Staphylococci and Diagnostic Tests

Staphylococci

• Staphylococci are a genus of Gram-positive bacteria
• They are spherical (cocci) and form in clumps/clusters
• They are facultative anaerobes
• Traditionally divided into two groups based on their ability to clot blood plasma (coagulase reaction/test)
  • Coagulase-positive staphylococci (e.g., S. aureus):
    • Colonize nasal passages and moist skin areas (axilla, groin)
  • Coagulase-negative staphylococci (>30 other species):
    • Common human skin commensals (e.g., S. epidermidis)

Coagulase Test

• A laboratory test used to differentiate S. aureus from other staphylococci (negative)
• Coagulase is an enzyme that causes blood clot formation (converting fibrinogen to fibrin)
• Allows bacteria to coat its surface with fibrin, potentially resisting phagocytosis

Catalase Test

• Used to differentiate staphylococci from streptococci (also a Gram-positive coccus)
• Presence of catalase (enzyme) is determined by the bacteria's ability to reduce hydrogen peroxide into water and oxygen, resulting in bubble production
• Staphylococci = Catalase +

Clinical Case 1

• A 19-year-old male presents with a painful, red lump on his neck
• The most likely diagnosis is a furuncle

Staphylococcus aureus (S. aureus)

• Found in moist skin folds, mucosal surfaces, nasopharynx
• 20–40% of healthy humans are colonized
• Increased colonization in individuals with diabetes mellitus, injecting drug use, or foreign bodies/implants
• Distinguished as methicillin-sensitive (MSSA) or methicillin-resistant (MRSA)
  • Resistance to usual treatments results from an alteration of penicillin-binding protein (PBP2a)
  • Usually a healthcare-associated infection; elderly and those with compromised immune systems at higher risk
  • Can also resist other antimicrobial classes
  • In hospitals, isolate and cohort patients with contact precautions

S. aureus: Pathogenesis (Portal of Entry)

• Ingestion
• Penetration
  • A break in the skin
  • Entry through mucous membranes, allowing access to adjoining tissues

S. aureus: Pathogenesis (Attachment to Cells)

• Surface proteins
  • Facilitate attachment
  • Capsule
    • Inhibits chemotaxis and phagocytosis
    • Facilitates adherence to implants
• Fibrin/fibrinogen binding proteins (clumping factor)
  • Attachment to blood clots and traumatized tissue
• Matrix-binding proteins
  • Promote collagen attachment (causing osteomyelitis or septic arthritis for strains)

S. aureus: Pathogenesis (Defeating the Immune System)

• Inhibition of phagocytosis while remaining within phagocytes
• Production of extracellular substances promoting invasion (Invasins and enzymes)

S. aureus: Pathogenesis (Damage to Host Cells)

• Direct damage to peptidoglycan cell walls
• Enzymes
• Exotoxins (Superantigens)
  • Toxic shock syndrome toxin (TSST-1)
  • Enterotoxins (A-E, G-I; cause food poisoning)
• Exfoliative toxins (cause scalded skin syndrome)
• Other cytotoxins (alpha, beta, leukocidin)

S. aureus: Pathogenesis (Spread)

• Person-to-person contact, skin carriage
• Environment (shedding onto surfaces)

Classification of Staphylococcal Infections

• Skin and soft tissue (e.g., boils, carbuncles, furuncles)
• Systemic/invasive infections (e.g., bloodstream infection [BSI], endocarditis, bone/joint infections, deep abscesses, pneumonia)
• Toxin-mediated infections (e.g., food poisoning, gastroenteritis, scalded skin syndrome, toxic shock syndrome)

S. aureus: Systemic Infections

• Bloodstream infection (BSI)
  • Usually secondary to BSI
  • Covered in more detail in Cardiovascular module
• Bone/joint infections (e.g., septic arthritis, osteomyelitis)
  • Refer to the lecture
• Deep abscesses (brain, spine, psoas muscle)
• Pneumonia
  • Risk factors: viral respiratory infections, cystic fibrosis, ventilation, aspiration
  • Covered in respiratory module

S. aureus: Toxin-mediated Infection - Scalded Skin Syndrome

• Spectrum of superficial blistering skin disorders
  • Localized blisters
  • Generalized exfoliation of entire body surface
  • Mucous membranes usually spared
• Exfoliative toxins spread hematogenously from a localized source
• Split intracellular bridges in the skin layer (middle layers)
• Is most common in children under 6, but seen in neonates, immunocompromised adults, and those with renal failure
• Contagious
• Mortality: Children <3%, Adults up to 60%

S. aureus: Toxin-mediated Infection - Toxic Shock Syndrome (TSS)

• TSS toxin-1 acts as a superantigen
• Massive cytokine release
• Historically associated with high-absorbency tampons
• Rapid, dramatic, and fulminant onset
  • Pyrexia, hypotension, rash (with desquamation)
• Other organ involvement
  • Renal failure
  • CNS (disorientation without focal neurological signs)
  • Muscular (severe myalgia, increase in CK)

Laboratory Diagnosis

• Day 0: Patient specimen (e.g., blood culture)
• Incubate at 37°C
• Day 1+: Gram stain (bunches of grapes), lab team review, potentially PCR for confirmation
• 24 hours later: Read agar culture plates, coagulase test, and phone team/patient review
• 24 hours later: Alter empiric antibiotic therapy as appropriate, if MRSA, infection control precautions and decolonization

Management of S. aureus Infection

• History (especially timing and presentation of symptoms)
• Clinical examination
  • Focus on a possible source/evidence of spread if systemically unwell
  • Skin (IV sites, indwelling devices, surgical wounds, soft tissue infections or abscesses); remember that scars can be a sign of implanted devices
  • Cardiovascular System (CVS): evidence of known or new/changed murmur
  • Musculoskeletal (new bone or joint pain, reduced range of motion or limp, swelling, loss of function)
• Investigations (depends on clinical presentation):
• General blood tests (FBC, U&E, CRP, lactate)
• Microbiology (specimen depends on infection site)
  • Abscess - pus or tissue biopsy
  • BSI: blood cultures
  • Pneumonia: sputum, bronchoalveolar lavage (BAL)
  • Septic arthritis: joint fluid
  • TSS: next in the slide
  • Food poisoning: food (not stool)
• Other imaging (e.g., CXR, CT or MRI scans, echocardiogram [ECHO])

TSS - Investigations & Management

• Investigations (if febrile or systemically unwell):
  • Blood cultures (rarely positive)
  • Wound swab if skin lesion
  • Other swabs (abscess, cervix/vagina)
• Management:
  • Quickly recognize and treat for resuscitation and critical care
  • Rapid IV antimicrobials
  • Source control (debriding infected or necrotic wounds, drain abscesses etc.)

Antibiotic Treatment

• Choice, route, and duration depend on site and complexity of infection
• Mild infections (e.g., boil, folliculitis): no treatment
• Skin/soft tissue infections and respiratory tract infections: 7 days
• Bloodstream infection: 14 days
• Complicated infections (e.g., endocarditis, septic arthritis, osteomyelitis): at least four weeks, potentially longer
• Flucloxacillin if susceptible (MSSA)
• 1st-generation cephalosporin (e.g., cefazolin) - alternative for options
• Vancomycin or Teicoplanin (glycopeptides) if MRSA
• Alternatives: daptomycin, linezolid, tetracyclines
• If bloodstream infection, consider the source; do ECHO and radiology
• Repeat blood cultures after commencing antimicrobials
• Confirm the blood cultures are now sterile

Epidemiology: S. aureus Bloodstream Infection in Ireland (2018–2022)

• Chart showing numbers of total and MRSA cases per year. 2022 shows the highest number of total cases.

Community-Acquired MRSA

• Skin infections and necrotizing pneumonia
• Younger patients, healthier patients
• Less antibiotic resistant, but more virulent
• Certain strains are common in North America

Coagulase-negative Staphylococci

• Natural inhabitants of human skin and mucosa
• Less virulent than S. aureus; rarely cause infection in healthy individuals
• S. epidermidis is commonly associated with prosthetic devices (e.g., joint replacements, prosthetic valves, pacemakers)
• S. saprophyticus causes urinary tract infections
• Covered again in year 2 REGUB module

Staphylococcus epidermidis pathogenesis

• Bacteria adhesion to biomaterial
• Biofilm formation and tissue cell displacement

S. epidermidis infections (Devices)

• Bloodstream infection secondary to IV lines
• Endocarditis (prosthetic valves)
• Prosthetic joint infections
• Continuous ambulatory peritoneal dialysis peritonitis
• Ventriculitis/shunt-associated meningitis

Clinical Case 2

• 65-year-old male with fever and rigors 16 days post-small bowel resection
• Central venous catheter (CVC) in situ for parenteral nutrition
• Erythematous skin around CVC insertion site
• Two sets of blood cultures sent for culture and susceptibility testing

Diagnosis & Management of CoNS

• Often patient is not particularly systemically unwell
• History and examination
• Blood cultures (at least two sets)
• Source control (often the prosthesis must come out for effective treatment)
• Culture prosthetic material
• Coagulase-negative staphylococci are often antibiotic resistant (including methicillin and flucloxacillin)
• Vancomycin is the usual empiric treatment
• Indication and duration of treatment depend on the location of infection and if the prosthetic material can be removed

Clinical Case 3

• 72-year-old female with fever and rigors six weeks post-total knee replacement
• Blood cultures shows:
  • Microscopy: Gram-positive cocci in clusters
  • Catalase: positive
  • Coagulase: negative
• Most likely causative pathogen is S. epidermidis based on the results - note the lack of coagulase activity

Understanding Laboratory Results

• Microscopy: Gram-positive cocci in clusters
• Catalase: positive
• Coagulase: positive, indicating S. aureus

Preventing Staphylococcal Infection

• Prevent transmission from patient-to-patient
  • Hand hygiene
  • Environmental and equipment hygiene
  • Transmission-based precautions (isolate infected patients, use gloves, aprons/gowns) for MRSA

Summary: Staphylococci

• Coagulase +ve (S. aureus)
  • 20-40% of population carries it without infection
  • Common cause of skin and soft-tissue. systemic, and toxin-mediated infections
  • Virulence factors (treatment: flucloxacillin for MSSA, vancomycin for MRSA)
• Coagulase −ve (S. epidermidis and S. saprophyticus)
  • Often normal flora
  • Rarely pathogenic
  • S. epidermidis: infections are often associated with prosthetic devices, requiring empiric vancomycin treatment
  • S. saprophyticus: causes UTIs

Virulence Factors of S. aureus

• Summary of S. aureus virulence factors, with enzyme descriptions, biological roles, and relevant details provided (Table-like format)

Staphylococcal Biofilm Formation

• CoNS and MRSA are exceptionally proficient at attaching to surfaces and producing biofilms