Overview
Staphylococci are Gram positive cocci that occur in grape-like clusters. They are ubiquitous and are among the most common causes of localized pyogenic lesions in humans, with a well-known ability to develop resistance to penicillin and other antibiotics.
First observed by Von Recklinghausen, and named by Sir Alexander Ogston after "staphylo" (bunches of grapes) and "kokkos" (a berry), the genus includes the highly pathogenic coagulase-positive Staphylococcus aureus and the largely commensal coagulase-negative species such as S. epidermidis and S. saprophyticus.
Learning Objectives
After this lesson you will be able toβ¦- Classify Staphylococcus into its major species.
- Describe the morphology of Staphylococcus.
- Discuss the cultural, biochemical, and resistance characteristics of Staphylococcus.
- Describe the virulence factors and pathogenicity of Staphylococcus aureus.
- Describe the laboratory diagnosis of Staphylococcus, including coagulase testing.
Clinical Story
Why This MattersA hospitalized patient with an intravascular catheter develops sudden fever and rigors. Blood cultures grow Gram positive cocci in clusters. Is this a dangerous, invasive Staphylococcus aureus bacteremia, or a common catheter-related coagulase-negative Staphylococcus epidermidis contaminant? The coagulase test β performed in minutes β makes all the difference to how urgently the patient is treated.
Core Concepts
Staphylococcus aureus β coagulase-positive, ferments mannitol, mostly pathogenic.
Staphylococcus epidermidis β coagulase-negative, does not ferment mannitol, mostly non-pathogenic.
Morphologically, they are spherical cocci approximately 1 ΞΌm in diameter, arranged in grape-like clusters, non-motile, non-sporing, and uniformly Gram positive. Pyogenic lesion isolates typically produce yellow (golden) colonies; normal skin isolates produce white colonies.
Staphylococci are aerobes that grow readily on ordinary media between 10β42Β°C (optimum 37Β°C, pH 7.4β7.6). On nutrient agar, colonies are large, circular, convex, smooth, opaque and easily emulsifiable; most strains produce a golden pigment optimally at 22Β°C. Selective media containing 8β10% NaCl (salt-milk agar, salt broth, lithium chloride, tellurite) help isolate S. aureus from mixed specimens like faeces.
Biochemically, staphylococci ferment many sugars producing acid but no gas, and are catalase positive. S. aureus characteristically ferments mannitol, produces clear hemolysis on blood agar, a golden yellow pigment, liquefies gelatin, and produces phosphatase.
Staphylococci are among the most resistant non-sporing bacteria, retaining viability for 3β6 months and withstanding 60Β°C for minutes (thermal death point 62Β°C for 30 minutes). Most strains tolerate 10% NaCl, some up to 15%.
Penicillin resistance develops via three mechanisms: (1) beta-lactamase (penicillinase) production splitting the beta-lactam ring, (2) altered surface receptors reducing beta-lactam binding β producing Methicillin Resistant Staphylococcus Aureus (MRSA), and (3) tolerance, where the organism is inhibited but not killed. Because methicillin itself is unstable, cloxacillin is used for routine sensitivity testing. Vancomycin remains useful against multi-resistant strains.
Cell surface proteins: Protein A binds the Fc portion of IgG, leaving the Fab region free β inducing platelet damage and hypersensitivity. The clumping factor (bound coagulase) causes visible clumping in the slide coagulase test.
Extracellular enzymes: lipases aid skin/subcutaneous invasion, hyaluronidase breaks down connective tissue, staphylokinase spreads infection, and a heat-stable nuclease is a characteristic marker of S. aureus.
Toxins: cytolytic hemolysins (alpha, beta, gamma, delta) and leucocidin; heat-stable enterotoxin causing food poisoning; Toxic Shock Syndrome Toxin (TSST) causing a potentially fatal multisystem disease; and exfoliative toxin causing Staphylococcal Scalded Skin Syndrome (SSSS) β severe forms are Ritter's disease (newborns) and toxic epidermal necrolysis (adults); milder forms are pemphigus neonatorum and bullous impetigo.
Common infections include skin/soft tissue (folliculitis, boils, abscess, wound infection, carbuncle, impetigo, paronychia), musculoskeletal (osteomyelitis, arthritis), respiratory (tonsillitis, pneumonia), CNS (abscess, meningitis), endovascular (bacteremia, endocarditis), and urinary tract infections (usually instrumentation/implant related). Staphylococcal lesions are characteristically localized, unlike the spreading nature of streptococcal infection.
Staph epidermidis is normal skin flora that can cause stitch abscesses and device-related bacteremia (artificial heart valves, catheters, prosthetics). Staph saprophyticus is normal periurethral flora that causes UTIs in sexually active young women; it is distinguished by novobiocin resistance.
Laboratory Principle
The coagulase test is the pivotal identification step: bound coagulase (clumping factor) on the bacterial surface binds fibrinogen directly, producing visible clumping in the slide test; free coagulase secreted into broth activates prothrombin, converting fibrinogen to fibrin and clotting plasma in the tube test. Because S. aureus almost uniquely produces both forms, coagulase positivity is the definitive marker separating it from the far more numerous coagulase-negative staphylococci that are usually non-pathogenic.
Equipment Required
Reagents & Materials
| Reagent / Material | Concentration / Grade | Purpose | Storage |
|---|---|---|---|
| Rabbit / human plasma | EDTA, oxalate, or heparin anticoagulated | Slide and tube coagulase tests | Refrigerated (2β8Β°C) |
| Saline | 0.85% NaCl | Emulsifying isolate for slide coagulase | Room temperature |
| Mannitol salt agar | 7.5% NaCl selective medium | Selective isolation and mannitol fermentation | Refrigerated plates, 2β8Β°C |
| Novobiocin disc | 5 ΞΌg | Differentiating S. saprophyticus (resistant) from S. epidermidis (sensitive) | Refrigerated, protect from moisture |
Step-by-Step Procedure
Collect specimen appropriate to the lesion (pus, sputum, nasal swab for carriers, food/faeces for food poisoning). Perform Gram staining directly on pus specimens β cocci in clusters may be readily seen.
Plate on blood agar; colonies appear after overnight incubation. For specimens where staphylococci are expected to be scanty, use selective media such as Ludlam's agar, salt-milk agar, or Robertson's cooked meat medium (10% NaCl).
Emulsify the isolate in a drop of saline on a slide, check for absence of autoagglutination, add a drop of plasma, and mix. Prompt clumping = positive for bound coagulase. Always set up positive and negative controls.
Add ~0.1 mL of a young broth or agar culture suspension to ~0.5 mL of plasma in a narrow test tube. Incubate at 37Β°C for 3β6 hours (up to 24 h if needed). A clot that does not flow on tilting = positive; this is the deciding test when slide and tube results diverge.
Confirm catalase positivity (differentiates from Streptococcus), then perform antibiotic sensitivity testing (including cloxacillin for MRSA screening) as a guide to treatment.
Flow Diagram
Quality Control
Run Staphylococcus aureus ATCC 25923 as a coagulase-positive control and Staphylococcus epidermidis as a coagulase-negative control with every batch of plasma tested. Discard any plasma lot that fails to clot with the positive control.
Participate in external proficiency testing programs that periodically send blinded Staphylococcus isolates (including MRSA strains) to confirm accurate species identification and susceptibility reporting.
Reference Values
Species Characteristicsβ οΈ Reference reactions may vary between strains. Always confirm identification with positive/negative controls run in parallel.
Clinical Interpretation
| Finding | Possible Significance | Action / Follow-up |
|---|---|---|
| Coagulase-positive GPC in clusters from pus | Staphylococcus aureus β likely true pathogen | Proceed to cloxacillin/MRSA susceptibility testing |
| Coagulase-negative GPC from a single blood culture | Often skin contaminant (S. epidermidis) unless from a device or repeated cultures | Correlate with clinical picture; repeat culture if uncertain |
| Cloxacillin/methicillin resistant S. aureus | MRSA β resistant to all beta-lactams | Initiate contact precautions; treat with vancomycin or alternative MRSA-active agent |
| Coagulase-negative, novobiocin-resistant isolate from urine | Staphylococcus saprophyticus β common UTI in young sexually active women | Treat as urinary pathogen, not contaminant |
Common Errors & How to Avoid Them
Cause: Some strains clump spontaneously in saline even before plasma is added, mimicking a true positive.
Prevention: Always check for autoagglutination in saline alone before adding plasma; if present, proceed directly to the tube test.
Cause: Methicillin itself degrades quickly and can give unreliable results.
Prevention: Use cloxacillin (a stable penicillinase-resistant penicillin) as the standard surrogate marker for MRSA screening.
Cause: Assuming any coagulase-negative isolate is a skin contaminant can miss true infections associated with prosthetic devices, catheters, and heart valves.
Prevention: Correlate with clinical context β repeated positive cultures, indwelling devices, or immunocompromise raise suspicion for true infection.
Laboratory Tips from the Bench
When the slide and tube coagulase results disagree, always trust the tube test β it is the definitive method for detecting free coagulase.
A novobiocin disc quickly separates the coagulase-negative species: S. saprophyticus is resistant, while S. epidermidis is sensitive β a useful shortcut in urine isolates.
"Grapes that clot the plasma" β Staphylococcus (grape clusters) that is coagulase positive = S. aureus, the true pathogen of the group.
Important Notes
Methicillin Resistant Staphylococcus Aureus (MRSA) causes serious nosocomial infections and requires prompt identification, isolation precautions, and treatment with vancomycin or other reserved agents.
Bacteriophage typing of Staphylococcus is of great importance in epidemiological studies, helping trace the source of hospital or community outbreaks by identifying the specific phage type responsible.
Interactive Quiz
Test Your KnowledgeFlashcards
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Clinical Case Study
Apply Your KnowledgeThree weeks after prosthetic valve replacement, presents with low-grade fever, malaise, and a new murmur. Three sets of blood cultures are drawn.
Coagulase-negative, novobiocin-sensitive staphylococcus growing repeatedly (3/3 sets) in a patient with a prosthetic valve strongly suggests true Staphylococcus epidermidis prosthetic valve endocarditis rather than a skin contaminant.
- βCoagulase-negative staphylococci are not always contaminants, especially with prosthetic devices.
- βMultiple positive blood culture sets increase confidence that an isolate is a true pathogen.
- βNovobiocin susceptibility helps distinguish S. epidermidis from S. saprophyticus.
Frequently Asked Questions
Sir Alexander Ogston named the genus from the Greek "staphylo" (bunches of grapes) and "kokkos" (berry) because the cocci divide in multiple, irregular planes, producing grape-like clusters when viewed under the microscope in culture.
Methicillin is chemically unstable and degrades under standard testing conditions, giving unreliable results. Cloxacillin, a stable penicillinase-resistant penicillin, is used instead as a reliable surrogate marker for MRSA.
Staphylococcal food poisoning is an intoxication, not an infection β it results from ingesting pre-formed, heat-stable enterotoxin in contaminated food (meat, fish, milk products), not from bacterial multiplication in the gut. Symptoms are rapid in onset and usually self-limiting.
Quick Revision
10-Minute ReviewKey Takeaways
- Staphylococci are spherical, non-motile, Gram positive cocci in grape-like clusters.
- Coagulase positivity is the defining feature separating S. aureus from other staphylococci.
- S. aureus produces a wide range of toxins and enzymes that drive its virulence.
- MRSA arises via altered PBP2a receptors and is a major cause of nosocomial infection.
- Coagulase-negative staphylococci can be true pathogens in device-related infections.
- Laboratory diagnosis relies on Gram stain, culture, catalase, and coagulase testing.
Competency Checklist
Track Your MasteryReferences
- Collee JG, Fraser AG, Marmion BP, Simmons A. Mackie & McCartney Practical Medical Microbiology. 14th ed. Churchill Livingstone.
- Ananthanarayan R, Paniker CKJ. Textbook of Microbiology. 10th ed. Universities Press.
- National Institute of Open Schooling (NIOS). Microbiology β Staphylococcus, Module Notes.