Experiment 6,7 & 8: Hydrolytic Enzymes, Serial Dilutions & Quantification Of Bacterial Viable Counts, Bacteriophages & Plaque Assay

Front 1

What are hydrolytic enzymes

Back 1

use water to split complex molecules in a process called hydrolysis.

Front 2

Do all bacteria posses hydrolytic enzymes? What 4 hydrolysis test are used to determine bacterial identification?

Back 2

No; Starch, Casein, Gelatin and Lipid

Front 3

What enzyme breaks down starch? Casein? Gelatin? Lipid?

Back 3

Amylase; caseinase; gelatinase; lipases.

Front 4

What compound is added to the starch agar after incubation in order to test for starch? If starch is present, what color forms? If no starch is present, what color forms after iodine is added?

Back 4

Iodine; Clear halo; black

Front 5

What color will the bacterium produce if caseinase is present? Not present?

Back 5

Clear halo; no clear halo

Front 6

Which 2 experiments did we use test tubes for the procedure rather than nutrient agar plates?

Back 6

Lipids and gelatin

Front 7

In gelatin hydrolysis, we stabbed bacterium into gelatin, if the gelatinase is produced what occurs to the gelatin? If the gelatinase isn't produced?

Back 7

It turns into liquid; remains a solid

Front 8

What are lipids composed of?

Back 8

glycerol with 3 fatty acids

Front 9

Which portion of the lipids is cleaved and used by the cell?

Back 9

Glycerol and fatty acids are cleaved; glycerol is used in the cell

Front 10

What reagent do we use to test for lipid hydrolysis in bacterium?

Back 10

Tween 80 Aka Polysorbate 80

Front 11

What is the composition of Polysorbate 80?

Back 11

Fatty acids and polyethylene sorbitol ester

Front 12

What does lipase hydrolyze in tween 80 into?

Back 12

Oleic acid and polyoxyethylated sorbitol

Front 13

What color occurs if lipase is present? Not present?

Back 13

pink color; yellow/straw color(original color= no color change)

Front 14

What two bacterium were used in gelatin hydrolysis? What two bacterium were used in both the casein and starch hydrolysis test?

Back 14

S. aureus and E. coli; B. subtilis and E. coli

Front 15

What two bacterium were used in the lipid hydrolysis test?

Back 15

E. coli and M. smegmatis

Front 16

Pathogens that can cause disease present in food?

Back 16

food poisoning

Front 17

Microorganisms present on food that cause it to be inedible?

Back 17

food spoilage

Front 18

Are all microorganisms in food products harmful?What four foods are discussed in lab to be easily contaminated?

Back 18

No; Fruits, meats, milk and processed foods

Front 19

Why do we quantify the concentration and types of bacteria in a sample by dilution?

Back 19

To acquire a readable amount

Front 20

What does serial dilution reveal?

Back 20

number of bacteria per unit of volume of the original sample (I.e. number of bacteria in one ml of sample)

Front 21

Serial dilution consists of diluting a fixed amount of the original stock ________ each time.

Back 21

tenfold

Front 22

The stock is diluted in _______ that contain a specific amount of buffer or water. What is the standardized volume of the blank?

Back 22

blank; 1 ml stock solution and 9 ml of buffer

Front 23

This technique involves plating a small amount from the dilution blank onto an agar dish?

Back 23

Spread plate technique

Front 24

What does CFU stand for? How do we determine the dilution factor?

Back 24

colony forming unit; df = volume of sample added/ total volume (sample + blank)

Front 25

If you dilute 1 ml of the sample into a 9ml blank what is the DF?

Back 25

DF = 1ml / 1ml + 9ml = 1/10 or 0.1

Front 26

To determine the dilution of the original stock, multiply the dilution factor of the previous dilution and _______?

Back 26

new dilution produced

Front 27

What 4 food products were used in the quantification experiment?

Back 27

strawberries, spinach, potato and milk

Front 28

What size colony is statistically relevant? What acronym is given to a colony with too many and too few colonies?

Back 28

30-300 colonies; TNTC (too many too count) and TFTC (too few to count)

Front 29

Note: The procedure states bacteria present per 1ml, but we only used 0.1 ml onto the plate**

Back 29

CFU/ml = CFU/ (volume plated)(df from the tube plated)

Front 30

If we count 125 colonies on the 10(^-3) plate what does CFU/ml equal?

Back 30

125/ (0.1ml)(10(^-3) = 1.25 x 10(^6)

Front 31

Noncellular elements that rely on other cells to replicate?

Back 31

Viruses

Front 32

A type of virus that infects bacteria? Are these viruses non-specific and infect any phage?

Back 32

bacteriophage; no, they are specific for particular bacteria

Front 33

Which virus cycle immediately replicates and new phage are synthesized? Which cycle consists of the phages DNA becoming integrated into the hosts genome called a prophage and is then replicated along with the bacterium's genome?

Back 33

Lytic cycle; lysogenic cycle

Front 34

While in the lysogenic cycle, an environmental factors or other stress on the cell triggers this latent cell to activate? Once activated what cycle does the cell undergo?

Back 34

prophage; lytic cycle

Front 35

What phage ONLY replicates by the lytic cycle and is considered a virulent phage?

Back 35

Phage T4

Front 36

These type of phages reproduce by both lytic and lysogenic cycle?

Back 36

temperate phages

Front 37

Can phages by grown randomly?

Back 37

No, ONLY if they've infected an appropriate host

Front 38

Bacteria form colonies on an agar plate, viruses form what?

Back 38

plaques

Front 39

Clearing in a lawn of bacterial growth?

Back 39

plaques

Front 40

What does PFU stand for? What technique is used to perform a plaque assay?

Back 40

plaque forming units; pour plate technique