Lab Exam 1 – Flashcards
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Unlock answersSupplemented thioglycollate |
used to assess aerotolerance but has added nutrients for growing bacterial cultures |
Kirby Bauer Test |
tests antibiotic sensitivity -uses standardized conditions of 1/4 inch thick Mueller-Hinton agar plates; pH between 7.2-7.4; lawn made from .5 McFarland turbidity standard; antibiotic discs have a standard amount of antibiotic in them -procedure: Label plate; innoculate plate to create lawn; apply antibiotic discs without overlapping zones of inhibition; press discs firmly into agar so they stick; invert plates and incubate. |
broad spectrum antibiotic |
effective against both gram positive and gram negative cells -used when specific bacteria causing infection is unknown |
narrow spectrum antibiotic |
effective against either NOT BOTH gram positive and gram negative cells -used when cause of infection is known - prevents killing off good bacteria in the body |
S10 |
Streptomyocin -gram negative narrow |
C30 |
Chloramphenicol -broad spectrum |
CF30 |
Cephalothin -broad spectrum |
TE30 |
Tetracycline -broad spectrum |
P10 |
penicillin -not effective |
NB 30 |
Novobiocin -gram positive narrow |
S10 |
Streptomyocin -gram negative narrow |
E15 |
Erithromyocin -gram positive narrow |
VA30 |
Vancomyocin -gram positive narrow |
E. coli gram ... |
negative |
S. aureus gram ... |
positive |
Antiseptic |
used on living tissue -longer contact time (time it continues to work) -iodine |
Disinfectant |
-used on inanimate objects -decreases number of bacteria, but not sterilizing -bleach |
confluent growth |
continuous bacterial growth without discrete colonies |
lawn on agar plate |
covering the entire surface of a plate with bacteria to grow -start in middle and do one half, back and forth; then start in middle and do other half; rotate 90 degrees; start in middle and do one half; then start in middle and do other half |
gram negative bacteria are more resistant to antibiotics because |
of their outer membrane |
Gram positive cells are more or less susceptible to antibiotics than gram negative cells? |
Gram positive cells are more susceptible |
Zone of inhibition |
-area of no bacterial growth around an antibiotic disc -measured in mm |
resistance OR susceptibility to antibiotic shown through zone of inhibition |
must compare measurement to standardized chart |
aseptic technique |
-used to prevent contamination/keep cultures pure -procedure: use inner blue cone on bunsen burner; sterilize loop; remove cap of tube with little finger; flame tube; cool loop on tube; flame tube; close tube; innoculate culture with loop; sterilize loop |
streak plate technique |
-purpose: to isolate a single colony -procedure: streak loop across plate in 1/4 of plate; sterilize loop; rotate plate; cool loop on agar; drag through previous section 2-3 times and then streak another 1/4 of plate; sterilize loop; rotate plate; cool loop on agar; drag through previous section 2-3 times and then streak another 1/4 of plate; sterilize loop; rotate plate; cool loop on agar; drag through previous section 2-3 times and then streak remainder of plate; sterilize loop |
errors in streak plate technique |
-digging loop into agar -too many streaks through last quadrant before moving on to next quadrant -overlapping streak sections creating confluent growth |
BSL |
Bio Safety Level |
precautions used for BSL 1 and 2 |
-goggles -gloves -lab coats -close toed shoes -long pants -hair pulled back -disinfectant for tables -sterilization of tools -autoclave contaminated materials |
biohazard disposal |
-double bag anything that goes in biohazard bin -biohazard glass or sharps go in red biohazard sharps container -gloves inside out in biohazard bag |
Broth is used for: |
?? |
slant is used for: |
-using less media than a plate (cost) ?? |
plate is used for: |
-identifying growth patterns (shape, margin) -isolating pure colonies |
aseptic |
pure (not sterile) |
sterile |
no living organisms |
Thioglycollate |
prepared using autoclave to remove all O2 -as it cools, the O2 reenters, creating an O2 gradient from aerobic to completely anaerobic in bottom of tube |
Obligate aerobe |
needs O2 to survive -only on top of tube |
Obligate anaerobe |
needs no O2 to survive -only on bottom of tube |
Facultative anaerobe |
prefers O2, but can adapt to grow without it -grows anywhere, better on top |
microaerophile |
loves a LITTLE O2 -grows in a band about 1/3 way down the tube |
Aerotolerant anaerobes |
prefers no O2, but can tolerate growth with O2 -grows anywhere, but better on bottom |
maximum temperature |
highest temperature for cells to be able to grow |
minimum temperature |
lowest temperature for cells to be able to grow |
optimum temperature |
best temperature for cells to be able to grow - highest growth rate |
cardinal temperatures |
range defined by the three temperatures in the range: -optimum -maximum -minimum |
temperature range |
the temps between the highest and lowest that an organism will tolerate to grow |
psychrophile cardinal temperatures |
Range: -5 to 15C Optimal: 10C icecap |
psychrotroph cardinal temperatures |
Range: -3 to 35C Optimal: 25C Santa Cruz |
mesophile cardinal temperatures |
Range: 15 to 45C Optimal: 40C rainforest; human pathogens |
thermophile cardinal temperatures |
Range: 40 to 80C Optimal: 70C hot spring |
extreme thermophile cardinal temperatures |
Range: 65 to 110C Optimal: 95C hot spring |
procedure for determining temperature category of microorganism |
-inoculate multiple specimens and grow at several different temperatures -compare growth on all plates to rule out temperature ranges where there was no growth -choose temperature category that includes all temperatures where there was some growth in medium |
hypotonic |
-fewer ions outside cell -low osmotic pressure -water moves into cell |
hypertonic |
-more ions outside cell -high osmotic pressure -water drawn out of cell, so plasma membrane shrinks away from cell wall (plasmolysis) |
isotonic |
-same amount of ions (osmotic pressure) inside and outside cell -fluid moves at same rate into and out of cell -0.85% osmotic pressure |
plasmolysis |
plasma membrane shrinks away from cell wall due to cell being in hypertonic solution |
halophile |
loves hypertonicity |
facultative halophile |
range: 0.85% to 15% -can grow with hypertonicity) |
obligate halophile |
range: must have over 3% |
extreme halophile |
range: must be over 15% up to 20% |
halotolerant |
can grow up to 5% but not well |
non-halophile |
grows most abundantly at 0.85% |
osmotic pressure order of organism tolerance |
non-halophile halotolerant facultative halophile obligate halophile extreme halophile |
order of organism tolerance (temperatiures) |
psychrophile psychrotroph mesophile thermophile extreme thermophile |
which osmotic pressure is closest to the average biological cell? |
0.85% |
purpose of including water as a sample in disinfectant experiment |
control |
procedure for preparing bacterial smear slide from broth culture |
-label slide -use transfer technique to apply bacteria to slide (3-4 loops) -heat fix bacteria using hot plate and/or flaming slide |
procedure for preparing bacterial smear slide from plate/slant culture |
-label slide -add a drop of DI water to slide -use transfer technique to apply small amount of bacteria to slide -heat fix bacteria using hot plate and/or flaming slide |
what does heating a slide after applying bacteria do? |
-fixes the bacteria to the slide so they don't wash away -kills the bacteria |
simple stain |
single stain applied to cells which makes them visible under the microscope -allows determination of cell size, shape and arrangement |
procedure to prepare a simple stain |
-begin with prepared slide -drop stain onto smear and let sit for 1 minute -rinse with DI water -blot w bibulous paper |
gram staining tell us what? |
-differential stain gives you information about whether a cell is gram positive or gram negative (has thick peptidoglycan VS outer membrane) |
differential stain |
allows for detection of differences between organisms or structures within one organism |
stains adhere through |
ionic bonding |
gram staining steps |
-primary stain - crystal violet -mordant - iodine -decolorizer - alcohol -secondary stain (counterstain) - safranin |
general differential staining steps |
-primary stain -mordant -decolorizer -counterstain/secondary stain |
gram stain pink results |
gram negative (outer membrane) -E coli - short bacillus |
gram stain purple/black results |
gram positive (thick peptidoglycan) -S aureus - coccus -B subtilis - long bacillus |
Microscope parts |
-on/off switch -rheostat (dimmer switch) -light source (bulb) -condenser lens -condenser lens adjustment knob -iris diaphragm adjustment lever -specimen stage -specimen stage positioning knobs -coarse and fine focus knobs -objective lens -occular lenses -diopter adjuster in left ocular lens -interpupillary distance adjuster |
focal plane |
plane in space at which the lens is in focus (parfocal lenses have the same focal plane) |
how do you calculate magnification of an objective lens? |
multiply number on lens by 10 (for ocular lens magnification) |
parfocal objectives |
have the same focal plane; when they are exchanged they are very close to being in focus |
oil immersion improves resolution because |
it minimizes light refraction by removing air from the light path |
resolution |
sharpness of image |
magnification |
size of image |
Colony Morphology Shape (3) |
-circular -irregular -punctiform (pin-point) |
Colony Morphology Margin (4) |
-entire -undulate -lobate -filamentous |
entire |
smooth edge colony morphology margin |
undulate |
wavy edge colony morphology margin |
lobate |
bumpy edge colony morphology margin |
filamentous |
hairy edge colony morphology margin |
colony or slant morphology texture (3) |
-moist -mucoid -dry |
colony or slant morphology optical properties (2 pair) |
-opaque/translucent -dull/shiny |
slant morphology growth pattern (3) |
-filiform -spreading -friable |
friable |
crusty |
filiform |
-growth restricted to inoculation site -edge is clear and defined |
Broth cultures morphology (4) |
-uniform fine turbidity -pellicle -sediment -flocculent |
Colony or slant Morphology Pigment |
Any |
Uniform fine turbidity |
uniform cloudiness |
Pellicle |
raft floating on broth |
sediment |
accumulation on bottom |
Plate morphology categories (5) |
-shape -margin -texture -optical properties -pigment |
Slant morphology categories (4) |
-texture -optical properties -pigment -growth |
procedure for loop transfer of culture (10 steps) |
-sterilize loop -remove tube cap -flame tube mouth -cool loop on tube -dip loop -remove loop w/o flicking -flame tube mouth -close tube cap -innoculate plate, slant or broth -flame loop |
how to get single colonies on a streak plate (3 steps) |
-4 quadrant -flame loop between each quadrant -drag cool loop through previous quadrant before moving into next |