Lab 22 - Antibiotic Sensitivity Flashcards
what is an antibiotic?
a drug naturally made by certain fungi or bacteria against competing microbes
what is an example of an antibiotic?
penicillin made by Penicillium mold
nowadays, what are two things that antibiotics include?
Nowadays, the term includes semi-synthetic and synthetic drugs as well
what are sensitive bacteria?
knocked out by antibiotics, aka susceptible
what are resistant bacteria?
they survive an attack by antibiotics
in a non-resistant cell, what happens when antibiotics enter?
the antibiotics enter the cell and bind to the target
in a resistant cell, what happens when antibiotics enter?
- Increased elimination: the bacterial cell pumps out the antibiotic after it enters the cell.
- Decreased uptake: The porin proteins prevent antibiotic entry into the cell, so the bacteria don’t allow entry.
- Alteration in target molecule: The antibiotic cannot bind to the bacteria/target.
- Antibiotic-inactivating enzymes: the enzymes modify the antibiotic, inactivating it.
what are three tests to determine antibiotic sensitivity? AKA what are the 3 antibiotic sensitivity assays?
- Broth Dilution test.
- E-test
- Kirby-Baur Disk Diffusion test
what is the broth dilution test?
it exposes bacteria to higher and higher concentrations of antibiotics, and no growth means the antibiotic is effective
what is MIC?
Minimum inhibitory concentration - which is the lowest antibiotic concentration that kills or inhibits the growth of a microbe
what is the E-test?
tests the MIC as well
what is the Kurby-Bauer Disk Diffusion Test?
-When you streak the desired bacterial species all over the plate (3x) to get a lawn.
-Place paper disks with specific antibiotics on the plate.
-Antibiotics diffuse out from the disks creating a concentration gradient - which is higher near the disk
what happens in the Kirby-Bauer test as antibiotics diffuse out? where is the MIC?
-As antibiotics diffuse out, they kill or inhibit the growth of bacteria forming clear “zones of inhibition”.
-The concentration at the edge of the zone is the MIC
when comparing the diameter of the zones of inhibition to the chart, we can determine what?
we can determine if the bacteria are sensitive to the antibiotic
what do the zones of inhibit size in mm mean for the bacteria Cefalothin? (eg. if it is sensitive, intermediate, or resistant) REMEMBER: THESE MEASUREMENTS ARE SPECIFICALLY FOR THE CEFALOTHIN BACTERIA
Cefalothin:
Sensitive: if the zone is bigger or more than 18 mm (this means it can be used in treatment)
Intermediate: if the zone is 15-17 mm (this means it MAY be successful in treatment)
Resistant: if the zone is smaller or less than 14 mm (this means it cannot be used in treatment)
can you compare zones of inhibition of two different antibiotics?
No, you cannot because each antibiotic has a different size and chemical properties, so each diffuses at a different rate across the agar
do disks of antibiotics contain different concentrations or the same concentration of each antibiotic?
the disks contain different concentrations of each antibiotic
what is the purpose of the antibiotic sensitivity lab?
to test different bacteria against several antibiotics and see which ones they are sensitive or resistant to
what test did we use for the antibiotic sensitivity lab? what is the medium that we used? what did we streak on the medium?
-Kirby-Bauer Disk Diffusion Test
-Medium: Mueller-Hinton Agar (a special agar)
-We used a sterile swab and streaked the bacteria evenly on the plates, using horizontal, vertical, and diagonal tight streaks.
what did we do after streaking the bacteria onto the plate?
we used sterile forceps to place antibiotic disks on the plates. We made sure they are are equidistant from each other, and we only used 5 disks.
what did we incubate the plates at?
37 degrees Celsius for 24-48 hours
why did the Pseudomonas aeruginosa plate turn green?
due to a diffusible green pigment made by the bacteria
where does the zone of inhibition fall in respect to the two circles for sensitive bacteria?
the zone of inhibition falls outside the second circle
what does it mean when the disk has no zone?
that means the bacteria is resistant to the antibiotic
if bacterial colonies are present within a zone of inhibition, what does that mean?
that means they are resistant mutants in the culture because they grew where others died. (they are numerous, small white dots)
what culture was our table given for the lab?
Staphylococcus epidermidis (G+)
what are two reasons a bacteria could be resistant from our lab?
- Innate:
- Mycoplasma
- E. coli - Acquired:
-Mutations
-Gene transger
what are narrow-spectrum antibiotics? an example?
Antibiotics that only work for specific bacteria. An example is Penicillin
what are broad-spectrum antibiotics? an example?
Can kill more than one type of bacteria. An example is Ampicillin
which bacteria was most resistant to antibiotics?
pseudomonas was most resistant to antibiotics
explain why a bacterial colony might grow in a zone of inhibition.
these represent mutants because they grew where others died. it developed a mutation to resist antibiotics
how did we place the antibiotics onto the plate?
by placing the forceps in alcohol, then putting it over the fire, then picking up the antibiotic disk
