Subject: Microbiology Topic: Bacteria Associated w/ skin Infections 1 Lecturer: Dr.

Eleanor Padla Date of Lecture: July 5, 2011 Transcriptionists: Blue Bolt, Aquamarine, Broken Editor: Blue Blink Pages: 8

STAPHYLOCCOCUS Gram (+) cocci in grape like cluster Facultative anaerobe Some are capsulated Non-motile Non-sporeforming

Antigenic Structures of Staphyloccocus Capsule Slime layer Protein A Peptidoglycan-techoic acid complex

Virulence Factors of Staphylococcus Capsule o Produced by some S. aureus strains o antiphagocytic Slime layer o Produced by S. epidermidis o Involved in adherence, persistence on foreign body Protein A o Surface protein; covalently bound to peptidoglycan o Regularly present in human strains of S. aureus; not found in coagulase (-) staph o Binds non-specifically with Fc portion of IgG Prevents Fc-mediated opsonization Induces complement activation Used in antigen detection (coagglutination) Polysaccharide A (ribitol techoic acid) o Peptidoglycan-techoic acid complex A. Peptidoglycan Provides rigid exoskeleton Has interpeptide bridge connecting oligoglycine peptides S. aureus L-lysineglycine 5-6 S. epidermidis L-lysineglycine 3-5 L-serine

CLASSIFICATION OF STAPHYLOCOCCUS 1. S. aureus 4 biotypes (A-D) o Biotype A most commonly found in humans Involved : a.) in superficial (local) infection b.) deep (systemic) diseases c.) toxigenic diseases 2. S. epidermidis Nosocomial bacteremia Endocarditis associated with valvular prosthesis Intravenous catheter infection Peritonitis in patients undergoing COPD Ventricular shunt infections

SY 2011-2012

 S. saprophyticus L Lysine-glycine 5-6 Lserine B. Techoic Acid Water soluble polymers of ribitol glycerol P04 o Ribitol techoic acid (polysaccharide A) Cell wall techoic acid Found in S. aureus Antiphagocytic o Glycerol techoic acid Cell membranes techoic acid Found in S. epidermidis and saprophyticus Extracellular enzymes o Hyaluronidase Hydrolyzes hyaluronic acid in connective tissues o Lipase Splits fats and oils o Staphylokinase (fibrinolysin) Dissolves fibrin clots o Coagulase (free) Lays down fibrin barrier during abscess formation

Alpha toxins o Forms transmembrane pores o Has lethal, dermonecrotic and leucocidal activities o Formed by majority of human strains of S. aureus o Not formed by coagulase (-) staph

Toxins o Leucocidin Consists of 2 protein components (F=fast; S = slow) Destroys polymorphonuclear leucocytes o Hemolytic toxins 4 major types (alpha, beta, gamma, epsilon) Damage red cell membrane Produce hemolytic zones on BAP

o Exfoliative toxins Proteins Produces intraepidermal splitting of tissues and necrosis seen in SSS; blisters in bullous impetigo Associated with S. aureus phage group II

2 types ETA Gene on Chromosome ETB Gene on plasmid Heatlabile;stable in EDTA

S. epidermidis i. Spread to normally sterile sites as a result of implantation of medical devices ii. Person-to-person spread (e.g in nosocomial setting) C. Diseases caused by S. aureus 1. Superficial (local) infection Impetigo o Honey colored scabs (on arms, legs, face) o 90% is caused by S. aureus, 10% by GAS o Bullous is caused staph while non bullous can be caused by staph or strep o More common in school children; occur in warm, humid environments (coz u see bacteria likes warm, most areas) o Highly contagious; spread by direct contact with lesions or with nasal carriers

o Pyrogenic exotoxins Family of secreted proteins Act as superantigens 1. Entorotoxins a. Seven antigenic types (A, B, C1, C2, D, E F) b. Enterotoxin A and D food posining c. Enterotoxin B necrotizing enterocolitis 2. Toxic Shock Syndrome Toxin-1 (TSST-1) a. Synonymous with Enterotoxin F PATHOGENICITY A. Habitat S. aureus i. Anterior nares, skin, nasopharynx, perineum S. epidermidis i. Skin, mucous membrane, anterior nares S. saprophyticus i. Mucosa of GUT, skin B. Transmission S. aureus i. Spread to normally sterile sites by traumatic ii. Person-to-person spread (e.g. in nosocomial setting)

Folliculitis o Follicular-based pustule (in areas of irritation) o Can occur through shaving, scratching, or with an injury to the skin o Most commonly superficial (affecting upper part of hair follicle; or deep (affecting the whole hair follicle)

3.

Toxigenic Diseases Staphylococcal scaled skin syndrome Staphylococcal food poisoning Toxic shock syndrome

STAPHYLOCOCCAL SCALDED SKIN SYNDROME (SSSS / RITTERS DISEASE) Stripping of superficial layers of the skin from the underlying tissue Mostly frequently involves infants and children < 5 years Majority caused by S. aureus phage group II (mostly commonly, type 71) LABORATORY DIAGNOSIS FURUNCLES (boils) o Starts as hard, tender red nodule surrounding a hair follicle -> abscess -> pus discharged from center -> resolve o Arise in hair- bearing areas where there is friction, occlusion, perspiration o Lesions may be isolated , or multiple CARBUNCLES o Start as smooth, domeshaped, acutely tender, painful lesion -> develop into swollen, painful area discharging pus from several sites (individual boils clustered together) o Often occur at the nape of the neck, the back or thighs o Deeper, more severe, develop and heal more slowly 2. Deep (Systemic) Diseases Acute osteomyelitis Acute endocartitis Pneumonia
4

1. GRAM STAIN Gram (+) cocci in clusters 0.5- 1.5 micrometer Gram-variable within neutrophils, in resolving lesions, in the presence of antibiotics; in old cultures 2. CULTURE Grows well on unenriched media Grows in 7.5% - 10% NaCl; in 40% bile Media : BAP, selective media (MSA, PEA)

Figure 1. Media: Mannitol Salt Agar

Figure 2. Colonies on blood Agar Plate

 Form smooth, raised colonies; mucoid if capsulate Colonial pigment variable o s. aureus golden yellow to white o s. saprophyticus white to pale grey o s. epidermis white Hemolytic reaction variable o S. aureus- usually beta-hemolytic o S. epidermis and s. saprophyticus usually nonhemolytic

Control and Prevention: Suppurative Infections: o Cleanliness, hygiene, and meticulous handwashing o Aseptic management of lesions o Isolation of persons with open lesions o Suppression/cure of nasal carriage

STREPTOCOCCI General Properties: Gram (+) cocci in chains Facultative anaerobe Some are encapsulated Some are flagellated Non-sporeforming

3. DIFFERENTIAL TESTS A. Catalase Test Staphylococci (+) Streptococci (-) B. Coagulase Test Slide Test (bound coagulase / clumping factor) Produced exclusively by S. aureus forming free coagulase Tube Test (free coagulase) Clotting of citrated plasma in the presence of coagulase reacting factor (CFR) For suspected S. aureus that fail to produce bound coagulase 4. Susceptibility Aids in choice of systemic drugs Epidemiologic tool 5. Serology (antigen detection) Phage Typing For epidemiologic tracing of infection Depends on differing susceptibilities to lysis by phages

Classification: A. Hemolytic Patterns Alpha hemolytic Incomplete hemolysis Greening reaction

Beta hemolytic Complete hemolysis Enhanced by AnO2 conditions Gamma hemolytic No hemolysis

B. Lancefield System Based on group-specific CHO antigen (C-CHO)

Amino sugar determines serologic specificity GAS = rhamnose-Nacetylglucosamine GBS = rhamnose-glucosamine GDS = glycerol teichoic acid Streptococci groups: A-H, K-V Human pathogens belong to Grps A to G Group A - possess Lancefield group B antigen - synonymous with S. pyogenes Group B - possess Lancefield group B antigen - Synonymous with S. agalactiae Group D - possess Lancefield group D antigen - includes Enterococcus spp. Viridans streptococci - lacks group-specific cell wall antigen - referred to as oral streptococci Streptococcus pneumonia - lacks group-specific cell wall antigen - referred to as pneumococcus -

Group A streptococci (GAS) Pharyngitis (sore throat/tonsillitis) Impetigo (pyoderma) Erysipelas Cellulitis Necrotizing fascilitis Scarlet fever Puerperal fever (childbed fever) TSSL (toxic shock syndrome-like) Acute bacterial endocarditis Non-suppurativeSequelae acute rheumatic fever acute glomerulonephritis

Group B Streptococci (GBS) bacteremia, pneumonia, meningitis in newborns Group D Streptococci (GDS) bacteremia, endocarditis, nosocomial infections

Viridans Streptococci dental plaque/carries subacute bacterial endocarditis

Streptococcus Pneumoniae
Figure 3. Lancefield Test

lobar pneumonia meningitis otitis media

GROUP A STREPTOCOCCI Group Characteristics: possess the Lancefield group A antigen rhamnose-N-acetylglucosamine

C. Biochemical Reactions For untypable species Used to speciateviridans streptococci

synonymous with S. pyogenes Antigenic Structures: capsule pili protein antigens (M,F,G,T proteins) C5a peptidase Group-specific polysaccharides (CCHO)

Figure 4. Biochemical reaction

Clinical Diseases

Virulence Factors: 1. Capsule - composed of hyaluronic acid - not immunogenic - antiphagocytic 2. Lipoteichoic acid - component of pili - together with M protein, mediates oral/pharyngeal, skin epithelial attachment 3. M protein - major virulence factor of GAS - component of pili - anti-phagocytic (prevents alternative Cpathwayopsonization) - dominant in binding to epidermis - strongly immunogenic - >80 antigenic types 4. F protein - fibronectin-binding protein - together with M protein, mediates nasopharyngeal adherence 5. G protein - Fc receptor (prevents Fcmediated opsonization) 6. C5a peptidase - surface-bound endopeptidase - anti-polymorphonuclear leucocyte chemoattractant (cleaves C5a) 7. Serum opacity factor - an alpha-lipoproteinase - produce opalescence in horse serum broth - adjunct to M typing 8. Pyrogenic exotoxins - protein in nature - 4 antigenic types (A-D) EXOTOXIN A Associated with rash in scaret fever; TSS-L A superantigen Produced only by lysogenized strains Basis of Dick/ SchultzCharlton tests

EXOTOXIN B Responsible for tissue destruction in necrotizing fascilitis 9. Hemolysins - responsible for beta-hemolysis on BAP - cytotoxic - 2 distinct types of hemolysins a. Streptolysin O - oxygen-labile - hemolytic in reduced form; responsible for subsurface hemolysis - formstransmembrane pores - strongly immunogenic b. Streptolysin S serum soluble, oxygen-stable responsible for surface hemolysis non-immunogenic

10. Spreading factors Hyaluronidase - splits hyaluronic acids in connective tissue Streptokinase (fibrinolysin) dissolves fibrin clots DNAse (streptodornase) depolymerizes viscous DNA in pus strongly immunogenic

Pathogenicity A. Habitat Inhabits URT, skin of humans

B. Transmission Persons to person by direct contact with mucosa or secretions Fomites C. Clinical Diseases Pharyngitis Impetigo (pyoderma) - non-bullous Erypselas (St. Anthonys fire)
7

peripherally spreading, hot, bright-red, edematous, wellcircumscribed, sharply marginated lesions most often affects the face, legs; spreads thru the lymphatic vessels common in infants, children, the elderly accompanied by severe constitutional symptoms Cellulitis Necrotizing fascilitis Scarlet fever Acute bacterial endocarditis Puerperal fever TSSL (toxic shock syndrome-like)

3. Identification 4. Antigen Detection Based on monoclonal antibodies which react with C-CHO (e.g. latex agglutination, coagglutination, ELISA) 5. Serology Anti DNAse B Determination To demonstrate previous or recent streptococcal infection Significant response in patients with pyoderma Prevention and Control: Early detection and antimicrobial therapy DOC: penicillin treating carriers
End of transcription
Do not be anxious about anything, but in every situation, by prayer and petition, with thanksgiving, present your requests to God. And the peace of God, which transcends all understanding, will guard your hearts and your minds in Christ Jesus. - Philippians 4:6-7 To God be the glory!

Necrotizing fasciitis (flesh- eating bacteria; streptococcal gangrene) toxin-mediated, highly destructive and potentially lethal infection of soft tissued (fatty tissues, fascia, muscles) always follows trauma Scarlet fever (scarlatina) Scarlet-colored (boiled-lobster) body rashes (sandpaper-like); fever; strawberry-like appearance of tongue Exotoxin mediated Evolves from tonsillar / pharyngeal focus Laboratory Diagnosis: 1. Gram stain

2. Culture Media BAP chocolate agar selective media (CAN, PEA) greyish white, transparent to translucent matt or glossy beta hemolytic