Staphylococcus aureus and CONS Dr Ashna Ajimsha PG resident Department of Microbiology SGMC and RF Venjaramoodu Kerala

1. Staphylococcus

2. History
◦ 1897 – Von Recklinghausen – first observed in pus
◦ 1880 – Louis Pasteur – First cultured in liquid
medium
◦ Named by Alexander Ogston (1880)
-Staphyle (Greek) – Bunch of grapes
-Kokkos – berry
◦ Rosenbach – 2 species based on pigmentation
-Staph.aureus(golden yellow colonies)
-Staph.albus(white colonies)

4. General Features
Gram positive
cocci in grape-
like clusters
Catalase
positive

5. Staphylococcus aureus

6. MORPHOLOGY
◦ Catalase +
◦ Coagulase+
◦ Facultative anaerobe
◦ Non motile, non sporing
◦ Occasionally capsulated
◦ Spherical cocci, approximately 1ɥm in diameter
◦ Arranged in grape-like clusters
◦ Cluster formation – cell division in multiple planes, daughter cells remain
attached together
◦ Under influence of penicillin – change to L forms

7. Virulence Factors
Cell wall associated factors
Peptidoglycan Rigidity,shape,inflammatory response,Endotoxin
Teichoic acid-ribitol phosphate polymers Adhesion to mucosal surfaces, Inhibit opsonisation
Cell surface adhesins
-Clumping factor-Fibrinogen binding adhesin
-Fibronectin binding adhesin
Collagen binding adhesin
Adhesion, Clumping
Protein A Anti-complementary Chemotactic, Mitogenic, Inhibit
opsonisation
Induction of platelet damage
IgG (Fc region) Co-agglutination
Microcapsule Inhibit phagocytosis

8. Toxins
Membrane active toxins
 Hemolysins-α,β,γ,δ
 Leucocidin F & S toxins + γ hemolysin hemolytic and
leucocidal (Synergohymenotropy)
Epidermolytic toxin Staphylococcal scalded-skin syndrome
Enterotoxins Serotype A- MC, Superantigen
Self limiting food poisoning IP<6 Hrs
Preformed toxin vagus stimulation
Toxic shock syndrome toxin Super antigen
Risk - abscesses, osteomyelitis, post-surgical
wound infection

9. Hemolysins
-hemolysin Inactivated at 70C reactivated at 100C
Lethal, leucocidal, dermonecrotic, cytotoxic and
neurotoxic
Lyses rabbit RBCs, but less active against sheep
and human RBCs.
Β-hemolysin Sphingomyelinase
Lyses sheep RBC, but not human or rabbit RBC
Exhibits hot-cold phenomenon
-hemolysin Act together with leucocidin for hemolytic
activity.
Lyses rabbit, sheep and human RBCs
-hemolysin Surfactant action
Lyses rabbit, sheep, horse & human RBCs
Lethal, leucocidal and dermonecrotic

10. EXTRACELLULAR ENZYMES
Coagulase + CRF prothrombin to thrombin
Tube coagulase test
Heat stable thermonuclease specific to S.aureus
Deoxyribonuclease specific to S.aureus
Lipase & phospholipase Break down lipids
Protease
Staphylokinase(fibrinolysin) Breaks down fibrin clot---spread of infection
Hyaluronidase Breaks down connective tissue network

11. Pathogenesis
Metastatic spread- hematogenous
Evasion of host defence mechanisms
Anti-phagocytic -microcapsule and Protein A
- Inhibition of leukocyte migration
- Intracellular survival -formation of small colony variants
Invasion- with help of enzymes
Introduction into the tissue- minor abrasions or instrumentation adhere to tissue
Colonization-anterior nares, axilla and perineal skin

12. Clinical
Manifestations
◦ Skin & Soft tissue
infections
◦ Folliculitis,
Furuncle
◦ Carbuncle,
Impetigo
◦ Mastitis and breast
abscess
◦ Surgical site wound
infections
◦ Cellulitis
◦ Hidradenitis
suppurativa
◦ Botryomycosis

15. Musculoskeletal
Infections
◦ Septic arthritis
◦ Osteomyelitis
◦ Pyomyositis in HIV
◦ Psoas abscess
◦ Epidural abscess
Respiratory Tract
Infections
◦ Ventilator associated pneumonia
in adults
◦ Septic pulmonary emboli
◦ Post viral pneumonia
◦ Empyema and Pneumothorax
◦ Pneumatocele in neonates

16. Bacteremia and
its Complications
◦ Sepsis, septic shock
◦ Central line associated blood
stream infection
◦ Metastasis - kidney, joints, bone
and lung
◦ Native-valve endocarditis
◦ Prosthetic-valve endocarditis
◦ Intravenous drug use associated
endocarditis

17. UTI-secondary to
bacteremia
Toxin-Mediated
Illnesses
◦TSS
◦Food poisoning
◦SSSS
Infections Associated
with CA-MRSA
◦Necrotizing pneumonia
◦Purpura fulminans
◦Necrotizing fasciitis

18. Staphylococcal Gastroenteritis
Enterotoxin - expressed by 50% of S.aureus
Preformed heat stable toxin and resistant to gastric juice
Serotypes(A–E, G–I, R-T and V). Type A – Most common
Incubation period – 1-6 Hrs
C/F- nausea, vomiting and occasionally diarrhea, hypotension, and dehydration. No fever. Self limiting
within 8–10 hours.
Source - food handler
Food items - milk products, bakery food,custards, potato salad, or processed meats.

20. Staphylococcal scalded-skin syndrome (SSSS)
◦ Epidermolytic/exfoliative toxin –
◦ Two proteins- ET-A (chromosomal and heat stable), ET- B (plasmid coded, heat labile).
◦ More common in newborns and children
◦ Clinical Features-
- vary from localized tender blisters& bullae formation to exfoliation & separation of outer
epidermal layer leaving denuded underlying skin (Nikolsky's sign).
◦ Milder form - pemphigus neonatorum & bullous impetigo
◦ Ritter’s syndrome - Severe form in newborns
- Fever, lethargy, and irritability,poor feeding
- S.aureus bacteriophage group II

21. SSSS

22. TOXIC SHOCK
SYNDROME
◦ Toxic shock syndrome toxin (TSST)-TSST-1
& TSST-2
◦ TSST-1- Enterotoxin F/Pyrogenic exotoxin
C a superantigen
◦ Risk factors- vaginal tampons, abscesses,
osteomyelitis and post-surgical wound
infection.
◦ Pathogenesis- TSST absorbed into
circulation - non-specific T cells
stimulation - excessive cytokine
production -multisystem disease.

23. ◦ Clinical features- Fever, hypotension, mucosal hyperemia, vomiting, diarrhea,
confusion, myalgia, abdominal pain and erythematous rashes  rapid
involvement of liver, kidneys, GIT and/or CNS
◦
◦ Diagnosis-
- Detection of TSST - latex agglutination test and ELISA
- TSST genes 1 and 2 – PCR
◦ Treatment- Clindamycin (MSSA), Vancomycin (MRSA)

25. LABORATORY
DIAGNOSIS
Infection Specimen
Suppurative lesion Pus, wound swab
Respiratory
Infections
Sputum
Urinary tract
infection
Mid stream urine
PUO, Bacteremia Blood
Food poisoning Feces, vomitus, food
Carriers Nasal & perianal swab

26. Microscopy
Direct smear microscopy
Gram staining -Gram positive cocci
in clusters with pus cells

27. CULTURE
◦ Nutrient agar – 1-3mm
-Golden yellow non diffusible
pigments (Beta carotene)
- Circular,smooth,convex,opaque
◦ Blood agar – colonies same as NA
+beta hemolysis

28. ◦ MacConkey agar- Small pink LF colonies
◦ Liquid medium - uniform turbidity
◦ Selective media
- Mannitol salt agar – yellow colonies -mannitol
fermentation 
- Salt milk agar
- Ludlam’s medium

29. BIOCHEMICAL
TESTS
S.aureus positive and
CoNS mostly negative
 Coagulase test
o Tube coagulase
o Clumping factor
 Heat stable thermo nuclease test
 DNase test
 Phosphatase (also produced by
S.epidermidis)
 Golden yellow pigmentation
 Hemolysis on blood agar
 Mannitol fermentation
 Black coloured colonies on
potassiumtellurite agar
 Gelatin liquefaction
 Protein A detection

30. Tube coagulase Slide coagulase
Due to coagulase enzyme Due to clumping factor
Requires CRF in plasma Does not require CRF in
plasma
Done in tube Done in slide
Positive if clot is formed Positive if clumps are
formed
Coagulase enzyme has eight
serotypes
Clumping factor has one
serotype
S.lugdunensisgives a
negative result
S.lugdunensisgives a
positive result
Both tube and slide coagulase positive for S.aureus,
S.hyicus, and S.intermedius

31. Typing of
S.aureus
◦Phenotypic methods
- Bacteriophage typing
- Antibiogram typing.
◦Genotypic methods -
PCR-RFLP

32. DRUG RESISTANCE IN S.aureus
1)Production of β lactamase/Penicillinase
enzyme
Cleave β lactam rings--- resistance to
◦Plasmid coded, transduction
◦Overcome by adding BLI’s- Sulbactam/
Clavulanic acid
β lacatam Antibiotics

33. 2) By alteration of
PBP – MRSA
◦ mec A gene – chromosomal
– PBP-2a - resistant to all β
lactam antibiotics

34. MRSA
Detection of MRSA
- Disc diffusion test - oxacillin/cefoxitin
discs
- Oxacillin screening agar 6Micro gm
/ml Oxacillin and 2-4% NaCl to MHA –
incubated @ 300C For 24 hrs
- PCR detecting mecA gene
- Latex agglutination for detecting
PBP2a

36. Treatment of MRSA
- Vancomycin - DOC
- Alternative drugs - teicoplanin, linezolid, daptomycin and
quinupristin/dalfopristin
- Nasal carriers- 2% Mupirocin ointment
- Other non-beta-lactams if sensitive by AST
3. Resistance to vancomycin (VRSA and VISA)

37. Control
measures
Proper hand washing
Screening of MRSA carriers
Treatment of carriers
Stoppage ofantibiotic misuse
Infection control measures

38. COAGULASENEGATIVE
STAPHYLOCOCCUS(CONS)

39. STAPHYLOCOCCUS
EPIDERMIDIS
Most common CONS (75-80%)
Normal flora - skin, oropharynx and vagina
Risk - prosthetic devices
Pathogenesis-Initial adhesion
- Colonization (Biofilm)
Infections: Prosthetic-device related infections endocarditis
with insertion of valvular prosthesis and ventricular shunt
infections
-Stitch abscess

40. Staphylococcus
saprophyticus
-UTI in sexually active
young women
--
Unlike other
Staphylococci resistant to
novobiocin
S.lugdunensis S.schleiferi
-Native-valve endocarditis
and osteomyelitis
-Enhanced pathogenesis
due to virulence factors- -
Clumping factor & lipase
(absent in other CoNS)