Staphylococcus aureus is a common human pathogen that can cause localized or disseminated infections. It is a Gram-positive coccus that grows in clusters, producing pigmented colonies. S. aureus virulence factors include cell surface proteins like protein A and coagulase, as well as exotoxins like hemolysins and enterotoxins. It is an opportunistic pathogen capable of causing skin and soft tissue infections as well as more serious conditions if it spreads systemically. Resistance to antibiotics like penicillin and methicillin has increased its importance as a human pathogen.

1. By – Dr.D.W.Deshkar
Assistant Professor Dept.of Microbiology
D.Y.Patil Medical College, Kolhapur.

2. • Staphylococci are Gram positive cocci arranged in grape like clusters.
• They are ubiquitous.
• > 45 known species of which many form part of normal commensal
flora of the humans.

3. • Are first observed in human pyogenic lesions by von Recklinghausen (1871)
• Sir Alexander Ogston named it as “Staphylococcus” { Staphyle in Greek
means ‘bunch of grapes’ & kokkos means berry} (1880).
• Rosenbach (1884) named TWO species based on the pigmentation of
colonies as Staphylococcus aureus ( golden yellow colonies) & Staphylococcus
albus( white colonies)
• Passet (1885) named a THIRD species as Staphylococcus citreus (lemon
yellow colonies).
S.aureus S.albus S.citreus

4. • S.aureus – Most imp. Human pathogen. Commonly causes localized suppurative
lesions in humans. Can also cause disseminated infections.
• Their ability to develop resistance to Penicillin & other antibiotics increased its
importance as human pathogen.
• Other clinically imp. Species include S.epidermidis, S.lugdenensis, S.haemolyticus,
S.hominis – they mostly cause health care associated infections. S.saprophyticus –
causes U.T.I. in young women. ( Collectively called “Coagulase negative
Staphylococci – CONS).
• S.aureus – Are Coagulase positive.

5. • MORPHOLOGY :
• Gram positive ,Spherical cocci 1m in diameter ,
characteristically arranged in grape like clusters.
• Cluster formation – due to cell division occurring in three
planes & daughter cells tending to remain in close proximity.
• Are non sporing & non motile.
• Few strains have microscopically visible capsule.
• Many non capsular strains have small amount of capsular
material on their surface.
• In presence of penicillin they may change to L forms.

6. • CULTURAL CHARACTERS :
• Specimens are inoculated on various media , incubated
at 370C aerobically overnight.
• Nutrient Agar – Colonies are 1 -3 mm in size, circular
smooth, convex , opaque & easily emulsifiable. Most
strains produce non diffusible golden yellow pigment
(made up of  - carotene ). Pigmentation can be
enhanced by prolonged aerobic incubation at 20 – 250C
or by incorporation of 1% glycerol in the medium
(Tween agar). Grown anaerobically, colonies are smaller
& grayish in colour.

7. • CULTURAL CHARACTERS :
• Nutrient Agar Slope : It produces confluent
growth, looks like oil paint appearance.
• Blood Agar : Colonies are similar to that on
nutrient agar, in addition surrounded by a
narrow zone of  haemolysis ( best observed in
sheep blood agar).
• MacConkey Agar : Small pink colonies are
produced due to lactose fermentation.

8. • CULTURAL CHARACTERS :
• Liquid Medium : (e.g. peptone water) – It
produces uniform turbidity.
• Selective Media : Useful when Staphylococci
are scanty.
1. Mannitol Salt Agar : Contains nutrient agar
with 7.5% of NaCl & phenol red as indicator. All
Staphylococci can grow at 7.5% salt, S.aureus
produces yellow coloured colonies, due to
mannitol fermentation.

9. • CULTURAL CHARACTERS :
• Selective Media : Useful when Staphylococci
are scanty.
2. Salt Milk Agar : Contains nutrient agar, 6.5%
NaCl & 10% skimmed milk.
3. Ludlam’s Medium : Contains lithium chloride &
tellurite - Produce small black colonies.
4. Phenolphthalein Phosphate Agar : S.aureus
produces phosphatase enzyme which liberates
free phenolphthalein & produce pink color
when exposed to NH3 vapors.

10. • RESISTANCE :
• Are uniformly resistant to lysozymes.
• Are susceptible to Lyostaphin – a mixture of enzymes produced by strains of S.epidermidis.
• Were uniformly sensitive to penicillin in pre – antibiotic era with few strains producing
• enzyme penicillinase.
• Soon after the clinical use of penicillin, resistant strains began to emerge, first in hospitals
& then in community.

11. • TYPE OF RESISTANCE :
1. BETA – LACTAMASE – MEDIATED – It is mediated through
production of  - lactamase or penicillinase which inactivates the
beta lactam ring . These are inducible enzymes & are plasmid
mediated. They can be transmitted through transduction or
conjugation. Penicillinase producing strains remain sensitive to
penicillinase – resistant penicillines such as methicillin, cloxacillin.
While the beta lactamase producing strains are sensitive to beta
lactamase inhibitor combinations like amoxicillin – clavulinic acid.

12. • TYPE OF RESISTANCE :
2. ALTERED TARGET SITE PBP2a : Alterations in the penicillin – binding – protein PBP2a &
changes in bacterial surface receptors reduces binding affinity of beta lactam antibiotics
to cells. This mechanism imparts resistance to all beta lactam antibiotics & has been
named methicillin – resistant S.aureus (MRSA) as it was found to be resistant to
penicillins like methicillin & oxacillin. These strains also show resistance to
erythromyecins, tetracyclines, aminoglycosides & cause outbreaks of hospital infection.

13. • TYPE OF RESISTANCE :
• MRSA : Regulated by a set of chromosomal genes called “ staphylococcal cassette
chromosomal mec genes (SCC mec), especially the mec A gene. Based on the type of
these genes, the MRSA strains are divided as
• HOSPITAL ACQUIRED MRSA ( HA MRSA) - SCC mec has Type I, II & III mainly & are multi –
drug resistant.
• COMMUNITY – ACQUIRED MRSA ( CA MRSA) – SCC mec is Type IV mainly. These strains
are less resistant, more likely to produce PVL toxin & more transmissible.
• TOLERANCE TO PENICILLIN – The bacterium is only inhibited but not killed by the
antibiotic. It is demonstrated by large difference in minimum inhibitory conc. (MIC) 7
minimum bactericidal conc.(MBC) of penicillin in vitro.

14. • TYPE OF RESISTANCE :
• VANCOMYCIN RESISTANCE (VRSA) : The strains that have resistance to vancomycin are
emerging in some parts of world. These strains possess both vanA & mec A genes. Such
strains remain susceptible to linezolid only.
• VANCOMYCIN INTERMEDIATE RESISTANCE (VISA) : These strains have decreased
susceptibility to vancomycin by in vitro microdilution tests. They do not carry any
resistance genes, but have a thickened cell wall. They are isolated from patients who are
on prolonged vancomycin treatment but show failure to treatment with vancomycin.

15. • PATHOGENICITY & VIRULENCE :
• Staphylococci produce two types of diseases : infections & intoxications.
• In infection – the cocci gain access to damaged skin , mucosal or tissue sites, colonise by
adhering to cells or extracellular matrix, evade host defense mechanisms & multiply &
cause tissue damage.
• In intoxications – the disease is caused by the bacterial toxins produced either in the
infected host or preformed in vitro.

16. • VIRULENCE FACTORS :
• CELL ASSOCIATED FACTORS –
• Cell associated polymers –
1. Peptidoglycan : Cell wall polysaccharide peptidoglycan confers rigidity & structural
integrity to the bacterial cell. It activates the complement & induces release of
inflammatory cytokines.
2. Teichoic acid : It is an antigenic component of cell wall which helps adhesion of the cocci
to the host cell surface & protects them from complement mediated opsonisation.
3. Capsular polysaccharide : surrounding the cell wall inhibits opsonisation.

17. • VIRULENCE FACTORS :
• CELL ASSOCIATED FACTORS –
• Cell Surface Proteins –
1. Protein A : Present in most strains of S.aureus. It has numerous biological properties
including chemotactic, antiphagocytic, & anti – complementary effects. It also induces
platelate damage & hypersensitivity. Protein A binds to the Fc terminal of IgG molecules
(except IgG3),leaving Fab region free to combine with its specific antigen. Protein A
bearing staphylococci coated with any IgG antiserum will be agglutinated if mixed with
the corresponding antigen. This procedure is called co – agglutination. It has many
applications such as streptococcal grouping & gonococcal typing.

18. • VIRULENCE FACTORS :
• CELL ASSOCIATED FACTORS –
• Cell Surface Proteins –
2. Clumping Factor : It is bound coagulase which is responsible for slide coagulase test.
3. Protein Receptors : Staphylococci possesses protein receptors for many mammalian
proteins such as fibronectin, fibrinogen, IgG & C1q. These help in adhesion of staphylococci to
host cells & tissues.

19. • VIRULENCE FACTORS :
• EXTRACELLULAR ANTIGENS –
1. Coagulase : This is an enzyme which brings about clotting of human or rabbit acts with
plasma. It acts with a coagulase releasing factor (CRF) present in plasma, binding to
prothrombin and converting fibrinogen to fibrin. It is the basis of tube coagulase test.
2. Lipid hydrolase : Staphylococci produce a no. of lipases which help them infect the skin &
subcutaneous tissues
3. Hyaluronidase : It breaks down the connective tissue. Staphylokinase( fibrinolysin), fatty
acid modifying enzymes & proteases help in initiation & spread of infection.
4. Nuclease : A heat stable DNase is a characteristic component of S.aureus which helps in
the identification of the organism.

20. • VIRULENCE FACTORS :
• TOXINS – Cytolytic toxins are membrane – active substances, consisting of four hemolysins
& a leucocydin :
1. Alpha Hemolysin – Most imp. It is a protein inactivated at 700 C, but reactivated
paradoxically at 1000 C. This is because at 60 - 700 C, the toxin combines with a heat labile
inhibitor which is denatured at 1000 C, leaving the toxin free. Alpha toxin lyses rabbit
erythrocytes, but less active against sheep & human red cells. It is also leucocidal,
cytotoxic, dermonecrotic, neurotoxic & lethal. It is toxic to macrophages, lysosomes,
muscle tissues, the renal cortex & the circulatory system.

21. • VIRULENCE FACTORS :
• TOXINS :
2. Beta Hemolysin – Is a sphingomyelinase,hemolytic for sheep cells but not for human or
rabbit cells. It exhibits a hot – cold phenomenon, the hemolysis being initiated at 370C, but
becoming evident only after chilling.
3. Gamma Hemolysin – Is composed of two separate proteins, both of which are necessary
for hemolytic activity.
4. Delta Hemolysin – It has detergent like effect on the cell membranes of erythrocytes,
leucocytes, macrophages, & platelates.

22. • VIRULENCE FACTORS :
• TOXINS : PANTON – VALENTINE LEUCOCIDIN (PVL) : It is also a two component toxin, like gamma lysine. It is
composed of the S & F components. It is grouped as synergohymenotropic toxins. It is being associated with
CA MRSA.
• ENTEROTOXIN – Responsible for Staphylococcal food poisoning – nausea, vomiting & diarrhea 2 – 6 hrs. after
consuming food contaminated by the preformed toxin. The toxin is relatively heat stable, resisting 1000C for
10 – 40 min. depending on the conc. of the toxin & nature of the medium. About ⅔ strains of s. aureus
growing on carbohydrate & protein foods, secrete the toxin. Meat & fish or milk & milk products cooked &
left at room temp. after contamination with Staphylococci, for enough time for the toxin to accumulate are
the common items responsible. Source of infection – food handler who is carrier. Recovery in a day or so.
Eight antigenic types of enterotoxins – A,B,C1 – 3 , D,E,& H. Are formed by toxigenic strains . Toxin acts on ANS
to cause the illness.

23. • VIRULENCE FACTORS :
• TOXINS : TOXIC SHOCK SYNDROME TOXIN (TSST) : A potentially fatal multisystem disease
presenting with fever, hypotension, myalgia, vomiting, diarrhea, mucosal hyperemia, & an
erythematous rash which desquamates .Associated with infection of mucosal sites by toxic
shock syndrome toxin producing S.aureus belonging to bacteriophage group – 1 , TSST – 1.
Staphylococcal enterotoxins & TSST – 1 are superantigens, which are potent activators of T
Lymphocytes. These toxins without any help from MHC molecules directly bind to V
domain of T cell receptors and stimulate large no. of T cells releasing large quantities of
cytokines that causes disregulated immune response. With release of IL – 1 & IL -2 , tumor
necrosis factor & interferon  there ensues multisystem failure .

24. • VIRULENCE FACTORS :
• TOXINS : EXFOLIATIVE (EPIDERMOLYTIC) TOXIN : Also K/a ET or exfoliatin is responsible
for the staphylococcal scalded skin syndrome (SSSS) an exfoliative disease in which the
outer layer of the epidermis gets separated from the underlying tissues.
• Severe form of SSSS – k/a Ritter’s disease.
• Milder forms of SSSS – are pemphigus & bullous impetigo.

25. • BIOCHEMICAL REACTIONS –
• Ferment glucose, maltose, lactose, sucrose &
mannitol.
• Are catalase positive.

26. • BIOCHEMICAL REACTIONS – Coagulase test – Two methods
- Tube coagulase test – detects free coagulase ( extracellular
product) of S.aureus. 0.1 ml of broth culture or agar culture
suspension of the isolate is added to 0.5 ml of human or rabbit
plasma in a narrow test tube. EDTA, oxalate or heparin – used
as anticoagulant for preparing the plasma. Citrate is not used
as it is utilized by some organisms causing false positive results.
Positive & negative controls set up. The tubes are incubated in
water bath at 370C for 3 – 6 hrs. If positive the plasma clots &
does not flow when the tube is tilted. Continued incubation is
not recommended as the clot may be lysed by the fibrinolysin
formed by some strains

27. • BIOCHEMICAL REACTIONS –
• Coagulase test – Two methods
- Slide coagulase test – detects bound
coagulase ( clumping factor) – The isolate
is emulsified in a drop of saline on a slide.
After checking for absence of
autoagglutination, a drop of human or
rabit plasma is added to the emulsion &
mixed. Prompt clumping of the cocci
indicates a positive test. Positive &
negative controls are set up.

28. • BIOCHEMICAL REACTIONS –
• DNase Test : On DNA agar, a clear halo is produced
surrounding the colonies of S.aureus, due to its
ability to digest DNA.
• Phosphatase Test : This test is positive for S.aureus,
S.epidermidis . Organism inoculated on
phenolphthalein diphosphate agar & later the
colonies grown are exposed to ammonia vapor-
S.aureus – splits phenolphthalein diphosphate in the
media – releases free phenolphthalein – reacts with
ammonia vapor – colonies turn pink .

29. • CLINICAL MANIFESTATIONS :
• Skin and Soft tissue infections :
1. Folliculitis – Infection of hair follicles
2. Furuncle (Boil) – Painful pustular lesion in moist regions due to
infection of hair follicles.
3. Carbuncle : Severe, painful lesion in the lower neck region,
extending to deeper subcutaneous tissue

30. • CLINICAL MANIFESTATIONS :
• Skin and Soft tissue infections :
4. Impetigo – Occurs in children ,appear as red sores on the face that
bursts & develop s into honey colored crusts.
5. Surgical site infection -
6. Cellulitis (Infection of skin & subcutaneous tissue.

31. • CLINICAL MANIFESTATIONS :
• Skin and Soft tissue infections :
7. Hidradenitis suppurativa - A recurrent follicular infection in areas
rich in apocrine glands such as axilla.
8. Botryomycosis - It is mycetoma like condition, characterized by
subcutaneous swelling, sinuses, & discharge containing granules.

32. • CLINICAL MANIFESTATIONS :
• Musculoskeletal Infections :
1. Septic arthritis involving Knee, shoulder, hip joints & phalanges.
2. Osteomyelitis – In children – long bones & in adults – vertebrae.
3. Pyomyositis ( Skeletal Muscle infection) – In tropics & in HIV
infected people.
4. Abscess – Psoas abscess & epidural abscess.

33. • CLINICAL MANIFESTATIONS :
• RESPIRATORY TRACT INFECTIONS :
• Ventilator associated pneumonia in adults
• Septic pulmonary emboli
• Post viral pneumonia
• Empyema & Pneumothorax
• Pneumatocele
• BACTEREMIA –
• Septic shock, central line associated infection, Infective endocarditis.

34. • CLINICAL MANIFESTATIONS :
• URINARY TRACT INFECTION :
• TOXIN MEDIATED ILLNESS –
- Toxic shock syndrome
- Food poisoning
- Staphylococcal scalded skin syndrome
• INFECTIONS ASSOCIATED WITH CA MRSA –
- Necrotizing pneumonia
- Fulminant sepsis – Waterhouse – Friderichsen syndrome.
- Necrotizing fascitis