A case of S. aureus infection

8/11/2019 A case of S. aureus infection

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CASE NO. 5A CASE OF Staphylococcus aureus

INFECTIONEVANGELISTA | FABROS | FARILLAS

GASTARDO | ILAGAN


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CASE PRESENTATION

An 18 month old female infant was rushed to theemergency room because of difficulty of breathing .1 week ago, she was noted to have multipleabscess formation on her scalp. 3 days prior to

consult she had fever and cough. Consult was madein a nearby health center where carbocisteine andamoxicillin were prescribed. However cough andfever persisted until day of consult when she was

noted to have difficulty of breathing manifested aschest indrawing with flaring of the alae nasi. Chestxray done in the emergency room revealedpneumatocoele formation.


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Staphyloccocus aureus

1. What is the most likely etiologic agent

involved?


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Characteristics

Gram-positve cocci occuring in clusters

Non-motile, non-sporeforming, facultative

anaerobe

Fermentation of glucose produces mainly lactic acid

Ferments Mannitol (distinguishes from S.

epidermidis)

Catalase positive

Coagulase positive


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Characteristics

Golden yellow colony on agar

Normal flora of humans found on nasal passages,

skin and mucous membranes

Pathogen of humans

Causes a wide range of suppurative infections, as

well as food poisoning and toxic shock syndrome


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Morphology

Kayser, Medical Microbiology 2005 Thieme, p. 231


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2. What biochemical tests can be requested to

assist you in differentiating this microorganism

from the rest of its relative species?

Biochemical tests that may be requested

include:

Catalase Test

Coagulase Test

DNAse Test


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Catalase Test

This is performed to differentiate between

genus Staphylococcus and Streptococcus:

Place a drop of H2O2 on a clean glass slide.

Take a small portion of test organism using a

sterile wooden stick or plastic loop and mix with

the drop of H2O2.

Observe for immediate bubbling (gas production).


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Catalase Test

Negative (no bubbles)

Positive (bubbles)


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Coagulase Test

Coagulase

An extracellular protein which binds to

prothrombin in the host, forming

staphylothrombin

Clots blood plasma surrounding the cocci;

enabling bacteria to resist phagocytosis by the

hostsimmune system Good indicator of pathogenic potential ofS.

Aureus


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Two forms of Coagulase

Bound Coagulase/Clumping Factor

Bount to the cell wall

Adsorbs fibrinogen from the plasma and alters it

so it precipitates on the staphylocci, causing them

to clump (cell agglutination)

Detected by the slide coagulase test


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Two forms of Coagulase

Free Coagulase

Liberated by the cell wall

Reacts with a substance in plasma to form a fibrin

clot

Detected by tube coagulase test (together with

bound coagulase)


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Two Types of Coagulase Test

Coagulase Slide Method

Emulsify a colony of Staph. sp. From the mediaona drop of saline solution on a glass slide

Mix a loopful of fresh plasma with the emulsifiedspecimen

Observe in the next 5 seconds for the appearanceof white clumps on the slide, indicating a positive

result


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Two Types of Coagulase Test

Coagulase Tube Method

Place 0.5 ml diluted plasma into a sterile test tube

Inoculate a loopful of bacteria into the dilute plasma

Incubate at 35

o

C Observe for clotting at 1, 2 and 24 hour

*False Negatives can occur when clots were only

observedat the 24thhour because some strains thatproduce coagulase also produce fibrinolysin, which

dissolves clot.


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DNAse Test

This is done to detect production of DNAse enzyme

Inoculate a single colony of test organism on DNAseagar plate.

Incubate the plate overnight at 37

o

C. Cover the surface of the agar plate with HCl

(Hydrochloric acid)

Look for clear zones around the colonies within 5

minutes after adding the acid. If clear zones seen around the colonies the test is

DNAse +ve


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DNAse negative DNAse positive


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3. Cite the characteristics of the clinically significant

species in this genus in terms of hemolysis and tests

used for presumptive diagnosis.

Test(s) S. aureus S. epidermidis S.

saprophyticus

Pigment Gold-white White-yellow White-yellow

Hemolysis + - -

Catalase + + +

Coagulase + - -

Novobiocin Susceptible Susceptible Resistant

Mannitol + - -

Mannose + + -

Trehalose + - +


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3. Cite the characteristics of the clinically significant

species in this genus in terms of hemolysis and tests

used for presumptive diagnosis.

HEMOLYSISTEST FOR PRESUMPTIVE

DIAGNOSIS

S. aureus -hemolysis (+) Coagulase TestS. epidermidis hemolysis (-) Trehalose fermentation

S. saprophyticus hemolysis(-) Mannose fermentation

(-) Novobiocin


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4. Name the culture media to be used for

definitive diagnosis and describe its colony

characteristics

Gram staining

PRELIMINARY IDENTIFICATION


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Definitive Diagnosis

Inoculation in culture media

4. Name the culture media...


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MANNITOL-SALT AGAR (MSA) pH of 7.4

Contains the following:

Casein digest

Animal tissue digest Beef extract

D-mannitol

NaCl

Phenol Red - indicator

Agar


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MANNITOL-SALT AGAR (MSA) Selectively favors growth of S. aureus because of

the 7.5% NaCl

S. aureus can ferment mannitol while other

Staphylococcal organisms can not.

Color change from red to yellow indicates

fermentation of mannitol.


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MANNITOL-SALT AGAR (MSA)


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Blood Agar and Chocolate Agar Enriched bacterial growth medium.

Includes:

Base with protein source

Soybean protein digest NaCl

5% sheep blood

Casein

dH2O Agar


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Blood Agar and Chocolate Agar Enzymes like hemolysin diffuse outward and

cause complete or partial destruction of RBCs in

the medium and complete Hgb denaturation

within cell products

Chocolate agar has similar contents except it

contains lysed RBCs instead of whole RBCs


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Blood Agar and Chocolate Agar Most staphylococci grow on agar media

containing peptone

Growth within 24 hours at 35 to 37 degrees

Celsius

S. aureus colonies appear YELLOW and exhibit -

HEMOLYSIS on sheep blood agar, but these may

also occur with other Staphylococcal species.


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Blood Agar and Chocolate Agar


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Other DiseasesBacteremia

Empyema

Endocarditis

Meningitis

Osteomyelitis

Pneumonia

Toxic Shock Syndrome

5. Name other diseases that are associated with this

organism; Include toxin-related diseases.


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Functions of Virulence Factors Evade phagocytosis by macrophages

Mediate adherence to host tissues Promote tissue destructionProduction of

toxins and hydrolytic enzymes

6. Name virulence factors associated with

this organism.


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Structural Constituents in the Bacteria

BACTERIAL

CAPSULE

Inhibits chemotaxis and phagocytosis;

Prevents proliferation of polymorphonuclear leukocytes

PEPTIDOGLYCAN

Elicits the production of endogenous pyrogen IL-1 and

opsonic antibodies by monocytes

Also a chemoattractant to leukocytes promoting

abscess formation

TEICHOIC ACIDS Peptidoglycan component that bind to fibronectin to

allow adherence to host tissues

PROTEIN A Binds to IgG and effectively prevents antibody-

mediated clearance of the bacteria

VIRULENCE FACTORS OF S. aureus


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Toxins

CYTOTOXINS

-hemolysinheterogeneous protein that acts on a broad

spectrum of eukaryotic cell membranes

-toxindegrades sphingomyelin and can affect RBCs

-toxindisrupts biological membranes and may be involve

in diarrheal diseases

-hemolysincan interact with Panton-Valentine Leukocidin

to lyse WBCs

EXFOLIATIVE

TOXINS (ETA and ETB)

Epidermolytic toxins that contain serine proteases to splitintercellular bridges of stratum granulosum yielding to

desquamation on staphylococcal scalded skin syndrome

ENTEROTOXINS

(from A to R)

Superantigens that stimulate T cell proliferation and cytokine

release

Stimulate release of inflammatory mediators Increase intestinal peristalsis and fluid loss leading to nausea

and vomiting

TOXIC SHOCK

SYNDROME TOXIN -1

Prototypical superantigen that stimulates proliferation of T

cells and release of cytokines, leading to leakage and

endothelial cell destruction


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Hydrolytic Enzymes

COAGULASE Enzymatically activate and initiate fibrin

polymerization

HYALURONIDASE Spreading factorhydrolyzes hyaluronic acids inconnective tissue

FIBRINOLYSIN Dissolves fibrin clots

LIPASES Hydrolyze membrane lipids

NUCLEASES Hydrolyze DNA

VIRULENCE FACTORS OF S. aureus


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7. Create a schematic diagram for the

laboratory identification of this organism.


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7. Create a schematic diagram for the

laboratory identification of this organism.


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END. THANK YOU!

A case of S. aureus infection

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