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89 Cards in this Set

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Ingredients in Mannitol Salt agar
7.5% NaCl
Mannitol
Phenol Red
Ingredients in DNase agar
Emulsion of DNA
Peptides
Methyl green dye
How mannitol salt agar is used to differentiate among the different Staph species?
Differentiates by whether or not the bacteria are able to ferment mannitol
Which Staphylococcus species are positive for MSA fermentation?
S. aureus
S. xylosus
S. saprophyticus
Which Staphylococcus species are DNase-positive?
S. aureus
S. intermedius
Which Staph species are resistant to the antibiotic Novobiocin?
S. saprophyticus
S. xylosus
What is the coagulase test and how is it used to identify S. aureus?
It tests for the presence of the enzyme coagulase.
Reagent used - citrated plasma
What is the catalase test and how can it distinguish strep from staph?
Tests for enzyme catalase
Reagent used peroxide
Which streptococci are alpha-hemolytic?
S. agalactiae
S. bovis
S. salivarius
S. mutans
E. faecalis
Which streptococci are beta-hemolytic?
S. equi
S. pyogenes
S. zooepidermicus
Ingredients in Bile Esculin agar
Bile salts (oxbile)
esculin
ferric ammonium citrate
beef and gelatin extracts
Which strep species are able to hydrolyze esculin?
S. bovis
S. salivarius
E. faecalis
What is the only member of the strep group that can grow on mannitol salt plates?
Enterococcus
Which Streptococcus species does the CAMP test identify?
S. agalactiae
Basic principles behind the CAMP test.
Relies on the fact tthat most S. agalactiae strains produce a diffusible extracellular compound called the CAMP substance that will conjugate with a specific beta-hemylosin of S. aureus cause a increased hemolytic effect of RBCs in an agar medium.
Is MSA selective, differential, or both?
Both.
Selects for halotolerant
Differentates for fermentations
Ingredients in starch media
polysaccharide
What enzymes degrade starch?
amylase
maltase
What reagent needs to be added to the starch plate?
Iodine
What is the lipid substrate in lipid media?
Tributyrin
Name the enzyme(s) that degrade lipids.
lipase
Know the ingredients in casein media.
casein (major-milk protein)
What enzymes degrade casein?
casease and protease
What is the peptonization process?
Hydrolysis of proteins is called proteolysis.
In proteolysis of casein:
Protein is broken down into peoptones, into polypeptides, into dipeptides and finally into amino acids.
What is the name of the enzyme which hydrolyzes gelatin?
gelatinase
Why was media containing ampicillin used to select for transformants of E. coli?
xxx
Why did we streak before and after transformation on media without ampicillin and media with ampicillin?
xxx
Be able to calculate transformation efficiency.
xxx
What are the 7 ingredients in a TSI slant and what is the purpose of each?
1) 0.1% glucose - carb. substrate
2) 1.0% sucrose - carb. substrate
3) 1.0% lactose - carb. substrate
4) peptones - ???
5) phenol red - pH indicator
6) Sodium thiosulfate - if produces H2S will use this as a substrate and react with Ferrous sulfate to form a black precipitate
7) Ferrous sulfate - see above
Name two other types of media that can be used to test for hydrogen sulfide production, and give an example of an organism that produces hydrogen sulfide.
SIM Deep Tube, Lysine Iron Agar
What is the name of the enzyme that can hydrolyze urea?
Urease
What are the end-products of urea hydrolysis?
Ammonia, water and carbon dioxide
What is the pH indicator in the urea broths?
Phenol red
Give an example of an organism which is urease-positive.
Proteus mirabilis
What are the components of a SIM agar deep tube?
Amino acid tryptophan, peptones, sodium thiosulfate and ferrous sulfate
What 3 things does this type of agar test for?
Indole production, H2S production and motility
What must be added to the tube following incubation in order to perform this test?
Kovac's reagent
What is the MRVP test?
Methyl Red Vouges-Proskaur
MR tests for fermentation of glucose with mutiple acid-end products
VP tests for fermentation of glucose with only 1 acid-end product ... usually acetic acid
What are the components of the MRVP broth, and what is the purpose of each component?
Peptones
glucose -substrate
phosphate buffer - lowers pH
What reagent is added for the MR test?
Phenol red
What reagents are added for the VP test?
Barritt's reagents A and B
Can an organism be positive for both MR and VP?
No
Give an example of an organism that is MR positive and one that is VP positive.
MR+ Morganella morganii
VP+ Klebsilla oxytoca
What is the purpose of the citrate test, and what are the ingredients in the citrate agar slant?
To determine if an organism can use citrate as a carbon source.
The slant contains sodium citrate, brom thymol blue (pH), sodium and water.
Give an example of an organism that is citrate positive.
Proteus mirabilis
What are the ingredients in nitrate broth?
Beef extract
peptone
nitrate
What enzymes reduce nitrate? Is this an aerobic or anaerobic process?
Nitrate reductase
Anaerobically
What reagents need to be added to the nitrate broth following incubation?
Nitrate reagent A
Nitrate reagent B
What does a positive result following addition of these reagents mean?
Color changes to red, means + for nitrate reduction
When would you need to add zinc to the tube?
If no color change after Nitrate Reagent B is added, then add zinc.
What does a positive result following addition of zinc mean?
Still no color change - then + for nitrate reduction because indicates nitrates were reduced beyond nitrites to ammonia or molecular nitrogen
What does a negative result following the addition of zinc mean?
Color changes, means - for nitrate reduction, because the red color means the zinc reduced the nitrates to nitrites, not the bacteria.
Give an example of an organism that is positive for nitrate reduction.
E. coli
What are the ingredients in LIA slants and what two biochemical reactions do these slants test for?
Lysine, glucose, peptones, brom cresol purple, sodium thisulfate and ferric ammonium citrate.
Tests for lysine decarboxylation and lysine deamination
Which reaction is aerobic? Which is anaerobic?
Deamination is aerobic
Decarboxylation is anaerobic
What are the gram-negative tests used for idenitification?
TSI
Urease
MRVP
SIM Deep - Indole Prod.
Citrate
Nitrate
LIA
What test would you use on your gram-positive organism to distinguish Streptococcus from Staphylococcus?
Catalase test
Which set of tests would you set up to identify a Strep?
MSA growth
Blood Agar (for hemolysis and antibiotic testing (Bactracin and SXT))
Bile Esculin
Know the principles behind the rapid Strep test, and be able to identify a positive and negative result.
Uses an immunoassay to detect group A strep antigens. The dipstick contains a membrane strip that has been coated with colored anti-Strep A antibodies.
Know the principles behind the rapid Staph test and be able to identify a positive and negative result.
Based on the fact that most pathogenic strains of S. aureus produce a bound coagulase and contain Protein A on their cell surface.
Be able determine using a microscope whether an acid-fast stain of a sputum sample is positive or negative.
Look for acid-fast rods and watch for symptoms
Know the basic principles behind the acid-fast stain.
Differential stain which stains bacteria, such as Mycobacterium and Nocardia, which have mycolic acids in their cell walls.
Be able determine using a microscope whether a gram stain of a patient specimen is positive or negative, and the morphology and arrangement of the stained organisms.
look for clusters, rods, chains, cocci, etc. to help with diagnosis
Know the basic principles behind the gram stain.
Differential stains which stains bacteria based on the amount of peptidoglycan in their cell walls.
Name the 2 bacteria used as active cultures in making yogurt.
Streptococcus thermophilus
Lactobacillus bulgaricus
Know the general process of yogurt production.
Boil milk
Cool to 55 degrees Celsius
Add scoop of yougurt
incubate
Refrigerate
What plates did we use to test for bacterial contaminants on raw chicken and beef?
Salmonella-Shigella Agar plates (SS Agar)
What ingredients do SS Agar plates contain, and what is the purpose of each ingredient?
Lactose - Carb. substrate that allows the differentiation between lacotse and non-lactose fermenters.
Bile salts - make agar more selective for gram- enteric bacteria by inhibiting the growth of gram+.
Ferric citrate - indictor of H2S production
Neutral red - pH indicator for when lactose is fermented.
What is the name of the broth media we used to test water purity?
Single strength lactose broth with Durham tube and double strength lactose broth with Durham tube
What is the purpose of the Durham tube?
XXX
Why is this called a presumptive test?
Because after incubation, if fermentation plus gas is seen in the tubes then the water tested is PRESUMED to contain fecal coliforms
Which is DNase? Is it a positive or negative rxn.?
The bluish area, positive
Alpha
Blood agar
Name alpha, beta and gamma hemolysis. What type of media is this?
Bile esculin slant

Tube 1: Positive for esculin hydrolysis in the presence of bile. Note black precipitate in > 50% of tube.

Tube 2: Negative for esculin hydrolysis in the presence of bile.
What type of media is this? Which is a positive rxn?
CAMP test
The arrows pointing to the "arrowhead effect" as hemolysins from S. aureus interact with hemolysins from S. aureus to produced an enhanced zone of beta-hemolysis.
What type of test is this?
MSA plate
Both organisms growing on the plate above are halotolerant. The organism (Staphylococcus aureus) indicated by the orange arrow is capable of mannitol fermentation, signified by the color change in the surrounding media. The organism (Staphylococcus epidermidis) indicated by the blue arrow is not capable of mannitol fermentation.
What type of media is this? Which is = or -?
Citrate test
Tube 1: Negative for citrate utilization. Note that there is no color change, and there is no growth on the slant itself.

Tube 2: Positive for citrate utilization. Note that the color has changed to "Caribbean blue", and there is also growth on the surface of the slant.
What type of media is this? Which is - or +?
Water Purity test
Media: Single (and Double)strength lactose broth with Durham tube
Tube #1: Positive for fecal coliforms. Note (1) the gas present in the Durham tube and (2) the color change from red to yellow as acid end-products react with the pH indicator.

Tube #2: Negative for fecal coliforms. Note the absence of gas in the Durham tube. Even though the pH indicator has changed from red to yellow, gas must be produced for a positive result.

Tube #3: Negative for fecal coliforms.
Name the media/test. What are the results?
Extracellular Enzymes:
This plate is positive for starch and lipid hydrolysis, and weakly positive for casein hydrolysis. Note the zones of clearing in the media around the bacterial growth. (Iodine has been added to the starch sector to aid in seeing starch hydrolysis.)
Name the media/test. Is this a - or +?
Extracellular Enzymes
This plate is negative for starch and lipid hydrolysis, and positive for casein hydrolysis. Note the absence of a zone of clearing in the starch and lipid sectors.
Name the media/test. Is this a - or +?
Gelatin Hydrolysis
Tube #1 is negative for gelatin hydrolysis.

Tube #2 is positive for gelatin hydrolysis.
Name the media/test. Which is + or -?
Urease Test
Tube 1: Positive

Tube 2: Negative
Name the media/test. Which is + or -?
Indole prod. - SIM deep tube
Tube 1: (1) Positive for H2S, (2) Negative for Indole production, (3) Positive for motility

Tube 2: (2) Negative for H2S, (2) Positive for Indole production, (3) Undetermined motility
Name the media. What does this test for? Results?
TSI slant
Tube 1: Acid slant/ Acid butt/ Gas/ no H2S

Tube 2: Alk slant/ Acid butt/ Gas/ H2S

Tube 3: Alk slant/ Alk butt/ no gas/ no H2S

Tube 4: Alk slant/ Acid butt/ no gas/ no H2S
Name the media. Results?
Lysine Iron Agar Slant
Tube 1: Positive decarboxylation (butt), negative deamination (slant)
Tube 2: Negative decarboxylation (butt), positive deamination (slant)

Gas production - bubbles
H2S prod. - black precipitate
Name the media/test. Results?
Which set of tests would you set up to identify a Staph? What do positive/negative reactions look like for each test?
Tri-plate
Beta, blood agar
Name hemolysis, and media
Gamma, blood agar
Name hemolysis and media
Salmonella-Shigella (SS) plates are used to detect bacterial contamination on raw meat. Salmonella-Shigella agar plates contain lactose, bile salts, ferric citrate, and neutral red. The bile salts select for Gram-negative bacteria. The other components differentiate among Gram-negative bacteria.

Coliform bacteria such as E. coli will ferment the lactose in the media, resulting in bacterial growth with a pink color. (They do not produce any hydrogen sulfide.) Members of the genus Salmonella will not ferment lactose, but do product hydrogen sulfide gas. The resulting bacterial colonies will appear colorless with black centers. Members of the genus Shigella do not ferment lactose or produce hydrogen sulfide gas, so the resulting colonies will be colorless.
Name the media. Which organisms are on each?