Lec 4: Staphylococci and Related Bacteria

GRAM POSITIVE STAPHYLOCOCCI: FEATURES

  • Gram-positive cocci organized in pairs, tetrads, short chains (3–4 cells) and “grapelike” clusters

  • Facultatively anaerobic, non-motile

    • Produce catalase

  • Able to grow in the presence of salt

    • Food contamination risk linked to salt tolerance and survivability in foods

  • Colony appearance on blood agar: large white, cream, yellow to orange raised colonies

  • Stain purple with Gram stain (Gram-positive cocci)

GRAM POSITIVE STAPHYLOCOCCI: PHYSIOLOGY & STRUCTURE

  • Polysaccharide capsule (virulence factor) protects from phagocytosis

  • Slime layer: facilitates easier adhesion/colonization

  • Cell wall rich in peptidoglycan

  • Penicillin-binding proteins (PBPs): enzymes that catalyze construction of peptidoglycan

    • PBPs are targets for modified penicillins binding (serine proteases)

EPIDEMIOLOGY

  • Staphylococci are ubiquitous (EVERYWHERE); humans are the reservoir (skin)

  • S. aureus colonization is common; nasal carriage in healthy adults (15%)

  • Contaminated fomites (towels, clothing, dry surfaces)

STAPHYLOCOCCUS SPECIES (2 TYPES)

  1. S. aureus

    • Toxins and enzymes:

      • Leukocidins – pore-forming toxins that lyse leukocytes (kills them)

        • multiple forms (α, β, δ, γ, etc.) with specific activities:

          • α-toxin: septic shock and dermonecrosis

          • β-toxin: sphingomyelinase activity

          • δ-toxin: activity on RBCs and WBCs

          • γ-toxin: tissue necrosis

        • PVL (Panton-Valentine leukocidin):

          • 3–5% of strains (approx.) carry PVL

            • less than 5% of clinical isolates contain gene

          • Associated with necrotizing pneumonia and severe skin infections

          • The gene is carried by a lysogenic phage; information can be acquired by Staph during infection

      • Enterotoxins – vomiting & diarrhea (STAPH FOOD POISONING)

        • (GI effects):

          • Cause disruption of intestinal mucosa, emesis, diarrhea (STAPH FOOD POISONING)

          • Act as superantigens in the GI tract; lead to large IL-1 and IL-2 release and cytokine storm in enteric context

          • Enterotoxin A (toxin A): preformed in food

          • Enterotoxin F (toxin F): similar to TSST in mechanism

          • Food poisoning incubation: typically 1-8 hours after ingestion

          • Food might appear normal/taster-like; toxin heat-stable; bacteria may be killed by heating, but toxin remains

          • Disease is SELF-LIMITING: usually resolves in

      • Toxic-shock syndrome toxin (TSST-1) – superantigen causing toxic shock

        • Toxic Shock Syndrome Toxin (TSST-1):

          • Superantigen that stimulates T cells by binding to MHC class II on macrophages

          • Potent inducer of TNF and IL-1

          • stimulates IL-2 release from CD4+ cells

          • In the population, 5%–25% of strains are highly stimulatory to the immune system

      • Exfoliatin – epidermolytic protease; causes epidermal separation

        • serine proteases that cleave desmoglein in desmosomes, leading to epidermal separation and scalded skin syndrome

      • Enzymes (coat and invade): coagulase, catalase, other secreted factors

        • Coagulase: converts fibrinogen/thrombin to fibrin, walling off infection

          • production by S. aureus (and some others; CoNS typically negative)

        • Beta-lactamase: plasmid-encoded; confers penicillin resistance; >9090 % of strains have beta-lactamase

        • Mutant penicillin-binding proteins (PBPs) – methicillin resistance (MRSA):

          • mecA gene codes for a novel PBP (PBP2) with low affinity for beta-lactams

          • PBP2 retains enzymatic activity but is not bound by beta-lactams

          • This provides resistance to methicillin and related penicillins

    • Structural/auxiliary factors:

      • Protein A – binds Fc region of IgG; blocks complement pathway and opsonization

      • Teichoic acids – mediate adherence

      • Lipoteichoic acids – induce cytokine release

      • Polysaccharide capsule – poorly immunogenic; complicates vaccine development

      • Peptidoglycan – endotoxin-like properties

      • Staphyloxanthin – pigment that helps inactivate oxygen-dependent killing within neutrophils

  2. Coagulase-negative Staphylococcus (CoNS)

    • S. epidermidis – “outer skin”

    • S. saprophyticus – “saprophyte”

    • S. haemolyticus & S. hominis – axilla (apocrine glands)

    • S. lugdunensis – Lyon, France

    • S. capitus – sebaceous glands

CLINICAL IMPORTANCE: S. AUREUS AS A PATHOGEN

  • Clinical prominence: #1 causative organism for several conditions

    1. Bacteremia and sepsis

    2. Osteomyelitis (trauma/hematogenous spread)

    3. Infective endocarditis

    4. Acute endocarditis (rapid onset)

      1. (native and prosthetic valves; risk of biofilm formation on prosthetic material)

  • Skin and soft tissue infections (SSIs) aka Pyogenic diseases:

    1. Furuncles, boils, cellulitis, impetigo (STAPH)

    2. Paronychia, conjunctivitis, burn and wound infections

    3. Bullous impetigo vs impetigo (board keyword):

      • bullous form is a localized, scalded-skin–type presentation; blisters may be culture-positive

    4. Pneumonia and empyema:

      • Aspiration of oral secretions or hematogenous spread

        • Can cause necrotizing pneumonia; post-viral lobar pneumonia (notably after influenza)

    5. Toxin-mediated diseases:

      1. Scalded skin syndrome (Ritter’s disease)

        • Abrupt onset of localized erythema that spreads over body

        • Blisters followed by desquamation of epithelium

        • Clear fluid – no organisms or leukocytes

          • These blisters are usually culture negative vs. bolus impetigo which is usually culture positive

          • Most often occurs in young children

      2. Food poisoning

        1. most common cause in the U.S.

          • Contamination of food by human carriers; high-salt foods favor S. aureus growth

          • Common vehicles: processed meats (high salt), custard-filled pastries, potato salad, ice cream

          • Bacteria introduced by person (sneeze, hand); food left at room temperature allows growth and toxin production

          • toxin is heat-stable; food may not taste tainted

            • ingestion of enterotoxin A

          • Symptoms: nausea, vomiting, watery non-bloody diarrhea

          • Onset: ~4 hours after ingestion

          • Course: SELF-LIMITING; resolves in 24-48 hours

          • incubates for 1-8 hours

      3. Toxic Shock Syndrome (TSS)

        1. Associated with tampon use (hyperabsorbent tampons) and vaginal colonization

          1. 5% of women with vaginal colonization

          2. Dramatic increase in disease seen with the advent of hyperabsorbent tampons

          3. Bacteria multiply rapidly in tampons and release toxin

        2. Strains that produce TSST-1 (5%-25%)

          1. High fever, sunburn-like rash, hypotension

          2. Desquamation of skin, palms and soles

COAGULASE-NEGATIVE STAPHYLOCOCCI (CoNS)

  1. Staphylococcus epidermidis

    • Clinical associations:

      • Bacteremia and sepsis in nosocomial settings: accounts for 75-90% of patients with nosocomial bacteremia

        • Nosocomial / iatrogenic infections: catheters, IV lines, prosthetics, pacemakers, hip joints, CSF shunts

          • introduced by putting device into body

            • Common in immunocompromised and neutropenic patients; ability to form biofilms on devices

        • Subacute endocarditis often from bacteremia; often involves abnormal or prosthetic valves

    • Virulence factors:

      • Polysaccharide slime layer enables adhesion and resistance to phagocytosis

      • Beta-lactamase (plasmid-encoded) and mutant PBPs contribute to penicillin resistance

      • Methicillin resistance (MRSE) also noted

        • E stands for epidermis

  2. Staphylococcus lugdunensis

    • Can cause endocarditis; more commonly native valve infections

    • Resembles S. aureus but is coagulase-negative

    • If it is hemolytic but coagulase-negative, this points to lugdunensis

      • Higher drug resistance compared with some other CoNS

  3. Staphylococcus saprophyticus

    • Clinical: Urinary tract infections (UTIs)

      • Pyelonephritis or cystitis

      • Most cases show pyuria and urinary symptoms

      • Commonly affects healthy, young, sexually active women

      • Non-gonococcal urethritis (NGU) in males

LAB DIAGNOSIS: KEY IDENTIFICATION TESTS

  • CATALASE TEST: breaks hydrogen peroxide into water and molecular oxygen , allowing for the differentiation between staphylococci (catalase positive) and streptococci (catalase negative).

    • Positive in Staphylococcus spp. (bubbles with H2O2)

    • Staphylococcus spp. are catalase-positive;

    • Streptococcus & Enterococcus are catalase-negative

  • HEMOLYSIS ON 5% SHEEP’S BLOOD AGAR:

    • Alpha (incomplete) hemolysis: green/brown discoloration due to partial hemolysis of red blood cells, commonly observed with Streptococcus pneumoniae.

    • Beta (complete) hemolysis: clear zones around colonies indicating complete lysis of red blood cells, typically seen with Streptococcus pyogenes.

      • S. aureus

    • Gamma (no hemolysis): no discoloration or lysis of red blood cells, characterized by the absence of any change in the agar surrounding the colonies, often associated with Enterococcus faecalis.

  • COAGULASE TEST:

    • S. aureus: Coagulase-positive

    • CoNS (e.g., S. epidermidis, S. lugdunensis, S. saprophyticus): Coagulase-negative

  • MANNITOL FERMENTATION TEST (on Mannitol Salt Agar, MSA):

    • High salt tolerance (7.5% NaCl) supports growth of Staphylococcus;

    • mannitol fermentation yields acid, changing indicator color

    • S. aureus ferments mannitol (yellow around colonies on MSA)

    • S. epidermidis is phenol red negative, meaning it does not ferment mannitol and does not produce acid, which is reflected in the lack of color change on the MSA plate.

  • NaCl TOLERANCE TEST:

    • Growth in 6.5% NaCl is a supportive test for Staphylococcus genus

  • OTHER IDENTIFICATION DETAILS:

    • S. lugdunensis: positive ornithine decarboxylase; additional tests often needed since it is CoNS

    • S. epidermidis: ferments glucose but not mannitol; grows on MSA with no color change; novobiocin-susceptible

    • S. saprophyticus: novobiocin-resistant (distinguishing from epi)

SUMMARY OF LAB IDENTIFICATION FOR COMMON SPECIES

  • STAPHYLOCOCCUS AUREUS

    • Catalase: Positive (strept neg)

      • also a virulence factor that degrades hydrogen peroxide and limits ability of neutrophils to kill them

    • Hemolysis: Beta

    • Coagulase: Positive

    • Mannitol fermentation: Positive (MSA)

    • 6.5% NaCl growth: Positive

  • STAPHYLOCOCCUS EPIDERMIDIS

    • Catalase: Positive

    • Hemolysis: None

    • Coagulase: Negative

    • Mannitol fermentation: Ferments glucose but not mannitol; grows on MSA with no color change

    • Novobiocin: Susceptible (S. saprophyticus resistant)

  • STAPHYLOCOCCUS LUGDUNENSIS

    • Catalase: Positive

    • Hemolysis: Beta

    • Coagulase: Negative

    • Ornithine decarboxylase: Positive

    • Requires additional testing due to CoNS and beta-hemolysis

  • STAPHYLOCOCCUS SAPROPHYTICUS

    • Catalase: Positive

    • Hemolysis: None

    • Coagulase: Negative

    • Novobiocin: Resistant

    • Distinguishes from S. epidermidis (which is novobiocin-sensitive)

NUMERICAL REFERENCES AND KEY STATISTICS (EMPHASIS ON NUMBERS)

  • Nasal carriage of S. aureus in healthy adults: 1515 %

  • PVL gene presence in clinical isolates: <5<5 %

    PVL-associated risk: severe necrotizing pneumonia and lung abscesses (3–5% carriers can cause severe disease in some contexts)

  • TSST-1-producing strains: 55 -2525 % of strains

  • MRSA resistance mechanism: mecA gene encoding PBP2 (mutant PBP) with reduced beta-lactam binding

  • Beta-lactamase production: >9090 % of strains

  • Endocarditis: rapid onset when involving abnormal or prosthetic valves (biofilm formation)

  • Immune evasion: Protein A blocks IgG Fc, preventing opsonization and complement activation

  • CoNS bacteremia: 75–90% of nosocomial bacteremia cases involve CoNS when devices are in place

  • MSA salt concentration: 7.57.5 % NaCl inhibits most bacteria; supports Staphylococcus growth and differentiates mannitol fermentation

  • Bacterial incubation/poisoning timelines:

    • Food poisoning incubation: 1–8hours1{–}8{ hours}

    • Food poisoning course: resolves in 24–48hours24{–}48hours

    • TSS symptom onset and progression: rapid with TSST-1–producing strains in susceptible individuals

REAL-WORLD RELEVANCE & IMPLICATIONS

  • Healthcare-associated infections: CoNS, especially S. epidermidis, on devices and prosthetics; device-related bacteremia

  • Antibiotic resistance trends: MRSA and MRSE due to mecA and mutant PBPs; beta-lactamase widespread

  • Public health: nasal carriage of S. aureus as a reservoir for infection and spread; high-salt food preparation and storage as risk factors for food poisoning

  • Immunological interactions: Protein A and teichoic acids influence host-pathogen interactions; PVL and TSST-1 drive severe systemic responses

  • Treatment considerations: choose antibiotics based on coagulase status, mecA presence, and local resistance patterns; MRSA coverage may be necessary in severe infections

CONNECTIONS TO FOUNDATIONAL PRINCIPLES

  • Host-pathogen interactions: evasion of phagocytosis via capsule and slime; immune evasion via Protein A and toxin-mediated hyperactivation

  • Microbial virulence concept: toxins and enzymes as direct mechanisms of tissue injury and immune modulation

  • Antibiotic mechanism of action vs resistance: beta-lactams target PBPs; resistance via mecA (PBP2) and beta-lactamase

  • Laboratory microbiology: differential testing (catalase, hemolysis, coagulase, mannitol fermentation, salt tolerance) to distinguish Staphylococcus species