Micro Bio 3150

A bacterial cultured is diluted 10,000 times through serial dilutions. 0.1 ml of the diluted culture are plated on a plate. After incubation, 150 colonies are observed growing on the plate. How many organisms were in the original (undiluted) culture?

1.5x107 CFU/ml

Assume that a DRT value for autoclaving a culture is 1.5 min. How long would it take to kill all the cells if 106 cells were present?

10.5 min

The top magnification of a light microscope is approximately

1000x

The best objective to determine the morphology of prokaryotes

100X

Place the steps of the Gram stain in the correct order: 1-Alcohol-acetone; 2-Crystal violet; 3-Safranin; 4-Iodine.

2,4,1,3

Place the following reactants in their proper order for the ELISA test we did 1 = enzyme-linked secondary antibody 2 = primary antibody 3 = antigen 4 = substrate

3-2-1-4

The table below shows the results of a hypothetical experiment. What is the thermal death time for this species at 50°C. 2 min 5 min 10 min 30 min 37°C +++ +++ +++ +++ 50°C ++ + + - 60°C + + - - 100°C + - - -

30 minutes

In this experiment you used 10 µl of pGLO plasmid at a concentration of 0.08 µg/µl. If you plated 200 µl out of the 500 µl of the solution and you got 100 colonies in the +pGLO LB+amp plate. What is the transformation efficiency?

312 CFU/µg of DNA

The Control of Microbial growth Moist and Dry Heat Sterilization: Thermal Death Point and Thermal Death Time 6. Know the physical agents that affect bacterial growth (temperature, UV, drying/osmotic pressure). 7. Know the effect of moist heat (denaturation of proteins) in controlling bacterial growth 8. Know how to determine the minimal amount of heat necessary for complete destruction of bacteria for a fixed amount of time (thermal death point) 9. Know how to determine the minimum amount of time required for bacterial death at a given temperature (thermal death time) 10.Know how to determine the decimal reduction time (DRT): the amount of time necessary to destroy 90% of bacteria (10% survive) 11.Know how to sterilize heat-sensitive liquids 12.Know at least one reason why some strains show a high resistance to heat

6) Temperature, Ultra-violet light, Drying, and Osmotic pressure are all physical agents that can effect bacterial growth. 7) Moist heat denatures protein, destroying 3-dimensional form, rendering it useless 8) Determining the temperature required to kill bacteria in 10 minutes is discovered only via a graph. 9) Thermal Death time is seen via a graph as well. Look for the first time where the bacteria died. ( see problem 33) 10)

What is the thermal death point for the species shown in the table in the previous question?

60

What does aseptic mean?

Absence of contamination

What is an advantage of the pour-plate technique over the streak-plate technique?

Allows to calculate bacteria concentration (CFU/ml) in the original sample

According to our rules, you should disinfect your bench at a minumum:

At the end of the lab

Laboratory media is most commonly sterilized by:

Autoclaving

Why might Gram positive bacteria incorrectly appear to be Gram negative after the Gram staining procedure

Because the culture was older than 24 hours Because the alcohol was left too long (more than 20 sec typically) on the slide only A and B

Match the antibiotic with its mode of action

Chloramphenicol-prevents peptide bond formation during translation Ciprofloxacin (a quinolone)-interfers with DNA replication Trimethoprim (a sulfa drug)-inhibits purine and pyrimidine synthesis Penicillin- inhibits crosslinking of cell wall's petidoglycan

What is the key test that separates Staphylococcus aureus from other staphylococci?

Coagulase Test

Wich of the following are considered virulence factors?

Coagulase, and DNAse

In moist heat, the microbial proteins undergo ____________, a process in which the three-dimensional form of the protein is destroyed and the protein loses its function.

Denaturation

What is the primary use of agar deeps?

Determine motility

In our pGLO lab experiment, which of the following will fluoresce under UV light?

E. coli (+pGLO) on LB amp/ara plates

Staphylococcus and Streptococcus are gram negative

False

Your partner streaks a plate from a broth culture. After incubation, you see confluent growth in all sectors. Which of the following errors could NOT explain this result?

He did not pick a big enough sample from the broth

The inoculating needle is used for:

Inoculating agar deeps

How would you sterilize a heat-sensitive growth medium containing thermoduric bacteria?

Membrane filtration

What agar is used on Kirby-Bauer Tests?

Mueller Hinton Agar

In the previous question, what would happen if you stop the heating process at 9 min?

One cell would be alive

What is a bactericidal antibiotic?

One that kills bacteria.

What is a broad spectrum antibiotic?

One that kills many different bacteria species

In our experiment, which antibiotic was narrow spectrum?

Penicillin

Which of the following behaviors is acceptable in the microbiology laboratory?

Placing a laboratory notebook on the bench

In a molecular biology laboratory, a student obtained competent E. coli cells and used a common transformation procedure to induce the uptake of plasmid DNA with a gene for resistance to the antibiotic kanamycin. The results below were obtained. On which petri dish do only transformed cells grow?

Plate II ( LB agar with kanamycin + Kan plasmd)

Which of these plates will provide a control to make sure that the normal (nontransformed) E. coli can grow under the conditions used for this lab?

Plate III ( LB agar, no plasmid added)

What is the explanation for the results in the previous question?

Plate III has antibiotic agar, but E. coli that has been transformed to be resistant to tetracycline can grow.

Which of the plates is used as a control to show that nontransformed E. coli will not grow in the presence of kanamycin?

Plate IV ( LB agar with Kanamycin, no plasmid added)

What is the most serious pathogen within the genus Staphylococcus?

Staphylococcus Aureus

All of the following are true about coagulases, except:

Staphylococcus epidermidis is coagulase positive

Which species of bacteria appears as spheres in chains?

Streptococcus lactis

A student has forgotten which antibiotic plasmid she used in her E. coli transformation. It could have been kanamycin, ampicillin, or tetracycline. She decides to make up a special set of plates to determine the type of antibiotic used. The plates below show the results of the test. Which antibiotic plasmid has been used?

Tetracycline

What is known as the "zone of inhibition"?

The clear zone around the disk where growth has been inhibited

The following statements are false EXCEPT:

The final dilution in a serial dilution is the product of all the dilutions that led up to it.

What is the most diagnostic species characteristic of Staphylococcus aureus?

The production of Coagulase

Suppose you don't get any colonies on plates II and IV. How would you explain this?

The transformation didn't work.

The lowest temperature at which all microorganisms are killed in 10 minutes is

Thermal Death Point

DNase positive strains can be distinguish because

They show a distinct clear zone around the streak where the DNA in the agar was broken down by bacterial DNAse

What is the purpose of the arabinose?

To bind to and inactive the repressor protein

Which species of bacteria is spiral?

Treponema pallidum

How do you determine when the loop is cool?

Trial and error.

Staphylococcus aureus is DNAse positive while S. epidermidis is DNAse negative

True

In the indirect ELISA, what might cause some positive results to be lighter in color than others?

Weak positive results may be an indication that the patient's blood serum carries few antibodies against the disease-causing agent The patient's exposure to the pathogen may be recent and the body may not have launched a full immune response yet (low antibody titer) The infection may have occurred long ago, and the level of antibodies in the patient's bloodstream is declining all of the above

If the resolving power of your microscope is 500 nm. Which of the following can NOT be seen with it?

a 200 nm virus

The direct ELISA test requires

a known antibody absorbed to the wall of the microtiter well

The inoculating loop should be flamed:

before an after use

You have a patient that has a spreading blood infection. You need to determine whether it is caused Staphylococcus or Streptococcus. A rapid test you could do is to

catalase test

The time for destruction of 90 percent of the microbial population is the

decimal reduction time

Cross contamination can result in false negative results

false

Mannitol salt agar is selective for staphylococci because of the ______________

high salt conc

In which growth phase is an organism most sensitive to an antimicrobial agent?

log phase

The purpose of streaking a plate is to

obtain isolated colonies in order to prepare a pure culture

What reasons can account for a false negative result?

patient's antibody level is too low, you forgot substrate, you forgot to add secondary antibody; conjugated.

The thermal death time at 100°C will be _____________ the one at 60°C.

shorter than

Inoculating loops are used for

streaking plates Correct Answer a, b, and c only Inoculating agar slants You Answered Inoculating broths

In our experiment in the lab, we determined

the Thermal Death time

In the indirect ELISA test the enzyme-linked antibody will attach to

the patient antibody

The picture below shows the results of a direct ELISA for HIV testing. Reactions are done in duplicate, left to right are the positive control, negative control and two patient samples: A (following the negative control) and B to the far right

the patient has antigens for HIV virus

In our lab, why did you perform three identical tests for each control and patient sample?

to ensure reproducibility of the results

In the indirect ELISA test the development of color means the patient has the antibody being tested for.

true