1

Case Study 8

Student

Professor

Institution

Date
 2

Part I —Return to Sender

1. Using light microscopy, you examine the soil samples and the “goo” from the

degraded polyurethane. Will this approach allow you to observe all microorganisms

present in the samples? Why or why not? What are the limitations of this

approach?

Light microscopy examines tiny specimens using various kinds of light. Bright-field microscopes

are the most frequent. When gazing through the microscope, this lit the backdrop. For 'goo' made

from deteriorated polyurethane, this method will work (Sofos, 2015). It would be a compound

microscope with a spinning noise piece and numerous lenses with various magnifications which

could zoom in on the specimen up to 1000x. As a result, the sample contains a large number of

bacteria. "To examine dead stained specimens and natural colorant living specimens; also used to

enumerate microorganisms," bright-field microscopes are employed. This method has limitations

such as too much light flowing through and too little contrast, making it difficult to view the

specimen. Due to the available wavelengths of electrons, the electron microscope has a

significantly higher resolving capacity (0.01nm to 0.001nm). Because it has a shorter, longer

wavelength, it has a higher resolution. "The capacity to identify items that are close together" is

defined as "resolution." The soil includes many specimens that are incredibly close together.

Thus an electron microscope would be preferable, although optical microscopes can still be used.

2. You use phase-contrast microscopy to observe a wet mount of a soil sample (the first

picture below) and a "goo" sample (the second image below) from the ELVIS. In

what ways are the potential ET microbes similar to microbes previously

characterized on Earth? In what ways are they different? How could you determine
 3

whether the microbes in the soil or goo samples are phylogenetically similar or

distant from known microorganisms on Earth?

Some prokaryotic cells compare to ET microorganisms. You may compare the forms of

prokaryotic cells' morphology, which includes "genetically encoded phenotypic that reflect the

growing environment, cell wall, and membrane components, as well as specific structure

defining proteins." Some of the microorganisms in a sample of soil seem to be coccus-shaped,

filamentous bacteria-shaped, rod-shaped, and coccobacillus-shaped (Stewart et al., n.d.) In

addition, the samples are 1um in size, whereas a prokaryote cell is generally 0.5um in size. When

it comes to differentiating cells, size makes a significant difference. For example, if the

specimens were closer to 10um, one could assume it was a eukaryotic cell, typically that size.

The RNA nucleotide sequence may be used to determine if the bacteria are phylogenetically

comparable or far from recognized microbes on the universe. During microorganisms’ similarity

evaluation, the 16S RNA sequence and structure are critical. "Due to the quantity of secondary

structure in the sequence, roughly two-thirds of it is base-paired, assuring positional homology."

The more similar the pattern and structure, the more similar the microbes are.

3. Your boss has done a little reading about microorganisms, but he finds it all pretty

complicated. “It’s like a foreign language!” he complains. “I have to face the press

to explain our idea that microbes might be responsible for all the damage to ELVIS.

I think that it will help my press conference presentation a lot if I can use some

visual aids. What I want to do is explain to my audience what bacteria look like.

You know… the functional architecture of bacteria and how they might be able to

degrade polyurethane. I think that eukaryotes might be too complicated for this
 4

audience, so I just want to show them what Gram-negative bacteria look like in a

schematic diagram. I’ve put together this diagram of a typical Gram-negative cell.

Take a look at it and make any corrections you think are necessary. Notice that I

not only labeled the features, I also indicated the major biochemical composition

and function(s) of each main feature. Oh yeah … this figure will probably make it

into lots of newspapers, magazines, and web sites, so it needs to be scientifically

correct. We wouldn’t want to make DuPont look stupid, would we? I’ve already

done most of the work. Just proofread it and make any necessary corrections.”

Bacteria are prokaryotes, for starters. They do not have a nucleus or any membrane-bound

organelles. Thus they do not have a nucleus. As a result, the label D is incorrect. It ought to be

called a nucleoid. Circular DNA with no histone protein is found in this area. In addition, the

cytoplasmic membrane is the innermost layer of the bacterial cell membrane, followed by the

cell wall, and finally, the capsule (Stewart et al., n.d.). The cytoplasmic membrane is B, the cell

wall is E, and the capsule is G. On the figure, C is incorrectly shown. The cytoplasm is

represented by the letter C. The periplasm or periplasm space, on the other hand, "contains the

peptidoglycan and periplasm." As a result, the bacteria's peptidoglycan layer would detect it. The

designation F for the pilus is likewise incorrect. A pilus is a kind of fimbria that is longer than

fimbriae but shorter than flagella. They employ them in a process known as recombination to

transfer DNA from one cell to another." It appears to be a flagella rather than a pilus, based on

the structure. Cilia (label A) is likewise incorrect. Cilia are smaller and more abundant than

flagella and expand the cell's surface. Cilia are not seen in bacterial cells." The structure that

looks like cilia is most probably pili, which also aids in cell navigation. The peptidoglycan wall

must also be thinner because it is effective at inhibiting.

Part II —Suspicious Minds

1. How would you go about isolating your pure culture?

To isolate the pure culture, I should start with diluting sequenced. The collaborative culture

would indeed be placed onto solid media using an inoculating loop to separate the different

microorganism cells from one another (Stewart et al., n.d.). I would utilize the aseptic

method during the operation to guarantee that I do not compromise the culture. I would study

the colonies that developed after the first streaking, noting variations in morphology. Each

colony with a distinct morphology would be scattered onto its fresh solid medium plate to

create pure cultures. This method would be repeated as needed until only colonial

possessions with the same morphology remained in the medium.

If your goal is to characterize the ETPUM, whose results are more informative: yours

or your boss’s? Why? What do your results indicate about the nature of this microbe?

Does its biochemical composition most closely resemble that of a prokaryote or a

eukaryote? Gram-positive or Gramnegative? Do you agree with your boss’s conclusion

that the ETPUM is a prokaryotic-eukaryotic hybrid? Why or why not?

My findings are more illuminating than ETPUM when it comes to describing the ET

polyurethane-degrading microbe. Because my supervisor chose to analyze the soil sample

immediately, which may contain more than one microorganism, the testing findings on the soil

sample are misleading (Stewart et al., n.d.). Aside from the ETPUM, the soil sample might

contain many microorganisms, indicating that the bacterium in question had both eukaryotic and

prokaryotic characteristics. Because my experiments were conducted on a pure isolated colony,

they are more valuable in identifying a single bacterium.

My analysis shows that the bacteria are motile because it contains flagella basal cellular

proteins. The presence of pilus proteins implies that the bacteria may transmit DNA to certain

other microbes. The microbe's biochemical makeup is similar to that of a prokaryotic cell since it

has 70S ribosomes, circular DNA, LPS, flagella body temperature proteins, and pilus peptides

(Stewart et al., n.d.). These features are exclusive to prokaryotes. The 80S ribosomal, linear

DNA, associated nucleotide proteins, and chromatin proteins, all found in eukaryotes, were not

found in my sample. The microorganism is Staphylococci because of the presence of LPS but not

lipopolysaccharide.

In my perspective, ETPUM is not a unicellular organism’s hybrid. My employer's sample

may have had a mixture of prokaryotic and eukaryotic germs because it was not pure cultivation

(Sofos, 2015). When comparing my pure product to the elements found in my owner's soil

sample, it's evident that I included not all of the chemicals included in the soil specimen in my

pure sample. According to my pure sample, ETPUM is a prokaryotic cell.

2. Come up with at least two possible alternative explanations for the “amazing”

redistribution of the ETPUM on the Lycra-sealed slide. Both of your explanations

should consider how the microbes “sensed” the presence of polyurethane. One of

your answers should not involve flagella.

The ETPUM's redistribute on the Lycra-sealed slides is likely caused by two factors: taxis and

the ETPUM's motility. A taxi is an automobile that responds to external stimuli by moving. The

ETPUM experienced the most excellent chemotaxis when it recognized Lycra, which contains

polyurethane, a portion of food for the bacteria. The ETPUM targeted the nutrient. The creature

also has flagellum, per the tests done on the sample produced. The flagellum directs the

microbe's movement; cell receptors provide information to the flagella, which changes the
 7

flagella's velocity and distance. The ETPUM microorganism used its flagella to disperse itself on

the Lycra-sealed slide.

Part III —All Shook Up

1. Which medium would you consider to be “complex” and which “defined”? Which is

“rich” and which is “minimal”? Explain your answers.

Because Medium 1 comprises 5 grams of yeast extract and 20 grams of trypticase extract, a

glucose yeast extract, it isn't straightforward. A complex media contains yeast, animal, or plant

tissues, as well as unknown compounds (Stewart et al., n.d.). Because it contains a carbon source

(glucose), inorganic ions, and water, Medium 1 is also a minimum medium. Medium 2 is a

defined medium since all of its constituents are pure biochemicals with precise measurements.

It's also a rich medium since it provides just the proper nutrients for organisms to thrive.

2. Given that polyurethane is a considerable polymer (MW >>100,000, Daltons), why

is it essential that the polyurethane is a secreted enzyme?

Because polyurethane is required to break down polyurethane, it must be a secreted enzyme.

Because polyurethane is a vital source of energy for the bacterium, it must be digested to

develop. The polyurethane enzyme is then secreted, degrading the polyurethane and releasing

citrate.

Suppose we assume that the polyurethane is the source of energy for the organism.

How can material (carbon atoms) from it find its way into the central metabolic

pathways of this microbe?

If polyurethane is used as a source of energy, it must first be broken down by bacterial

exoenzymes. Then, by enhanced diffusion, it would be absorbed into the cell.

What is the “entry point”?

Carbon enters the organism through the membrane and is catabolized.

What happens after it enters the metabolic pathway?

The organism can grow.

3. Why does the growth of ETPUM in Medium #$ require oxygen? Think about this in

terms of how ETPUM can generate a net gain in ATP by processing polyurethane.

Remember that the degradation of polyurethane by polyurethanes does not expend

ATP. To answer this question, address each of the following questions in your

answer:

a) Is there a net gain or loss of ATP during the transport of the citrate?

There is a net loss of 2 ATP.

b) Consider the ATPs that can be generated via substrate-level phosphorylation. Will

glycolysis help generate any ATPs during growth on polyurethane?

Because glycolysis provides a net gain of ATPs, it may be used to generate ATP. Per cycle,

one glucose produces two ATP. Organic molecules can be employed as the primary electron

donor once ETPUM has gone through yeast fermentation.

How many ATPs can be generated via TCA?

2 ATPs will be generated through TCA.

Is this enough to support growth (is there a net positive in the ATP tally)?

Yes, there is enough net positive ATP tally to support growth.

c) Now consider how else ETPUM can generate ATPs (if not by substrate-level

phosphorylation). Can this process generate a net positive in the ATP tally?
 9

Using oxygen during ATP synthesis is more efficient than just using other kinds of

respiration. Its function as an electron acceptor on the ETC permits the substrate to go

through the glycolysis process.

d) Now explain the importance of oxygen as it relates to the ATP tally.

Anaerobic respiration produces 2 ATP from a six-carbon glucose molecule, whereas aerobic

metabolism produces 38 ATP from a six-carbon glucose molecule. The respiratory system in the

mitochondria, which produces ATP, is a redox process. In the adenosine triphosphate process,

oxygen serves as the electron acceptor (Stewart et al., n.d.). This process is catalyzed by ATP

synthase. In the glycolysis process, both aerobic and anaerobic degradation of glucose molecules

generates pyruvate and two ATPs. In aerobic cellular respiration, pyruvate is transformed to

Acetyl-CoA, which then enters the Krebs cycle and the adenosine triphosphate process, yielding

36 ATP. The pyruvate is transformed to lactate in anaerobic respiration, but no further

respiration happens.

Part IV —A Little Less Conversation

1. At a press conference announcing your company’s successful isolation and

characterization and cultivation of ETPUM, a reporter raises an important

question: “How do you know that this microbe actually came from NPIP (the

Nearby Previously Invisible Planet) and not from Earth? Could this microbe really

be an Earth microbe that was present in/on ELVIS before it was launched into

space?” “it is impossible,” your boss answers. “Prior to launch, we wiped down the

entire ELV and ELVIS with the disinfectants ethanol and triclosan. In addition we

used plastics impregnated with disinfectant chlorhexidine. I had our chemists check!

See chemicals do all kinds of nasty things to cells: coagulate proteins and destroy
 10

membranes. No microbes could have survived that.” What do you think? Were

these treatments adequate to rule out the possibility raised by the reporter?

For bacteria, standard disinfection and aseptic measures are sufficient, but for spores, they

are not. Spores are resistant to high temperatures and chemicals and may readily dodge them.

2. You are a consultant for a second ELVIS mission to NPIP. Describe what physical

and chemical treatments you would require before the lift to minimize the

opportunity for contamination of the ELV (the landing module of the spacecraft)

and ELVIS (a new version of the robot) by Earth microbes. HINT: Remember the

following: ELVIS's skin cannot withstand temperatures above 90°C; the ELV itself

is approximately the size of a minivan and includes many metal parts. Use scientific

terminology to discuss this answer: e.g., are you trying to achieve disinfection or

sterilization? Are you recommending the use of antiseptics or disinfectants? Specify

how each of your treatments would achieve the killing of microbes.

Disinfectants would not be used since disinfectants only stop germs from growing as long as

they are present. The most effective technique would be sterilization. Sterilization kills all

live bacteria (Stewart et al., n.d.). Therefore I advocate using physical and chemical

sterilization procedures to keep as many microorganisms from developing as possible.

Ethylene Oxide, Ozone, Bleach, Glutaraldehyde, and Formaldehyde are examples of

chemical sterilizers. Radiation and other physical sterilizers are examples of physical

disinfectants.

3. In your opinion, is EPTUM an earthly isolate or an extraterrestrial isolate? Some

things to think about: Have bacteria been isolated from outer space to date? What is
 11

this field of study? What agencies fund this field? What is “Planetary Protection”?

Are there earthly microbes that have not yet been isolated and grown in culture? If

so, what is the predicted percentage? Are there methods to study microbes that

cannot be cultured?

I assume it is an earthly isolate microorganism based on the info that I supplied. Because we're

dealing with alien microorganisms, nothing is considered out. We do not even know about 75%

of the microorganisms on EARTH, let alone in space. It is possible to achieve anything.

References

Sofos, J. N. (2015). Microbial Growth - an overview | ScienceDirect Topics. Sciencedirect.com.

https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-

biology/microbial-growth

Stewart, R., Smith, A., & Shields, P. (n.d.). ELVIS Meltdown! Microbiology Concepts of

Culture, Growth, and Metabolism Part I-Return to Sender. Retrieved September 26,

2021, from https://sciencecases.lib.buffalo.edu/files/elvis.pdf