13.2 Using Physical Methods to Control Microorganisms

Heat-Based Methods for Controlling Microbial Growth

Heat is the most widely used physical method for controlling microbes. It works by denaturing proteins and nucleic acids, which destroys cell structures and stops metabolic processes. Different heat methods achieve different levels of microbial control depending on the temperature, pressure, and duration used.

Pasteurization

Pasteurization reduces microbial load in liquids without completely sterilizing them. Two common approaches:

• Batch (LTLT) pasteurization heats liquid to 63°C for 30 minutes
• Flash (HTST) pasteurization heats liquid to 72°C for 15 seconds

Pasteurization kills most vegetative bacteria (like Salmonella), many viruses, and some fungi, but it does not destroy endospores. It's the standard treatment for milk, juice, beer, and other beverages.

Autoclaving

The autoclave is the gold standard for sterilization. It uses pressurized steam at 121°C and 15 psi for 15–20 minutes, which is hot enough to kill all microorganisms, including bacterial endospores like those of Clostridium. Autoclaves are used to sterilize laboratory media, surgical instruments, and medical devices.

The key here is that pressure raises the boiling point of water, allowing steam to reach temperatures above 100°C. Without that pressure, you can't achieve true sterilization with moist heat.

Boiling

Boiling (100°C at sea level) kills most vegetative bacteria, such as E. coli, and inactivates many viruses. However, boiling does not kill endospores or certain heat-resistant viruses. That makes it useful for disinfection (e.g., making drinking water safer) but not for sterilization.

Key Heat Concepts

• Thermal death time (TDT): The minimum time required to kill all microorganisms in a sample at a given temperature.
• Decimal reduction time (D-value): The time required to kill 90% of the microorganisms at a specific temperature. This is useful for calculating how long a heat treatment needs to last to reach a target level of microbial reduction.

Both values depend on the specific organism and the temperature used. More heat-resistant organisms have longer TDT and D-values.

Physical Methods for Inhibiting Microbial Growth

These methods don't necessarily kill microbes. Instead, they slow or stop microbial growth, often by limiting water availability or reducing metabolic activity.

Desiccation

Desiccation removes moisture from materials. Without available water, most microbes can't grow or reproduce. This is why dried foods like beef jerky, raisins, and rice have long shelf lives. Keep in mind that desiccation is bacteriostatic, not bactericidal. Many microbes survive in a dormant state and can resume growth once moisture returns.

Freezing

Lowering the temperature below 0°C slows metabolic processes dramatically and reduces available liquid water. Freezing is commonly used to preserve foods like vegetables, meats, and prepared meals. Like desiccation, freezing generally inhibits growth rather than killing all microbes. Some organisms survive freezing and can become active again upon thawing.

High-Pressure Processing (HPP)

HPP applies extreme pressures (100–1000 MPa) to food products. This disrupts cell membranes and denatures proteins, inactivating vegetative microbes without using heat. It's used commercially for products like guacamole, salsa, and oysters where heat would change the texture or flavor. HPP does not reliably destroy endospores at lower pressures.

Lyophilization (Freeze-Drying)

Lyophilization freezes a sample and then removes water by sublimation (converting ice directly to vapor under vacuum). This preserves microorganisms and biological materials for long-term storage. It's widely used in research labs to maintain reference cultures and in the pharmaceutical industry to stabilize vaccines and other biologics.

Radiation and Filtration for Sterilization and Disinfection

When heat isn't an option, radiation and filtration provide alternative ways to eliminate or remove microbes.

Radiation

Radiation controls microbes by damaging DNA and other cellular molecules. The two main categories differ in energy level and penetrating power:

• Ionizing radiation (gamma rays, X-rays) has enough energy to knock electrons from atoms, creating reactive ions that break DNA strands and destroy other cellular components. Because it penetrates deeply, ionizing radiation is used to sterilize pre-packaged medical devices (syringes), pharmaceuticals, and some food products.
• Non-ionizing radiation (primarily UV light at ~260 nm) causes thymine dimers in DNA, which block replication. UV light has poor penetrating power, so it only works on exposed surfaces, air, and shallow water. It's commonly used for disinfecting lab benches, hospital rooms, and water treatment systems. UV is considered a disinfectant, not a sterilant, because it can't reach shielded microbes.
• Cold plasma is an ionized gas that generates reactive oxygen and nitrogen species. It can inactivate microorganisms on surfaces and in liquids at low temperatures, making it useful for heat-sensitive materials.

Filtration

Filtration physically removes microbes by passing liquid or air through a material with pores small enough to trap them. No killing is involved; organisms are simply separated out.

• Membrane filters (0.2–0.45 μm pore size) remove most bacteria, protozoa, and fungi from liquids. They're essential for sterilizing heat-sensitive solutions like antibiotics and certain culture media. A 0.2 μm filter is the standard for producing sterile liquids. Note that viruses and some very small bacteria (like Mycoplasma) can pass through these filters.
• HEPA filters (High-Efficiency Particulate Air) capture 99.97% of particles $\geq$ 0.3 μm in diameter. They're used to maintain sterile air in biosafety cabinets, operating rooms, and pharmaceutical clean rooms. HEPA filters trap most bacteria and fungal spores but are less effective against individual virus particles.

Microbial Control Terminology

These terms come up constantly in microbiology, and they have precise meanings:

• Sterilization: Complete elimination of all living microorganisms, including bacterial endospores. Autoclaving and ionizing radiation achieve sterilization.
• Disinfection: Elimination of most pathogens from inanimate surfaces, but not necessarily endospores or all microbial forms. Chemical disinfectants and UV light are common disinfection tools.
• Antisepsis: Reduction of microbes on living tissue (like skin). Antiseptics are milder than disinfectants because they need to be safe for human cells.
• Bacteriostatic vs. bactericidal: Bacteriostatic methods inhibit growth without killing (like refrigeration), while bactericidal methods actively kill bacteria (like autoclaving). This distinction matters when choosing a control strategy.