Lab 6 Streptococci All are Gram Positive Cocci arranged in chains short or long All are Catalase ve ve ve Gram Stain Staphylococci Streptococci Culture Fastidious organism ID: 927007
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Slide1
Streptococcus
Basmah almaarik Lab # 6
Slide2Streptococci
All are Gram Positive Cocci arranged in chains (short or long)
All are Catalase -
ve
+
ve
-
ve
Slide3Gram Stain
StaphylococciStreptococci
Slide4Culture
Fastidious organism no growth on NAGrows well on BAPFacultative anaerobe
Grow at 37C need 24 to 48 hour .
Slide5Streptococcus
ClassificationAlpha hemolyticS. viridansS.
pneumoniae
Beta hemolytic:
Group A, B, C, G & D
Lancefield grouping according to antigens in their cell wall
Non-hemolytic:
Enterococcus (formally known as Group D)
Slide6Hemolytic Streptococci
Alpha hemolysis on BAP (partial hemolysis).Beta hemolysis on BAP(complete hemolysis).
No hemolysis
on BAP
Slide7Beta Hemolytic Streptococci
Lancefield grouping depending on antigens in their cell wall, they are classified into groups : A, B, C, D, F, and G.Facultative anaerobes, grows at 37˚C for 24h.Need 5-10% CO2
Slide8Gram Stain
Slide9Beta Hemolytic Streptococci
Group A (S. pyogenes).BacitracinGroup B (
S. agalactiae
)
CAMP test.
Hippurate
hydrolysis.Group D (Enterococcus)
Grow in high salt conc.Esculin hydrolysis
Tolerate high tempreture
Slide10Beta Hemolytic Streptococci
S. Pyogenes(Group A Strep)S. Agalactiae(Group B Strep)
Catalase –ve
.
Bacitracin
sensitive.
Catalase
–
ve.Bacitracin resistant .
Slide11S. agalactiae(Group B Strep)
CAMP test:Streak a Staph.aureus across a blood agar plate.
Inoculate test organism at right angles to it. The tested organism must not touch the staphylococcal
inoculum
.
37˚C X 24 hr
Slide12S.
agalactiae(Group B Strep)S.aureus produce beta-lysin.
S.agalactiae produce extracellular protein CAMP factor. Both react together
synergeticly
to produce enhanced
haemolysis
arrow shape haemolysis.
Slide13CAMP test +ve and -ve
Slide14S. agalactiae(Group B Strep)
Hippurate hydrolysis test : hippuratehydrolase enzyme hydrolizehippurate
benzoic acid + glycine
We add
ninhydrin
reagent which will react with
glycine
to give us purple color.
Slide15S. agalactiae(Group B Strep)
Bacterial suspenstion in 1% sodium hippurateIncubate 2 hours at 37˚CAdd ninhydrine
reagent.Incubate for 10 min.
Purple color
+
ve
No purple color -
ve
Slide16Enterococcus (Group D Strep)
Grow at wide range of temperature 10˚C - 45˚C.Mainly non hemolytic on blood agar but can show alpha or beta-haemolysis.
Non-fastidious can grow on nutrient agar.
Slide17Grow in high slat concentration
Inoculate the organism in a tube of broth containing 6.5% NaCl.Incubate overnight at 37oC.Turbid growth can tolerate high salt conc
+ve
Clear no growth can’t grow in high salt conc. -
ve
Slide18Esculin Hydrolysis
The organism can hydrolyze esculin and produce black color in the media. It can also grow in the presence of 40% bile salt.
Slide19Esculin Hydrolysis
Black color
esculin
hydrolysis +
ve
Slide20Growth in high heat.
Make a suspension of the organism.Inoculate on a blood agar (0 time) Incubate organism suspension in water bath 45˚C X 10 min sub-culture to BAPRepeat step 3 tow more times.
Then incubate the BAP 37 ˚C x 24 hr.
Slide21Rapid identification test for beta-heamolytic
streptococci
Slide22YOUR WORK FOR TODAY
Do gram stain and show it to the instructorDo camp test with group A and BDo bacitracin sensitivity with group ADo esculin hydrolysis test with group D
Do salt tolerance test with group D
See instructors
hippurate
test (
Demonestration)See instructors rapid identification kit for Beta-heamolytic stryptococci
(Demonestration)