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microbio lab final


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who does rosie love?
who does chetho love?
who's anniversary is on 3-21-05?
chetho n rosie
who does capital cool love?
conqueress o melee
who does conqueress o melee love?
capital cool
culture medium (definition)
a solution or suspension of nutrients that provides an appropriate physical and chemical environment to support the growth of microorganisms
first media used?
broths (hay infusion and meat infusions)
-provided way to grow large #'s in single container
-difficult to isolate
how to isolate cells
-grow on solid surface (agar-does not melt at boiling temp and stuff cant break it down)
what is defined (synthetic) medium?
medium in which the exact chemical composition is known
what is undefined (complex) medium?
extracts or digests are used, exact composition not known. provides widest array of nutrients, more used
sterilized ?
freed from all life forms (use autoclave=chamber using steam under pressure)
What is the Germ Theory of Disease?
developed by Robert Koch that diseases are caused by specific microorganisms
tiny droplets scattered in the air
previously touched objects that harbor microorganisms
(doorknobs, money, sickroom contents)
aseptic technique?
methods taken to prevent contamination
what are wet mounts for
to observe live cells, to observe motility
example of motile bacterium?
Pseudomonas aeruginosa
brownian movement?
vibration of cells when struck by water
movement of cells caused by flowing of water or medium used to make slide
one flagellum
many flagella located at opposite ends
many flagella located at one end of the bacteria
many flagella located all over (like hair)
in a gram stain,
purple is Gram ____ and red is Gram _____
purple=gram +
red = gram - (usually rods)
why are gram positive stains purple?
because they have thick, tightly cross-linked peptidoglycan cell membrane wall and it holds the crystal violet
why dont gram negative hold the purple?
they have only a little peptidoglycan in their cell wall so crystal violet can be washed out
in a gram stain, what is the primary stain?
crytal violet
in a gram stain, what is the mordant and its purpose?
iodine, helps the color to bind further
what is the alcohol in a gram stain for?
to decolorize the primary crystal violet stain
what is the counterstain in a gram stain?
safranin (red), it just stains whatever is decolorized by the alochol
what is the acid-fast stain for?
used to identify members of the genus Mycobacterium (Tuberculosis and leprosy)
why use the acid-fast?
because bacteria have mycolic acid in their cell wall so Gram stain can't penetrate
what color acid-fast positive cells? negative?
acid-fast positive=bright pink
what is a capsule?
slimy layer of polysaccharides, proteins or both that surround the cell, and inhibits phagocytosis
if a capsule is present what does does the cell look like when stained? example?
there is a clearing around the cell. example=Kebsiella pneumoniae has capsules
what are endospores?
thick coated survival structures that help a cell survive pH changes, heat, cold dessication, lack of nutrients, etc.
examples of cells that form endospores?
Bacillus and Clostridium (look like green circle inside red mother)
selective media?
has something added that will inhibit the growth of all but the desired type of organism (dyes, salt concentration)
differential media?
containes substances that will allow you to visually distinguih colonies with certain characteristics (carbohydrate fermentation or substrate break down)
differential and selective?
select for a desired group of organisms and allow you to distinguish which among that group has the metabolic ability you are seeking
crystal violet agar
selective or differential?
inhibits growth of Gram +
only gram - will grow
blood agar
selective or differential?
what is it for?
what is alpha hemolytic?
organisms used for?
related disease?
categorizes bacteria by how it breaks down red blood cells
alpha=partially break down rbc's
beta=completely break down
gamma=not broken down at all
--used for Streptococcus
-strep throat
Mannitol Salt Agar
Selective or differential?
grows which bacteria?
turns yellow if what? example?
remains pink if what? example?
Both S and D
yellow if ferments mannitol, example fermenter = s. aureus
pink if cannot ferment
example of no ferment= s. epidermis
MacConkey Agar
s or d?
any pH indicator?
shows which metabolic activity?
group of bacteria?
both s and d
neutral red
inhibits gram + (crystal violet)
if organism can ferment lactose, acid produced reacts with neutral red turns deep red
strong fermenters =dark pink (e. coli)
Family Enterobacteriacea
Eosin-Methylene Blue (EMB)
group of bacteria?
s or d?
tests for what?
family enterobacteriacea
both s and d
tests for Gram -, and identifies e. coli as strong lactose fermenter
-strong lactose fermenter (e.coli) turn metallic green
lesser fermenters=pinkish purple (e. aerogenes) non fermenters= colorless
cold loving ~5 degrees C fridge
heat loving ~>50 C volcanoes, hot springs
room temp ~20 C
mesophile in little colder temp (fridge)
optimum temp?
best temp for organism to grow
included in media to make up for the pH changes caused by the organisms' own waste
obligate aerobes?
must have oxygen to survive
do not need oxygen
aerotolerant anaerobes?
ignore oxygen with no harm to themselves
obligate anaerobes?
killed by oxygen
facultative anaerobes?
grow best with oxygen but can still grow without oxygen
only grow if provided with small amount of oxygen
Gas-pak? how does it work?
species that would grow in it?
uses water activated packet to chemically release hydrogen to bind with the oxygen in the sealed jar and also release carbon dioxide and replace the oxygen
works 100% of except 2 hour it takes to rid jar of oxygen
clostridium (obligate anaerobe) would grow
starch plate
checks for break down of what?
which shows what enzymatic activity?
break down starch into sugars shows amylase activity (clearing around growth)
ability to break down lipids
lipase activity
skim milk plate
ability to break down casin into amino acids = caseinase actiity
nutrient gelatin
ability to break down gelatin into amino acids =gelatinase activity
carbohydrate fermentation
(glucose, lactose, sucrose, mannitol)
contain phenol red
show ability to break down specific carb
if can break down=acid (pyruvic) produced during fermentation turns media yellow
if gas produced=CO2 in durham tube
IMViC tests for what
Enterobacteriacea, Gram -, faculative anaerobic rods
1% tryptone broth
tests for ability to produce indole (tryptophanase activity)
kovac's reagent gives red ring
hydrogen sulfide, indole, and motility (used for hydrogen sulfide)
2 causes of UTI's
cause of stomach ulcers
what tests for these?
proteus vulgaris and proteus mirabilis (and e. coli)
helicobacter pylori
urea test=tests oranisms that hydrolyze urea into ammonia and water by urease
what is triple sugar iron agar for?
identify enterobacteriacea
litmus milk detects what? 2 purposes?
detects metabolic activities that occur in milk
serves as pH indicator and redox indicator
what does catalase do?
splits hydrogen peroxide into water and oxygen
what are the 3 ways genetic recombination can occur in prokaryotes?
conjugation, transformation, and transduction
the direct passage of DNA into another cell
taking up and incorporating naked DNA from the environment
viruses inject genetic material into the cell
cell that has fertility plasmid (transfer F plasmid via conjugation)
do not require threonine, leucine, and thiamine
resistant to streptomycin
cell that does not have fertility plasmid (only has chromosome)
do require t,l,t
resistant to streptomycin
HFr (high frequency recombinant)
has fertility plasmid incorporated into chromosome (transfers F plasmid and chromosomal DNA via conjugation)
do not require t,l,t
sensitive to streptomycin
competent cell?
example of naturally competent?
cell that has ability to take up naked DNA from environment
example= Bacillus subtilus and Streptococcus pneumoniae
ways to make cell competent?
1. electroporation=shock cells with electricity to put holes in membrane
2. CaCl2- treat cells with calcium chloride then treat with cold and heat shock (we used)
plasmid extraction
-to prove that the cell actually took up the plasmid and not just a mutation
1. lyse cell using lysozyme (also in tears and sweat), necessary for G (+) cells
2. Disrupt membrane using sodium hydroxide (base) and SDS (detergent). Detergent disrupts lipids of membrane rupturing membrane. High pH of NaOH disrupts base pairing in chromosomal DNA causing denaturation.
3. Plasmid reanneals using potassium acetate. Also, chromosomal DNA, lipids and proteins get trapped in complex (plasmid in supernatant and DNA, lipids, and proteins in pellet at bottom of microcentrifuge tube)
4. Supernatant added to isopropyl alcohol to remove water and precipitate plasmid
Restriction endonuclease
naturally occurring enzymes in bacterial cells that cut nd destroy foreign DNA (cells protect their own DNA by methylation)
restriction enzymes
recognize pallindromic sequences
gel electrophoresis
movement of charged fragments through a gel due to an electrical current
DNA is ____ charged so moves how?
negatively charged so moves toward positive electrode
what is tracking dye for?
added to sample so movement can be visualized
what is staining dye for?
added to gel while liquid so DNA can be viewed uner UV
to check for S. aureus in food what do you do?
streak on Mannitol salt agar plate
to check for fungi?
streak on Sabouraud dextrose agar plate (sab)
check for spores in bacillus and clostridium?
heat and spread on NA plates
(clostridium is anaerobe so put in gas pak)
check for e. coli?
eosin-methylene blue plate (EMB)
what do acids and alcohols do in fermentation?
act as preservatives and prevent growth of other microbes
what is non-iodized salt used for in saurkraut
iodine would inhibit microbial growth
fermentation aerobic or anaerobic?
microbial succession?
different microbial species acting sequentially
1. leuconostoc and lactobacillus then 2. streptococcus (acid-loving)
production of wine
used what organism?
why regulate temp in initial stages?
used saccharomyces cervisiae
regulation of temp in initial stages essential since fermentation could be stopped before all the sugar has been fermented
protozoe eukary or prokaryotes?
2 stages in life cycle?
what is the one foundin cat feces and can cross the placenta?
eukaryotes (have nucleus)
motile trophozoite and non-motile cyst
fungi, eukary or prokaryotes?
fungal infection
body of fungus
filaments of fungus

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