Chapter 7: The Control of Microbial Growth

Introduction

The control of microbial growth is essential in healthcare, food safety, and laboratory settings. This chapter explores the terminology, mechanisms, and methods used to control or eliminate microorganisms, including both physical and chemical approaches.

Terminology of Microbial Control

Key Terms and Definitions

• Sepsis: Refers to bacterial contamination.

• Asepsis: Absence of significant contamination; aseptic techniques prevent microbial contamination of wounds.

• Sterilization: Removal and destruction of all microbial life.

• Commercial Sterilization: Killing Clostridium botulinum endospores in canned goods.

• Disinfection: Destruction of harmful microorganisms on inanimate surfaces.

• Antisepsis: Destruction of harmful microorganisms from living tissue.

• Degerming: Mechanical removal of microbes from a limited area (e.g., skin before injection).

• Sanitization: Lowering microbial counts on eating utensils to safe levels.

• Biocide (Germicide): Treatments that kill microbes.

• Bacteriostasis: Inhibiting, not killing, microbes.

The Rate of Microbial Death

Patterns and Factors Affecting Microbial Death

Microbial death occurs at a logarithmic rate when exposed to control agents. The effectiveness of treatment depends on:

• Number of microbes present

• Environmental factors (organic matter, temperature, biofilms)

• Time of exposure

• Microbial characteristics (e.g., endospores, cell wall structure)

It takes longer to sterilize a larger population of microbes than a smaller one, even if the rate of killing is the same.

Actions of Microbial Control Agents

Effects on Cellular Structures

• Damage to Plasma Membrane: Causes leakage of cellular contents and interferes with cell growth.

• Damage to Proteins (Enzymes): Denaturation or inactivation of essential enzymes.

• Damage to Nucleic Acids: Prevents replication and normal cellular function.

Physical Methods of Microbial Control

Heat

• Heat Denatures Enzymes: Leading to cell death.

• Thermal Death Point (TDP): Lowest temperature at which all cells in a liquid culture are killed in 10 minutes.

• Thermal Death Time (TDT): Minimal time for all bacteria in a liquid culture to be killed at a given temperature.

• Decimal Reduction Time (DRT): Minutes to kill 90% of a specific population of bacteria at a given temperature.

Moist Heat Sterilization

• Coagulates/denatures proteins.

• Methods: Boiling, free-flowing steam, autoclaving.

• Autoclave: Steam under pressure (121°C at 15 psi for 15 minutes) kills all organisms (except prions) and endospores. Steam must contact the item’s surface.

• Large containers require longer sterilization times; test strips indicate sterility.

Pasteurization

• Reduces spoilage organisms and pathogens in milk and juices.

• High-temperature short-time (HTST): 72°C for 15 seconds.

• Thermoduric organisms survive but are unlikely to cause disease or spoil refrigerated milk.

Ultra-High-Temperature (UHT) Treatments

• Sterilizes milk, creamer, and juice for storage without refrigeration (140°C for 4 seconds, followed by rapid cooling).

Dry Heat Sterilization

• Kills by oxidation (flaming, incineration, hot-air sterilization at 170°C for 2 hours).

Filtration

• Passage of substance through a screenlike material; used for heat-sensitive materials.

• HEPA filters remove microbes >0.3 μm; membrane filters remove microbes >0.22 μm (some as small as 0.05 μm).

Other Physical Methods

• Low Temperature: Bacteriostatic effect (refrigeration, deep-freezing, lyophilization).

• High Pressure: Denatures proteins, alters carbohydrate structure.

• Desiccation: Absence of water prevents metabolism.

• Osmotic Pressure: High concentrations of salts and sugars create a hypertonic environment, causing plasmolysis.

Radiation

• Ionizing Radiation: (X-rays, gamma rays, electron beams) ionizes water to create reactive hydroxyl radicals, damaging DNA by causing lethal mutations. Used for sterilizing food, pharmaceuticals, and medical supplies.

• Nonionizing Radiation: (Ultraviolet, 260 nm) damages DNA by creating thymine dimers; effective for surfaces but does not penetrate deeply.

• Visible Blue Light (470 nm): Kills a wide range of bacteria by forming singlet oxygen.

• Microwaves: Kill by heat, not especially antimicrobial.

Chemical Methods of Microbial Control

Principles of Effective Disinfection

• Concentration of disinfectant

• Presence of organic matter

• pH

• Temperature

• Time of exposure

Evaluating Disinfectants

Use-Dilution Test

• Standard method for evaluating disinfectants.

• Metal cylinders dipped in test bacteria, dried, exposed to disinfectant, then transferred to culture media to check for survival.

Disk-Diffusion Method

• Filter paper disks soaked in chemical agents are placed on a culture; zones of inhibition indicate efficacy.

Types of Chemical Disinfectants

• Phenol and Phenolics: Injure plasma membranes; remain active in presence of organic matter. Example: O-phenylphenol (Lysol®).

• Bisphenols: Disrupt plasma membranes. Examples: Hexachlorophene (skin infections), Triclosan (antimicrobial soaps, discontinued except in toothpaste).

• Essential Oils: Plant extracts (e.g., tea tree oil, pine oil) with broad-spectrum activity, especially against gram-positive bacteria.

• Halogens: Iodine (impairs protein synthesis, alters membranes; used as tincture or iodophor), Chlorine (oxidizing agent; used in bleach and water disinfection).

• Alcohols: Denature proteins and dissolve lipids; ineffective against endospores and nonenveloped viruses. Ethanol and isopropanol require water for activity.

• Heavy Metals: Oligodynamic action; denature proteins (e.g., silver nitrate for newborn eyes).

• Surfactants (Quats): Surface-active agents; disrupt membranes.

• Chemical Food Preservatives: Sulfur dioxide, organic acids, nitrites/nitrates prevent spoilage and endospore germination.

• Peroxygens: Oxidizing agents (hydrogen peroxide, peracetic acid) used for sterilizing surfaces and equipment.

Microbial Characteristics and Microbial Control

Microbial Resistance to Control Methods

• Gram-negative bacteria: More resistant to biocides due to outer membrane lipopolysaccharide.

• Mycobacteria: Resistant due to waxy cell wall; require special disinfectants.

• Bacterial endospores: Highly resistant to many biocides.

• Nonenveloped viruses: More resistant than enveloped viruses.

• Prions: Extremely resistant; require harsh sterilization (NaOH and autoclaving).

Effectiveness of Chemical Antimicrobials Against Endospores and Mycobacteria

Summary Table: Physical and Chemical Methods of Microbial Control

Additional info: These notes synthesize and expand upon the provided lecture slides and textbook content, ensuring a comprehensive, exam-ready summary of microbial control methods.