MICROBIOLOGICAL TECHNIQUES-Mini project (Bioteknologi)-

GROUP NAME : Faradila Bt. Zakaria E20122005522

Nur Hana Bt. Syamsul E20122005474

Nur Faika Bt. Yusof E20122005520

N. Farahzatul Fariza Bt. Md. Hussin E20122005514

•Roky was cycling and suddenly fell off into the drain, thatcauses his lip swelled up and develop a fever. Roky still havea fever even though, he already prescribed cephalothin, anantibiotic related to penicillin, to cure Roky’s fever, but threeday’s later Roky still was in the hospital with a fever of39.4ºC, coughing up blood, and trouble in breathing. So thedoctor’s decided to check the MRSA and found that thereason why the fever not cool down. MRSA is any strain ofStaphylococcus aureus that has developed, through theprocess of natural selection, resistance to beta-lactamantibiotics, which include (methicillin, penicillin, cephalothin)So the Doctor’s give the Roky’s different type of antibioticthat is vancomycin. As a result Roky’s fever quickly betterafter that. So Haikal as a researcher laboratory was given a

task to investigate measuring antibioticresistance of s. aureus and MRSA

Introduction

MRSA

MRSA is the abbreviation for methicillin-resistant Staphylococcus aureus.

can cause a multitude of diseases as a result of infection of various tissues of the body.

Within one to two years, Staphylococcus aureus bacteria (S. aureus) started to be isolated that were resistant to methicillin. These S. aureus bacteria were then termed methicillin-resistant. MRSA usually show resistance to many antibiotics.

In the majority of cases, the colonizing bacteria do not cause disease. However, damage to the skin or other injury may allow the bacteria to overcome the natural protective mechanisms of the body and lead to infection; because of its ability to destroy skin, it is also one of the types of bacteria that has been termed a "flesh-eating bacterium."

• To investigate the degree of antibiotic

resistance in bacterial species using a Kirby-Bauer test.

• To developing and implementingprevention strategies that reduce the incidence of nosocomial acquired( originating or taking

placed in a hospital acquired in a hospital especially to an infection) MRSA.

• Kirby-Bauer antibiotic testing (also called KB testing or

disk diffusion antibiotic sensitivity testing)

• Function : antibiotic-containing wafers or disks to test

whether particular bacteria are susceptible to specific

antibiotics

• PROCEDURE

1) First, a pure culture of bacteria

is isolated from the patient.

2) Then, a known quantity of bacteria are grown

overnight on agar (solid growth media) plates in the

presence of a thin wafer that contains a known amount of a relevant

antibiotic.

* If the bacteria are susceptible to

the particular antibiotic from a wafer, an area of clear media where bacteria

are not able to grow surrounds the wafer, which is known as the zone of inhibition. Because the concentration

of antibiotic that diffuses into the media decreases with increasing

distance from the source, a larger zone of inhibition around an

antibiotic-containing disk indicates that the bacteria are more sensitive to

the antibiotic in the disk.

Material that we

used?

Broth cultures of MRSA and

S.Aureus & Mueller-Hinton

agar

Forceps

Bunsen burner

Discs (methicillin, vancomycin,

penicillin, cephalothin)

Whatmanfilter paper

Sterile cotton swab

Methodology:Antimicrobial Analysis Techniques

Step Procedure Justification

1 Do the experiment in the laminar flow. To prevent

contaminations.

2 Used the sterile cotton swab to create a lawn of

MRSA on one of the plates of Mueller-Hinton agar

aseptically.

To transfer broth culture

to agar plate.

Because the cotton swab

were sterile.

3 Used sterile forceps, pick discs (methicillin,

vancomycin, penicillin, cephalothin) and placed

them on opposite sides of a petri dish containing

MRSA, with the code side facing up. Make sure,

label the date and the bacteria. Then tap them gently

with sterile forceps to stick them to the agar.

Sterile forceps to prevent

contamination.

Label the date and

bacteria for make sure

that the result not

changed.

Step Procedure Justification

4 Wrap parafilm around all four plates and place its

on their lids and in the incubator at 37ºC for 48

hours.

Repeat step one until four using the plates of

S. Aureus but incubator at 37ºC 24 hours.

To prevent people who

handle them from

contamination by bacteria.

For possible pathogens

5 After 48 hours in the incubator, check for the

presence of antibiotic activity. This is done by

looking for a clear area, called a zone of inhibition,

surrounding a paper disc. Remember to never open

the dish for a better view.

To determine the affectivity of an antibiotic,

measure the diameter, in millimetre, of the zone of

inhibition.

To know which one is

very susceptibility of the

bacteria

Expected result