Lab Practical #2

Question 1

Fluid Thioglycollate Media Broths

Answer 1

FTM; medium used for both aerobic and anaerobic organisms

Question 2

GasPak Anaerobic Jar

Answer 2

Provides an oxygen-free incubation environment for petri plates of anaerobic agar; hydrogen is generated, which removes the oxygen from forming water

Question 3

Brewers Anaerobic Agar

Answer 3

A solid medium excellent from culturing anaerobic bacteria in petri plates; contains thioglycollate (reducing agent) and resazurin (oxidation/reduction indicator); must be incubated in an oxygen-free environment

Question 4

Resazurin

Answer 4

An indicator for oxygen:
- O2 present: pink/lavender
O2 not present: clear/yellow

Question 5

Obligate Aerobes

Answer 5

Bacteria that require oxygen for growth because they carry out respiratory metabolism in which oxygen functions as a terminal electron acceptor; at the top in FTM

Question 6

Microphiles

Answer 6

Bacteria that prefer to grow in oxygen concentrations of 5-10%; limited in their ability to carry out respiration or because they may have oxygen-sensitive proteins and enzymes; band in the middle in FTM

Question 7

Facultative Anaerobes

Answer 7

Bacteria that can grow aerobically or anaerobically depending on the culturing conditions; respiration in the presence of oxygen, fermentation if absent; scattered throughout in FTM

Question 8

Obligate Anaerobes

Answer 8

Bacteria that cannot tolerate oxygen and must be cultivated under conditions in which oxygen is removed, otherwise they are killed; carry out anaerobic respiration, in which inorganic compounds such as sulfate or nitrates replace oxygen as the terminal electron acceptor; at the bottom in FTM

Question 9

Psychrophiles

Answer 9

Bacteria with an optimal growth temperature between -5 and 20 degrees C

Question 10

Mesophiles

Answer 10

Bacteria with an optimal growth temperature between 20 and 50 degrees C; most bacteria fall into this class

Question 11

Thermophiles

Answer 11

Bacteria with an optimal growth temperature between 50 and 80 degrees C

Question 12

Hyperthermophiles

Answer 12

Bacteria with an optimal growth temperature above 80 degrees C

Question 13

Halophiles

Answer 13

Bacteria that require high concentrations of sodium chloride to grow

Question 14

Halotolerant

Answer 14

Bacteria that are capable of growth in moderate concentrations of salt

Question 15

Osmophiles

Answer 15

Bacteria that are able to grow in environments where sugar concentrations are excessive

Question 16

Fermentation Experiment

Answer 16

Experiment that helps determine if bacteria can ferment a certain type of sugar (glucose, lactose, mannitol) and if these fermentation reactions produce gas as a product (bubble is present in Durham tube); positive turns yellow

Question 17

MR(acid)-VP(alcohol) Experiment

Answer 17

Two experiments in one broth; determines if bacteria produce acids of many varieties during fermentation that strongly influence the ph of the broth or produce alcohols that do not influence the pH; bacteria cannot carry out both types of reactions; positive for acid turns red, for alcohol turns pink

Question 18

Catalase Production Experiment

Answer 18

Experiment that determines if the bacteria produces an enzyme capable of detoxifying hydrogen peroxide; if a few drops of hydrogen peroxide along a colony exhibits an immediate bubbling (a release of oxygen gas with in 10 seconds), then test is positive

Question 19

Citrate Utilization Experiment

Answer 19

Experiment that determines if bacteria have the enzyme citrase to convert citrate to pyruvate, which can then be used in fermentation; Simmons agar slant will turn from green to blue

Question 20

Hydrolysis

Answer 20

Bacteria using enzymes to do catabolic chemical reactions

Question 21

Starch Hydrolysis Experiment

Answer 21

Experiment to determine if bacteria have the enzyme amylase that breaks down starch to sugar; flood starch agar plate with Gram’s Iodine, which will cause starch to turn blue or a dark color; if there is a “zone of clearing” (lighter coloration) around the colony then test is positive

Question 22

Urea Hydrolysis

Answer 22

Experiment to determine if bacteria have enzyme urease to break down urea into ammonia; broth contains phenol red indicators, which turn the broth red or slighly purplish (pH driven above 8) in a positive test

Question 23

Tryptophan Hydrolysis

Answer 23

Experiment to determines if bacteria have the enzyme tryptophanase to break down the amino acid tryptophan into indole and pyruvate; tubes cannot be shaken; 10-12 drops of Indole reagent added, if red layer forms at the top of the tube where indole collects, then it is present (positive test)

Question 24

Kliger’s Iron Agar

Answer 24

Experiment that determines if bacteria are capable of conduction acidic fermentation with glucose and lactose, and if they produce hydrogen sulfide gas from the amino acid cysteine; inoculate deep media using needle (not loop)

Question 25

Possible Results of Kliger’s Iron Agar

Answer 25

• If the bottom of the tube turns yellow, but the top of the tube remains reddish, then glucose was used during fermentation; solid media will be split apart if gas was produced
• If the entire tube turns yellow, then the organism uses both glucose and lactose during acidic fermentation; solid media will be split apart if gas was produced
• If the bacteria can degrade the amino acid cysteine into pyruvic acid, then the hydrogen sulfide will have a dark (black) percipitate

Question 26

Bergey’s Manual of Systematic Bacteriology

Answer 26

A large volume of books that are the most used comprehensive guide to identifying unknown bacteria; contains a massive amount of information (gram reaction, motility, endospore producers, presence of capsules, etc.) pertaining to many species and varieties of bacteria

Question 27

Dichotomous Key

Answer 27

Always has two answers for each question asked

Question 28

Diagnostic Key

Answer 28

Simple tests used to determine the identity of an unknown bacteria; dichotomous key

Question 29

Thermal Death Point (TDP)

Answer 29

Temperature required to destroy a population of bacteria in 10 minutes

Question 30

Thermal Death Time (TDT)

Answer 30

Time required to destroy a population of bacteria at a specific temperature