Microbiology Lab Week 10

How are agar plates inoculated for disk diffusion testing of antiseptics and disinfectants?

A swab is used to inoculate the entire surface of the plate, producing a lawn of growth.

How is an antiseptic different from a disinfectant?

An antiseptic is used to remove or kill microorganisms on tissue, whereas a disinfectant is used to remove or kill microorganisms on inanimate objects.

Which of the following DNA sequences would be the most likely to be damaged by UV radiation?

CTTGAATTAA

T/F A large zone of inhibition around a disk containing disinfectant generally indicates that the bacteria being tested is resistant to the disinfectant. Group starts

False

For the Disk Diffusion Assay, what would cause a larger clearing size around the filter disk? What would cause NO clearing around a filter disk?

For the Disk Diffusion Assay, more effective disinfectants would cause a larger clearing size around the filter disk. No clearing would mean that the bacteria is not susceptible to the disinfectant.

From the virtual lab experiment, did the length of UV exposure time affect the growth of microbes? Describe the correlation and explain why.

In the virtual lab experiment, the longer the exposure time, the growth of microbes decreased. Because the longer the exposure, the more mutations will occur in the microbial DNA which will more likely lead to cell death.

Why is it important to spread the bacteria over as much of the surface as possible when inoculating your plate for the disk diffusion test? How would this impact your results?

It is important to spread the bacteria over as much of the surface as possible when inoculating your plate because you don't want growth to overlap each other. You need room for the zone of inhibition for each bacteria and if they are too close together, it could lead to unreliable results.

If after performing Lab # 24, you see no growth of bacteria on the entire plate after exposing to UV light after 5 minutes, even underneath where it was covered with material that blocks UV light. What would be your conclusion regarding the experiment? Is the data reliable? Why or why not?

The experiment was under the UV light for too long. The data is not reliable because the control (under the cover) did not grow either.

For the UV light experiment, what effect on bacterial growth would you expect if you increase the time the agar plate/bacteria is exposed to UV light treatment? Explain your reasoning.

The longer the exposure, the more damage to DNA occurs and the less bacterial growth. Because the longer the exposure, the more mutations will occur in the microbial DNA which will more likely lead to cell death.

For Lab #25 Effects of UV light, what was the purpose of covering half of the plate prior to exposure? What would you conclude if nothing on the plate grew?

The purpose of covering half of the plate prior to exposure was so that we can have a control group that we can compare the experimental group to. If there was nothing on the plate that grew, the bacteria was exposed to the UV light too long.

Explain how the size of the Zone of Inhibition relates to the effectiveness of a disinfectant/antibiotic. Explain the correlation?

The zone of inhibition is a clear zone that will appear around the discs where the chemical agent was able to inhibit the growth of the inoculated organism. The correlation between the zone of inhibition and the effectiveness of the disinfectant is that the larger the zone of inhibition, the more sensitive the microbe is to the disinfectant in the disk.

When exposing your bacteria to UV light, how might the results differ if a UVA lamp compared to a UVC lamp?

UVA exposed plate would have more bacterial growth

Which range of the UV spectrum is the most biocidal?

UVC

Which of the following infectious agents is least likely to be damaged by UV exposure? - protozoa - prions - bacteria - fungi - viruses

prions

UV light has ______ wavelengths than visible light waves, and its peak energy is at a wavelength of ______.

shorter; 254nm

Which of the following would be appropriately sterilized using UV radiation? - stainless steel surface in biological safety cabinet used for media preparation - flask of nutrient broth - stack of plastic petri dishes, prior to pouring media - nutrient agar plates with lids

stainless steel surface in biological safety cabinet used for media preparation

What might happen if you forget to remove the cover of the petri dish before exposing it to UV light for Lab #25- Effects of UV light?

the cells would not be affected by UV light treatment and will all grow normally because UV cannot easily penetrate the plastic cover

When two adjacent thymine molecules are joined by a covalent bond as a result of exposure to UV light, this structure is referred to as a

thymine dimer.

The clear area devoid of bacterial growth around an antiseptic or disinfectant disk is referred to as the

zone of inhibition.

What two factors increase the effectiveness of a disinfectant on microorganisms? - Concentration of disinfectant - Time of exposure - Material used to apply disinfectant - Corrosiveness of the disinfectant

- Concentration of disinfectant - Time of exposure

Which structures are less susceptible to UV damage? - Fungal spores - Endospores - Protozoan cysts - Vegetative bacterial cells

- Endospores - Protozoan cysts

Which of the following would affect the size of a zone of inhibition around an antiseptic or disinfectant disk? - The susceptibility of the microorganism to the chemical - The concentration of chemical in the disk - The cellular target of the chemical - How quickly the chemical diffuses through the agar - The depth of the agar

- The susceptibility of the microorganism to the chemical - The concentration of chemical in the disk - How quickly the chemical diffuses through the agar - The depth of the agar

Which of the following are limitations of using UV radiation for disinfection? - UV radiation does not penetrate through paper, plastics, or liquids - UV radiation damages plastics and other nonmetal materials - UV radiation destroys bacteria but not viruses - UV radiation can damage the skin and eyes

- UV radiation does not penetrate through paper, plastics, or liquids - UV radiation can damage the skin and eyes

Place these types of bacteria in order of their susceptibility to chemical disinfection, from least susceptible to most susceptible. - Bacterial endospores - Gram-positive bacteria - Gram-negative bacteria - Acid-fast bacteria

1. Bacterial endospores 2. Acid-fast bacteria 3. Gram-negative bacteria 4. Gram-positive bacteria

Place these steps in the order in which they will be performed when setting up a disk diffusion assay to test the effectiveness of disinfectants and antiseptics. - Using a swab, inoculate a suspension of the bacteria being tested onto an agar plate, spreading the inoculum over the entire surface. - Saturate sterile filter paper disks with the test chemical (disinfectant or antiseptic) and place onto the inoculated agar plate. Repeat with each chemical being tested. - Incubate inoculated agar plates overnight to promote bacterial growth. - Observe plates to determine the presence of a zone of inhibition around each antiseptic or disinfectant disk.

1. Using a swab, inoculate a suspension of the bacteria being tested onto an agar plate, spreading the inoculum over the entire surface. 2. Saturate sterile filter paper disks with the test chemical (disinfectant or antiseptic) and place onto the inoculated agar plate. Repeat with each chemical being tested. 3. Incubate inoculated agar plates overnight to promote bacterial growth. 4. Observe plates to determine the presence of a zone of inhibition around each antiseptic or disinfectant disk.