STAPHYLOCICCI
Lecture # 3
Staphylococcus sp.
Morphology: Gram positive cocci. In clusters
Culture: Facultative anaerobes Incubation 37ºC x 24 hr
Staphylococcus sp.
Coagulase +ve: Staphylococcus aureus.
Coagulase –ve: Staphylococcus saprophyticus. Staphylococcus epidermidis.
All Staphylococci are CATALASE +VE
CULTURE
Blood Agar: Creamy colonies 1-2 mm Beta-heamolysis
Blood Agar: Grayish white colonies No heamolysis
Coagulase +ve
Staphylococcus aureus
Coagulase -ve Staphylococci
CULTURE
MacConkey Agar: Pink colonies (lactose
fermenting)
MacConkey Agar: Only some are give Pink
colonies (lactose fermenting)
Coagulase +ve
Staphylococcus aureus
Coagulase -ve Staphylococci
CULTURE
Mannitol Salt Agar: Growth Change color from Red to
Yellow (Mannitol fermenting)
Mannitol Salt Agar: Growth No yellow color no
fermentation.
Coagulase +ve
Staphylococcus aureusCoagulase -ve Staphylococci
Differential: Staph
aureus can frement mannitol and give diffused
yellow color
Selective: Containing very high salt conc only staph can grow
Biochemical Tests
Biochemical Tests are:
1- Catalase
2- Coagulase
3-DNase
1. Catalse: 2H2O2 O2 + 2H2O
Streptococci vs. Staphylococci
Coagulase Test
2. Coagulase test: Fibrinogen in plasma Fibrin
A. Free (tube) coagulase:1. 1 ml pepton + 1 ml EDTA plasma2. Emulsify loop full of organism3. Incubate 30 min x 37ºC4. If negative incubate at RTº overnight
Coagulase Test
Coagulase Positive Coagulase Negative
Coagulase Test
B. Bound (slide) coagulase: 1. One drop of saline on clean slide2. Emulsify loopful of organism in saline3. Add one drop of plasma4. Mix rocking for 2 min
Coagulase Test(Latex agglutination test)
• Several latex agglutination test kits are available to identify Staphylococcus aureus.
• Latex particles coated with fibrinogen.• Rapid clumping of latex particles occurs
when mixed with colony material of S. aureus.
Biochemical Tests
3. DNase Test:DNase enzyme hydrolyze DNA
4. Spot-inoculate organism on a DNA containing media
5. Incubate 37ºC x 24 hr6. Flood plate with 1 mol/ml HCL sol
HCL will precipitate unhydrolyzed DNA4. Tip off the excess acid5. Clearing around colonies DNase positive strain6. No Clearing around colonies DNase negative
strain
MRSA (methicillin resistant Staphylococcus aureus)
• These strains are resistant to methicillin and related penicillins and are particularly difficult to treat because they are also resistant to most other common antibiotics.
• MRSA strains cause hospital infections, particularly wound infection and septicaemia.
• Vancomycin is often needed to treat MRSA infections.
Coagulase Negative Staphylococci
Novobiocin Disk:
Staphylococcus saprophyticus Resistant Staphylococcus epidermidis Sensitive
Micrococcus
Gram Positive Cocci in pairs and tetrads. Catalase +ve. Generally harmless, but can cause
disease in immunocompromised host.
Micrococcus
Blood agar or chocolate agar:
Small, non-hemolytic, variably pigmented (white, orange, yellow pink) colonies.
MacConkey:
no growth.
Your Work For Today.
1. Prepare Gram Stain, show it to the instructor.
2. Perform Catalase test.
3. Perform Coagulase test.
4. Perform DNA’s test.
5. Do subculture of the organism to:
Blood agar plate.
Mannitol Salt Agar.