Sensitivity test

Antimicrobial Sensitivity test

An important function of diagnosis in lab of microbiology, it is, help the physician to select effective antimicrobial agents for specific therapy of infectious disease. It's means the ability of antibiotics or other antibacterial agent to inhibit bacterial growth in vitro.

He observed inhibition of staphylococci on an agar plate contaminated by a Penicillium mold

Antibiotics 

Are biochemical substances produced by some microorganisms that inhibit the growth or kill other microorganisms. Antibiotics can be classified into 1-Natural antibiotic agents: Metabolic products produced by microorganisms (bacteria , fungi)

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2-Semi-synthetic antibiotic agents: chemically modified natural agents (large group of modern antibiotics), penicillin V 3-synthetic antibiotic agents: Chemically related to natural antibiotics but completely industrially manufactured (sulfonamide, quinolones)

Mechanisms of action of the most antibiotics:

1. Inhibition of cell wall synthesis, some antibiotics, e.g. penicillin, and cephalosporin interfere with the cell wall synthesis and cause bacteriolysis. 2. Inhibition of cytoplasmic membrane function, some antibiotics cause disruption of the membrane and leakage of cellular proteins and nucleotides leading to cell death, e.g. nystatin, polymyxin.

Mechanisms of action of the most antibiotics:

3-Inhibition of nucleic acid synthesis (DNA or RNA). e.g. metronidazole, nalidixic acid 4. Inhibition of protein synthesis, many antimicrobial agents block protein synthesis by acting on the 30s or 50s subunits of the bacterial ribosome, e.g. chloramphenicol, tetracycline, erythromycin, gentamycin, streptomycin. 5. Antimetabolite drugs like sulfonamide and trimethoprim, these analogues compete with the metabolites in the metabolic processes because of their similarity.

Mechanism of action of the most antibiotics

The selection of antimicrobial agent for treating disease depends on several factors including : 1.Antimicrobial agents should have selective toxicity that means drug kills harmful microbes without damaging the host. 2.Be stable when stored in solid or liquid form 3.Most antimicrobial agents have some side effects, produce allergic reactions.

Antibiotic are classified according to their application and spectrum of action

Narrow spectrum antibiotics. broad spectrum antibiotics. Depending on their activity against range of Gram positive bacteria and Gram negative bacteria & other different microorganisms

bacteriostatic

bactericidal
without

Resistance to antimicrobial drugs

1-The organism produces enzymes that destroy the drug. e.g. production of beta – lactamases. 2-The organism changes its permeability to the drug, by modification of protein in the outer cell membranes, e.g. resistance to polymyxins.

Resistance to antimicrobial drugs

3-The organism develops an altered receptor site e.g., resistance to aminoglycoside is associated with alteration of a specific protein in the 30s subunit of the bacterial ribosome. 4-The organism develops an altered metabolic pathway e.g. resistance to sulfonamide.



Q 1 why most antimicrobial agents have selective toxicity for microbial cells but not mammalian cells?

Measurement of Antimicrobial activity :A-Disc diffusion method, Kirby-Bauer Test:Procedure :
1-Using the spread plate technique by sterile swab spread the culture of bacteria. leave(5-10)min to dry the surface of agar. 2-By using forceps add to the disk to the surface of an agar plate. 3- invert the plates & Incubate at 37Co for 18–24 hours. 4-Measure the diameters of inhibition zone in (mm) and compared for size values listed in a standard chart.

Procedure

0.1ml
m.o
inoculate
Aug
Az
Van
Cef
Az

incubate

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=
26 mm
Inhibition zone

B. Dilution method

This procedure establishes the concentration of antibiotic that is effective in preventing the growth of the pathogen. Standardized microbial inoculums is added to tubes containing serial dilution of an antibiotic the growth of microorganism is monitored as change in turbidity.

By this procedure we can determined the lowest concentration of an antibiotic that inhibits the growth of microorganism.
Minimum inhibitory concentration ( MIC )

Procedure :

1.6
No antibiotic
No inoculums
MIC
MBC
0.4ml broth +0.1ml E.coli
Con. of antibiotic

3.2 mg/ml

1.6 mg/ml
0.8 mg/ml
0.4 mg/ml
0.2 mg/ml
0.1 mg/ml
Sub-culture to agar medium
MIC = 0.8 mg/ml
Minimal Inhibitory Concentration (MIC) vs. Minimal Bactericidal Concentration (MBC)
con. of antibiotic

Thank you