Download Clinical Case Example - Montana State University Extended University

LABORATORY STANDARD OPERATING PROCEDURES
1. Personal Protective Equipment including: gloves, eye protection, and a
lab coat, must be worn when working with human cells or bacteria. Do not
wear personal protective equipment outside of the lab. Open-toed shoes
may not be worn in the lab.
2. No food or beverages in lab. Food is stored outside the work area in
cabinets or refrigerators designated for this purpose only.
3. Disposable labware is to be discarded in biohazard bags. Samples
containing ≤ 1 mL volume can be discarded directly into biohazard bags.
Petri dishes containing cultures are discarded directly into biohazard bags.
4. Sink area must be kept reasonably clean throughout the day. Eyewash
station must be readily accessible. Sink must be free of all labware prior to
leaving for the night.
5. Lab benches where work has been conducted must be decontaminated
at the completion of work or at the end of the day and after any spill or
splash of viable material with 70% EtOH, 10% Bleach, or Lysol.
6. If a spill occurs spray Lysol, or 70% EtOH, or a 10% Bleach solution on
spill and place a paper towel on top of the spill to prevent further spreading.
After ≥ 30 min remove paper towel and thoroughly decontaminate area
using more of the disinfectant. If you are unsure about how to handle a
spill contact Dr. Voyich or a science assistant.
7. Personnel must wash their hands after they handle viable materials,
after removing gloves, and before leaving the laboratory. Bacteria can be
easily passed on by human contact, and is commonly spread by the hands.
P a g e | A1
P a g e | A2
Activity 1: Identify microbiota in your nose and throat
Goal: Students will identify normal flora microorganisms from both nose and throat and look for
Staphylococcus aureus and Streptococcus species.
Day 1
Throat swab:
1. Get on your lab coat, goggles, and gloves.
2. Obtain an aliquot of sterile phosphate buffered saline (PBS), and sterile swabs. Obtain 2 plates each of
Blood Agar (BA), Tryptic Soy Agar (TSA) and MacConkey Agar (MAC), and Mannitol Salt Agar (MSA).
3. Wet the swab with PBS. Say “Ahhhhhh.” Swab both of your tonsils. Place swab in sterile 15 mL tube
containing a small amount of PBS. Discard swabs in biohazard.
4. Take the swab containing your partner’s normal microbiota and streak for isolation (page A3) on BA, put the
swab back in the 15 mL tube with the PBS, repeat the procedure and streak for isolation on TSA, CNA, and
MAC. Follow the directions for streaking for isolation on the board and ask for help if you need it. Discard all
materials used for streaking for isolation in the biohazard.
5. Let the science team know when you are done and one of them will put your plates in the incubator.
Nasal swab:
1. Get on your lab coat, goggles, and gloves but you don’t want a partner for this one!
2. Obtain an aliquot of sterile PBS. Swab one of your nostrils as demonstrated by the science team and
streak for isolation on MSA, followed by TSA, CAN, and MAC.
3. Let the science team know when you are done and one of them will put your plates in the incubator.
P a g e | A3
Streaking for Isolation
1. After dipping your nasal or throat swab into sterile PBS (in the 15 mL conical tube labeled PBS), move it
rapidly over the full-width of the plate as indicated below in Circle 1. Place your swab in the 15 mL conical.
2. Turn the plate 90°, use an inoculating loop and start your streak at the end of the primary inculation zone.
Make 3- 4 passes over the primary inoculation zone as shown in Circle 2. Dispose of your inoculating loop in
the biohazard bag.
3. Get a new inoculating loop, turn the plate again 90° and repeat, this time make sure you have a portion of
the streak that does not re-enter the previous streaks (see Circle 3).
4. Invert and incubate your plates overnight (science assistants will take your plates to the incubator).
P a g e | A4
Day 2
Use the charts below to record your observations from the nose and throat experiments.
For the recording growth use the plus system. This can be estimated by determining the zone where you
ended up with single colonies (indicated on the Circle diagram page A3).
1 – 2+ = growth in the initial 1/3 to 1/2 of the plate.
3+ = growth in the middle 1/3 of the plate
4+ = growth in all zones
For the recording of the characteristics refer to the sheets in your notebook demonstrating growth
characteristics of the different agars. Record color changes and types of hemolysis plus any other
observations you note (different types of bacterial colonies shapes and sizes).
NASAL
TSA
MSA
BA
MAC
EMB
CNA
Are the
colonies all
the same
size?
Did the
colonies
ferment
mannitol?
What kind of
hemolytic
reaction did
you observe?
Did the
colonies
ferment
lactose?
Did the
colonies create
a green sheen?
Are the colonies
all the same size?
TSA
MSA
BA
MAC
EMB
CNA
Are the
colonies all
the same
size?
Did the
colonies
ferment
mannitol?
What kind of
hemolytic
reaction did
you observe?
Did the
colonies
ferment
lactose?
Did the
colonies create
a green sheen?
Are the colonies
all the same size?
Characteristics
of growth on
plate
Amount of
Growth (plus
system)
Throat
Characteristics
of growth on
plate
Amount of
Growth (plus
system)
P a g e | A5
Activity 2: Methods to determine if a bacterial isolate has antibiotic resistance.
Antibiotic resistance is a growing problem. When a bacterium has antibiotic resistance it limits the
treatments we can use to successfully eradicate an infection. Therefore, with some microorganisms
that have a high likelihood of harboring genes that confer antibiotic resistance (like Staphylococcus
aureus) we do tests to look for specific genes. For today’s activity you will isolate DNA and look for a
specific gene, mecA, that confer antibiotic resistance in Staphylococcus aureus.
Goal: To understand basic techniques for determining if a pathogen has antibiotic resistance.
Day 1
1. Get on your lab coat, goggles, and find a partner.
2. Obtain an aliquot of lysed bacteria. Also obtain all reagents for DNA isolation (science team will assist).
Record the number that identifies your bacterial aliquots in your lab notebook. Both aliquots are from
Staphylococcus aureus but one of the aliquots has antibiotic resistance while the other does not. Since the
bacteria are lysed they are already dead and are therefore not infectious. Do all procedures for both
samples.
3. Add 25μl Proteinase K, vortex.
4. Add 200μl Buffer AL from DNeasy kit and give samples to a science team member to incubate it for 30min
@ 56°C.
5. Add 300μl 100% ethanol (EtOH), vortex.
6. Transfer sample to DNeasy mini spin column, spin @ max RPM for 1min, and discard flow through (in the
bottom tube there will be liquid, pour it in the appropriate container labeled “DNA Discard”).
7. Place column in clean collection tube, add 500ul Buffer AW1, spin @ max RPM for 1min, discard flow
through (in the bottom tube there will be liquid, pour it in the appropriate container labeled “DNA Discard”).
8. Place column in clean collection tube, add 500μl buffer AW2, spin @ max RPM for 3min.
9. Obtain a snap capped tube (1.5 mL microcentrifuge tube), and label the tube with the number of the sample
and your initials. Transfer the column to your snap capped tube and add 40μl buffer AE (elution buffer).
Incubate 1min @ RT and spin for 1min @ max RPM. THIS IS THE DNA. Discard the column.
P a g e | A6
Day 2
Determining the presence of mecA by polymerase chain reaction (PCR)
1. Get two PCR tubes each containing a bead. Label one tube with a “G” and one tube with a “M”.
2. Into the tube labeled “G”, pipette 10 µl of gyrB forward primer and 10 µl of gyrB reverse primer.
3. Into the tube labeled “M”, pipette 10 µl of mecA forward primer and 10 µl of mecA reverse primer.
4. Into each PCR tube, pipette 5 µl of your isolated DNA.
5. Place the lid on each tube and hand it to the science mentor.
6. The science mentor will run the PCR on your samples and run the samples through an agarose gel to
separate the pieces of DNA in your sample (gel electrophoresis). You will receive a photo of your gel so you
can determine if you did your procedure correctly. This will be indicated by amplification of gyrB a gene
present in all Staphylococcus aureus strains. You will also determine if your sample had the mecA gene which
confers presence of antibiotic resistance in Staphylococcus aureus.
Tape your photo here and indicate your results.
P a g e | A7
Activity 3: Solve a microbiology case study
Goal: To use your knowledge gained in the previous activities to identify the microorganism from a
mixed culture and present your diagnosis to the other students.
Day 1
1. Obtain a mystery sample and a case study from the science team. The mystery may contain Gram positive
or Gram negative microorganisms (or both).
2. Using a swab streak your unknown on each agar and using an inoculating loop streak for isolation on BA,
MSA, TSA, MAC, EMB, and CNA. Also streak on the agar that contains an antibiotic (science team will
explain).
3. Give your plates to the science team to incubate overnight.
Day 2
1. Prepare a Gram stain of your mystery sample.
A. Draw 3 circles on a slide using a wax pen.
B. In circle 1 take a single colony from the Staphylococcus plate this will be your control demonstrating
a Gram-positive result (ask a science team member for help).
C. In circle 3 take a single colony from the Escherichia coli plate this will be your control demonstrating
a Gram-negative result (ask a science team member for help).
D. In circle 2 take a single colony from your unknown sample.
E. Follow the steps on the Gram Stain Procedure Handout (following page).
?
2. Perform catalase test (page A8).
3. Write down your results and discuss your findings with your team.
4. Report the most likely culprit based on laboratory results and based on the clues given in the case study
(i.e. how did the person get infected, what area of the body is experiencing the illness).
P a g e | A8
Catalase Test
Some bacteria such as Staphylococcus aureus and Staphylococcus epidermidis, contain the enzyme catalase
which reacts with hydrogen peroxide and forms water and bubbles of oxygen. Other types of bacteria, such as
Streptococci species do not contain catalase. Thus, a drop of hydrogen peroxide on a sample of bacteria can
help to differentiate what type of bacteria is present.
1. Obtain a single culture and pick up the culture with an inoculating loop. Spread the colony on a glass
slide.
2. Ask for a science mentor to come and assist you with the catalase test. You will pipette a small amount
of hydrogen peroxide onto your sample. Record the results.
P a g e | A9
Use the chart below to record your observations from your case study.
For the recording growth use the plus system. This can be estimated by determining the zone where you
ended up with single colonies (indicated on the Circle diagram page A3).
For the recording of the characteristics refer to the info sheets in your notebook demonstrating growth
characteristics of the different agars. Record color changes and types of hemolysis plus any other
observations you note (different types of bacterial colonies shapes and sizes).
Unknown #
TSA
MSA
BA
MAC
EMB
CNA
Are the
colonies all
the same
size?
Did the
colonies
ferment
mannitol?
What kind of
hemolytic
reaction did
you observe?
Did the
colonies
ferment
lactose?
Did the
colonies create
a green sheen?
Are the colonies
all the same size?
Characteristics
of growth on
plate
Amount of
Growth (plus
system)
Additional observations:
Gram stain results:
Catalase Results:
Clues from the case study:
Diagnosis:
P a g e | B1 P a g e | B2 Mannitol Salt Agar (MSA) A selective and differential agar which contains 7.5% salt to select for certain Gram‐positive bacteria such as Staphylococci. The high salt concentration selects for halophilic organisms (organisms that like salt). It also contains mannitol sugar and phenol red (a pH indicator) which will select for organisms that ferment mannitol. If an organism can use mannitol as its energy source, the agar will turn yellow due to a drop in the pH which causes a color change. For example, both Staphylococcus aureus (section A) and Staphylococcus epidermidis (section B) will grow on MSA but only Staphylococcus aureus can ferment mannitol. Section C demonstrates no growth. Blood Agar (BA) A nutrient rich medium that contains 5% sheep blood. This agar will grow most organisms and will help you identify what bacteria is growing based on the ability of the bacteria to lyse blood cells. Different bacteria have different types of hemolysins: α, β, and δ hemolysin. The blood agar will look different depending on what type of hemolysins the bacteria have. P a g e | B3 MacConkey Agar (MAC) A selective and differential medium used to select Gram‐negative bacteria. It can differentiate bacteria based on their ability to ferment lactose. If bacteria can use lactose, they will turn a pink color. If bacteria cannot use lactose, they will remain colorless or yellow. E. coli will ferment lactose but Salmonella sp. cannot. Eosin Methylene Blue Agar (EMB) A selective and differential medium that allows growth of Gram‐negative bacteria. The agar contains lactose and two dyes: eosin and methylene blue. When a bacteria is able to ferment lactose, it produces acid and turns the bacterial colony a dark purple to black. Colonies that do not use lactose will be colorless. P a g e | B4 Colistin‐Nalidixic Acid Agar (CNA) A selective medium containing the antibiotics colistin and nalidixic acid which selects for Gram‐
positive bacteria. S. aureus, S. epidermidis, and Group A Streptococcus will grow on this medium. Tryptic Soy Agar (TSA) A supportive medium which contains glucose and amino acids for the growth of most microorganisms, it will not select for Gram‐positive or Gram‐negative (both groups will grow on this agar). Bacterialogical Media , http://faculty.mc3.edu/jearl/ML/ml‐8.htm C1
Gram‐negative rod
Ferments lactose
Catalase positive
Clinical Presentation
oWatery or bloody diarrhea
oUrinary Tract Infection (UTI)
oSeptic Shock
Pathobiology
In normal GI flora
Animal sources of infection
Transmits via fecal‐oral
E. coli on MacConkey
agar
Clinical Case Example
Patients in a small town visit the hospital complaining about bloody diarrhea, fatigue, and confusion. After interviewing the patients, the doctors discover that each patient frequents the same fast‐food burger joint. The doctors identify the causative agent using serological testing and stool cultures.
C2
Gram negative rod Motile by flagella
Produces hydrogen‐sulfide
Does not ferment lactose
Catalase positive
Clinical Presentation
oGastroenteritis
Pathobiology
Carried in animals and humans
Transmits fecal‐oral
S. enterica on MacConkey
Agar
Clinical Case Example
A veterinary student complains to the doctor of diarrhea and abdominal tenderness. He also had nausea and vomited the day before. He admits he recently played excessively with his turtle.
C3
a.k.a. Group A Streptococcus
Gram positive cocci in chains
Typically infects throat or skin
Β‐hemolytic on blood agar
Catalase negative
Clinical Presentation
oPharyngitis (Strept throat)
oImpetigo
oCellulitis
Pathobiology
Transmits human to human via respiratory droplets, saliva
Trauma introduces bacteria into skin
S. pyogenes on blood agar
Clinical Case Example
A young child presents with a fever and skin rash localized around the lips and on his arms. The rash appears to have pustules with yellow crusts. Cultures from the skin show Gram pos. cocci and are β‐hemolytic. The doctor administers penicillin G. C4
Gram‐positive grows in clusters
Catalase positive
Coagulase negative
Clinical Presentation
oInfection on indwelling medical devices such as catheters or prosthetic joints
Pathobiology
Normal flora on skin
Forms biofilms and adheres to medical device
S. epidermidis on Mannitol
Salt Agar (MSA)
Clinical Case Example
Ten days after undergoing chemotherapy for cancer a, a middle‐aged man develops a fever. On exam, he has erythema and tenderness at the insertion site of the IV catheter. Blood cultures are positive for Gram‐positive bacteria. The original catheter is removed and the patient is started on antibiotics.
C5
Gram positive clusters
Catalase positive
Coagulase positive
Antibiotic resistance is a problem (MRSA = methicillin‐
resistant S. aureus) Clinical Presentation
o Skin/subcutaneous: impetigo, cellulitis, boils
o Sepsis, Endocarditis
Pathobiology
Colonize skin or pharynx and immune system responds and causes inflammation and abscess development
Entry into blood via ruptures in skin
S. aureus on Mannitol Salt Agar (MSA)
Clinical Case Example
A college basketball player presents to the clinic with several red painful purulent boils on his upper arms. Gram stain of the purulent material reveals Gram‐positive clusters. Culture is resistant to penicillin and methicillin. C6
P a g e |D 1 P a g e |D 2 Tips for Your Project
Read this over before the ID virtual lab meeting. During the
virtual meetings we will answer questions about testing a
hypothesis and on your experimental design.
1. You can work in groups or work by yourself.
2. Develop a hypothesis to be tested. Think of a topic you are interested in learning
more about. For example, have you ever wondered if hand sanitizers actually work? Or
how fast they actually work? Do they work better than plain old soap and water? Figure
out what is known about the topic – perform an internet search, read an article about it,
interview an expert. Develop an educated guess i.e. a hypothesis, based on your
knowledge.
I hypothesize that 60 seconds of rubbing my hands with waterless hand sanitizer
will reduce bacterial numbers and diversity on my hands compared to washing my
hands with soap and water for 60 seconds.
3. Write down your experimental procedure. How will you test your hypothesis? What
will the variables be? How many plates do you need? Since we are shipping supplies in
October you can tell us if you need more plates and or swabs to test your particular
hypothesis. Of course, there is a limit on how large you can get these experiments but
we can accommodate a few extra materials.
Experimental Procedure:
A. Identify your control. How do you obtain a baseline of bacteria from your
hands? Should you wash your hands first before beginning the experiment?
Should the test be done on the same day or on a different day? What is the more
controlled experiment? The control will be compared to the experimental groups
to assess how the treatment altered the outcome.
B. Design a procedure where the hands will be as equally dirty. Make sure you
are consistent with how you dirty your hands! Some possible methods include
washing your hands with soap and water (to keep the baseline bacteria on your
hands similar between tests) and then:
1. touching raw chicken for 60 seconds.
2. brushing your horse without gloves for 5 minutes.
3. typing on a public computer for 5 minutes (at a library or computer lab
at school).
P a g e |D 3 C. Decide how to sample the bacteria on your hands. Will you swab each finger?
Will you swab 3 fingers on each hand – on one hand only? Will you combine the
swab material? If so how – on the petri dish? Make sure you are consistent with
how you collect samples!!
D. Standardize how you “wash” your hands. Your hypothesis says 60 seconds
but doesn’t specify how vigorous you’ll be washing. Probably want to make sure
the motion is very similar between washing with the waterless hand sanitizer and
with soap and water. Now again you need to sample the bacteria on your hands –
do this exactly as you did after touching the contaminated material.
4. Record exactly how you are doing your experiment so you can determine if you have
introduced more variables that are complicating your results.
5. Record results, take pictures, and fill out table for growth on media: how much
growth (use the plus system from the module, 1-2 +, 3+, 4+), and which plates supported
growth (diversity of the microorganisms)?
6. If possible, repeat!
7. Was your hypothesis correct? What are the implications of your findings? How does
this support or add to the information that is already available on soap and water versus
waterless hand sanitizers? If you were to perform your experiment again what would you
change?
8. Put together a presentation. You can either have a poster or oral presentation for our
meeting at Montana State University in January 2013.
P a g e |D 4 Use this Outline to Design your Experiment
Hypothesis:
Background Facts:
Methods:
Variables:
Results:
P a g e |D 5 Which media supported growth? How much growth did you find? Review your materials
from the hands-on module at Montana State University.
TSA
MSA
BA
MAC
EMB
Additional observations:
Implications of your findings:
What would you do differently if you had the chance to re-do the experiment?
CNA
P a g e | 1 P a g e | 2 Abscess: a collection of pus on the body that causes pain, swelling, and inflammation around it;
typically due to an infection
Aliquot: a portion of a solution
Antibiotic Resistance: a microorganism that is able to survive exposure to an antibiotic
Antibiotics: a substance or compound that slows down or prevents the growth of bacteria
Bacteria (Eubacteria): a domain of microorganisms, only micrometers in length, with a
primitive nucleus
Buffer AE: a solution that binds to DNA in order to remove purified DNA from a column
Buffer AL: a solution that disrupts protein structures
Buffer AW1: a solution that denatures (destroys) proteins in order to purify DNA
Buffer AW2: a solution that contains ethanol, which removes cellular salts from a sample in
order to purify DNA
Carrier: an individual who is colonized (infected) with a pathogen, but free of disease, who is
capable of acting as a source of infection for others
Catalase: an enzyme that degrades hydrogen peroxide into hydrogen and water
Catheter: a tube that can be inserted into a body cavity, duct, or vessel that allows for
drainage, administration of fluids or gases, or access by surgical instruments; an example is a
urinary catheter inserted in the bladder to drain urine
Cellulitis: a common skin infection caused by bacteria, symptoms commonly include: redness,
inflammation, and soreness of the skin; also can cause fever-like symptoms
Centrifuge: a piece of equipment that puts an object in circular motion, causing denser
substances to settle to the bottom of the tube- “the pellet”- while the less dense constituents
remain on top- “the supernatants”
Coagulase: a protein produced by some microorganisms that converts fibrinogen to fibrin,
resulting in clumping of blood; used to distinguish between different types of Staphylococcus
species
Colonization: establishment of a microbial population in the animal/human host
Colony: a visible group or cluster of bacteria derived from one bacterium, you count colonies on
an agar plate to quantify bacterial growth
Crystal Violet: a dye used in Gram staining; it remains inside bacterial walls containing higher
amounts of peptidoglycan
Deoxyribonucleic acid (DNA): found in all cells and contains the genetic instructions used for
development and function; consist of two sugar backbones (containing deoxyribose sugar) with
linked nucleotides in between; the nucleotides can be one of four bases and the sequence of
these bases determines genetic make-up of the organism
P a g e | 3 Disease: a condition that is accompanied by an impaired body function
Echocardiogram: a type of ultrasound that images the heart; can provide an assessment of the
state of heart tissue
Endocarditis: an inflammation of the inside lining of the heart chambers and heart valves
Endotoxin: a type of toxin found on Gram-negative bacteria that is made up of the
lipopsaccharide chain found on the outer membrane of the bacterium
Erythema: redness of the skin occurring with skin infection, injury, or inflammation
Ethanol: an alcohol used for many biological purposes; can be used to disrupt protein structure
and remove salts to help purify DNA; can be also used as a disinfectant due to its ability to
penetrate bacterial cell walls and is used to decolorize bacterial cells in the Gram staining
procedure
Exotoxin: a potent toxin secreted by some forms of bacteria
Flora: a population of microbes that inhabit the internal and external body surfaces of healthy
humans and animals (see: microbiota)
Furuncles: a boil; results in a painful swollen area on the skin filled with pus and dead tissue
Gastroenteritis: a medical condition characterized by inflammation of the GI tract, including the
stomach and small intestine; results in a combination of diarrhea, vomiting, abdominal pain, and
cramping
Gel Electrophoresis: the separation of DNA, Ribonucleic Acid (RNA), or protein, based on size
and charge; electric currents are applied to move molecules through the matrix
Gram Stain: a method of differentiating bacterial species into Gram-positive and Gram-negative
bacteria based on the amount of peptidoglycan in their cell walls
Gram-negative: bacteria that do not retain the crystal violet stain due to a low amount of
peptidoglycan in the cell wall; these bacteria typically stain red during Gram staining (they retain
the color of safranin, the counter-stain)
Gram-positive: bacteria that retain the crystal violet stain due to a high amount of
peptidoglycan in the cell wall; these bacteria typically stain purple during Gram staining
gyrB: a gene present in all Staphylococcus aureus; used as a control during PCR to confirm
DNA was isolated from Staphylococcus aureus
Hemolysis: from the Greek word meaning “release of blood”, is the rupturing of red blood cells
followed by the release of hemoglobin into surrounding medium, there are three types of
hemolysis: alpha (α) reduces the iron in the blood (bacteria colonies form “a green halo” on
blood agar), beta (β) completely ruptures the blood cells (bacteria colonies form a “clear halo”
on blood agar), and gamma (γ) is a lack of hemolysis (colonies often appear grey with no halo
on blood agar)
Hypothesis: a proposed explanation for a phenomenon/occurrence, often based on previous
observations that can be tested with a set of experiments
P a g e | 4 Impetigo: a common skin infection characterized by a single, or many, blisters filled with pus
and, when broken, leaves a reddish, raw-looking base
Infection: results from a microbe that penetrates body surfaces, gaining access to tissues and
multiplying which then induces a host response
Iodine: a trapping agent that binds to crystal violet making it a larger molecule to trap the dye in
the peptidoglycan
Lactose: a sugar found in milk formed by galactose and glucose
Lipopsaccharide chain: found on the outside of the outer envelope of a Gram-negative
bacteria, often called endotoxin
Lyse: to burst or cause dissolution or destruction of cells
Lysostaphin: an antibacterial enzyme that can cleave components of Staphylococcus aureus
cell wall; this is used in DNA extraction to release the genetic makeup (DNA) out of bacteria
Mannitol: a sugar alcohol, typically of a lower (acidic) pH; component of Mannitol Salt Agar
(MSA); colonies of bacteria that can ferment mannitol appear yellow on the plate while those
that cannot appear pink
mecA: a gene found in bacteria that confers resistance to a large class of antibiotics including
penicillin and methicillin; Staphylococcus aureus strains that have this gene are called
methicillin-resistant Staphylococcus aureus (or MRSA)
Media: a liquid or gel designed to support the growth of microorganisms or cells
Meningitis: a bacterial infection of the membranes covering the brain and spinal cord
Microbiota: a population of microbes that inhabit the internal and external body surfaces of
healthy humans and animals (see: flora)
Microorganism: a microscopic organism that can consist of a single cell, cell cluster, or a
multicellular complex organism; includes bacteria, viruses, algae, fungi, etc.
Morphology: the form and structure of organisms and their specific features
MRSA: methicillin-resistant Staphylococcus aureus a form of Staphylococcus that has
developed resistance to a wide spectrum of antibiotics (see mecA)
Opportunistic Pathogen: microorganism that is free living or a part of the host’s normal
microbiota but may become pathogenic under certain circumstances, such as when the immune
system is compromised
Pathogen: a microorganism capable of producing disease in a healthy animal or human host
Pathogenicity: ability to cause disease
Peptidoglycan: a major component of the bacterial cell wall that contributes to the shape of the
bacteria; amount of peptidoglycan is used to differentiate bacteria in the Gram stain (see Grampositive and Gram-negative)
Pharyngitis: inflammation of the throat; also known as a sore throat
P a g e | 5 Pharynx: the throat
Phosphate Buffered Saline (PBS): a buffer solution used in biological research; a water-based
salt solution that is non-toxic to cells
PI Buffer: a buffer used in the isolation of DNA that prevents cellular clumping and degrades
the RNA that is present
Polymerase Chain Reaction (PCR): a technique used to amplify DNA in order to generate
thousands to millions of copies of a specific sequence; may be used for DNA cloning, identifying
evolutionary ancestors, diagnosing genetic diseases, and identifying genetic fingerprints used in
forensic science labs
Primer: a start point for DNA synthesis and replication; primers are typically short strands of
nucleic acid (the building blocks of DNA) that are used to bind a specific sequence of DNA in
order to amplify it during a PCR reaction
Proteinase K: destroys proteins that degrade DNA and RNA
Reagents: a substance or compound added to a system in order to cause a reaction
Ribonucleic acid (RNA): like DNA, found in all cells; consists of one sugar backbone
(containing ribose sugar) with nucleotides bound to the backbone; RNA is created based on a
DNA template and is then used to make proteins necessary for cellular function
Safranin: a biological stain used as a counterstain in Gram staining
Sepsis: a medical condition characterized by a whole body inflammatory response to an
infection; also known as blood poisoning
Serological Testing: a test used to determine the presence of a microorganism in the blood
Supernatant: the liquid above a solid residue (pellet) after centrifugation
Virulence: attributes of a microbe that enhance its pathogenicity