Counting colonies on a plate
Direct and viable
Counting colonies using spectrophotometer
Indirect and total
As transmittance increases, turbidity:
Decreases
If you take 1ml of a culture and add it to 9 ml of media, what kind of dilution is this?
1:10
Would you use a plate with 500 colonies or one with 120 colonies to determine the
concentration of bacteria in your original culture?
The one with 120 colonies
If your organism grew throughout a thioglycollate tube but benefited from oxygen, how would you classify it?
Facultative anaerobe
If an organism grows at low oxygen, what is it called?
Microaerophile
How would you classify an organism that grows optimally at 37 degrees C?
Mesophile
What is the optimal growth for Serratia marcescens and how
would you classify this organism?
25 degrees C, psychrotroph
What color are the colonies of Serratia marcessans at 25 degrees C?
Red or pink
An extreme or obligate halophile grows at what salt concentration?
15%
Organisms that can grow at salt concentrations up to 10% are called ____________.
Facultative halophiles
Why are Bacillus most resistant to heat than other organisms?
They have endospores
What is to the time to kill all cells in a culture at a given temperature?
Thermal death time
What is the time, in minutes, to kill 90% of a population at a given temperature?
Decimal reduction time
What does UV irradiation do to the DNA?
It creates thymine dimers
Is UV radiation ionizing or nonionizing?
Nonionizing
For the UV experiment, the side of the plate covered with the index card, had more or less colonies?
More
When you place an antibiotic disc on a bacterial plate, do you measure the radius or diameter of the zone of inhibition the next day?
Diameter
What are some reasons for antibiotic resistance?
Using antibiotics too often or when not needed
Plasmids with antibiotic resistance genes
Bacteria can pump out antibiotics
Is the m-Staph broth, selective, differential or both?
Selective
Are the MSA plates selective, differential or both?
Both
SHOULD Novobiocin be sensitive towards
Staphlocococcus aureus, Staphylococcus epidermidis or both?
Both
How does Staphyloslide work?
The antibodies in Staphyloslide bind to the antigens on Staph. aureus
Which organism(s) tests positive for coagulase?
Staphylococcus aureus
Bacterial enumeration
The measurement of the number of bacterial cells per milliliter, gram, or cubic meter of a sample (the units depend on the nature of the sample)
Direct Counts
...means actually counting cells
More accurate, but takes a lot longer
Indirect Counts
mean estimating the number of cells based on turbidity in liquid or dry weight.
Less accurate, but quick
Total count
All the cells in a culture
Viable count
Only live cells in a culture
obligate anaerobes
Cannot tolerate oxygen
Obligate aerobes
Require oxygen
Facultative anaerobes
Can grow with ot without oxygen
Microaerophiles
Grow at low concentrations of oxygen
Aerotolerant
Cannot use oxygen, but tolerate it well
Oxygen test uses semi-solid media called thioglycollate broth.
there is a pink indicator to demonstrate
that oxygen is present in the broth. When oxygen is used, it will disappear.
If an organism can only grow on the 1% or 5% salt plates only, it is
classified as
Non-halophile
If the organism can also grow on the 10% plate, it is
classified as
a Facultative halophile
If it can grow on the 15% salt plates, it
is called
an Extreme halophile
Hypotonic
Less salt in the environment and more inside the cell (cell will lyse)
Hypertonic
More salt in the environment and less inside the cell (cell will shrink)
Isotonic
Equal level of salt in the environment and inside the cell
Osmosis
How the water moves into the cell...by osmosis
Psychotrophile
Temp range -10 to 20 degrees C (Optimal growth 10 degrees C)
Psychotroph
Temp range 5 to 30 degrees C (Optimal growth 25 degrees C)
Mesophile
Temp range 10 to 45 degrees C (Optimal growth 37 degrees C)
Thermophile
Temp range 40 to 75 degrees C (Optimal growth 55 degrees C)
Hyperthermophile
Temp range 65 to 120 degrees C (Optimal growth 90-100 degrees C)
++ double-positive
Lots of growth
+ positive
Some growth
- negative
No growth
Broths that produced a pinkish color in temp lab meant that ______________.
it grew very well. (The darker the pink, the better the growth).
Indole test
Performed to help differentiate species of Enterobacteriaceae. Tests bacteria's ability to produce indole.
Indole Media and reagents
Tryptone broth (contains tryptophan). Kovac's reagent-HCL, dimethylamoniobenzaldehyde, and amyl alcohol.
Indole results
Positive- Kovac's reagent reacts to indole and creates a red color at the top part of the test tube. Negative remains yellow.
Methyl Red/ Voges Proskauer (MR/VP)
Both tests Performed to help differentiate species of Enterobacteriaceae
MR
Tests for endproducts from glucose fermentation
VP
Tests for acetoin production from glucose fermenation
MR/ VP Media and reagents
Glucose broth. Methyl Red indicator for acid. Voges Proskauer reagents: 5% Alpha-Napthol and ethanol; Potassium Hydroxide and Deionized water.
MR result
Positive result is red (indicating a pH below 6); Negative result is yellow (indicating no acid production)
VR result
Positive result is red after VP reagents added (Indicating presence of acetoin); Negative result has no color change.
Citrate
Performed To help differentiate species of Enterobacteriaceae. Selective for bacteria that consumes citrate as its sole source of carbon, and ammonium as its nitrogen source.
Citrate Media and reagents
Simmon's citrate Agar (sodium citrate as carbon source), ammonium ion (as nitrogen source), ans pH indicator-bromthymol blue.
Citrate result
Positive-Blue color-( indicates bacteria metabolized citrate and produced acid as an end product). Negative result- Green.
H2s Production in SIM
Used to differentiate species of the family Enterobacteriaceae. Determines bacteria's ability to reduce sulfer into H2S.
H2S in SIM Media and reagents
SIM media-contains sulfer, cysteine, sodium thiosulofate, and peptonized iron or ferrous sulfate.
H2S in SIM results
Positive result-contents of tube turn black; Negative results- Does not turn black.
Urea Hydrolysis
Done to determine a bacteria's ability to hydrolyze urea to make ammonia using enzyme urease.
Urea Media and Reagents used
Urea broth (Yeast extract, monopotasium phosphate, disodium phosphate, urea) Phenol red indicator.
Urea results
Positive: Bright pink-indicates ammonia is made. Negative: Yellow (no color change)
Motility test
Performed to help differentiate species of bacteria that are motile.
Motility Media and Reagents
Motility media contains tryptose, sodium cholride, agar, and a color indicator.
Motility results
Positive: Red line will be growth going out from the stab line. Negative: red color will be straight down.
Lactose fermentation
Tests ability of a bacteria to ferment lactose
Lactose Media and reagents
Lactose broth: Beef broth, geletin peptone, and lactose. Phenol red indicator added to indicate acid production from fermentation.
Lactose results
Positive: Yellow after indicator is added; Negative: Reddish, no color change
Sucrose Fermentation
Tests ability to ferment sucrose and production of acid end product
Sucrose Media and reagents
Sucrose broth: Beef broth extract, gelatin peptone, and sucrose. Phenol red indicator detects acid end-product.
Sucrose results
Positive: Yellow; Negative: No color change or reddish.
Glucose Fermentation and Gass Production
Tests bacteria's ability to ferment glucose and produce gas and/or acid end product
Glucose Media and reagents
Glucose broth: Beef extract, gelatin peptone, and glucose. Phenol red indicator to indictae acid end product. Durham Gas tube added to indicate gas production.
Glucose results
Positive: Yellow indicating presence of acid and glucose fermentation; Positive result for gas is a bubble in Durham's tube; Negative: No color change, or reddish color and no bubble.
BAP with Novobiocin Test
Determines if organism is resistant or sensitive to novobiocin, and determines if organism can lyse red blood cells.
BAP Media and reagents
Nutrient agar w/ 5% sheep's blood. Novobiocin disk added after inoculation.
BAP results
Positive: Hemolysis- Occurs if media surrounding colonies turns translucent or green in color.
MSA (Manitol Salt Agar)
Tests bacteria's ability to tolerate 7% salt concentration and ferment mannitol. Selective--selects for salt tolerant bacteria. Differential---because it differentiates salt tolerant organisms on their ability to ferment mannitol.
MSA Media and reagents
MSA media--nutrient agar, mannitol, 7% sodium chloride and phenol red indicator.
MSA results
Can ferment mannitol: yellow zones around the colonies. Cannot ferment mannitol: Media will remain pink. Organism will grow if it can tolerate salt. Will not grow if it can't.
Mstaph Broth
Tests bacteria's ability to tolerate 7% salt concentration and ferment mannitol. Selective, because it selects salt tolerant Staphlyococcus.
Mstaph Broth Media and Reagents
Contains nutrients appropriate for growing Staphylococcus and 7% salt.
Mstaph results
Positive: Cloudy, will grow--tolerates salt; Negative: Clear (yellow), will not grow--does not tolerate salt.
Coagulase
Tests for bacteria's ability to clot blood plasma using enzyme coagulase.
Coagulase Media and reagents
Rabbit plasma dissolved in buffer.
Coagulase results
Positive: Clumps Negative: No clumps
Zone of inhabition
Measured in mm. I dentify S (Sensitive), I (Intermediate), or R (Resistant).