Physical and Chemical Controls of Microorganisms

Overview and Learning Objectives

This chapter explores the principles and practices of controlling microbial growth in the environment, focusing on both physical and chemical methods. Students will learn key terminology, mechanisms of action, and the effectiveness of various control strategies.

• Aseptic procedures: Techniques to prevent contamination by microorganisms.

• Microbial control: Methods to reduce or eliminate microorganisms from surfaces, environments, and materials.

Terminology of Microbial Control

Key Terms and Definitions

Understanding the terminology is essential for discussing microbial control methods. The following table summarizes important terms:

Primary Targets and Resistance of Microbes

Microbial Resistance Hierarchy

Microbial control methods must account for varying resistance among different types of microorganisms. The following list ranks microbes from most resistant to most susceptible:

• Prions (most resistant)

• Bacterial endospores

• Mycobacteria

• Cysts of protozoa

• Active-stage protozoa (trophozoites)

• Most Gram-negative bacteria

• Fungi

• Nonenveloped viruses

• Most Gram-positive bacteria

• Enveloped viruses (most susceptible)

Factors Affecting Microbial Control

Effectiveness of Antimicrobial Treatments

The effectiveness of microbial control depends on several factors:

• Number of microbes: Higher populations require longer treatment times.

• Environment: Presence of organic matter, temperature, concentration, and biofilms can affect treatment.

• Time of exposure: Sufficient exposure is necessary for effective killing.

• Microbial characteristics: Structural features (e.g., cell wall, glycocalyx) and resistance mechanisms influence susceptibility.

Modes of Action of Antimicrobial Agents

How Agents Work

Antimicrobial agents target key cellular structures and functions:

• Cell wall: Blocking synthesis, digesting, or breaking down the wall makes cells fragile and prone to lysis.

• Cell membrane: Disruption leads to loss of integrity and cell death.

• Protein and nucleic acid synthesis: Binding to ribosomes or DNA prevents transcription and translation; mutagenic agents cause irreversible damage.

• Protein denaturation: Changes in shape or blocking active sites render enzymes inactive.

Physical Methods of Microbial Control

Heat Methods

Heat is a widely used method for microbial control, with moist and dry heat applications:

• Dry heat: Includes flaming, incineration, and hot-air sterilization. Kills by oxidation.

• Moist heat: Includes boiling, pasteurization, and steam under pressure (autoclaving). More effective than dry heat for most microbes.

• Autoclave: Uses steam under pressure (15 psi, 121°C) for sterilization.

Pasteurization

Pasteurization is used to disinfect beverages by applying heat to kill pathogens and spoilage organisms while preserving flavor and nutritional value. Methods include:

• Flash method: 71.6°C for 15 seconds

• Batch method: 63–66°C for 30 minutes

• Ultra-high-temperature: 140°C for less than 1 second

Radiation Methods

Radiation is used to kill microbes by damaging their DNA:

• Ionizing radiation: Gamma rays, X-rays, electron beams; cause DNA mutations and are used for sterilizing pharmaceuticals and medical supplies.

• Nonionizing radiation (UV): Causes thymine dimers in DNA, used for disinfecting surfaces and air.

• Microwave radiation: Kills by heating water in cells; less effective for dry materials and endospores.

Filtration Methods

Filtration removes microbes from air and liquids by passing them through filters with defined pore sizes:

• HEPA filters: Remove microbes larger than 0.3 µm from air.

• Membrane filtration: Removes microbes larger than 0.22 µm from liquids.

Other Physical Methods

Additional methods include:

• Low temperature: Refrigeration, deep-freezing, and lyophilization inhibit microbial growth.

• High pressure: Denatures proteins.

• Desiccation: Prevents metabolism by removing water.

• Osmotic pressure: Causes plasmolysis in cells.

Chemical Methods of Microbial Control

Criteria for Choosing Microbicidal Chemicals

Effective chemicals should act rapidly, be stable, have broad-spectrum activity, penetrate surfaces, resist inactivation, and be affordable and safe.

Major Groups of Chemical Agents

The following table summarizes chemical methods of microbial control:

Phenols and Phenolics

Phenol (carbolic acid) and its derivatives are used as disinfectants and antiseptics. They denature proteins and disrupt cell membranes. Examples include Lysol, pHisoHex, and Pine-Sol.

Alcohols

Alcohols such as ethanol and isopropanol are used for disinfecting surfaces and skin. They denature proteins and disrupt membranes. Optimum concentration for ethanol is 70%.

Halogens

Halogens such as iodine and chlorine are effective disinfectants. Iodine is used in tinctures and iodophors (e.g., Betadine), while chlorine is used in water treatment and bleach.

• Iodine: Denatures proteins by breaking disulfide bonds.

• Chlorine: Forms hypochlorous acid in water, a strong oxidizing agent.

Biguanides

Chlorhexidine is a biguanide used for skin antisepsis and disinfection. It targets cell membranes and protein structure, is bactericidal for both Gram-positive and Gram-negative bacteria, and is inactive against spores.

Peroxygens (Oxidizing Agents)

Peroxygens such as ozone and hydrogen peroxide oxidize cellular components and disrupt membranes and proteins. Hydrogen peroxide is used as an antiseptic and disinfectant, and benzoyl peroxide is used in acne medications.

Surfactants (Detergents)

Surfactants reduce surface tension and disrupt cell membranes. Cationic detergents are more effective than anionic detergents. Soaps are weak microbicides but gain value when combined with other agents.

Heavy Metals

Heavy metals such as silver, mercury, and copper denature proteins and are effective in small amounts. Silver nitrate was used to prevent eye infections in infants, and copper sulfate is used to control algae.

Aldehydes

Aldehydes such as formaldehyde and glutaraldehyde inactivate proteins by forming covalent crosslinks. Formalin is used for preserving specimens, and glutaraldehyde is a sterilizing agent for hospital instruments.

Gaseous Sterilants

Gaseous agents such as ethylene oxide denature proteins and are used for sterilizing heat-sensitive materials.

Enzymes

Enzymes such as lysozyme digest peptidoglycan in bacterial cell walls. Prionzyme is used to remove prions from medical instruments.

Dyes as Antimicrobial Agents

Aniline dyes (e.g., crystal violet, malachite green) are active against Gram-positive bacteria and fungi. Acridine dyes are used for wound treatment but have limited applications due to staining and narrow spectrum.

Acids and Alkalis

Extreme pH can destroy or inhibit microbial cells but is limited in application due to corrosive and hazardous nature.

Evaluating Disinfectants

Disk-Diffusion Method

The disk-diffusion method is used to evaluate the effectiveness of disinfectants. Microbial species are tested, and the zone of inhibition is measured to determine efficacy.

Summary and Take Home Points

• Know the terminology associated with microbial control.

• Understand the physical methods of control discussed.

• Know the different targets, pros, and cons of the chemical agent groups discussed.