Control of Microbial Growth

Control of Microbial Growth

• Control is achieved either by killing microorganisms or inhibiting their growth.
• Typically involves using physical or chemical agents to either kill or prevent the growth of microorganisms.
• Cidal agents kill cells, while static agents inhibit growth without killing cells.

Sterilization and Disinfection

• Sterilization involves removing or killing all living microorganisms, including bacteria and their spores.
• Disinfection involves removing or killing disease-causing microorganisms but not necessarily all microorganisms.
• Sterility is the absence of living organisms.
• Sterilization is the process of achieving sterility.

Official Sterilization Methods

• Moist heat
• Dry heat
• Filtration
• Gaseous ethylene oxide
• Ionizing radiation

Aseptic Techniques

• Necessary to prevent contamination of sterile surfaces.
• Involves a combination of protocols that collectively maintain sterility.

Resistance Levels of Microorganisms

• Prions (Creutzfeldt-Jakob Disease) are the most resistant, requiring prion reprocessing.
• Bacterial spores (Bacillus atrophaeus) require sterilization.
• Coccidia (Cryptosporidium) require disinfection.
• Mycobacteria (M. tuberculosis, M. terrae) have high resistance.
• Nonlipid or small viruses (polio, coxsackie) have intermediate resistance.
• Fungi (Aspergillus, Candida) have intermediate resistance.
• Vegetative bacteria (S. aureus, P. aeruginosa) have low resistance.
• Lipid or medium-sized viruses (HIV, herpes, hepatitis B) are the most susceptible.

Factors Influencing Antimicrobial Treatment

• The number of microbes present affects the time it takes to eliminate the population.
• Endospores are very difficult to destroy.
• The susceptibility of vegetative pathogens can depend on factors such as their environment and genetic factors, leading to significant variations in resistance to antimicrobial agents.
• Other factors include population size and composition, concentration or intensity of antimicrobial agent, duration of exposure, temperature, and local environment.

Physical Methods of Microbial Control

• Heat denatures enzymes and other proteins. Heat resistance varies among microbes.
• Moist heat kills microorganisms by coagulating their proteins
• Moist heat is generally more effective than dry heat.

Methods of Sterilization

Heat Sterilization

Dry Heat: Red heat, flaming, incineration, hot air oven

Moist Heat: Pasteurization, boiling, autoclaving

Heat to 100°C during Boiling

• Kills vegetative forms of bacterial pathogens, almost all viruses, and fungi and their spores within 10 minutes or less.
• Endospores and some viruses are not destroyed quickly. Brief boiling will kill many vegetative cells are actively growing and metabolizing bacteria that are sensitive to harsh environmental conditions and antimicrobial agents. Unlike endospores, which can resist extreme heat and desiccation, vegetative cells can be easily destroyed by methods such as heating, disinfectants, and antiseptics. These cells represent the reproductive and active form of bacteria, playing a crucial role in infection and disease transmission. They can also rapidly multiply under favorable conditions, leading to an increase in microbial population.

Virus Facts

• Hepatitis virus can survive up to 30 minutes of boiling.
• Endospores can survive up to 20 hours or more of boiling.

Autoclave

• Requires temperatures above the boiling point of water for reliable sterilization with moist heat.
• Filled with hot steam under pressure.
• Preferred method of sterilization, unless the material is damaged by heat or moisture.
• Steam reaches 121°C at twice atmospheric pressure.
• All organisms and endospores are killed within 15 minutes.
• Prions are resistant to sterilization; prolonged high temperatures are needed for inactivation. 138 degrees celsius for 18 min

Items That Can Be Autoclaved

• Cultures and stocks of infectious material
• Culture dishes and related devices
• Discarded live and attenuated vaccines
• Contaminated solid items like petri dishes, eppendorf tips, pipettes, gloves, and paper towels

Items That Cannot Be Autoclaved

• Chemical, chemotherapeutic, or radioactive waste
• Certain kinds of plastics
• Items containing corrosives (acids, bases, phenols)

Dry Heat Sterilization

• Kills by oxidation
  • Flaming
  • Incineration: Waste destruction in a furnace by controlled burning at high temperatures
  • Hot-air sterilization

Incineration

• Disposal and sterilization of contaminated materials achieved at 870-980°C.

Hot Air Oven

• Protein denaturation and oxidation damage to cytoplasm occur.
  • Holding Times
    • 160°C for 2 hours
    • 170°C for 1 hour
    • 180°C for 30 minutes

Materials Sterilized by Hot Air Oven

• Glassware
• Powders like dusting paraffin
• Fats
• Grease
• Glassware should not be opened until 2 hours after cooling.

Hot Air Sterilization Overview

• Dry heat transfers heat less effectively than moist heat.
• 160°C for 2 hours is equivalent to 121°C for 15 minutes in an autoclave.

Other Physical Methods

• Filtration removes microbes.
• Low temperature inhibits microbial growth.
• High pressure denatures proteins.
• Desiccation prevents metabolism.
• Osmotic pressure causes plasmolysis.
• Radiation damages DNA.

Filtration

• Removes microbes by passing a liquid or gas through a screen-like material with small pores.
• Used to sterilize heat-sensitive materials like vaccines, enzymes, antibiotics, and some culture media.

Membrane Filters

• Uniform pore size, used in the industry and research.
• 0.22 and 0.45 μm pores are used to filter most bacteria but do not retain spirochetes, mycoplasmas, and viruses.
• 0.01 μm pores retain all viruses and some large proteins.

HEPA Filters

• High Efficiency Particulate Air Filters
• Used in operating rooms and burn units to remove bacteria from the air and in biosafety cabinets.
• Removes 99.97% of particles with a size of 0.3 μm from the air

Low Temperature

• Effect depends on the microbe and treatment applied.
• Refrigeration: Temperatures from 0 to 7°C provide a Bacteriostatic effect.
• Reduces the metabolic rate of most microbes so they cannot reproduce or produce toxins.

Freezing

• Temperatures below 0°C.
• Flash freezing does not kill most microbes.
• Slow freezing is more harmful because ice crystals disrupt the cell structure.
• Over 1/3 of vegetative bacteria may survive for 1 year.

Dessication

• In the absence of water, microbes cannot grow or reproduce, but some may remain viable for years.
• After water becomes available, they start growing again.

Osmotic Pressure

• High concentrations of salts and sugars in foods increase osmotic pressure, creating a hypertonic environment.
• Plasmolysis occurs; as water leaves the cell, the plasma membrane shrinks away from the cell wall.
• Cells may not die, but usually stop growing.

Radiation

• Three types of radiation kill microbes: ionizing, nonionizing, and microwave radiation.

Ionizing Radiation

• Includes gamma rays, X-rays, electron beams, or higher energy rays with short wavelengths (< 1 nm).
• Dislodges electrons from atoms and forms ions, causing mutations in DNA and producing peroxides.
• Used to sterilize pharmaceuticals and disposable medical supplies; the food industry is interested in its use.
• Disadvantages include penetration of human tissues and potential for genetic mutations.

Non-Ionizing Radiation

• Ultraviolet light has a longer wavelength than 1 nanometer.
• Damages DNA by producing thymine dimers, which cause mutations.
• Used to disinfect operating rooms, nurseries, and cafeterias.
• Disadvantages include damage to skin and eyes and failure to penetrate paper.

Microwave Radiation

• Wavelength ranges from 1 mm to 1 meter.
• Heat is absorbed by water molecules.
• May kill vegetative cells in moist foods.
• Bacterial endospores contain no water and are not damaged.
• Solid foods are unevenly penetrated.

Rate of Microbial Death

• Dependent on:
  • number of microbes present
  • type of microbes
  • environmental influences
  • Exposure duration