Microm 302 Quiz 3

UW

What technique is used to count bacteria in a sample?

Plate count

What is the name of a group of pure cells made from one cell?

Colony

What can happen if too many cells grow in the agar?

Confluent growth

How do you prevent too much cell growth in a sample?

Dilution of a sample

Explain why it's impossible to grow a mixture of cells?

Can't design medium that allows all bacteria to grow/form colonies due to all different growth requirements

What is the label of the types of bacteria that are able to be grown in a lab?

Colony-forming unites (cfu)

How many colonies is on a countable plate?

30-300 colonies

What's the deal with melted TSY agar?

Melts at 100C, solidifies at room temp, stays melted at 50C but cannot re-melt in 50C

How do you mix agar with a cell sample?

Figure eight pattern 2-3 times

When is a mixed agar + bacteria plate ready to be incubated?

When the agar has solidified after at least 5 minutes, then can be inverted and incubated (37C)

What is it called when there are more than 300 colonies in a sample to count?

Too Numerous To Count (TNTC)

Why can't we trust a sample that has less than 30 colonies in it?

Random sampling is too great

What is the first step to identifying an unknown bacterium?

Determine it's shape and Gram stain reaction.

What are the different mediums of samples and which one is the most concentrated?

A solid culture is more concentrated, while a broth sample is less concentrated.

How much of a colony do you need when making a smear for a Gram stain?

Only a light touch, as you will use the same one for other procedures

Why are colonies re-streaked?

To ensure that you have a pure culture

Why should you save plates from an ongoing experiement?

In case you need to re-do an experiment if a step goes wrong, or until you know a new inoculated culture has grown well.

What is a saline suspension for?

To ensure a uniform and light inoculum in a biochemical test

What will happen if you have an organism makes an enzymes able to hydrolyze gelatin?

It will cause the gelatin to liquify due to the solidifying properties to be lost. A positive result is if gelatin remains liquid when left in a cold bath for at least 10 mins

What does a positive result in tryptone broth look like and why?

A colored indicator (red at the top when Kovac's reagent is added) can show a positive result due to some bacteria able to break down tryptophan (amino acid) into indole. Brown-yellow color is a negative reaction

What does a positive result in a sugar ferment look like and why?

A pH indicator will change color due to the sugar fermenting, being more acidic, and if gas is trapped in the Durham vial during fermentation.

What is catalase?

enzyme that breaks down hydrogen peroxide into water and oxygen

What does a positive test for a catalase test look like?

Bubble formation

What does a catalase test show?

Can define streptococci from other Gram-positive cocci

What organisms usually make catalase?

Organisms that respire aerobically

Why don't we perform the catalase test on a colony on an agar plate?

Yeast-extract, blood, and other media components are catalase-positive and could mess up the results

How are sulfonamides selectively toxic?

They can be more harmful to bacteria than humans through their interference with the folic acid synthesis

Why do bacteria and humans need folate?

Forms cozyme F which is used for nucleotide and amino acid synthesis

How do bacteria get folate?

They make it from a precursor called PABA (para-aminobenzoic acid), PABA affects them

How do humans get folate?

The can't make it so it needs to be consumed through their diets, PABA doesn't affect them.

What does bacteriostatic mean?

Inhibit without killing cells.

What does bactericidal mean?

kill bacterial cells

Can enzymes distinguish between PABA and sulfa?

No, and can bind to sulfa instead of PABA and then PABA out-competes for the active site of the enzyme

What is competitive inhibition?

It competes for the active site of an enzyme

How can sulfa effects be reversed and what is this process called?

Adding excess PABA, which will outcompete sulfa for enzyme active site OR add nucleotides/amino acids that coenzyme F makes. These processes are both called non-competitive inhibition

What is the folate pathway?

PABA -> Folate -> Coenzyme F -> Amino acid/nucleotide biosynthesis. Sulfa can inhibit PABA

Is sulfa bactericidal or bacteriostatic?

Bacteriostatic by using inhibition, not killing bacteria.

Do all microorganisms have the same O2 requirements?

No, they can vary depending on the organism

What are obligate aerobes?

Cells with an absolute requirement for O2 and produce catalase enzyme

What are obligate anaerobes?

Don't require O2 but are not killed/inhibited by it and they do not produce catalase

What are facultative anaerobes?

Can use O2 for respiration if available but can also ferment to produce O2. These produce catalase

What are microaerophiles?

Require O2 but in limited amounts

What are aerotolerant anaerobes?

grow in O2 but don't need it for metabolism

What is an agar shake tube?

Agar deep that has been boiled to get rid of any gas within it before inoculation. O2 will only be present in the very top part

Why are agar shake tubes boiled?

To get rid of any gas within the agar deep before inoculation.

How can you mix agar shake tubes without reintroducing air?

Rolling it gently between your palms

What does an agar shake tube look like if bacteria produce gas?

May displace some of the solid material due to gas moving it

Why couldn't an agar plate work the same as an agar shake tube?

A shake tube provides a varying amount of oxygen while a plate allows for maximum O2 exposure.

What is a vegetative cell?

Actively grows and does not produce spores

What is an endospore?

Very resistant cell form.

Why are endospores made and what is this process called?

It happens when there is a big change in conditions and this is called sporulation

What is germination?

Endospores return to a vegetative state and happens in more favorable conditions

What does it mean for something to be equilibrated?

Having something in equilibrium with something else (equal)

How long do you have to leave something for it to equilibrate?

~10 mins

Why are controls important?

To make sure that the treated samples have an effect.

Why are broth tubes incubated?

To check for viable cells by looking for growth

What is UV light used for?

For disinfecting

What does UV do and how?

destroys airborne organisms by causing mutation in DNA and alters the bonds. This can be lethal to cells

When does UV work?

When there is no plastic/glass obscuring the site that it is used on

How should you use UV?

With caution, with goggles and no hands in the area when it is on