Biochemical Testing Flashcards
TSIA - purpose
- Used to differentiate Enterobacteriaceae
- Differentiates them from other Gram-negative rods such as Pseudomonas
All other sugar must be converted to glucose before they enter the EMP. (T/F)
True. All other sugar must be converted to glucose before they enter the EMP.
Glucose utilization occurs anerobically on the slant where O2 is not available and on the butt where there is aerobic conditions. (T/F)
Glucose utilization occurs aerobically on the slant where O2 is available and on the butt where there is anaerobic condition.
Incubation period for TSIA
6 hours
Describe different processes occurring in TSIA medium for:
- Lactose/sucrose fermenters
- Non lactose/sucrose fermenters
- Gas and/or H2S producers
Lactose/sucrose fermenters
1. Once the glucose fermenting bacterium has reduced all of the available glucose to pyruvate, it will further metabolize pyruvate via TCA cycle to produce acid end products.
2. The acid in the medium cause the ph indicator phenol red, to change its colour to yellow. Both the slant and butt with a glucose fermenter will appear yellow.
3. After the depletion of glucose, the organism (that has the correct enzymes) starts to degrade lactose/sucrose
and continues to make acid end products.
Non lactose/sucrose fermenters
4. If the organism cannot use lactose/sucrose in the medium, it must shift to protein utilization as energy source.
5. As the protein breakdown occurs only in aerobic condition, the byproduct of protein and amino acid metabolism i.e NH3 changes the pH environment of the medium from the neutral to alkaline in the slant region where the condition is aerobic.
Gas and H2S producers
6. Gas production can be detected when holes are formed or the medium is broken into several fragments.
When do we call a TSIA rxn acid/acid?
– Butt and slant that still remain yellow for lactose and sucrose fermentors (degrades) after 18 hours of incubation and the reaction is called acid/acid.
When do we call a TSIA rxn alkali/acid
– The colour of slant appears red & is called alkali/acid while the colour of the butt is still yellow due to anaerobic glucose breakdown.
Production of what causes blackening of medium in TSIA?
– H2S production by organism results in the blackening of the medium.
H2S is produced from an ingredient in TSIA medium which is?
Sodium thiosulphate
An ingredient called sodium thiosulphate is contained in TSIA medium, what does it combine with to form ferrous sulphide?
This colour is due to the production of H2S from an ingredient of the medium, sodium thiosulphate,
- which then combines with another component of the medium, ferrous ammonium sulphate resulting in the formation of black, insoluble compound, ferrous sulphide.
What black, insoluble compound is formed as a result of H2S production in TSIA?
Ferrous sulphide
Red/yellow colour in TSIA rxn indicates?
Alkali/Acid
Utilizes Glucose only and peptone
Yellow/Yellow colour in TSIA rxn indicates?
Acid/Acid
Utilizes Glucose, Lactose and sucrose
Red/Red colour in TSIA rxn indicates?
Alkali/Alkali
No sugars are utilized, utilize peptone
H2S gas production is indicated by ?
Blackening of media
Gas production is indicated by?
Bubbles in butt
E. coli gives acid/Acid with gas production without H2S production. What does this mean?
This means E. coli utilizes all sugar anaerobically with production of gas during fermentation.
Pseudomonas spp gives alkaline/alkaline (Red/Red), what does this mean?
Pseudomonas spp does not utilize any sugar
Escherichia, Klebsiella, Enterobacter
Acid slant/ Acid butt (A/A)
Gas +, H2S -
Shigella, Serratia
Alkaline slant/Acid butt (K/A)
Gas -, H2S -
Salmonella, Proteus
Alkaline slant/ Acid butt (K/A)
Gas +, H2S +
Pseudomonas
Alkaline slant/ alkaline butt (K/K)
Gas -, H2S -
Urease test - purpose
Urease test is used to ID which organisms?
- Allows for Proteus to be distinguished from non-lactose-fermenting members of the Enterobacteriaceae.
- Rapid test on gastric biopsy samples to detect the presence of H. pylori.
The urea test is part of the battery of tests to identify the following:
Gram-negative enteric pathogens, including Yersinia spp.
Fastidious
Gram-negative rods—Brucella, Helicobacter pylori, and Pasteurella.
Gram-positive rods—Corynebacterium and Rhodococcus spp.
Yeasts—Cryptococcus spp.
Urease test - principle and controls
- Differentiates organisms based on their ability to hydrolyze urea with enzyme urease, utilizing urea as sole source of nitrogen
Product = ammonia
Positive test: Proteus mirabilis.
Negative test: Escherichia coli.