Study guide for Quiz 2 lab PDF

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Jennifer Manista ...

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Study Guide for Quiz 2: Media Chapter 7-10 Quiz 2 will focus on the different media that we learned about this semester. The media that will be covered in the quiz are: TSA, TSB, BAP, MAC, MSA, SAB, UREASE, INDOL, CITRATE AND BILE ESCULIN. Quiz 2 will be worth 15% of your lab grade. Question format will be: multiple choice, matching, true/false corrections, fill-in-the-blank and short answer. GENERAL ●

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Four Different Categories of Media ○ Complex: is media that contains a simple food source that promotes most bacterial growth. (NANutrient agar, TSA- tryptic soy agar, TSB- tryptic soy broth) ■ non-selective means most bacteria will grow on these media ○ Enriched: is a type of complex media that contains special ingredients that can help distinguish different types of bacteria and/or grow bacteria that cannot grow on complex media alone. ■ BAP, chocolate agar, MAC, MSA, indole, citrate, urease, bile esculin, O/F ○ Selective: is a type of enriched agar that contains dyes or toxic substances which will inhibit the growth of certain bacteria while promoting the growth of other bacteria. ■ MAC and MSA ○ Differential: is a type of enriched media which differentiates or distinguishes between different types of bacteria based on differences in appearance of growth or color change ■ Both selective and differential ■ BAP, MAC, MSA, urease, citrate, indole, bile esculin, O/F Describe the steps of inoculating a broth tube-can be inoculated with a loop or a needed, depends on the test you’re performing (TSB, urease, tryptone), before inoculation, the media is a clear, tan color. And after inoculation, you see bacterial growth is making the broth cloudy Describe the steps of inoculating an agar slant tube- Both tubes contain agar, can be inoculated with a loop or a needle, depends on the test you’re performing- (TSA, bile Esculin, O/F). before inoculation the media is tan color and after inoculation, you can see bacterial growth on the agar surface Describe the steps of preparing a streak plate○ Flame and cool loop ○ Take loop and get a loopful bacteria of source ○ Agar plate gently grad loop across first quadrant with bacteria ○ Flame and cool loop ○ Rotate plate ○ Start with loop in first quad and streak across second quad ○ Flame and cool loop ○ Rotate plate ○ Starting in second quad dip in and streak third quad ○ Flame and cool loop ○ Rotate plate ○ Starting third quad streak across the fourth quad or with what’s left ○ Incubate place agar side up at 37 degree Celsius for 24-48 hours ○ Common streak plate errors■ using too much bacteria on the first quad ■ Going back over the source more than once ■ Forgetting to cool the loop after flaming ■ Forgetting to flame the loop between quadrants ■ Going back into previous quadrants ■ Jabbing the agar with the loop ■ Going too fast, not taking my time Explain the purpose of a streak plate- A technique that involves streaking a loopful of bacteria across a solid medium (agar) with the goal of obtaining isolated colonies. Diluting out the bacteria.

TSA & TSB ● ● ● ●

Categories: Selective? Only complex media- this is non-selective and most bacteria will grow on these types of medias What will grow: Most bacteria will grow on TSA unless they need a special diet What will not grow: most Everything will grow What do we use this media for? We use this media for general growth of bacteria

TSA

Blood Agar (BAP) ● ● ● ● ●

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Categories: Differentiate by differ strep. and enrich What will grow: Hemolysis, intacted RBC – page 60 in book What will not grow: nothing What do we use this media for? We use BAP for a general growth media and to observe the different HEMOLYSIS properties of bacteria. Define and describe the three hemolysis we can see on a BAP plate ○ Alpha- “green hemolysis” or incomplete hemolysis. Bacteria produce hydrogen peroxide, changing color of hemoglobin to green ○ Beta- also known as complete hemolysis because RBCs are completely lysed and produced a clear zone around the bacteria colonies ○ Gamma- also called non-hemolytic. No lysis of RBCs. No change in agar. If given a picture, be able to identify the three different hemolysis

MacConkey Agar (MAC) - example in the video ● ● ● ● ● ● ●

Categories: enrich, complex Selective AND differential agar What will grow: Gram negative bacteria (GAMMA hemolytic on BAP) What will not grow: Gram positive bacteria What do we use this media for? We use MAC to observe if the bacteria can ferment _lactose Fermenters will grow & change the media to a __pink colonies_________ color Non-Fermenters will grow & change the media to a __ it will appear colorless_ If given a picture, be able to identify a fermenter and a non-fermenter

Mannitol Salt Agar (MSA) ● ●

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Categories: Selective for gram positive and differential medium between bacteria that can ferment mannitol and cannot ferment mannitol What will grow: Al lSt aphyl ococcus( S.aur eusandS.epi der mi di s) ,onl ysomest r ept ococc us, Gr am posi t i ve ○ S.sapr ophyt i cus ○ S.aur eus ○ Somet i mesent er ococcusf aecal i s What will not grow: high salt content, cannot grow on it ○ Escherichia coli ALL GRAM NEGATIVES WON’T GROW ○ Strep. agalactiae ○ Strep. Pyogenes What do we use this media for? We use MSA to observe if the bacteria can ferment __Mannitol?__This is an acidic byproduct that is formed causing the phenol red in the agar to turn yellow______. Fermenters will grow & change the media to a ___Yellow______ color Non-Fermenters will grow & and change the media to a ___Pink_____ color. If given a picture, be able to identify a fermenter and a non-fermenter

Sabouraud Dextrose Agar Agar (SAB) ● ● ● ●

Categories: selective for mold and fungus What will grow: mold and yeast will grow What will not grow: The acidity of the agar and the presence of antibodies that inhibits bacteria from growing on agar while selecting for fungus What do we use this media for? We use SAB to grow molds, fungi and yeasts

Urease ●

Categories: Complex, enriched, selective, differential

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What will grow: gram negative like proteus mirabilis, E.coli, What will not grow: Gram positive What do we use this media for? We use Urease to see if bacteria can break down urea into ammonia Positive result: pink Negative result: remains orange If given a picture, be able to identify a positive and negative result

Indol ● ● ● ● ● ● ● ●

Categories: Enriched, Differential What will grow: Gram negative, like E.coli What will not grow: Gram positive What do we use this media for? We use Indol to see if bacteria can break apart tryptophan. Determines whether the microbe produces indole from the amino acid tryptophan Need to add __5-10 drops of Kovac’s reagent_________ after incubation in order to complete the test! Positive result: red/pink ring Negative result: Green ring If given a picture, be able to identify a positive and negative result

Citrate ● ● ● ● ● ● ●

Categories: Complex, enriched, selective, differential What will grow: Gram negative, salmonella typhi and proteus mirabilis, E. coli What will not grow: Gram positive What do we use this media for? We use Citrate to see if bacteria can utilize citrate as their sole carbon source are citrate positive and will change the media color to blue Positive result: blue Negative result: remains green If given a picture, be able to identify a positive and negative result

Bile Esculin ● ● ● ● ● ● ●

Categories: Complex, enriched, selective, differential What will grow: enterococcus and group D strep will produce positive results What will not grow: gram negative What do we use this media for? We use Bile Esculin to see if bacteria can hydrolyze esculin in the presence of bile Positive result: media turns black (differential) Negative result: media stays tan (differential) If given a picture, be able to identify a positive and negative result...