Antibiotics
– biochemical
substances produced by microorganisms that inhibit the growth of, or kill,
other microorganisms.
Many
drugs are now completely synthetic or the natural drug is manipulated to change
its structure called semisynthetics.
There is a large number of antimicrobial
agents available for treating diseases caused by microorganisms. Such drugs are
now an essential part of modern medical practice.
The antimicrobial agents used in medical
practice are aimed at eliminating the infecting microorganisms or at preventing
the establishment of an infection.
To be of therapeutic use, an antimicrobial
agent must exhibit selective toxicity;
it must exhibit greater toxicity to the infecting pathogens than to the host
organism. A drug that kills the patient is of no use in treating infectious
diseases, whether or not it also kills the pathogens.
As a rule, antimicrobial agents are of
most use in medicine when the mode of action of the antimicrobial chemicals
involves biochemical features of the invading pathogens not possessed by normal
host cells.
Characteristics
of an antibiotic
• It should be toxic to
the infecting organism while harmless to the host cells and the microbiota of
the host.
• It should stay in toxic
form for a sufficient amount of time to affect the infecting microorganism. If
it changes to another form or is broken down in the body, it may not be useful.
• Sufficient amounts of
it should reach the site of infection to kill the infecting agent.
• The infecting agent
should be sensitive to it.
Bacteria
respond in different ways to antibiotics and chemosynthetic drugs, even within
the same species. For example, Staphylococcus
aureus is a common normal flora bacterium found in the body. If one
isolated this bacterium from 5 different people, the 5 isolates would likely be
different strains, that is, slight genetically different.
It
is likely that if antibiotic sensitivity tests were run on these isolates, the
results would vary against the different antibiotics used.
The determination of
antibiotic susceptibility of a pathogen is important in selecting the most
appropriate one for treating a disease. There are several different procedures
used by clinical microbiologists to determine the sensitivity of microorganisms
to antibiotics.
Two such procedures are commonly
used. The first one (the Kirby-Bauer Disc Method) is used to determine which
antibiotic is the most effective against a certain pathogen. The second (MIC)
is used to determine the lowest concentration that is needed to kill the
pathogen at the site of infection.
The
Kirby-Bauer test for antibiotic susceptibility (also called the disc
diffusion test) is a standard that has been used for years.
First
developed in the 1950s, it was refined and by W. Kirby and A. Bauer, then
standardized by the World Health Organization in 1961. Now clinical labs use
automated tests. However, the K-B is still used in some labs, or used with
certain bacteria that automation does not work well with.
This
test is used to determine the resistance or sensitivity of aerobes or
facultative anaerobes to specific chemicals, which can then be used by the
clinician for treatment of patients with bacterial infections.
The
presence or absence of an inhibitory area around the disc identifies the
bacterial sensitivity to the drug
Kirby-Bauer Test to Measure Antibiotic Sensitivity.
In Kirby-Bauer testing, bacteria are placed on a
plate of solid growth medium and wafers of antibiotics (white disks) are added
to the plate. After allowing the bacteria to grow overnight, areas of clear
media surrounding the disks indicate that the antibiotic inhibits bacterial
growth. The concentration of antibiotic that diffuses into the media decreases
with increasing distance from the source.
Therefore, the more sensitive the bacteria are to a
given antibiotic, the larger the clear bacteria-free zone that forms around
the disk containing that antibiotic
The basics
The
bacterium is swabbed on the agar and the antibiotic discs are placed on top.
The
antibiotic diffuses from the disc into the agar in decreasing amounts the
further it is away from the disc.
If
the organism is killed or inhibited by the concentration of the antibiotic,
there will be NO growth in the immediate area around the disc-
the zone of inhibition
The
zone sizes are looked up on a standardized chart to give a result of sensitive,
resistant, or intermediate.
- Dip a sterile swab into the broth and express any
excess moisture by pressing the swab against the side of the tube.
- Swab the surface of the agar completely (do
not leave any unswabbed agar areas at all).
- Pictures below show what happens when the plate is not
swabbed correctly with uniformn coverage of the bacterium over the entire
agar.
- After completely swabbing the plate, turn it 90
degrees and repeat the swabbing process. (No need of re-moistening the
swab.)
- Run the swab around the circumference of the plate
before discarding it in the discard bag.
- Allow the surface to dry for about 5 minutes before
placing antibiotic disks on the agar.
- Use a pair of forceps to take an antibiotic disc from
the dispenser- the forceps have to be sterile. Place the forceps in
alcohol, flame the forceps until they catch on fire, let the flame go
out----sterile forceps.
- Lightly touch each disc with your sterile inoculating
loop to make sure that it is in good contact with the
agar surface.
- Incubate agar side of the plate down at 37o C.
Interpretation
- Place the metric ruler across the zone of inhibition,
at the widest diameter, and measure from one edge of the zone to the other
edge. Holding the plate up to the light is better to measure the zone.
- Zone diameter measured in millimeter units. If there is no zone at all, report it as 0---even
though the disc itself is around 7 mm.
- Zone diameter is reported in millimeters, looked up on
the chart, and result reported as sensitive, resistant, or
intermediate.
- Record the results.
|
Antibiotic (Antimicrobial
Agent) |
Resistant (<
or = mm) |
Intermediate
(mm) |
Susceptible (=
or > mm) |
|
Amoxicillin (other) |
<13 |
14-17 |
>18 |
|
Amoxicillin (Staph) |
19 |
|
20 |
|
Ampicillin (other) |
11 |
12-13 |
14 |
|
Ampicillin (Staph) |
28 |
|
29 |
|
Carbenicillin (other) |
17 |
18-22 |
23 |
|
Carbenicillin (Pseudomonas) |
13 |
14-16 |
17 |
|
Cefoxitin |
14 |
15-17 |
18 |
|
Chloramphenicol |
12 |
13-17 |
18 |
|
Ciprofloxacin |
15 |
16-20 |
21 |
|
Clindamycin |
14 |
15-20 |
21 |
|
Erythromycin |
13 |
14-22 |
23 |
|
Gentamycin |
12 |
13-14 |
15 |
|
Kanamycin |
13 |
14-17 |
18 |
|
Methicillin (Staph) |
9 |
10-13 |
14 |
|
Oxacillin (Staph) |
10 |
11-12 |
13 |
|
Streptomycin |
14 |
15-20 |
21 |
|
Sulfamethoxazole-trimethoprim |
10 |
11-15 |
16 |
|
Tetracycline |
14 |
15-18 |
19 |
|
Tobramycin |
12 |
13-14 |
15 |
|
Vancomycin |
9 |
10-11 |
12 |
Many
charts have a corresponding column that also gives the MIC (minimal
inhibitory concentration) for that drug.
The MIC is
currently the standard test run for antibiotic sensitivity testing because it
produces more pertinent information on minimal dosages.
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