IDENTIFICATION OF STAPHY Flashcards
demonstrate the presence of catalase
an enzyme that catalyzes the release of oxygen from
hydrogen peroxide (H2O2)
Catalase Test
To distinguish Staphylococci spp. And Micrococci
spp. (positive)
Catalase Test
percentage of hydrogen peroxide or H2O2 in catalase test used for the routine culture
3%
percentage of hydrogen peroxide in catalase test used for detection of catalase in an aerobe
15%
During tests bacteria protect themselves from the little effect of hydrogen peroxide which is
accumulated as an end product of aerobic
carbohydrate metabolism
Catalase Test
positive result of catalase test
bubble formation or effervescence
Best single pathogenicity test for staphylococcus
Coagulase (Slide or Tube Method)
is a biochemical test that is used
to differentiate staphylococcus aureus from other
staphylococci species like staphylococcus
epidermidis and staphylococcus saprophyticus
Coagulase (Slide or Tube Method)
a test used to detect the ability to produce the
coagulase. coagulase is an enzyme-like protein
and causes plasma to clot by converting
fibrinogen to fibrine
Coagulase (Slide or Tube Method)
2 methods of coagulase test
Slide Method (screening test)
Tube Method (confirmatory test)
specimen used in coagulase test
plasma (plasma, 0.5 mL rabbit’s plasma for tube)
among the 2 methods of coagulase test, which one detects BOUND COAGULASE (CLUMPING FACTOR)?
Slide Method (screening test)
positive results of slide method (screening test)
agglutination
among the 2 methods of coagulase test, which one detects UNBOUND COAGULASE (staphylocoagulase)?
Tube Method (confirmatory test)
incubation time of tube method coagulase test
2hrs, 4hrs, and another 24hrs
positive result of tube method coagulase test
coagulum/clot
medium used in O/F reactions
O/F glucose medium
O/F glucose medium is also called as
Hugh Leifson medium
a method for biochemical
differentiation of microbes it is used to detect the
oxidation or fermentation of carbohydrates by
bacteria.
Oxidation-Fermentation (O/F) Reactions
In O/F reactions, _____ferment glucose or
fermentative, whereas _____ fail to produce
acid under anaerobic conditions.
Staphylococcus , micrococci
sugars added in O/F reaction’s medium aside from sucrose, lactose, and glucose
cylos, mannitol, and maltose
In O/F reactions, ___
utilization of the carbohydrate will result in acid
production or color yellow in the open tube only
Oxidative
In O/F reactions, ____ utilization of the carbohydrate will
result in acid production or yellow in both open
and closed rooms
Fermentative
___ organism, meaning
absence okay it will not produce acid in either of
the tube
asaccharolytic
what is added in hugh leifson medium to serve as a barrier for oxygen
mineral oil
In O/F reactions, if the closed tube turns yellow it means?
it is fermentative
In O/F reactions, if the open tube turns yellow it means?
oxidative
In O/F reactions among the genera if both tube turns yellow, is it staph or micrococci?
staph
In O/F reactions, if only the open tube turns yellow. Is it staph or micococci?
micro
active chemical component of modified oxidase test
Tetramethyl-paraphenylene diamine dihydrochloride
is a reagent impregnated disk
recommended for use in qualitative procedures to
aid in the differentiation of staphylococcus and
micrococcus by the detection of the oxidase
enzyme
Microdase disk
a positive result in modified oxidase test
blue to purple to black complex (micro lang ang psoitive sa modified OXIDASE test)
medium used for mannitol fermentation
Mannitol Salt Agar
positive result of mannitol fermentation
yellow
negative result of mannitol fermentation
pink
a yellow positive result in mannitol fermentation indicated the presence of what
staphylococcus aureusq
indicator used in mannitol fermentation
Phenol red
principle behind the mannitol fermentation
If an organism can ferment mannitol, an acidic
byproduct is formed that will cause the phenol red
in the agar to turn yellow. Most pathogenic
staphylococci such as staphylococcus aureus will
ferment mannitol and most non-pathogenic
staphylococci will not ferment mannitol
medium used in DNAse test
DNAse Medium
indicator used in DNAse test
methyl green
what is added in DNAse test
1 N hydrochloric acid
positive result of DNAse test
zone of inhibition
a positive zone of inhibition in DNAse test identifies the presence of
staphylococcus aureus
a negative result in DNAse test identifies the presence of
Staphylococcus epidermidis
what percent of —- is used in gelatin medium for GELATIN LIQUEFACTION/HYDROLYSIS TEST
12%
principle behind dnase test
If the organism that
grows in the medium produces
deoxyribonuclease, it breaks down dna into
smaller fragments. When the dna is broken down
it no longer binds to the metal grid and the green
color fades and the colony is surrounded by a
colorless zone.
Gelatin is a protein derived from the connective
tissues of vertebrates that is ___
collagen
positive result in Gelatin Liquefaction/Hydrolysis Test
liquefaction
Presumptive identification of ______ is accomplished by testing for
novobiocin susceptibility
staphylococcus saprophyticus
Staphylococcus saprophyticus is
__to novobiocin whereas most other coNs
are susceptible
resistant
___ microgram of novobiocin disk is used in novobiocin disk
5
STEPS IN COLLECTION OF BACTERIAL SAMPLE
- Use sterile cotton swab to collect bacterial sample in the skin/nasal cavity
- Inoculate the sample in MSA
- Incubate at 37C for 18-24 hours
- Observe for growth and color development
STEPS IN OXIDASE TEST
- Using a sterile loop/applicator stick, rub a colony into the oxidase strip
- Observe for color change
(+) Purplish to black
(-) No color change
STEPS IN CATALASE TEST
- Place 1 drop of 3% hydrogen peroxide reagent in a glass slide
- Emulsify a colony using applicator stick/inoculating loop
- Observe
(+) bubble formation
(-) no gas bubbles
STEPS IN DNASE TEST
- Using bacteria from a plate, inoculate the DNAse medium in a straight line
- Incubate the DNAse medium for 18-24 hours
- After incubation, add a small amount of 1N HCl to the plate, flooding the colonies.
- Discard the excess acid and observe the media surrounding the colonies.
(+) Clearing surrounding the colonies
STEPS IN DNASE TEST
. Using bacteria from a plate, inoculate the DNAse medium in a straight line
2. Incubate the DNAse medium for 18-24 hours
3. After incubation, add a small amount of 1N HCl to the plate, flooding the colonies.
4. Discard the excess acid and observe the media surrounding the colonies.
(+) Clearing surrounding the colonies
STEPS IN COAGULASE TEST - SLIDE
- Divide the slide into two sides using a marking pen. Label the left portion “T” (for test)
and the right portion “C” (for control) - Place a drop of plasma in the T portion of the slide
- Place a drop of saline in the C potion of the slide
- Emulsify few colonies into the T portion
- Emulsify few colonies into the C portion
- Mix well for 5 seconds and observe for clumping within 10 seconds
(+) clumping
(-) no clumping
*negative control should show no clumping
STEPS IN COAGULASE TEST - TUBE
- Add 0.5 mL of plasma into a tube
- Inoculate a loopful of bacteria
- Incubate for 2 hours and observe for clot formation
- If no clot is observed, incubate for another 2 hours and observe for clot formation
- If no clot is observed after a total of 4 hours incubation, incubate further up to 18-24
hours
(+) Coagulum
(-) No coagulum
