Lab 32: Antibiotic Sensitivity

 Antibiotic= a natural substance (made by fungi or bacteria) that inhibits or kills microorganisms

-in common usage antibiotics are often equated with antibacterial drugs and synthetic drugs

      --in 1928, Alexander Fleming found a mold contaminant on an agar that inhibited growth of Staphylococcus aureus

-the mold was Penicillium chrysogenum and so the inhibitory chemical was named Penicillin

        -both Penicillin and Sulfas were developed during WWII, with derivatives also being developed 

         -antibiotics were thought to be the "magic bullet" to wipe out bacterial infections

         - However, bacteria are becoming resistant to traditional antibiotics 

Bacteria develop resistance through:

1. mutation 

2. acquisition of R plasmids 

        -resistance promoted from the misuse/overuse of antibiotics

Resistant Bacteria=

-S. aureus (from Flemings lab) is now major resistant bacteria 

-MRSA=methicillin resistant S. aureus

-VRSA= Vancomycin resistant S. aureus

       -Increased bacterial resistance, means that more antibiotics are needed, but drug companies don't want to produce        antibiotics because they are too expensive to make

                 -can take up to 10 years to make: must find, test, get approved, produce

                 -and after that bacteria can still develop resistance to drug

3 ways to treat infections

1. Trial and error approach= patient given a drug that may or may not work

2. giving them lots of antibiotics- ie kitchen sick= promotes drug resistance and has many side effects to          patient

3. Determining susceptibility is the best way to give patients the proper drugs in the proper dosage

Ways to Determine Susceptibility

1. Minimum Inhibitory Concentration (MIC)= the minimal concentration one is needed to inhibit growth of bacterium

used in:

a. Broth dilution methods-fill test wells with increasing concentration of antibioitic and with the same      concentration of bacterium, then look for turbidity, 1st well with no cloudiness= MIC

b. Kirby-Bauer Disk DIffusion test= small disks with standard concentrations of antibiotics are placed on agar with bacterium

-the antibiotics diffuse outward, creating a zone of inhibition (no growth) around disks

-the size of the zone cannot be directly compared due to the variable solubility and diffusion rates of the  drugs in the medium, so zones must be compared to a standardized chart to assess if the bacteria are sensitive or resistant

-ie. only tells you if bacteria is sensitive or resistant 

c. E test- small strips with quantified antibiotic gradients are used

-zones of inhibition form a teardrop (since high concentrations will diffuse farther than low concentrations)

-the smallest part of teardrop=MIC

-advantage of this test is that it is quantitative for MIC

In this Experiment- test selected antibiotics using the Kirby Bauer Disk Diffusion Test 

-ie. tells you if bacteria is sensitive/resistant and correct dose

-once we grew plates, we looked for zones of inhibition 

-then using a transparency, we overlaid the appropriate antibiotic disk diagram onto its antibiotic disk to determine if the bacteria are Sensitive (S), Intermediate (I) or Resistant (R) to each antibiotic

- The bacteria are S,I, or R if they are in the S,I, or R zone 

R-Resistant= Zone of inhibition with a diameter equal to or less than that of the inner circle

I- Intermediate=Zone of inhibition with a diameter greater than R, but less than that of the outer circle

S- Sensitive= Zone of inhibition with a diameter equal to or greater than that of the outer circle 

****note- Colonies growing WITHIN the zone of inhibition = bacterium resistant to antibiotic (Yikes!)

 -We had two plates

- antibiotic disks with NO ZOI are all Resistant to antibiotic

-so for #33 thats all but one disk 

Plate #16

 Sensitive

 Sensitive 

Sensitive

Resistant 

(I think some got Intermediate on this one)

 Plate #33 =Sensitive