Antibiotic Susceptibility Test

Introduction

The presence of antimicrobial agents at low concentration through leaching or continued usage may lead to the development of drug-resistant strains and Multiple Antibiotic Resistance (MAR) in bacteria which may result to resistance transfer to pathogenic bacteria and reduced efficacy of antibiotic treatment for human and animal diseases. Several studies have been done to investigate the possible consequences of the used of antimicrobials.

Antibiotic susceptibility pattern has long been utilized as useful epidemiological markers since it is very simple to perform and the results could be obtained in a ver y short time and are easy to interpret (Tenover et al., 1995). The existence of bacterial antibiotic resistance may be as a result of non-clinical used of antibiotics in both animal and humans.

Resistance to antimicrobial compounds can be conferred by innate structural features of microorganisms such as an impermeable outer membrane that resists penetration of antibiotics. Gram-negative bacteria have a thick lipopolysaccharide layer that acts as a barrier to limit diffusion of antibiotic molecules into the cell while gram-positive bacteria characteristically have lipophilic substances in their cells walls that retards penetrations of hydrophilic, cationic and antimicrobial compounds. In addition to barriers, microorganisms may possess a variety of other resistant mechanism. For example, the organism may lack a transport system necessary for antibiotic uptake or be lacking the biochemical target required for attachment and proper functioning of the antimicrobial compound (Volk et al., 1996).

The multiple antibiotic resistance index of the isolates is defined as a/b where ‘a’ represents the number of antibiotics to which the particular isolate was resistant and ‘b’ the number of antibiotics to which the isolate was exposed to (Krumperman, 1983).

Objectives

To expose student with the knowledge of the Antibiotic Susceptibility Test

Materials: 1. 2. 3. 4.

Mueller Hinton Agar Pure culture Sterile cotton swab Antibiotic discs

Methodology: 1. Briefly, organisms were grown at 37ºC in Mueller Hinton broth. 2. Ten microliters of the overnight culture were used to inoculate a fresh 3. Mueller Hinton broth followed by incubation at 37 ºC with shaking until a 0.5 McFarland turbidity standard (biomerieux, France) was obtained. 4. A sterile swab was dipped into this culture and used to inoculate the surface of a fresh Mueller Hinton agar plates and were allowed to dry for 2 to 5 minutes. 5. The antibiotics discs were spaced out onto the plates and were incubated at 37ºC for 24 hours. 6. The diameter of the clear zone or each antibiotic disc was interpreted as susceptibility or resistant categories according to the guideline recommended by the National Committee for Clinical Laboratory Standards (2004). Observation:

Antibiotics Discs Types of Culture

Streptomycin

Nycin

Kanamycin

Diameter of clear zone:

Diameter of clear zone:

Diameter of clear zone:

1.8 cm

2.0 cm

susceptible

susceptible

Diameter of clear zone:

Diameter of clear zone:

1.8 cm

2.2cm

susceptible

susceptible

Staphylococcus aureus

1.8 cm

susceptible Salmonella sp.

Diameter of clear zone: 1.8 cm

susceptible

Diameter of clear zone: 2.0cm

Diameter of clear zone:

Diameter of clear zone:

1.8 cm

2.2cm

susceptible

susceptible

Vibrio para susceptible

Discussion:

Antibiotic sensitivity is a term used to describe the susceptibility of bacteria to antibiotics. Antibiotic susceptibility testing (AST) is usually carried out to determine which antibiotic will be most successful in treating a bacterial infection in vivo. In the disk-diffusion susceptibility test, disks containing known amounts of an antimicrobial agent are placed on the surface of an agar plate containing a non selective medium that has been inoculated with a suspension of a strain. The antimicrobial agent diffuses into the medium, causing a zone of inhibition of growth of the strain around the disk corresponding to the susceptibility of the strain to the agent. Bacteria are not able to grow around antibiotics to which they are sensitive. Interpretative inhibition zone diameters have been established for susceptibility test results to permit classification of an isolate as being susceptible, intermediate (or exhibiting decreased susceptibility), or resistant to an antimicrobial agent. From the result obtained, we can see that kanamycin inhibit the growth of bacteria the most followed by streptomycin and nycin. Usually, according to National Committee for Clinical Laboratory Standards, susceptible categories is interpreted when the clear zone is ≥ 18 mm. From the result, we can be confirmed that all the bacteria were susceptibled to all 3 antibiotics.

Conclusion:

From this experiment, we learned to perform Antibiotic Susceptibility Test and observe the result on how certain bacteria sustained to certain antibiotics. We also learned to measure the diameter of the clear zone and how to classify the isolate as being susceptible, intermediate or resistant according to National Committee for Clinical Laboratory Standards (2004).

BCS 3224 MEDICAL MICROBIOLOGY

LAB EXERCISE 3 ANTIBIOTIC SUSCEPTIBILITY TEST

LECTURER: MISS INTAN FARAHA

NAME: FARHANNA MOHAMMED MATRIC NUM: 870131565028 PROGRAM: ISM INDUSTRIAL BIOTECHNOLOGY