Chapter 7
Control of Microbial Growth
• Boiling kills microbes by coagulating the proteins – endospores are not destroyed.
• Autoclaving - Temperature 121oC, 15 PSI, 15 min – even the endospores are destroyed.
• Dry heat – direct flaming is used to sterilize the inoculating loop.
• Refrigeration – slows down the growth of microbes. It does not kill the microbes.
• Osmotic environment – high concentration of salt – plasmolysis.
• UV light – mutation – inducing the formation of thymine dimers.
• Iodine - inactivates enzymes – skin infection, wound infection.
• 1% silver nitrate – ophthalmea neonatorum – eye infection of the newborn.
• Gas sterilization – ethylene oxide – mattresses, petri plates.
• Quaternary ammonium compounds – damage to the plasma membrane – mouthwash.
• Sodium nitrite – preserve meat products – prevents the germination of botulism endospores – used to preserve hot dogs.
• Sepsis refers to microbial contamination
• Asepsis is the absence of significant contamination
• Aseptic surgery techniques prevent microbial contamination of wounds
The Terminology of Microbial Control
The Terminology of Microbial Control
• Sterilization: Removing all microbial life
• Commercial sterilization: Killing C. botulinum endospores
• Disinfection: Removing pathogens
• Antisepsis: Removing pathogens from living tissue
The Terminology of Microbial Control
• Degerming: Removing microbes from a limited area
• Sanitization: Lowering microbial counts on eating utensils
• Biocide/germicide: Kills microbes
• Bacteriostasis: Inhibiting, not killing, microbes
• The usual definition of sterilization is the removal or destruction of all forms of microbial life;
Effectiveness of Treatment
• Depends on– Number of microbes– Environment (organic matter, temperature,
biofilms)– Time of exposure– Microbial characteristics
Actions of Microbial Control Agents
• Alteration of membrane permeability
• Damage to proteins
• Damage to nucleic acids
Moist Heat Sterilization
• Moist heat denatures proteins
• Autoclave: Steam under pressure
Figure 7.2
An Autoclave
Pasteurization
• Reduces spoilage organisms and pathogens
• Equivalent treatments– 63°C for 30 min
Hot-Air Autoclave
Equivalent Treatments 170˚C, 2 hr 121˚C, 15 min
Dry Heat Sterilization
• Kills by oxidation– Dry heat– Flaming– Incineration– Hot-air sterilization
Figure 7.4
Filtration
• Membrane filtration removes microbes >0.22 µm
Physical Methods of Microbial Control
• Low temperature inhibits microbial growth– Refrigeration– Deep-freezing– Lyophilization
• High pressure denatures proteins
• Desiccation prevents metabolism
• Osmotic pressure causes plasmolysis
Figure 7.5
Radiation
Principles of Effective Disinfection
• Concentration of disinfectant
• Organic matter
• pH
• Time
Figure 7.7c, d
Bisphenols
• Hexacholorphene, triclosan– Disrupt plasma membranes
Biguanides
• Chlorhexidine– Disrupt plasma membranes
Halogens
• Iodine– Tinctures: In aqueous alcohol– Iodophors: In organic molecules– Alter protein synthesis and membranes
• Chlorine– Bleach: Hypochlorous acid (HOCl)– Chloramine: Chlorine + ammonia– Oxidizing agents
Heavy Metals
• Ag, Hg, and Cu– Silver nitrate may be used to prevent
gonorrheal ophthalmia neonatorum– Silver sulfadiazine used as a topical cream on
burns– Copper sulfate is an algicide
• Oligodynamic action– Denature proteins
Chemical Food Preservatives
• Organic acids– Inhibit metabolism– Sorbic acid, benzoic acid, and calcium
propionate– Control molds and bacteria in foods and
cosmetics
• Nitrite prevents endospore germination
• Antibiotics– Nisin prevent spoilage of cheese
Gaseous Sterilants
• Denature proteins
• Use: Heat-sensitive material– Ethylene oxide
Chapter 18
Practical Applications of Immunology
History of Vaccines
• Variolation: Inoculation of smallpox into skin
(18th century)
• Vaccination: – Inoculation of cowpox virus into skin (Jenner) -
1790
Types of Vaccines
• Attenuated whole-agent vaccines– MMR
• Inactivated whole-agent vaccines Salk polio
• Toxoids– Tetanus
Types of Vaccines
• Subunit vaccines– Acellular pertussis– Recombinant hepatitis B
• Nucleic acid (DNA) vaccines– West Nile (for horses)– DNA injected into skin cells, protein made –
stimulates immune system.
The Development of New Vaccines
• Culture pathogen
• rDNA techniques– In plants
• Adjuvants
• Deliver in combination
Diagnostic Immunology
• Sensitivity: Probability that the test is reactive if the specimen is a true positive
• Specificity: Probability that a positive test will not be reactive if a specimen is a true negative
Monoclonal Antibodies (Mabs)
• Hybridoma: “Immortal” cancerous B cell fused with an antibody-producing normal B cell, produces
• Monoclonal antibodies
Monoclonal Antibodies
Figure 18.2
Monoclonal Antibodies
Figure 18.2
Monoclonal Antibodies
Figure 18.2
Monoclonal Antibodies (Mabs)
• Infliximab: For Crohn’s disease, blocks cytokine
• Ibritumomab + rituximab: For non-Hodgkin’s lymphoma, destroy cancer cells
• Trastuzumab: Herceptin for breast cancer
• Spread of cancer
Figure 18.5
Agglutination Reactions
• Particulate antigens and antibodies
• Antigens on a cell (direct agglutination)
Figure 18.7
Indirect Agglutination
Complement Fixation Test
Figure 18.10
Complement Fixation Test
Figure 18.10
Figure 18.11a
Fluorescent-Antibody (FA) Techniques
• Direct FA
Figure 18.11b
Indirect Fluorescent-Antibody Test
Enzyme-Linked Immunosorbent Assay
• Also called ELISA
• Enzyme linked to Ab is the indicator
Direct ELISA
Figure 18.14a
Indirect ELISA
Figure 18.14b
Serological Tests
Figure 18.13
Serological Tests
• Direct tests detect antigens (from patient sample)
• Indirect tests detect antibodies (in patient’s serum)
ELISA
• Enzyme liked immunosorbent assay
• Plastic plates with wells in them are used
• Direct ELISA – presence of antigen in a sample
• Indirect ELISA – presence of antibodies against a specific antigen in a sample.
Neisseriagonorrhoeae
Antibodies against HIV
Secondary antibodies
Against human Antibodies.
Enzyme linked antibodies
Substrate – colorlessProduct – blue