Lab exam 2 micro

Selective Media Learning Objectives

1. Distinguish between selective and differential media, and between defined and undefined (complex) media. Give an example of each. Can a medium be both differential and selective?

Give several examples of medium components that make the medium undefined.

• Differential media include ingredients that allow us to visually distinguish (or differentiate) one group of bacteria from another based on metabolic features. Selective media contain ingredients that prevent the growth of certain bacteria.

•  Defined media get their name from the fact that each organic and inorganic ingredient is precisely known and quantified, which means these media are accurately reproducible. complex media contain ingredients such as milk proteins, blood, or yeast extracts, less precisely defined organic nutrients.

• Ex differential = starch agar, ex selective = PEA agar, enriched (complex) = PEA, defined (synthetic) = MR-VP broth

• EMB is selective and differential

• Complex (undefined) media uses milk proteins, blood, or yeast extracts to make medium undefined.

2. What characteristic/feature of the medium makes it selective? Differential? Explain and give specific examples.

• inhibit growth of certain bacterial. Differentiates on groups of bacteria from other ones. Differential = nitrate broth, selective = MSA 

3. Describe the nature and purpose of indicators in media. What is a common type of indicator and how do they work? Give specific examples of indicators used in the media studied in lab.

• substances added to a culture plate or broth that visually change color or appearance when specific biochemical reactions occur, allowing researchers to differentiate between different types of microorganisms based on their metabolic properties. An example of an indicator is phenol red which works by changing color due to pH.

4. Create a table (landscape orientation) listing phenylethyl alcohol agar, mannitol salts agar, eosin methylene blue agar, and Hektoen enteric agar. For each indicate (a)Type, whether it is defined or undefined and selective and/or differential; (b) Application/purpose and (c) Mechanism of action/theory. 

• refer to exam 2 study guide

5. Which of the above media can be used to isolate gram-positive cocci? What are some examples of common gram-positive cocci in human samples? 

• MSA, s.epidermis and s. aureus

6. Which of the media in LO 4 is/are used to isolate gram negative rods? What are some examples of common gram-negative rods in human samples?

• HE (maybe EMB??), salmonella, and shigella

7. What is the significance of the bacterial family Enterobacteriaceae? Explain what is meant by the terms enteric and coliform.

• Enterobacteriaceae family members are increasingly resistant to carbapenems along with many other antibiotics.All the enteric bacteria belong to the family Enterobacteriaceae and can ferment glucose to produce acidic end products whereas, coliforms are a subgroup of enteric bacteria, which are ubiquitously present in the feces of the animals.

8. Give two reasons why clinical microbiologists would use selective media. Give a reason why environmental microbiologists would use selective media.

• EMB can be used to isolate human pathogens in a healthcare setting and in environmentally through water supply. MSA can be used to isolate different staphylococcus species that case disease.

9. Explain what is meant by a test’s sensitivity and specificity.

• sensitivity" refers to a test's ability to correctly identify individuals who have a disease (true positives), meaning it minimizes false negatives, while "specificity" refers to a test's ability to correctly identify individuals who do not have the disease (true negatives), minimizing false positives
  
  

Differential Tests- IMViC and SIM Learning Objectives

1. Explain what is meant by the IMViC acronym and give the general purpose of these tests.

• I = indole test, M = methyl red test (MR), V = voges-proskasaur test (VP), C = citrate test. MR-VP differentiate bacterial species based on the fermentation pathway used to metabolize glucose. Indole test allows us to assess bacteria for indole production. Citrate test reveals if a bacterium can use citrate as its sole carbon source.

2. Explain mixed acid fermentation and identify which test is used to confirm its occurrence.

• In mixed acid fermentation, the pyruvate made in glycolysis is metabolized to form a variety of end-products such as lactate, formate, acetate, succinate, ethanol, carbon dioxide (CO₂), and hydrogen gas (H₂). MR-VP is used to confirm this occurrence.

3. Create a table including the 4 IMViC tests and for each (a) identify what indicates a positive result (include reagent where applicable) and (b) what a positive result indicates (interpretation).

• Refer to exam 2 study guide

4. Identify two pH indicators used in IMViC tests and for each explain how it is used.

• MR-VP uses pH indicator methyl red which is used to indicate a positive test result if the pH is between 4.4. A negative result is indicated at a pH of 6.2 (yellow). The citrate media contains bromothymol blue in the citrate test. In a positive test at a 7.6 pH and negative at 6.9.

5. Describe the purpose of the citrate utilization test. 

• refer to question 1

6. Explain the basis (theory) for the citrate utilization test.

• The basis of the citrate untilization test is that if the bacteria is able to use citrate as it’s main source of carbon then they produce ammonium and ammonium hydroxide they make the media basic which leads to a positive result. if they cannot the media stays the same color (green).

7. Explain what is meant by the acronym SIM, giving the general purpose of each test.

• S = sulfur reduction, I = Indole test production from trypotophan, M = motility. refer to exam 2 study guide.

8. Describe the actions of cysteine desulfurase, thiosulfate reductase, and tryptophanase.

• cysteine desulfurase catalyzes the hydrolysis of the sulfur-containing amino acid cysteine to pyruvate, ammonia (NH₃), and hydrogen sulfide (H₂S) during putrefaction. thiosulfate reductase catalyzes the reduction of sulfur (in the form of sulfate) to H₂S at the end of an anaerobic respiratory electron transport chain.  tryptophanase can hydrolyze the amino acid tryptophan to pyruvate, ammonia (by deamination), and indole.

9. Describe how to perform and interpret sulfur reduction, indole production, and motility tests. Explain why assessing these characteristics is useful.

• Each of the respective media is stabbed with a needle containing bacteria. After that kovac’s reagents to yield results. 

10. Describe how E. coli and Enterobacter aerogenes differ in their IMViC reactions.

• Both are SIM motile, EC = Indole +, EA = indole -, EA = citrate + (growth), EC = citrate - (no growth), EC = MR +, VP -, EA = MR -, VP +

Differential Media: Respiration and Fermentation Learning Objectives

1. Identify the 3 components of phenol red broth and give the function of each component.

• A protein source, phenol red, and a carbohydrate (glucose, lactose, or surcose). Phenol red = indicates pH change, carbohydrate acts as the fermentable substrate, and provides essential nutrients for bacterial growth.

2. Describe the appearance of phenol red broth tubes with the following reactions: acid, acid plus gas, alkaline. Explain what each of these reactions indicates and give the standard symbols for each.

• Acid = yellow broth no bubble, acid plus gas = yellow broth + bubble, alkaline = pink broth + no bubble

3. Give the functions of catalase, cytochrome c oxidase, and nitrate reductase, including the substrate and product(s) of each.

• Catalase, an enzyme in aerobes, microaerophiles, and facultative anaerobes, converts hydrogen peroxide to water and gaseous oxygen. cytochrome c oxidase, a participant in certain aerobic electron transport chains (ETCs).  nitrate reductase to perform a single-step reaction in which a reduction reaction reduces nitrate to nitrite (NO₂^–).

4. Describe how hydrogen peroxide is formed during cellular respiration.

• as a byproduct of electron leakage from the mitochondrial electron transport chain, where oxygen is partially reduced to superoxide radicals, which are then converted to hydrogen peroxide by the enzyme superoxide dismutase

5. Explain the basis (theory) of the oxidase test.

• The basis of the oxidase test is to see if a bacteria produces cytochrome c oxidase which is found in the ETC. In theory, the test uses redox reactions and TMPD to change the color if the bacteria contains cytchrome c oxidase (purple/dark blue).

6. Explain why E. coli is oxidase negative even though it can carry out aerobic respiration.

• cells can still perform aerobic respiration even though they are oxidase negative because the test only determines that it lacks cytochrome c oxidase

7. Describe the two possible ways to observe a positive test for nitrate reduction.

• In nitrate reduction if after both reagents are added the sol’n is red you can get a positive result. However if there is no color change zinc can be added and if there is still no change in color (yellow) it is positive.

8. Describe the clinical applications of each of the following tests: phenol red broth, catalase test, oxidase test, and nitrate reduction test.

• Phenol red = helps identify pathogenic bacteria in urine, throat, blood etc.

• Catalase = differentiates staphylococcus positive and streptococcus negative species

• oxidase = Used to identify pathogenic oxidase positive species (Neisseria, Moraxella, Campylobacter, and Pasteurella genera)

• Nitrate = differentiate Neisseria gonorrhoeae