Biochemical Tests: Gram Negative Bacilli Flashcards
Why did we perform these specific tests?
bacteria are categorized upon reactivity/morphology;
these specified biochemical tests are used to differentiate between gram negative bacilli species: nutrition preferences (with pH indicators), confirmation of specific enzyme
Bacterial Organisms that are Gram-Negative Bacilli
E. coli, K. pneumoniae, E.cloacae, S. typhimurium, A.faecalis
Biochemical Tests (Gram-Negative Bacilli) Performed
phenol red
oxidase
TSI
decarboxylation
Phenol Red
testing nutrient preferences with different sugars and gas production
Oxidase
testing whether each one of the organisms contains the enzyme oxidase
TSI
nutrient preference (peptones vs glucose) and ga production
Decarboxylation
testing whether each one of the organisms has the enzyme decarboxylase (removes a carboxyl group from amino acids)
Phenol Red Results
purpose: to see if the organisms have a preference/ability to digest certain sugars
sugars: lactose, glucose, sucrose
organisms: E. coli, S. aureus, E. faecalis
procedure: inoculate an organisms into the three sugar broths;
results (pH indicator): color change and acids production (A), Gas production (G)
yellow color indicates acid production and air bubble is + gas production
Phenol Red Results
E. coli: glucose (A/G), lactose (A/G), sucrose (negative)
S. aureus: A for all three, no gas production
E. faecalis: negative for all three sugars
Oxidase Results
purpose: to see which organism contains the cytochrome c oxidase (protein used in electron chain transport)
organisms: E.coli and A. faecalis
procedure: take each organism using a wooden dowl and inoculate the oxidase strips
results: positive (purple color), negative (no color change or light coloring)
Oxidase Results
E.coli: negative (no color change production)
A. faecalis: positive (indigo color)
TSI (Triple Sugar Agar) Results
purpose: differentiates bacteria based on their fermentation of lactose, glucose, and sucrose and on the production of hydrogen sulfide
organisms: E.coli, A.faecalis, S.typhimurium, K. pneumoniae
procedure: take each organism and inolculate both the top of the agar and the bottom (using the loop and needle)
results: record both the slant and the butt reaction
A: yellow, glucose is the preferred nutrient source
K: Pink: peptones are a preferred nutrient source
H2S: Black: this molecule is present
Gas: production of bubbles like cracks or spaces in agar
NC: no color change
TSI Composition
Phenol Red; an added pH indicator that creates any sort of color change in the agar
Alkaline Reactions: if the bacteria are unable to digest the sugars, then they might rely on peptones; this will not cause a pH reaction and keep the agar red
Carbohydrate Source: glucose, lactose, sucrose
Sodium thiosulfate and ferrous sulfate: used to detect the production of H2S by the bacteria and turns agar black
TSI Results
E.coli: yellow slant and butt; facultative anaerobe; this is gas +
A.faecalis: red slant; NC in the butt; aerobic organism
S.typhimurium: K (red slant); A (yellow slant) and H2S (black production); facultative anaerobe
K. pneumoniae: yellow slant and butt; facultative anaerobe; gas+
Decarboxylase Results
E.cloacae: positive
K.pneumoniae: negative
Casein Hydrolysis Test
test whether a bacterium can synthesize the caseinase enzyme (found in milk protein)
DNA Hydrolysis Test
test whether a bacterium can hydrolyze DNA as a main source of carbon and energy for growth
Nitrate Reduction Test
test whether the bacteria can reduce nitrate (NO3-) to nitrite (NO2-)
Biochemical Tests
DNA Hydrolysis: E.clocae: positive, S.marcescens: white, poitive, S.aureus: positive, S.epidermidis: negative
Cansein: E.coli: negative, B.subtilis: positive
Nitrate Reduction: A.faecalis: true negative, E.coli: positive, P.areuginosa: positive
