Micro TA Review 1April 4, 2013

• Tips & hints• Memorization… this is the time!

• Flashcards, stories, whatever it takes.• You really do need to know this.

• Pathogen clinical presentation & vice versa• Questions: know all the answers• It goes fast – try to keep up as best you can!

• Resources• Clinical Micro Made Ridiculously Simple• Lippincott Micro Cards• TA PPTs• Firecracker

Bacterial Cell Walls

TRANSFORMATION:Ability to take up naked DNA of RELATED bacteria from environment

(especially S. pneumo, H. influenzae, and Neisseria -- SHiN)

TRANSPOSITION:Segments of DNA that can “jump” from one chromosomal location to

another via excision and reincorporation. Can also jump from chromosome to plasmid and vice versa.

TRANSDUCTION:Bacteriophage (virus that infects bacteria) transfers bacterial DNA from one bacteria to another.Can be generalized (lytic phage) or specialized (lysogenic phage).

Bacterial Genetics

Bacterial Genetics

CONJUGATION:“Bacterial Sex”– may involve UNRELATED bacteria, requires direct contact and transfer is UNIDIRECTIONAL. Two options:

1. F+ x F-: F plasmid contains genes for conjugation and it is transferred via pilus (no transfer of chromosomal genes).

2. Hfr x F-: F plasmid can become incorporated into DNA -- now called an Hfr cell and now may transfer plasmid + some flanking chromosomal genes.

Normal Bacterial Flora• Skin:– Normal: Staphylococcus epidermidis– Transient: S. aureus

• Upper Respiratory tract– Normal: Anaerobic cocci, Viridans Streptococci– Transient: Neisseria spp., Streptococcus pneumoniae, Haemophilus influenzae

• Gastrointestinal Tract– Anaerobic GNR (Bacteroides), Enterobacteriaceae (E. coli), Enterococci

• Genitourinary tract– Normal: Lactobacillus– Transient: GBS

• Ability to block arms of immune system (phagocytosis, antibodies, complement) (Staph aureus- Protein A; Strep pyogenes- M protein)

• Slime and biofilms (Staph epi, Strep viridans)

• Toxins (endotoxin vs. exotoxin) (Endotoxin: gram negatives; Exotoxin: Staph aureus, Strep pyogenes)

• Adhesion (Staph sapro)

• Flagella• Ability to survive phagocytosis (Strep pyogenes, hyaluronic acid capsule)

• Ability to survive in hostile environments

Virulence Factors

Exotoxin v. Endotoxin

Superantigens

Definition:• Superantigen toxins allow the non-specific binding of

MHC II with T cell receptors, resulting in polyclonal T cell activation (versus other toxins, which are just monoclonal activation so not as intense).

• Polyclonal T-cell population cytokine storm multisystem disease and shock.

Examples:• S. aureus: TSST-1• S. pyogenes: Exotoxin A

Antibiotic Targets

Antibiotic ClassesCell Wall Synthesis Inhibitors: Peptidoglycan Cross-LinkingPenicillins, Cephalosporins, Carbapenems (Beta-Lactams)• Mechanism: bind to transpeptidases (aka PBP) → inhibit cross-linking of

peptidoglycan no cell wall death by osmolysis• Resistance via beta lactamases, altered penicillin binding protein, altered porins.

Cell Wall Synthesis Inhibitors: Peptidoglycan SynthesisVancomycin• Mechanism: binds D-ala D-ala portion of cell wall precursors inhibits

peptidoglycan synthesis.• Resistance via amino acid change from D-ala D-ala to D-ala D-lac; gram-

negatives are intrinsically resistant because vanco can’t cross outer membrane.

Protein Synthesis Inhibitors Macrolides, Tetracyclines, Aminoglycosides

Inhibitors of DNA Polymerase ActivityFluoroquinolones, Metronidazole

Folic Acid Synthesis InhibitorsSulfonamides and Trimethoprim (TMP/SMX)

OVERVIEW!Do not be overwhelmed.

this week’s focus: gram+ cocci

Gram Positive Cocci

Also see the table in your lab manual!

Types of Hemolysis

Also: Staph aureus, GBS

Also: Strep viridans

Enterococci have variable hemolysis

IN THE LAB• Gram + cocci in clusters• Coagulase +, catalase +• Beta-hemolytic

EPIDEMIOLOGY• Nosocomial and community, all ages

CLINICAL• Pyogenic skin infxn (impetigo, cellulitis, abscesses)• Acute endocarditis (v. S. viridans subacute)• Osteomyelitis• Septic arthritis• Toxin-related syndromes (see below)

VIRULENCE FACTORS• Protein A, biofilm, capsule, coagulase, catalase, hemolysins, penicillinase• Toxins: TSST-1 (superantigen, causes Toxic Shock Syndrome); enterotoxin (preformed

superantigen, causes gastroenteritis); exfoliative toxin (causes scalded skin syndrome)

TREATMENT• Most strains secrete penicillinase MSSA, treat with nafcillin• Some strains alter the PBP MRSA, treat with vancomycin

Staphylococcus aureus

impetigoscalded skin

cluster of grapes

IN THE LAB• Gram + cocci in clusters• Coagulase - (vs S. aureus), catalase +, novobiocin sensitive (vs S. sapro)• Gamma-hemolytic

EPIDEMIOLOGY• Nl. flora of the skin; nosocomial & iatrogenic

CLINICAL• Foreign body or device infxns (think prosthetics, IV lines, Foley catheters)• Common contaminant in blood cultures

VIRULENCE FACTORS• Biofilm: extracellular scaffold that mediates attachment to foreign devices

TREATMENT• Vancomycin & remove the infectious source (i.e., remove the device)

Staphylococcus epidermidis

Staphylococcus saprophyticusIN THE LAB• Gram + cocci in clusters• Coagulase - (vs S. aureus), catalase +, novobiocin resistant (vs S. epi)• Gamma-hemolytic

EPIDEMIOLOGY• Sexually active women 2nd most common cause of UTI (1st is E. coli)

CLINICAL• UTI (burning, urgency, frequency)

VIRULENCE FACTORS• Specialized mucosal receptors

TREATMENT• Folic acid synthesis inhibitors (TMP/SMX)

IN THE LAB• Gram + cocci in chains• Catalase – (vs Staphs), bacitracin sensitive (vs GBS)• Beta-hemolytic

EPIDEMIOLOGY• All ages

CLINICAL• Suppurative (bc of host neutrophil response): pharyngitis (“strep throat”), cellulitis (inc.

erysipelas), impetigo, necrotizing fasciitis• Non-suppurative (toxigenic): scarlet fever, Streptococcal Toxic Shock Syndrome• Delayed (immunologic): rheumatic fever, poststreptococcal glomerulonephritis

VIRULENCE FACTORS• M protein, superantigen, streptolysin S & O, streptokinase, SpeA, B, C, anti-C5a peptidase,

hyaluronidase (in capsule), T antigen, DNAase• Diagnose recent infxn with ASO titers (antistreptolysin O)

TREATMENT• Penicillin

Streptococcus pyogenes (GAS)

strep throat w/oropharyngeal petechiae

Streptococcus agalactiae

Group B is Bad for Babies!

Streptococcus agalactiae (GBS)IN THE LAB• Gram + cocci in chains• Catalase – (vs Staphs), bacitracin resistant (vs S. pyogenes)• Beta-hemolytic

EPIDEMIOLOGY• Neonates, peripartum women, diabetics

CLINICAL• Neonatal sepsis, pneumonia, meningitis• Maternal sepsis• Less commonly: UTI, endocarditis

VIRULENCE FACTORS• Not important

TREATMENT/PROPHYLAXIS• Beta-lactams (penicillin)

IN THE LAB• Gram + cocci in pairs or chains• Catalase – (vs Staphs)• Variable hemolysis (alpha or gamma)• Subdivided into Lancefield Group D and non-Lancefield Group D depending on carbs in

cell wall

EPIDEMIOLOGY• Nl GI flora; nosocomial infxn (esp VRE)

CLINICAL• UTI, subacute endocarditis, biliary tract infxn• Peritonitis• Nosociomal superinfxn bacteremia

VIRULENCE FACTORS• Dextran (glycocalyx) helps bind to heart valves

TREATMENT• Inherent antibiotic resistance to cephalosporins• Now have vancomycin resistance as well (VRE) treat with daptomycin or ampicillin +

aminoglycoside

EnterococcusEnterococcus (E. faecalis, E. faecium)

Strep viridans (S. mutans, S. sanguis)

foliage growing on mitral valve from oral cavity

IN THE LAB• Gram + cocci in chains• Catalase – (vs Staphs), optochin resistant (versus S. pneumo)• Alpha-hemolytic

EPIDEMIOLOGY• Nl oropharynx flora

CLINICAL• S. mutans dental caries (cavities)• S. sanguis subacute bacterial endocarditis (heart valve damage)

VIRULENCE FACTORS• Dextran (glycocalyx) helps bind to heart valves

TREATMENT• Penicillin (usually sensitive to lotsa antibiotics)

Streptococcus pneumoniaeIN THE LAB• Gram + lancet-shaped cocci in pairs (“diplococci”)• Catalase – (vs Staphs), optochin sensitive (versus S. viridans)• Alpha-hemolytic

EPIDEMIOLOGY• See later lectures

CLINICAL• Most common cause of MOPS: bacterial Meningitis in kids & elderly, Otitis media in kids,

community-acquired Pneumonia, Sinusitis.• Also, sepsis.

VIRULENCE FACTORS• Polysaccharide capsule prevents phagocytosis.

• Diagnose with Quellung reaction (capsular “swellung”)

TREATMENT• Penicillin• Some resistance high dose penicillin & cephalosporins• Lotsa resistance vancomycin