Lab Practical #2 Flashcards
Fluid Thioglycollate Media Broths
FTM; medium used for both aerobic and anaerobic organisms
GasPak Anaerobic Jar
Provides an oxygen-free incubation environment for petri plates of anaerobic agar; hydrogen is generated, which removes the oxygen from forming water
Brewers Anaerobic Agar
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
Resazurin
An indicator for oxygen:
- O2 present: pink/lavender
O2 not present: clear/yellow
Obligate Aerobes
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
Microphiles
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
Facultative Anaerobes
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
Obligate Anaerobes
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
Psychrophiles
Bacteria with an optimal growth temperature between -5 and 20 degrees C
Mesophiles
Bacteria with an optimal growth temperature between 20 and 50 degrees C; most bacteria fall into this class
Thermophiles
Bacteria with an optimal growth temperature between 50 and 80 degrees C
Hyperthermophiles
Bacteria with an optimal growth temperature above 80 degrees C
Halophiles
Bacteria that require high concentrations of sodium chloride to grow
Halotolerant
Bacteria that are capable of growth in moderate concentrations of salt
Osmophiles
Bacteria that are able to grow in environments where sugar concentrations are excessive
Fermentation Experiment
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
MR(acid)-VP(alcohol) Experiment
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
Catalase Production Experiment
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
Citrate Utilization Experiment
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
Hydrolysis
Bacteria using enzymes to do catabolic chemical reactions
Starch Hydrolysis Experiment
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
Urea Hydrolysis
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
Tryptophan Hydrolysis
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)
Kliger’s Iron Agar
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)
Possible Results of Kliger’s Iron Agar
- 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
Bergey’s Manual of Systematic Bacteriology
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
Dichotomous Key
Always has two answers for each question asked
Diagnostic Key
Simple tests used to determine the identity of an unknown bacteria; dichotomous key
Thermal Death Point (TDP)
Temperature required to destroy a population of bacteria in 10 minutes
Thermal Death Time (TDT)
Time required to destroy a population of bacteria at a specific temperature
