Lab Exam Flashcards
What consists of the frame of microscope
-arm and base
-arm for carrying
-base for support
How should the microscope be set when u are packing it away (thus when u receive it)
-cord wrapped around base or around cord caddy
-x4 objective lens should be in placed
Stage
Stage:
-between upper lens system and lower devices
-has a light that shines through hole
-mechanical stage holds light in place
-adjustment knobs that move stage vertically and horizontally
Use of stage
only the x4 or x10 objective lens should be used when placing or removing a slide on the stage
-dont need to move the stage out of focus since lots of space between x4 and x10
Use of ocular head
-dont apply pressure when turning head
-adjust the eyepieces to fit ur eyes
-to adjust eyepiece focus; focus right eye and right eyepiece on specimen then focus left
-focus on same spot using focussing ring
Name of objective lenses
4x: scanning
10x: low power
40x: high-dry
100x: oil immersion
-all are on a nose piece that rotates
what does the 100x require that the others dont
OIL
-needs immersion oil to enable the object to collect light at angles that would normally fall outside its collection window
What objective does the ocular lens piece have
-10x
How to calculate magnification
ocular lens x objective lens
ex: 40x high dry lens used (objective)
ocular is always 10x
40x10 =400
Condenser and Iris diaphram
light from lightbulb is directed to here
-placed under central opening of stage
-CONDENSER collects and concentrates light, directing up THROUGH any object on stage and can be adjusted by opening or closing iris diaphram
Raise condenser all the way then turn 1/4 down for best results (with condenser adjustment knob)
If too much light is sent through what happens?
-contrast decreases and depth of field becomes less that desired
what does lowering condenser do
decreases the amount of light that reaches the object
FOR 100x, make sure condenser up akk the way
Iris Diaphragm
-regulates the amount of light that passes through the condenser
-higher the objection, higher the intensity of light required
Start by having iris diaphram in
half position, if lacks contrast, adjust
What is used for critical focussing
fine adjustment knob
when stsrting to use microscope, what do u do to iris
half open (black adjustment with numbers)
Basic rule of focus
FOCUS AWAY FROM THE SPECIMEN
-bring specimen as close to lens as possible using fine adjustment (look not thorugh eyepiece)
-THEN look through eyepiece and bring it away from ituntil in foxus (turn knob other way)
-adjust light to comfortable level
-unfold or fold eyepieces to make one circle
Most bacteria exists in what shapes
Rods (bacillus), circle (coccus), corkscrew, spiral (spirochette or spirillium)
Stalked forms (caulobacter), club-shaped (corynebacterium), comma-shaped (vibrio)
Variations due to…
-age
-composition of culture medium
What are the characteristics
check table
How to calculate magnification of specimen in drawing
size of specimen in drawing (um) / actual size of image (um)
WHo was gram stain created by
Danish scholar Christian gram in 1884
What type of stain is a gram stain and who was it made by
Differential stain: used to differentiate bacteria types based on their ability to retain stain
Different from simple stain since that just helps u see bacteria, doesnt differentiate them
Christian gram in 1884
What are the stains used in gram stain
Primary stain: Crystal violet
Mordant( sticks stain to cells): Gram’s Iodine
Decolourizing agent: 95% ethyl alcohol
Counter-stain: Safranin
What are the results of gram stain
General:
Gram Pos: Purple bc wont be decolourized
Gram-neg: Pink bc will be decolourized and stained by safranin
What affects culture staining
1) Age of culture
2) PH of medium used to grow bacteria
gram pos may look gram neg or gram variable due to acid medium or old medium
gram neg may look gram variable due to same thing
Iodine Mordant
-acts as a glue to stick crystal violet to the nucleic acid of cells to form a complex which is not readily remov ed from gram pos in decolourzing step
Acid Stain
-differential stains
-used primarily for mycobacterium, turbucolosis bacillus and turbo leprae
-used bc these bac have waxy cell walls due to high levels of liposodal material (mycolic acids), lipid rich so imppermable to certain stains
Ziel-nielson Acid fast technique
-smear with carbolfuschin (dark red, with 5% phenol), heat used to penetrate dye
-decolourized using acid alcohol
-counterstain of methylene blue used
non-acid fast will be decolourized and will be blue
acid fast will resist decolourizing by acid alcohol so they will be red
Kinyon acid fast
-cold stain
-concentrations of phenol and carbolfuschin increased and detergent is added so that heating is not needed
Are all acid fast gram positive
Yes, all acid fast are gram postiive but NOT ALL GRAM POSITIVE ARE ACID
Results of Acid-fast
Staphyloccocus epidermidis: BLUE
Mycobacterium: RED
(spiral)
What are spores, why?
-when environmental conditions are harsh (ex, lack of important nutrients), to permit further vegetative growth, certain bacteria condense their vital cellular components into ENDOSPORES
-vegetative cell slows down, loses moisture, and withdraws its substance into one area which it surrounds with thick impermeable wall.
-slowly original cell reminants falls apart and the spore that is highly resistant to environmental conditions remains
-What is it highly resistant to?
-high heat, ionizing radiations, dessication, and chemicals
-Bacillus and clostridium genus do this
Spore staining
-spores resist staining of ordinary dyes such as meth blue, carbofuschin etc so specific dyes must be used (malachite green)
-decolourizing will also be resisted by spores
SO
Malachite green is the primary stain used, heat is used to drive it into it
then safranin is used to differentiate the vegetative cells (actively growing) from the spores
Size and placement of endospores in cell is a characteristic of spore forming bacteria
Schaeffer-fulton method: uses steam to drive malachite green into spore
Cold methed (Bartholomew and mittwer): increases concentratoon of malachite green and uses detergent to replace heating to drive stain into cells
Results of Spore Stain
Bacillus brevis: Yes, central endospores
Clostridium sporogenes: Yes,
-live in environments that cause them to need ot have a way to live, like digestive tract.Live in ocean sediment, high temp, high pressure
Negative Stain
-Another simple stain
-Uses Negatively charged dye (negatively charged chromophore) instead of positively charged dye like the other simple staining technique used
-Nigrosin is the dye used
-it repells the net negative charge of the cell surface
-chromophore repells cell surface so cell remains undyed and there is a deposit that forms around the cells that is clear, background dark
Advantages:
-doesnt use heat so cells cant be distorted
-natural size and shape of cells is maintained
-also used to visual bacteria that are hard to stain like mycobacterium
Result of negative stain
Bacillus brevis
-rod
Negative stain procedure
Place a dot of dye and place a loopful of bacteria on top, mix them gently
-if a solid culture of bac used, add water to nigrosin with the solid bac
-gently slide a second slide on top, creating a thin layer
-let it dry
Capsule Stain
-envelope of muciliaginous substances which can be referred to as a capsule, slime layer or glycocalyx
-consists of polysaccharides, peptides, and carbohydrate material which accumulate on the cell surface, giving structure an enlarged appearance.
-Alcian blue is the dye that is water soluble in nature, it forms linkages with acidic groups of mucopolysacchariedes.
-can aslo be identified by negative stain since polysacharides are water-soluble so normal simple staining dyes will not stick to it, this technique allows for the cell and background to be stained
-most staining techniques stain the cell and the background, leaving only the capsule unstained
Capsules info
-size various sizes dependent on species and strain of species
-more virulent bacteria have thicker (heavy capsule) as these bacteria are harder to destroy by phagocytic cells
-identification of things such as pneumonia is therefore used by capsule stain
Capsule stain results
Klebsiella pneumonaie: purple background, purple cell, qhite halo like capsule around cell
most common ranges for micropipettes
0.5 to 10ul
10-100 ul (P100)
100 to 1000 ul (P1000)
p20 also used for 2-20ul
-they must be used within their range to have accucuracy achieved
Tips for the micropipettes are
autoclaved and sterile
How to eject tip
either push ejector button or push to third stop on plunger
p100 settings
100|0 is 100
050|0 is 50
p20 settings
05|0 is 5
10|0 is 10
20|0 is 20 ul
Culture medium
material in which or on which bacteria are grown
-used for cultivation of microorganism
-can be liquid or solid or semi-solid
-materials in medium can be limited to well-defined inorganic or organic compounds
Chemically defined medium
medium whose chemical constituents are known
Not clearly defined medium
-medium whose components are not known, contain complex materials such as animal tissue extract, blood serum, etc)
-usually for nutritionally fastidious organisms such as human pathogens
Nutrient broth or nutrient agar
most used medium
-supports growth of many organisms
-also known as All purpose medium or general purpose medium
Solid medium is made up of?
Agar
-agar made up of complex carbohydrates, galactan which is extracted from the marine algae of genus gelidum
-these are components not used for growth but a solidiifying agent
-melts when heated to 100 and remains liquid until about 43
-cause liquification by reheating again to 100
Selective media
-contain specific chemicals which do not affect the growth of some organisms (you wish to isolate) while discouraging and preventing the growth of other groups of microorganisms
ex: encorporating sodium azide selectively isolates lactic acid bacteria such as a species of streptococcus
-lack cytochrome system so are not affected since sodium azide binds tightly to a ring on cytochrome
-other examples are dyes (Crystal violet), high conc of nacl (halophile bacteria isolated), antibiotics, specifics sugars.
Differential media
contain chemicals or dyes that allow the obsrver to differentiate between types of bacterial colonies that are present after incubation
Example
-Eosin Methylene Blue Agar (EMB)
Used int the detection of enterobacteriacae (prouce light pink colony, dark center, which rarely show metallic sheen) and related coliform rods such as e.coli (produce dark centers)
NOTE: EMB may be considered differential and selective because it contains lactose so it can allow the growth of orgs that can produce b-galactosidase
-also possesses dyes (eosin y and methylene blue) that surpress the growth of various gram-pos species
Enriched media
-used for fastidious microorgansims (have specific requirements)
-enriched with certain vitamins and other growth-promoting subsatncfes
examples: blood, serum, extracts of plant and animal tissue
Results of agar plate experiment
Tryptic soy agar plate (enrichment):
E.coli- alot
enterobacter aerogenes; alot
staphylococcus epidermis; alot
entercoccus faecalis: alot
EMB Agar (Differential):
E.coli: +
Enterobacter aerogenes: +++
Staphylococcus epidermis: None
Enterococcus faecalis : + to none
KF Stretococcal Agar (Selective, isolates fecal streptococci/ enterococci found in food, milk, etc))
E.coli: none
Enterobacter aerogenes: none
Staphyloococcus epidermis: none
ENtercoccus faecalis: ++++
-found in gut and liver so makes sense for fecal
Pure culture
only containing one type of microorganism
Three dilution methods to obtain a pure culture from material obtained in nature
THREE DILUTION METHODS
streak plate
pour plate
spread plate
Why do we need to do dilution methods to obtain a pure cultyre
because cultures / specimen obtained from nature contain a mixed population of micro-organisms
Streak plate tecnique
do it to a quarter, sterilize, rotate 90, streak one line or 2 through previous streak and streake anothr corner, sterilizde loop, rotate 90 and do same thing (few times over the previous then dilute), DO NOT STERILIZE LOOP, rotate 90 and cut across 1-3 times again and spread only in area that has not been touched, dont make contact with other streaks
Pour Plate Idea
-isolate colonies by diluting specimen into a series of cooled (43-50) liquid agar mediums which are then poured into empty petri dishes
-immediately after pouring, plate is rocked,
after incubation it is grown in or around
-necessary to make several dilutions so that you could get to one that is countable since population of microbes are not known
methoon next card
Pour Plate general steps
one drop of bacterial suspension into tube 1 agar
2 drops of tube 1 agar into tube 2 agar
pour tube 1 into plate
3 drops of tube 2 agar into tube 3 agar
pour tube 2 and 3 into plate
Pour Plate Results
Most growth on tube 1 plate in small dots all around
white yellow colour
second tube had bigger colonies butless overall growth
white in colour
last plate had least growth, countable colonies
white
Sterile and Sterilized mean
-free of all life including viruses
-if a culture media is to be used for microbiologically studies, it must be sterile condition
-storing non-sterile media in fridge merely DELAYS the growth of microorganisms
What is the principle lethal agent in steam prep?
HEAT
Microorganisms are killed when their cellular proteins and enzymes are denatured and hence irreversably destroyed
The higher the temp the ______ the time for sterilization
shorter
Autoclave is
an instrument used to sterilize bacteriological media and surgical equipment
-based on same thing as pressure cooker, increases temp uses build ip of steam pressure, thus preventing boiling
Normal steam sterilization: Temp kept at 121 (15-16 pounds pressure per square inch) for 15 - 20 inches
-this is usually the average used since increasing more might alter the medium and maybe make it inusable for biologixal things
results of cultivation media sterilization
Sterile: Clear
NOn-sterile tsb: CLOUDY
When transfer content of the same thing do u need to change tip? (think sterile water transfer)
NO, jsut make sure it doesnt touch anything else
Standard Plate Count (SPC)
-reveals info only related to viable organisms as u count the colonies seen on the plate after incubation (living organisms)
-determines number of organisms present in water, malk and food
-can be used to determine number of living organisms in a culture
-uses the idea that each viable cell will develop into a colony so each colony represents one cell from the orginal culture
Colony forming unit
-since a clump of cells may also give rise to a colony (and it is hard to break up clumps into single cells) this term came about
-colony forming unit is used in replacement of coloinies in a quantitative plate count
SO when number of bacteria are counted it is reported as “number of bacteria colonies/mL or number of CFU/ml
How to calculate colony forming units?
number of colonies/ dilution xamount plated (mL added)
OR
number of colonies / dilution factor
THIS DILUTION REFERS TO THE DILUTION OF THE SAMPLE BEING PLATED
Motility
-possess flagella (long extensions that create current in water)
-non-motile bacteria lack flagella
-we observed motility using tubed cukture media and direct observation under microscope
True motility
cells move across microscopic field, usually zig-zagging its way along
-true independent movement
Brownian motility
(false motion)
-observed as organisms srtay in one place but vibrate or shake
-brownian motility is due to water molecules bombarding the organism
-not a characteristic of true independent motion
Motility agar test
Stab a semisolid medium with a innoculating needle that has bacteria on it, keep it straight
-if growth observed only on the line, it is non-motile
-if spreading or doffise of bacterial growth is seen, that is motile
Semi-solid medium is softer cuz it contains less agar, making it easier for them to move
Hanging drop presentation
-add loopful of water to slide and mix small amount of bacteria on it
-ring edges of depression slide with petroleum jelly, place the depression slide over the coverslip droplet,
-examine with low power objective , then higher, focus on edfges as this is where most organisms are drawn due to tension
,for oil immersion, donr use bright light as it will make it harder to see the bacteria
Result of motility tests
Lactobacillus plantarum: Non-motile in both agar adn hanging drop (just brownian movement so not motile)
Pseudomonoas fluroescens: Yes in agar and in hanging drop
Bacterial Flagella
-can be pertrichous or polar
-characteristic of taxonomic important
-Grays’ 1926 method was one of tje classeics used to study flaggella
_issue is, bacteria are beyond the limit of resolution of the light microscope
(15nm or 0.015 um), which are 100x smaller than most bacteria cells and 10 times smaller than the limit of resolution of light microscope (.2 um)
How can we observe flagella / mode of flagella insertion (peritrouch, polar) due to their size
-coat with a mordant such as tannic acid and potassium alum, and then staining them with silver nitrate (west method, named after marcia west) THIS INCREASES DIAMETER/ THICKNESS OF THE FLAGELLA
-the dye (silver nitrate) also allows for contrast whivh makes it more easier to see
-this precipitation of the mordant will occur on any cellular debris though, and the cell body so must have clean culture and slide
Bacterial flagella experiemnt
E.coli: peritrouchous flagella
Nutrition of microorganism Experiment Overview
Basic requirements of all living organisms are:
-water
-carbon
-energy (carbohydrates (glucose, starches), amino acids)
-nitrogen (need to manufacture protein molecules; many use amino acids or protein as their nitrogen source; some use ammonium phosphate, potassium nitrate, and other inorganix nitrogen compounds)
-minerals (sodium, potassium, calcium, etc)
-and growth factors
How to determine minimal growth requirements of a organisms
-try to grow it on a plate with everything except for one ex available nitrogen.
-if it cant grow, then add in nitrogen compounds singly to determine whether it does need it to grow
Nutrition of Bacteria Results
Types of Agar Use:
-Agar Only (no nutrients)
-Agar + minerals
-Agar +minerals + carbohydrate
-Agar+ minerals + Carbohydrates + Nitrogen
Agar Only
-B. Subtilis: NO
E.coli: NO
Pseudomonas Fluorescens: NO
Lactobacillus Plantarum: No
Agar+ Minerals:
B. Subtilis: NO
E.coli: NO
Pseudomonas Fluorescens: NO
Lactobacillus Plantarum: No
Agar +Min+CArb
B. Subtilis: Yes
E.coli: Yes
Pseudomonas Fluorescens: Yes
Lactobacillus Plantarum: No
Agar+Min+Carb+Nitro
B. Subtilis: Yes
E.coli: Yes
Pseudomonas Fluorescens: Yes
Lactobacillus Plantarum: YES
Metabolic activity of Bacteria
Fermentation of carbohydrates
Production of Indole
Activity of Urease
Production of hydrogen sulfide
Fermentation of Carbohydrates
Allow us to differentiate species on the basis of whether or not they can use a carbo as well as the products that may be formed in the fermentation reaction
use tubes that contain one specified carbohydrate and a durham tube inside to drap sny acidic gases ofrmed
-also have a colour indicator in the tube, like bromocresol purple, that help differentiate a change in pH if acid is produced
-change in colour indicates if the organism ferments that carbohydrate
-growth indicates if that organism can use that carbohydrate
-some can produce acid but no gas or some can peodue both
-if org dont ferment the carbo thenthey may produce alkanine results by using proteins in broth
Differences are caused by difference in environment
Production of Indole
-produced due to the breakdwon of amino acid tryptophan
-Can be detected by adding kovac;s reagent to the culture
-turns bright red
Activity of urease
-splitting of the urea molecule releasing carbon and ammonia is done by urease
-urea must be added to the culture as the substrate
-phenol red is used as a pH indicator
-due to ammonia being released when urease breaks down urea, this causes the pH to turn alkaline, leading to a change in colour (dark pinK)
since urease in common in proteus, it is a common test used to differentiatied between proteus species and others that resemble it
What is rapid urease a characteristic of
proteus species and very few enteric bacteria
Production of hydrogen sulfide
produced when amino acids containing sulfur are metabolized by orgsnisms
-if medium contains metallic ions such as l,ead =, bnismjuth or iron, (in. addition to the appropreiate amino acid), these metals combine with the hydrogen sulfide that is produced by the amino acid rto form a metallic sulfide that blackens the mediumn
SIM medium is convienient for this (contains iron and is semi solid)
Results of the metabollic activities of bacteria
FERMENTATION OF CARB:
Glucose:
E.coli:Yes (G)
Pseudomonas aeruginosa:Yes
B. Subtilis: Yes
Proteus mirabilis:Yes (G)
Lactose:
E.coli:Yes (G)
Pseudomonas aeruginosa: NO
B. Subtilis: No
Proteus mirabilis: NO
Sucrose:
E.coli:NO (G)
Pseudomonas aeruginosa: NO
B. Subtilis: YES
Proteus mirabilis: NO (G)
INDOLE PRODUCTION
E.coli: YES
Pseudomonas aeruginosa: NO
B. Subtilis:NO
Proteus mirabilis:NO
Urease Activity
E.coli: NO
Pseudomonas aeruginosa: NO
B. Subtilis: NO
Proteus mirabilis: YES (PINK)
Hydrogen Sulfide:
E.coli: NO
Pseudomonas aeruginosa: NO
B. Subtilis: NO
Proteus mirabilis: YES
WHat colour indicated acid was produced for fermentation of carbs? BAse?
Bromocresol purple
turned yello when acid
turned dark blue when base
Bacterial enzymes
-some have enzymes that can break down polysaccharides, proteins or lipids
these are extracellular and do this via hydrolysis
ex: Carbohydrases (such as amylase) hydrolyzse polysacchardies into sugars
Proteases: hydrolyze proteins to polypeptides and amino acids
Lipases: hydrolyze lipids (fats) to glycerol and fatty acids
Endoenzymes:
Catalase and oxidase
Catalsae: acts on hydrogen peroxide, which is formed as a oxidative end product of aerobic respiration, breaking it down to water and oxygen
-bubbles of o2 can be seen when 3% hydrogen peroxide is added to a catalase pos organism
Oxidase: activates oxidation of reduced cytochrome c by molecular oxygen in aerobic orgs during ETC
Oxidative cytochrome c transers O2 to tetramethyl-p-phenylenediamine when the reagent is added to oxidase pos org
Enzymes results
Starch Agar:
-amylase activity detected using Gram’s Iodine since iodine in presence of starch is Blue, if amylase was present, starch would have been hydrolyzed so a clear zone would appear
Ecoli: NO
B.subtilis:Yes
Pseudomonas aeruginosa: NO
Tween Agar:
-indicates presence of lipase activaty by having cloudy areas, hydrolysis of lipids means that a prepicipitate is formed so cloudy
Ecoli: NO
B.subtilis:Yes
Pseudomonas aeruginosa: Yes
Milk Agar:
-casein is nitrogen (protein) present in milk
-if clear zone present, means that casein was hydrolyzed and protease is present
Ecoli: NO
B.subtilis:Yes
Pseudomonas aeruginosa: Yes
Catalase:
Ecoli, b.subtilis and p. aeruginosa have
Oxidase:
Ecoli. NO
B.sub (variable, 6 seconds)
Pseudomonas Aeruginosa: Yes
Effect of temperature on growth
Micro have a wide range they can grow at but they also have am OPTIMAL GROWTH TEMPERATURE at which bichem rxns happen at maximum speed
Names of Bacteria temp categories
Psychrophiles: 0 degrees
Thermophiles: high temps, 75 degrees
Mesophiles: intermediates, 20-45
each type has a min, max, and optimum growth temp
Temperature experiment results
4-6
Staphyloccoccus Aureus: no
Serratia Marcesscnes:no
Pseudomonas flurpescens: +
Bacillus Stearothermophilus: ++
Saccharomyscyes cerevisae: ++
Aspergillus niger: no
20-25
Staphyloccoccus Aureus: +++ Yrllow
Serratia Marcesscnes: +++ red
Pseudomonas flurpescens: +++
Bacillus Stearothermophilus: no
Saccharomyscyes cerevisae: +++
Aspergillus niger: ++
37:
Staphyloccoccus Aureus: +++
Serratia Marcesscnes: +++ red
Pseudomonas flurpescens: +
Bacillus Stearothermophilus: ++
Saccharomyscyes cerevisae: +
Aspergillus niger: +++
55:
Staphyloccoccus Aureus: no
Serratia Marcesscnes:no
Pseudomonas flurpescens: no
Bacillus Stearothermophilus: +++
Saccharomyscyes cerevisae: no
Aspergillus niger: +
What were the optimal temps for the bacteria used
Staphyloccoccus Aureus: 20-37
Serratia Marcesscnes: 20-37
Pseudomonas flurpescens: 20-25
Bacillus Stearothermophilus: 55
Saccharomyscyes cerevisae: 20-25
Aspergillus niger: 37
pH and microbial growth
-dependent on hydrogen ion concentration of the org environment
-has greatest influence on growth aside from temp
-term pH is relative to the conc of ions
-there is optimum amount of pH that orgs grow best in
when do hydrogen ioon requirements become different
when other factors are not held constant such as temp, composition of medium, and osmotic pressure
pH results
checl
Oxygen requirements of microorgansism
Obligate aerobes: need oxygen to survive
Obligate anaerobes:canot grow in the presence of oxygen
Faculative orgs: can grow in the presence or absence of oxygen. grow better with oxuygen but can grow without. Will use oxygen or some alternate form of it but prefers oxygen over alternate
Faculatitive anerobe is the ones that spceifically grow better with oxygen
Microaerophiles: require small amounts of oxygen
Indifferents: show no preference for one or the other, will grow equally well in aerobic or anaerobic conditions
Media for aerobic or anaerobic bcteria
Anaerobic:
media containing sodium thioglycollate or cysteine lower oxi-red potential to favour anaerobic growth
-usually also contains agar, resazurn, yeast extract and proteins
Resazurin: dye used to help indicate presence of oxygen (becomes pink when o2 present so will be pink on top, colourless in middle and bottom)
Agar: helps localize the growth and favours anaerobic growth at the bottom
Even though this medium is better for anaerobic growth, aerobic will happen at the top where oxygen is higher
Results of oxygen requirements
Ecoli: Faculatative anaerobe
Staphylococcus aureus: obligate aerobe
Lactobacillus plantarum: aerotolerant anaerobe
Clostridium sporogenes:
microaerophile
What bacteria is not harmed by uV
Photosynthetic bacteria
What type of UV damages bacteria
(210-300 nanometers) Short UV wavelengths are the only ones that cause damage to non-photosynthetic bacteria
What is the peak lethality of UV
265 nanometers, produces germacidal effects
What does UV light do
-causes peroxides to form in medium, which turn into oxidizing agents
-exposure of nucleic acids to this cause mutations bc of abmnormal linkages forming between nitrogenous bases, SPECIFICALLY THYMINE DYMERS due to base thymine interacting with other thyymine groupings
AS a result abnormal protein molecules are produced, or none at all, which may prevent replication so LETHAL
Greater the amount of UV exposuer, the greater the damage, so good for sterilization
UV experiment results
organisms are tested to see the minimum amount of uV light that causes to kill 100% of the organisms
Staphylococcus Aureus; vegetative, nonspore forming:
0s: ++++
0.5min: ++++
1min: ++
2: +
3: –
3min w lid on: +++
Bacillus Megatareium: Spore forming:
0s: ++++
0.5min: ++++
1min: +++
2: ++
3: ++
3min w lid on: +++
B Megatarium in stationary phase will?
show best survival following exposure to UV radiation as SPORES FORM here cuz nutrients limited
Thermal death time
Time it takes to kill a suspension of cells or spores at a given temperature
Thermal Death point
temperature at which an organism is killed in 10 minutes
What affects the measurement of temperature things TDT and TDP
-pH, moisture, composition of medium and age of cells greatly influence TDT and TDP
Results of Lethal effects of temperature
Bacillus Megaterium:
60 degrees
15 +++
30 ++
45 ++
80 degrees:
15 +++
30 ++
45 ++
100 degrees:
15 +++
30 ++
45–
Ecoli:
60 degrees
15 +++
30 +
45 –
80 degrees:
15 ++
30 –
45 –
100 degrees:
15 –
30 –
45 –
Osmotic pressure definitions
Hypotonic: low solute content of solution
-for most except marine bacteria, not harmful
Hypertonic: high solute content of solution
-growth is probably inhibited, degree of inhibition dependent on conc of solute and nature of org
-cytoplasm becomes dehydrated and shinks away from the cell
PLASMOLYSZED cells, lack water inside but are able to bounce back to regular gunction when placed in isotonic solution
Other organisms may be irreversibly affected due to permanent inactivateion of enzymes
Osmotic pressures test results
check table
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