Faculty of Medicine and Health Sciences Microbiology Lab Second semester 2013-2014

1. Faculty of Medicine and Health Sciences Microbiology Lab Second semester 2013-2014 prepared by: Mohammad Al-Qadi E-mail: m.qadi@najah.edu

2. Assessment of antibacterial activity of disinfectants and calculation of the phenol coefficient  Lab # 9

3. Introduction Antiseptics: - Chemicals used on skin and mucous membranes to kill microorganisms -Not for use on inanimate objects Disinfectants: -Chemicals used to kill microorganisms on inanimate objects -Not for use on skin or mucous membranes

4. Introduction Sterilization: -Destruction or removal of all forms of microbial life, including endospores. Bactericidal: - Chemical agents capable of killing bacteria. Bacteriostatic: -Chemical agents that inhibit the growth of bacteria but do not necessarily kill them.

5. Introduction Sepsis: -bacterial contamination Asepsis: -absence of significant contamination Aseptic technique: -minimizes contamination

6. Methods of Microbial Growth Control Physical Methods Chemical Methods 1- Heat 2- Filtration 3- Low Temperature 4- Desiccation 5- Osmotic Pressure 6- Radiation -Chemical Agents used on living & nonliving surfaces -Few chemical agents achieve sterility Example of Chemicals: -Phenol -Alcohol -Chlorine -Iodine -Hydrogen Peroxide -Formaldehyde

7. Factors that affect Antimicrobial Treatment 1- The Number of Microbes The more microbes there are to begin with , the longer it takes to eliminate the entire population 2- Environmental Influences The presence of organic matter often inhibits the action of chemical antimicrobials. 3-Time of Exposure Chemical antimicrobials often require extended exposure for more resistant microbes or endospores to be affected. 4- Microbial Characteristics Microbial characteristicsaffectchemical and physicalcontrolmethods.

8. Assessment of Disinfectant 1- Phenol Coefficient 2- Dilution Test Methods 3- Disk - Diffusion Methods

9. Phenol coefficient: Phenol-coefficient: *Is a measure of the bactericidal activity of a chemical compound in relation to phenol. *Phenol-coefficient is stated as the ratio of the greatest dilution of the disinfectant killing the test organism in 10 min and not in 5 min to the highest dilution of phenol killing the test organism in the same period. *Since phenol is employed as standard for comparison  Disinfectants that are more effective than phenol have a coefficient > 1. Those that are less effective have a coefficient < 1.

10. Disk - Diffusion Methods Procedure: • Prepare a bacterial suspension in sterile water ( you may use a broth bacterial culture) • Using a sterile swab, inoculate a nutrient agar plate with the bacterium • A sterile filter paper that is soaked in a disinfectant or an antiseptic is then applied to the inoculated plate. • The disinfectant or the antiseptic will diffuse into the medium. The further the distance from the soaked paper the less the concentration of the disinfectant or the antiseptic in the medium. • Within a certain distance from the soaked filter paper, these chemicals will kill the bacteria so no growth will be seen next day within a circular area around the soaked filter paper. • This circular area of no growth around the soaked filter paper is known as the inhibition zone. • Upon comparing the diameters of the inhibition zones for different disinfectants or antiseptics, the larger the inhibition zone, the more effective the disinfectant or the antiseptic in killing the bacteria.

13. No Disinfectant is substitute for the following Procedures • Hand washing (hand hygiene). • The use of personal protective equipment (e.g. gloves) when handling blood, body substances. • Appropriate handling of patient care equipment and soiled linen. • The prevention of needle stick/sharpinjuries. • Environmental cleaning • Appropriate handling of waste • Taking care of yourself (e.g. immunization)

14. Do not forget to Wash Your Hands • Immediately on arrival at work & Before leaving work • Before and after examining each Person • After touching anything that might be contaminated • After handling specimens • Before putting on gloves for clinical procedures & after removing gloves

15. Good luck