Lab 2 Flashcards
How can we identify a negative motility from the SIM media test?
Negative test is indicated by growth being along the inoculation path. (well-defined inoculation path.)
What metabolic process could bacteria have if they are positive for S in SIM test?
Sulfur reduction
What metabolic process do bacteria have if they are positive for I in SIM test?
Indole production (the hydrolysis of tryptophan to indole, pyruvic acid, and ammonia)
What do bacteria have if they are positive for M in a SIM test?
They have a motility organelle. Most commonly a flagella.
What enzyme is responsible for urea hydrolysis?
Urease
What color indicates a positive urea hydrolysis test?
Bright pink (fuchsia)
What is the pH indicator used in the urea hydrolysis test?
Phenol red
What happens to the pH when urea is hydrolyzed?
The pH increases due to ammonia production
What enzyme mediates nitrate reduction?
Nitrate reductase
What does a red color indicate after adding reagents A and B in the nitrate reduction test?
Positive result for nitrate reduction (NO3 to NO2)
During nitrate reduction test; what does no color change after adding reagents A and B indicate?
Nitrate was not reduced or was fully reduced beyond nitrite
Why is zinc added in the nitrate reduction test?
To check if nitrate is still present by forcing its reduction to nitrite
During a nitrate reduction test; what does a red color after adding zinc indicate?
Negative result for nitrate reduction (nitrate was not reduced by the organism)
In a nitrate reduction test; what does no color change after adding zinc indicate?
Positive result for complete nitrate reduction (beyond nitrite)
What is the purpose of the nitrate reduction test?
To determine if bacteria can reduce nitrate (NO₃⁻) to nitrite (NO₂⁻) or other nitrogenous compounds using nitrate reductase.
What enzyme mediates nitrate reduction?
Nitrate reductase.
What does a red color indicate after adding reagents A and B in the nitrate reduction test?
A positive result, meaning nitrate was reduced to nitrite.
What happens if no color change occurs after adding reagents A and B, during a nitrate reduction test?
The bacteria either did not reduce nitrate or reduced it further beyond nitrite.
What is the purpose of adding zinc in the nitrate reduction test?
To determine if nitrate was still present in the broth, indicating a negative result if it turns red.
In a nitrate reduction test; what does it mean if no color change occurs even after adding zinc?
The bacteria reduced nitrate to nitrogen gas or other nitrogenous compounds, indicating a positive result.
What does “MR” mean from the MR-VP lab?
Methyl red
What does “VP” mean from the MR-VP lab?
Vogues-Proskaurer
What is the MR test testing for?
Mixed-acid fermentation pathway. Causing a significant decrease in the pH.
Why does the mixed-acid fermentation pathway significantly reduce the pH of the medium compared to other acid producing metabolic pathways?
Because it produces 4 moles of acidic products to every 1 mole of glucose fermented.
Why is methyl red used as a pH indicator in the MR test?
Because it is a less sensitive pH indicator. It changes color from yellow to red at a pH of 4.4 or below. While other pH indicators will pick up a pH change at a higher pH.
What is the VP test testing for?
If the bacteria ferement glucose via the butanediol pathway.
What color is a positive MR test?
Pink/Red
What does the butanediol pathway produce?
Produces Acetoin as an intermediate, which can be further reduced to 2,3-butanediol.
What are we testing for in the VP test?
Testing for the presence of Acetoin. Acetoin can be be oxidized by KOH (Reagent B) with α-naphthol (Reagent A) acting as a catalyst. Acetoin oxidized produces Diacetyl. Diacetyl reacts with the guanidine group in the medium to form a pinkish-red colored product.
Define bacterial transformation.
Uptake a free DNA (plasmid) by cells.
Define a plasmid
A small circular piece of double-stranded DNA
What type of agar was used in the Bacterial transformaton: pGFP?
LB agar: another type of nutrient agar.
How were we able to test if the the bacteria was able to receive the plasmid?
If the bacteria became antibiotic resistant, and is able to grow on the agar imbued with ampicilin
What does the GFP gene encode?
Encodes a protein called green fluorecent protein.
Is the GFP gene expression regulated?
Yes the GFP gene is regulated by a Lac operon.
What plates is the GFP protein expressed on and why?
+DNA/LB+Amp+IPTG. The GFP gene is a regulated gene by a lac operon. IPTG is a lactose isomer, and binds to the repressor causing the repressor to unlatch from the operon. Allowing the gene to be transcribed.
What is the name of the test in the Antimicrobial Susceptibility lab?
Kirby-Bauer disk diffusion susceptibility test.
What is the purpose of the antimicrobial susceptibility test?
Determine the bacterial resistance to antimicrobials.
Define the Zone of inhibition.
The area of no bacterial growth surrounding an antimicrobial drug disk.
Why does the zone of inhibition size have no meaning on its own
Because different antimicrobal agents have different diffusion rates. One my have a higher diffusion rate compared to their smaller zone of inhibition size meaning that antimicrobal agent is less affective against the bacteria.
Define a bacteriostatic antibiotic.
Antibiotics that inhibit microbial cell growth.
Define a bactericidal antibiotic.
Antibiotics that result in the antibiotic killing the bacterial cells.
