  some bacterial DNA may be packaged in viral capsid • Specialized – Lysogenic phage infects bacteria  viral DNA incorporated into bacterial chromosome  phage DNA excised with flanking genes • Transposition • Segment of DNA that “jumps” from one location to another • Plasmid to chromosome and vice versa

23. GRAM POSITIVES Clostridium Bacillus Corynebacterium Listeria Enterococcus Peptostrep S. viridans S. pneumo S. pyogenes(GAS) S. agalactiae(GBS) S. aureus S. epidermidis S. saprophyticus

24. Staphylococcus sp. – Staph aureus • Causes: • Inflammatory disease – skin infections, organ abscesses, pneumonia, etc. • Toxin-mediated disease – toxic shock syndrome (TSST-1), scalded skin syndrome (exfoliative toxin), rapid-onset food poisoning (enterotoxins) • MRSA – nosocomial and community acquired infections. Resistant to β-lactams due to altered penicillin binding protein • Vireulence factors

26. Staphylococcus sp. – Staph epidermidis • Infects prosthetic devices and IV catheters • Biofilm • Normal skin flora – contaminates blood cultures • Endocarditis in IVDA

27. Streptococcus sp. • Streptococcus pneumoniae– encapsulated, IgA protease • Most common causes of: • Meningitis • Otitis media (in kids) • Pneumonia • Sinusitis • Viridans group strep - α-hemolytic • Normal flora of oropharynx – dental carries • Subacute bacterial endocarditis

28. Streptococcus sp. • Streptococcus pyogenes(GAS) • Causes • Pyogenic – pharyngitis, cellulitis, impetigo • Toxigenic – scarlet fever, toxic shock syndrome • Immunologic – rheumatic fever, acute glomerulonephritis • ASO titer detects recent S. pyogenesinfection • Streptococcus agalactiae (GBS) • β hemolytic, bacitracin resistance • Causes – pneumonia, meningitis, and sepsis mainly in babies

30. Group D Strep • Enteroccocus • E. faecalisand E. faecium • Normal colonic flora • Penicillin G resistant • Cause UTIs • VRE - important cause of nosocomial infections • Streptococcus bovis • Colonizes gut • Causes – bacteremia and subacuteendocarditis • Colonizes 15% of colon cancer patients

31. Diphtheria • Corynebacteriumdiphtheriae • Gram positive Rod • Exotoxin inhibits EF-2 • Pseudomembranouspharyngitis with lymphadenopathy • Lab Dx – Gram - rods with metachromatic granules • Toxoid vaccine

33. Clostridium sp. • Gram positive rods, spore forming, obligate anaerobes • C. tetani • Tetanospamin (tetanus exotoxin)  tetanus, spastic paralysis • C. botulinum • Heat-labile toxin inhibits ACh release at neuromuscular junction  botulism • In adults  ingestion of preformed toxin • In babies  ingestion of spores (floppy baby syndrome)

34. Clostridium sp. • C. perfringens • α-toxin myonecrosis (gas gangrene) and hemolysis • C. difficile • Cytotoxin kills enterocytespseudomembranous colitis • Often 2° to antibiotic use especially clindamycin or ampicillin • Dx: check for C. diff toxin in stool • Tx: metronidazole

36. Anthrax • Bacillus anthracis • Spore forming, gram positive rod, produces anthrax toxin • Capsule contains D-glutamate • Cutaneous anthrax – contact, black eschar formation; can progress to bacteremia, death • Pulmonary anthrax – inhalation of spores; flu-like symptoms rapidly progress to fever, pulmonary hemorrhage, mediastinitis, and shock.

38. Listeriamonocytogenes • Gram positive rod, facultative intracellular • “actin rockets” to move cell to cell • Tumbling motility • The only Gram + with endotoxin • Ingestion of unpasteurized milk/cheese and deli meats (avoid in pregnancy) or by vaginal transmission • Dz: amnionitis, septicemia, spontaneous abortion, granulomatosisinfantiseptica; neonatal meningitis; meningitis in immune compromised; mildgastroenteritis in healthy individuals

40. Actinomyces vs. Nocardia • Gram positive rods in long branching filaments resembling fungi • Actinomycesisraelii • Anaerobe, normal oral flora • Oral/facial abscesses • Drain through sinus tracts in skin – yellow sulfur granules • Nocardiaasteroides • Weakly acid fast aerobes • In soil • Pulmonary infection in immunocompromised patients

42. TB • Mycobacterium tuberculosis • Primary TB – (non-immune host) • Ghon complex = Ghon focus (calcified scar in lower lobe) + lobar and perihilar node involvement • Healing with fibrosis (+ PPD), OR • Progressive lung disease (in HIV, etc) rarely death, OR • Severe bacteremia (miliary TB)  Death, OR • Pre-allergic lymphatic or hematogenous dissemination  dormant TB in organs  reactivation TB • Secondary TB – Reactivation TB • Fibrocaseouscavitary lesion in upper lobes • Extrapulmonary TB • CNS, Vertebral body (Potts disease), lymphadenitis, Renal, GI

44. Leprosy • Mycobacterium leprae • Acid-fast bacillus • Likes cool temps – infects skin and superficial nerves • US reservoir in armadillos • 2 forms of disease • Lepromatous – leonine facies = loss of eyebrows, nasal collapse, lumpy earlobes • Tuberculous – diffusely over skin, is communicable; lose sensation in fingers and toes (spontaneous amputations) • Treatment – long term oral dapsone; or rifampin + clofazimine + dapsone • 6 months to 2 years of antibiotics

46. Other Mycobacteria • M. kansasii • Pulmonary TB-like symptoms • M. avium-intracellulare • Often multidrug resistant • Disseminated disease in AIDS

47. Gram Negatives H. flu Pasturella Brucella Bordatellapertussis Neisseria Moraxella N. gonorrhoeae N. meningitidis Shigella Salmonella Proteus Pseudamonas Citrobacter Serratia Klebsiella E. Coli Enterobacter

48. Neisseria sp. • Gram negative cocci – diplococci • Gonococci • No capsule, maltose non-fermenter, no vaccine, STD • Causes gonorrhea, septic arthritis, neonatal conjunctivitis (give erythromycin eye drops at birth), PID, and Fitz-Hugh-Curtis Syndrome • Meningococci • Capsule, maltose fermenter, vaccine, spread by respiratory and oral secretions • Causes meningococcemia and meningitis, Waterhouse Friderichsen syndrome

49. Legionellapneumophila • Gram negative rod • Silver stain • Grows on charcoal yeast extract w/ cystein • Dx: antigens in urine • Transmission: aerosol from water source • Legionnaire’s disease = severe pneumonia and fever • Pontiac fever = mild flu-like syndrome • Treatment - erythromycin

50. Pseudomonas aeruginosa • Gram – rod, aerobic, blue-green pigment, grapelike odor • Associated with wound and burn infections • Also causes: • Pneumonia (CF) • Sepsis • External otitis (swimmer’s ear) • UTI • Drug use (endocarditis) • Diabetic osteomyelitis • Hot tub folliculitis • Malignant otitisexterna (in DM) • Tx: Aminoglycoside + extended spectrum penicillin