Take Two and Call Me in the Morning A Case Study in Cell Structure and Function

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Take Two and Call Me in the Morning A Case
Study in Cell Structure and Function

• by
• Peggy Brickman
• University of Georgia

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Part I - Mysterious Illness

• Well, Becky thought, being a dorm counselor for
  freshmen was not going to be that bad. She got a
  free room for the year and the food was plentiful
  - free steaks last week at an outdoor BBQ
  followed by a hay ride in a horse-drawn wagon in
  their welcome celebration.
• But, then again, it wasnt perfect she had ended
  up covered in bug bites some of the students got
  sick from eating steak that was burned on the
  outside and raw in the middle the horses had
  mucked up the courtyard and pigeons had roosted
  on the dorm roof.
• At least tonight the students were finally
  settling in and quieting down, she mused.

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Part I, continued

• The quiet was shattered a few minutes later when
  one of the other counselors, Ann, yelled through
  her door
• Becky, weve got a problem. One of the students
  found a homeless kitten, and the girl has been
  keeping her in her room. I only found out because
  the girl, Ellie, just came to my room complaining
  of being sick. I felt sick too when I saw the
  mess that kitten made. I thought cats were born
  housebroken, but I guess not.
• Anyway, now I think Ellie might really be sick,
  Ann continued. Shes feverish and says shes
  going to throw up.

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Part I, continued

• What do you want me to do? Becky asked.
• Im freaking out! Ann answered. Forget about
  the mess, just help me figure out what to tell
  them at the health center. I dont know what
  shes been exposed to. Or what weve been exposed
  to for that matter! This is the second girl this
  week with aches, fever, and nausea.
• My Mom sent me a bunch of medicine, Becky
  answered. Ill make a list of where weve been,
  what weve eaten, and what weve possibly been
  exposed to. Then we can start taking something
  right away to keep from getting it, too.

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Beckys Task

• The Health Center will be using the differences
  between organisms to diagnose and treat Ellie.
• In the next 2 minutes, list the clues in the
  story that help you identify how Ellie could have
  contracted a disease with flu-like symptoms. Come
  up with possible suspects (organisms) that could
  cause her to be sick.

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Your Task

• Becky did an Internet search and found 5 possible
  suspects that could be causing Ellies illness.
• During this class session we will investigate the
  differences between them.
• Organisms are usually distinguished by the
  characteristics you listed in your homework
  answers to Table 1. Add any details you missed so
  that when you hear the results of the Health
  Center tests you will be able to figure out what
  was making Ellie sick.
• Fill in possible drug treatments in Table 2.

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Ellies Diagnosis

• Initial Identification The Health Center
  collected blood samples from Ellie and observed
  her cells under a microscope. They identified
  foreign structures with DNA and outer membranes.
  The cells were gram negative and about 1/10 the
  size of her cells.
• Ah, ha! said Becky. That matches one of my
  suspects. I knew those were a health hazard. I
  just need to re-check the size thing. This
  internet chart compares our cells to viruses and
  stuff.

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Metric Review

• 1 meter (m) 3 feet
• 1 meter (m) 1000 millimeter (mm)
• 1 millimeter (mm) 1000 micrometer (µm)
  (smallest size distinguished by naked eye)
• 1 micrometer (µm) 1000 nanometer (nm) (only
  seen with light microscope)
• 1 mm poppy seed (1000 µm/mm) 1000 µm

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Beckys Internet Search Results List of Suspects
Suspect 1 Coxiella burnetii causes Q-fever.
Coxiella are often found in livestock and are
excreted in milk, urine, and feces. Infection
occurs 2-3 weeks after inhalation of barnyard
dust. They are 0.3-0.5 µm gram-negative bacterium
(prokaryotes) that must invade and reside inside
human cells to cause infection.
1µm
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Prokaryotes

• Unicellular
• Reproduce asexually
• Composition
• Protected interior (cytoplasm) that contains
  genetic material (one circle of DNA) as well as
  complexes of protein enzymes to carry out
  necessary functions of gathering energy,
  manufacturing proteins (ribosomes), etc.

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Prokaryotes

• Size
• 0.2-10 micrometer (µm)
• Composition
• Phospholipid membrane, many contain cell wall
  composed of peptidoglycan (positive for chemical
  Gram stain), those with little or no
  peptidoglycan called Gram negative (like
  Coxiella).

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CQ1 Thats great, Becky said. My Mom sent
me 3 different antibiotics to kill bacteria.
Given the description of Ellies test results,
which antibiotic will definitely NOT work

• A Amoxicillin, Penicillin, and other ß-lactams
• Blocks the enzyme that normally creates links in
  peptidoglycan molecules.
• B Streptomycin
• Blocks prokaryotic ribosomes.
• C Ciprofloxacin hydrochloride (Cipro)
• Blocks bacterial DNA gyrase enzyme needed to
  counteract excessive twisting of DNA that occurs
  when circles of DNA are unwound to be copied
  into DNA or RNA.

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CQ2 Wait a minute! Ann said. The doctor said
the blobs in Ellies blood were 1/10th the size
of her cells. Could they be Coxiella?
A Yes B No
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Part II Microscope Analysis
Becky and Ann talked together outside the
students room at the student health center the
next morning. Youre right! Becky exclaimed
after viewing photographs of Ellies blood up
close. I wish I hadnt started taking the
antibiotics. The little crescent shaped
structures that I thought were the bacteria may
not be. When you zoom in on them, they show up
clearly in the electron micrograph on the right.
They arent too big to be bacteria, but they
arent too small to be mitochondria or some kind
of protozoan parasite. Wait a minute, Ann
replied. The things on the right are the
pathogens? Look at their insides, they cant be
bacteria. Why not? Becky asked.
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Part II, Continued
1µm
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CQ3 Well, Becky admitted, there should be
differences between Ellies cells and the little
blobs they saw. Otherwise, it might mean one of
my other suspects is the cause. These are some of
the structures normally found in all
cells. No, Ann answered, one
isnt. Which structure is NOT found in all
cells?
A Cytoplasm B DNA C Outer phospholipid
membrane D Ribosomes E Membrane-bound organelles
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Eukaryotes Prokaryotes
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Beckys Internet Search Results List of Suspects

• Eukaryote - Suspect 2 Cryptococcus neoformans
• 2.5-10 µm encapsulated fungus found in decaying
  pigeon or chicken droppings. Inhaled as spores
  that eventually spread to the brain causing
  meningoencephalitis. Has a black pigmented layer
  that can be seen sometimes on bird seed.

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Beckys Internet Search Results List of Suspects

• Eukaryote - Suspect 3
• Toxoplasma gondii
• 4-6 µm single-celled protozoan parasite of
  mammals birds.
• Most likely acquired through ingesting cysts in
  undercooked meat.

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Beckys Internet Search Results List of Suspects

• Suspect 3
• Toxoplasma gondii
• Usually no symptoms, but can cause flu-like
  complaints.
• Sexual life cycle occurs in cats, so infection
  can follow contact with cat feces.

• See QuickTime movies
• Invasion 4 and Escape 2 at
• http//www.uvm.edu/microbiology/videos_ward.php?id
  23

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Eukaryotes

• Uni- or multicellular.
• Reproduce asexually
• sexually.
• Composition
• Genetic material (long linear strands of DNA
  chromosomes) especially isolated and enclosed in
  membrane (nucleus)
• Some have cell walls (plants have cellulose,
  fungi ß-glucan)

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Eukaryotes

• Size 10-100 (µm)
• Composition
• Phospholipid membrane outside, as well as inside.
• Interior membranes separate functions such as
  gathering and transforming cellular energy and
  manufacturing macromolecules.

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Eukaryotic Organelles
Animal Cell
Plant Cell
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Eukaryotic Organelles
Mitochondrion
Nucleus
Endoplasmic reticulum
Golgi
Chloroplast
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• CQ4 Beckys Anti-Eukaryotic Medicines
• Pyrimethamine, Sulfonamides Interfere with
  enzymes used to make the folic acid needed to
  make thymine and uracil nucleotides.
• Polyenes combine with a component of fungal and
  some bacterial membranes, disrupt and break them.
• One of these drugs specifically affects one of
  the two eukaryotic suspects. Which test of
  Ellies blood would help you tell which
  eukaryotic suspect she was infected with?

• A Presence of DNA.
• B Presence of ß-glucan-containing cell walls.
• C Presence of cellulose.
• D Presence of peptidoglycan cell walls.

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Part III Viruses

• Becky and Ann are back at the dorm waiting for
  the results of more tests.
• Ive also got some tamiflu, Becky volunteered.
  I mean, what if those cells in the picture
  arent really making her sick. Maybe she just
  has the plain old flu.
• What do you mean? Ann asked. Whats the
  difference?
• Flu is a virus, Becky answers. Ive actually
  got two suspects that are viruses. Theyre
  probably the most different from the prokaryotes
  and eukaryotes. Theyre not even cells.

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Beckys Internet Search Results List of Suspects

• Influenza Virus Spread primarily through
  respiratory droplets from sneezing or coughing.
  Virus has single strand of RNA surrounded by
  phospholipids/protein envelope (80-120nm).
• West Nile Virus Spread by mosquitoes that have
  previously fed on infected birds. 20 of
  infected people show symptoms. Single stranded
  RNA, phospholipid/protein envelope (50nm).

Electron micrographs
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Viruses

• Not cells
• Cannot reproduce alone
• hijacks a host cell to replicate itself.
• Composition
• Outer shell repetitive protein often inserted
  into a lipid envelope (responsible for
  recognition and infection of host cell.)

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Viruses

• Size
• Smallest Organisms (50nm)
• 100 times smaller than bacteria
• Composition
• Protected interior that contains genetic material
  (DNA or RNA) with important protein enzymes
  required for duplication.

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Virus hijacking host system
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Tamiflu Blocks neuraminidase enzyme made by all
influenza A strains (cause the flu and avian
flu.) Viruses are unable to remove sticky sialic
acid, and cant escape.
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Table 1

• Take a few minutes to complete the homework
  table comparing viruses, bacteria, and eukaryotes.

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CQ5 Match the description with the suspect.
Use your responses to Table 1 to identify the row
below that best describes West Nile Virus.
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Part IV DNA Analysis

• Well, it isnt viral, Becky said, closing her
  cell phone. No neuraminidase. But, they found
  some foreign DNA with the sequence AACGTGGTCGTT.
  The closest match is a gene used to make
  ribosomes (rDNA). They are searching a huge DNA
  database of sequences to find the organism that
  has the closest match.

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CQ6 Foreign DNA sequence isolated from Ellie
ACGTGGTCGTT. Which sequence is the best match
with this foreign DNA?

• A Ellies nucleus ATGGTCTCAATG
• B Ellies mitochondria TTGGTCCGTCAG
• C Coxiella bacteria TTGGTCGGTCAG
• D Toxoplasma nucleus AACGTGGTAGTT
• E Cryptococcus nucleus ATGGTGGCAATG

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Strange Similarities

• Whats weird is that Ellies mitochondrial DNA
  matches the Coxiella sequence so closely.
• Ellies mitochondria TTGGTCCGTCAG
• Coxiella bacteria TTGGTCGGTCAG
• That makes sense in a way, Ann answered.
  There is a lot of evidence that points to
  mitochondria being descendents of gram-negative
  bacteria just like Coxiella.
• What type of evidence? Becky asked.

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Eukaryotic Organelles Endosymbiosis

• Strange similarities
• Chloroplasts and mitochondria are the same size
  as prokaryotes.
• Both have circular DNA without histones with
  similar sequence to photosynthetic bacteria
  (cyanobacteria) and obligate intracellular
  rickettsia bacteria.
• Both divide like prokaryotes.

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Eukaryotic Organelles Endosymbiosis

• Strange similarities
• Have their own protein synthesis machinery
  (ribosomes) more like bacteria than eukaryotes
  (sensitivity to Streptomycin).
• Inner membrane of mitochondria contains unusual
  phospholipid characteristic of bacterial
  membranes.

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Similarities Used to Group Organisms
Single-celled eukaryotes 2 billion years ago
First prokaryotes 3.75 billion years ago
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CQ7 Which letter best defines the place
mitochondria would take next to their closest
relatives on this family tree of living organisms?
A B C D E
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Finale Ellies Prognosis

• Well, Ellies responding well to the
  pyrimethamines that the doctors prescribed,
  Becky commented to Ann while checking her email a
  few days later at the dorm.
• Yeah, and were lucky the cipro we took
  couldnt harm our cells, Ann replied. We were
  so wrong! Im never self-medicating again. Do
  you think we should warn the other students.
  They might have had contact with the kitten,
  too.
• We dont know if it was from cat poop, Becky
  answered. I learned that something like 25-40
  of American adults are already infected with
  Toxoplasma gondii, and not because of their
  catsusually its from eating raw meat. Plus,
  apparently the oocysts in fresh cat poop arent
  infectious for a couple of days. So, if you
  scoop the box right away you dont have to
  worry.
• So now I have to know how often the cat box is
  cleaned?! I dont think Im cut out for this
  job! Ann moaned.