Staphylococcus Notes

Gram-Positive Cocci

Learning Objectives

  • Classify gram-positive cocci.

  • Explain morphological, physiological, and growth characteristics of staphylococci in culture media.

  • Describe staphylococcal diseases, their pathogenesis, and transmission.

  • Describe antibiotic resistance in Staphylococci and its significance.

  • Explain diagnosis and treatment of staphylococcal infections.

Gram-Positive vs. Gram-Negative Cell Wall

  • Gram-positive: Thick peptidoglycan layer, no outer membrane.

  • Gram-negative: Thin peptidoglycan layer, outer membrane containing lipopolysaccharides (LPS).

Classification of Cocci

  • Gram-Positive Cocci:

    • Staphylococcus, Micrococcus, Peptococcus: Round in clusters and tetrads.

    • Streptococcus, Peptostreptococcus, Enterococcus: Oval shape in chains.

    • Corynebacterium, Listeria, Erysipelothrix, Mycobacterium, Propionibacterium: Club-shaped and/or in palisades.

    • Bacillus, Clostridium: Spore-bearing, large, uniform.

  • Gram-Negative Cocci:

    • Neisseria, Veillonella.

  • Gram-Negative Bacilli: (diverse morphologies and arrangements, branching, filamentous, coccobacillary, curved, uniformly bacillary, coiled & sphero-plastic)

Aerobic Gram-Positive Bacteria

  • Listeria, Bacillus, Corynebacterium, Nocardia (weakly acid-fast).

Anaerobic Gram-Positive Bacteria

  • Clostridium, Actinomyces (not acid-fast).

Streptococcus Differentiation

  • Hemolysis:

    • α-hemolysis: Partial hemolysis (e.g., S. pneumoniae, Viridans streptococci like S. mutans, S. mitis).

    • β-hemolysis: Complete hemolysis (e.g., S. pyogenes (group A), S. agalactiae (group B)).

    • γ-hemolysis: No hemolysis (e.g., Nonenterococcus S. bovis, Enterococci such as E. faecium, E. faecalis).

  • Tests:

    • Optochin sensitivity: Differentiates S. pneumoniae from Viridans streptococci.

    • Bacitracin sensitivity: Differentiates S. pyogenes from other β-hemolytic streptococci.

    • Growth in 6.5% NaCl: Differentiates Enterococci from Nonenterococcus.

Staphylococcus Differentiation

  • Catalase test: Positive for Staphylococcus.

  • Coagulase test:

    • Coagulase-positive: S. aureus.

    • Coagulase-negative: S. epidermidis, S. saprophyticus.

  • Novobiocin sensitivity:

    • Sensitive: S. epidermidis.

    • Resistant: S. saprophyticus.

Staphylococcus Overview

  • Found on skin, upper respiratory tract, lower digestive system, and urogenital system.

  • Opportunistic pathogen.

  • Can cause life-threatening systemic diseases, including skin/soft tissue, bone, and urinary tract infections.

  • Common cause of nosocomial infections (MRSA).

Staphylococcus Species

  • Genus consists of 45 species and 24 subspecies.

  • S. aureus: Most virulent and best-known member.

  • Other species: S. epidermidis, S. haemolyticus, S. lugdunensis, S. saprophyticus.

Important Staphylococcus Species in Humans

  • Staphylococcus aureus

  • Coagulase-Negative Staphylococci (CoNS)

  • Staphylococcus epidermidis

  • Staphylococcus saprophyticus

Characteristics of Staphylococcus

  • Gram-positive cocci in irregular, grape-like clusters.

  • Nonmotile.

  • Facultative anaerobic (fermentative).

  • Catalase positive.

  • Oxidase negative.

  • Grow in simple environments and high salt concentrations (e.g., 10% NaCl).

  • Optimal growth temperature: 30-37 °C (range: 10-42 °C).

  • Form pigmented colonies:

    • S. aureus: Golden yellow.

    • S. epidermidis (albus): White.

    • S. citreus: Lemon yellow.

Locations of Staphylococcus

  • Common in mammals, particularly in the nasal cavity, skin, and mucous membranes.

  • Temporarily found in the digestive tract.

  • Infections are usually endogenous but can also be nosocomial.

Colony Morphology

  • Staphylococci form S-type colonies.

  • Usually white and bright in color, up to 4 mm in diameter.

  • S. aureus strains form golden-colored colonies.

  • Some coagulase-negative staphylococci colonies are also pigmented.

  • Make homogeneous turbidity in liquid medium.

Hemolysis on Blood Agar

  • S. aureus typically exhibits β-hemolysis (complete hemolysis).

  • Colonies are surrounded by a wide zone of beta-hemolysis.

Antigenic Structure and Virulence Factors

  • Peptidoglycan:

    • Provides rigidity and structural integrity.

    • Exhibits endotoxin-like activity.

  • Teichoic Acids:

    • Species-specific.

    • Poor immunogens but stimulate a specific antibody response when bound to peptidoglycan.

  • Protein A:

    • Found in virulent S. aureus strains.

    • Surface adhesion protein.

    • Binds IgG through the Fc part, acting as an anti-phagocytic factor.

    • Dependent on cell wall or partly releasable.

  • Coagulase:

    • Surface adhesion protein.

    • Exists as free coagulase and clumping factor (bound coagulase).

  • Capsule and Slime Layer:

    • Protects bacteria by inhibiting phagocytosis.

    • Contributes to slime layer and biofilm formation.

  • Toxins and Enzymes: Contribute to virulence.

Virulence Factors of Staphylococcus aureus

  • Catalase-positive, Gram-positive cocci arranged in clusters.

  • Coagulase-positive.

  • Mannitol-positive.

  • Often Beta-hemolytic.

  • Sensitive to novobiocin.

Diseases Caused by Staphylococcus aureus

  • Toxin-mediated diseases (food poisoning, toxic shock syndrome, scalded skin syndrome).

  • Pyogenic diseases (impetigo, folliculitis, furuncles, carbuncles, wound infections).

  • Systemic diseases (pneumonia, osteomyelitis).

  • Hospital- and community-acquired infections with MRSA.

  • Methicillin-resistant Staphylococcus aureus (MRSA) is a threat due to the mecA gene.

Virulence Factors - Structural Components and Biologic Effects

  • Capsule: Inhibits chemotaxis and phagocytosis, inhibits proliferation of mononuclear cells.

  • Slime Layer: Facilitates adherence to foreign bodies, inhibits phagocytosis.

  • Peptidoglycan: Provides osmotic stability, stimulates endogenous pyrogen production, leukocyte chemoattractant, inhibits phagocytosis.

  • Teichoic Acid: Binds to fibronectin.

  • Protein A: Inhibits antibody-mediated clearance by binding IgG, IgG2, and IgG4 Fc receptors; leukocyte chemoattractant, anticomplementary.

Virulence Factors - Toxins and Enzymes

  • Cytotoxins: Toxic for various cells (erythrocytes, fibroblasts, leukocytes, macrophages, platelets).

  • Exfoliative Toxins (ETA, ETB): Serine proteases that split intercellular bridges in the stratum granulosum epidermis.

  • Enterotoxins (A-R): Superantigens that stimulate T cell proliferation and cytokine release, leading to inflammatory responses.

  • Toxic Shock Syndrome Toxin-1 (TSST-1): Superantigen causing T cell proliferation and endothelial cell damage.

  • Coagulase: Converts fibrinogen to fibrin.

  • Hyaluronidase: Hydrolyzes hyaluronic acid in connective tissue, promoting spread.

  • Fibrinolysin (Staphylokinase): Dissolves fibrin clots.

  • Lipases: Hydrolyzes lipids.

  • Nucleases: Hydrolyzes DNA.

Infections Caused by Staphylococcus aureus

  • Skin and soft tissue: Folliculitis, furuncles, abscesses, wound infections, carbuncles, impetigo, paronychia.

  • Musculoskeletal: Osteomyelitis, arthritis, bursitis, pyomyositis.

  • Respiratory: Tonsillitis, pharyngitis, sinusitis, otitis, bronchopneumonia, lung abscess, empyema, rarely pneumonia.

  • Central nervous system: Abscess, meningitis, intracranial thrombophlebitis.

  • Endovascular: Bacteremia, septicemia, pyemia, endocarditis.

  • Urinary: Associated with instrumentation, implants, and bacteremia.

Toxin-Mediated vs. Suppurative Infections

  • Toxin-Mediated:

    • Scalded Skin Syndrome: Disseminated desquamation of epithelium in infants, blisters without organisms or leukocytes.

    • Food Poisoning: Rapid onset of vomiting, diarrhea, and abdominal cramping after consuming contaminated food.

    • Toxic Shock Syndrome: Multisystem intoxication with fever, hypotension, and a diffuse rash; high mortality without treatment.

  • Suppurative Infections:

    • Impetigo: Localized cutaneous infection with pus-filled vesicles.

    • Folliculitis: Impetigo involving hair follicles.

    • Furuncles (Boils): Large, painful, pus-filled cutaneous nodules.

    • Carbuncles: Coalescence of furuncles extending into subcutaneous tissues.

    • Bacteremia and Endocarditis: Spread of bacteria into the blood.

    • Pneumonia and Empyema: Consolidation and abscess formation in the lungs.

    • Osteomyelitis: Bone destruction.

    • Septic Arthritis: Painful joint with purulent material.

Staphylococcal Scalded Skin Syndrome (Ritter Disease)

  • Presents 48 hours after birth; rare in children older than 6 years.

Methicillin-Resistant Staphylococcus aureus (MRSA)

  • mecA gene alters PBP-2 to PBP-2A, resulting in loss of target affinity to methicillin.

  • Estimated 323,700 cases in hospitalized patients in 2017, with 10,600 deaths and $1.7B in healthcare costs.

Epidemiology

  • Normal flora on human skin and mucosal surfaces.

  • Organisms can survive on dry surfaces for long periods.

  • Person-to-person spread through direct contact or contaminated fomites.

  • Risk factors: Foreign bodies, prior surgery, antibiotic use.

  • Patients at risk: infants (scalded skin syndrome), young children (impetigo), menstruating women (TSS), patients with catheters (bacteremia, endocarditis), patients with compromised pulmonary function (pneumonia).

  • MRSA is a common cause of community-acquired skin and soft-tissue infections.

Treatment, Prevention, and Control

  • Localized infections managed by incision and drainage; antibiotic therapy.

  • Empiric therapy includes antibiotics against MRSA: trimethoprim-sulfamethoxazole, doxycycline/minocycline, clindamycin, or linezolid.

  • Vancomycin is drug of choice for intravenous therapy; daptomycin, tigecycline, or linezolid are alternatives.

  • Symptomatic treatment for food poisoning.

  • Prevention involves infection control measures and avoiding prolonged tampon use.

Coagulase-Negative Staphylococci (CoNS)

  • Catalase-positive, coagulase-negative.

  • Relatively avirulent.

  • Slime layer allows adherence to foreign bodies and protects from phagocytosis and antibiotics.

  • Infections: subacute endocarditis, foreign body infections, urinary tract infections.

Staphylococcus epidermidis

  • Normal human skin flora.

  • Causes bacteremia, IV catheter infections, CSF shunt infections, endocarditis, UTI.

  • Mannitol (-), coagulase (-), novobiocin sensitive.

Staphylococcus saprophyticus

  • Normal flora colonizing the perineum, rectum, urethra, cervix, and GI tract.

  • Second most common cause of cystitis in young women.

  • Mannitol (+ or -), coagulase (-), novobiocin resistant.

CoNS Infections

  • Wound infections: Erythema and pus at traumatic or surgical sites, especially with foreign bodies.

  • Urinary tract infections: Dysuria and pyuria in sexually active young women (S. saprophyticus), and in patients with urinary catheters.

  • Catheter and shunt infections: Chronic inflammatory response to bacteria coating a catheter or shunt.

  • Prosthetic device infections: Localized pain and mechanical failure of the device.

Staphylococcus saprophyticus - Details

  • Colonizes the rectum, urethra, perineum, and anus.

  • Second most common cause of UTIs.

  • Can form biofilms on medical implants. The biofilm, composed of EPS (extracellular polymeric substance), makes it difficult for antibiotics to reach the bacteria.

CoNS Epidemiology

  • Normal human flora on skin and mucosal surfaces.

  • Organisms survive on dry surfaces.

  • Spread through direct contact or contaminated fomites.

  • Risk factors: Foreign bodies.

  • Ubiquitous with no geographic or seasonal limitations.

CoNS Treatment, Prevention, and Control

  • Antibiotics of choice: Oxacillin/penicillinase-resistant penicillin or vancomycin for oxacillin-resistant strains.

  • Removal of foreign body is often required.

  • Prompt treatment for endocarditis or shunt infections is necessary.

Laboratory Diagnosis - Specimen Collection

  • Pus from pyogenic lesions.

  • Blood from septicemia.

  • Cerebrospinal fluid from meningitis.

  • Sputum from respiratory infection.

  • Suspected food, vomit, or feces from food poisoning.

  • Mid-stream urine in urinary tract infection.

  • Anterior nasal swab from suspected carriers.

Laboratory Diagnosis - Microscopic Examination

  • Gram stain: Gram-positive cocci.

  • Arrangements: Single cells, pairs, short chains, but predominantly in grape-like clusters.

Laboratory Diagnosis - Culture Characteristics

  • On blood agar:

    • S. aureus: β-hemolysis (clear zone around colonies).

    • S. epidermidis: White-creamy colonies, no hemolysis.

    • S. saprophyticus: White-yellow colonies, no hemolysis.

  • On mannitol salt agar:

    • Differential medium for mannitol fermenters; contains mannitol, phenol red indicator, and 7-10% sodium chloride.

    • S. aureus: Yellow colonies with yellow zones (mannitol fermenter).

    • Other Staphylococci: Small pink or red colonies with no color change (non-mannitol fermenters).

Hemolytic Reactions on Blood Agar (Streptococci Classification)

  1. β-Hemolysis: Complete lysis of red cells.

  2. α-Hemolysis: Partial or “Greening” hemolysis associated with the reduction of red cell hemoglobin.

  3. γ-Hemolysis: Nonhemolytic colonies.

Additional Information on Staphylococcus Differentiation

  • Coagulase Test: Differentiates S. aureus from other Staphylococci.

  • Catalase Test: Positive for staphylococcus, negative for streptococcus

  • Novobiocin Sensitivity: Differentiates S. epidermidis and S. saprophyticus.

Preventing Antibiotic-Resistant Infections

  • Infection control actions combined with every patient interaction.

  • Prevent infections from catheters and after surgery.

  • Prevent bacteria from spreading.

  • Improve antibiotic use.