Bacterial Growth and Reproduction

• Bacteria reproduce asexually through binary fission
• The average bacterial division time is 1-3 hours

Phases of Bacterial Growth

• Lag phase: Bacteria adapt to the medium with no major population change
• Log phase: Bacteria exhibit exponential, logarithmic growth and are most vulnerable
• Stationary phase: The bacterial population stabilizes, showing little or no net growth
• Death phase: Nutrients deplete, leading to a decline in the bacterial population

Optimal Bacterial Growth Conditions

• Bacteria have minimum, maximum, and optimum temperatures for growth
  • Psychrophiles thrive in cold environments
  • Thermophiles thrive in extremely hot temperatures
• Most bacteria favor a pH between 6.5 and 7.5
  • Molds and yeasts prefer slightly more acidic conditions
• Optimal osmotic pressure, or isotonic conditions, is vital for bacterial survival
  • High concentrations of salt or sugar prevent growth through hypertonicity

Controlling Nosocomial Infections

• The field of controlling hospital-acquired infections is relatively new with advances in the last century
• Ignatz Semmelweis was a pioneer, noting high post-childbirth sepsis mortality
  • Semmelweis mandated hand washing with chlorinated lime
• Joseph Lister recognized high post-operative mortality
  • Lister treated surgical sites with phenol (carbolic acid)

Aseptic Surgical Technique

• Developed techniques aim to prevent surgical site contamination

Terminology for Microbial Control

• Sterile: All forms of microbial life, including bacterial endospores, have been destroyed on equipment and surfaces
• Sterilization: An absolute state, it is either sterile or it is not with no degrees of the condition
• Most common method to sterilize involves pressurized steam
• Other methods include ethylene oxide gas or liquid glutaraldehyde

Destruction of Pathogens

• Disinfect: Only the vegetative forms of pathogens are destroyed, not endospores, this process removes infectious particles
  • Refers to treating inanimate surfaces with chemicals, not sterilization
• Antisepsis involves chemical disinfection of living tissue
• Sanitize reduces microbial numbers to safe levels, mainly with kitchen utensils
  • Uses high temperature washing or chemical disinfectant dips

Methods of Killing Microbes

• "Cidal" agents outright kill microbes
  • Germicides are chemical agents with classifications such as: Bactericides, Sporicides, Fungicides, Virucides, and Amoebicides
• "Static" agents halt growth
  • Bacteriostatic agents stop multiplication but do not kill, putting bacteria in suspended animation

Further Definitions for Microbial Control

• When static agents are removed, organisms resume growth
• Fungistatic: Halts the growth of fungi
• Asepsis: No significant microbial contamination on an object or area
• Aseptic technique: Prevents contamination through steps during surgery, IVC use, catheterization, etc

Influencing Microbial Control - Temperature

• Low temperatures are bacteriostatic, not bactericidal, and slow or stop growth via refrigeration or freezing
• High temperatures are bacteriocidal, making disinfectants more effective when warmed
• Safe storage temperatures are either less than 40° F or greater than 140° F
• The danger zone for temperatures is between 40 F and 140F

Influence Factors - Types of Microbes

• Microbes exhibit natural resistance to chemicals
• Gram-negative bacteria are generally more resistant due to the protective lipid layer
  • Pseudomonas
    • Common environmental organism
    • Naturally resistant to many chemicals
    • Can thrive in some disinfecting solutions, like quaternary ammonium compounds ("quats")

Influence Factors - Physiological State of Microbes

• Enveloped viruses are easier to kill than non-enveloped viruses
• Actively dividing organisms are the most fragile
• Vegetative stages are next most susceptible
• Endospores are the most resistant form
  • Clostridium botulinum spores can survive boiling
  • Anthrax spores can remain viable for decades
• Protozoal cysts are highly resistant
  • Toxoplasmosis and Giardia are difficult to eliminate

Influence Factors - Environment

• Organic matter (debris) protects from disinfectants, inactivating some
  • Always clean instruments before sterilization: wash then disinfect
  • Washing alone with soap removes 99% of bacteria

Rate of Microbial Death

• Microorganisms die at a constant rate when using disinfectants correctly
• The initial number of microbes determines the contact time needed for effective disinfection
• Physical scrubbing to remove as many organisms as possible is crucial

Physical Methods of Microbial Growth Control - Osmotic Pressure

• Early civilizations used desiccation and osmotic pressure (drying and salting)
• High concentrations of salt or sugar draw water from bacterial cells, inhibiting growth (osmosis)
  • Curing meats and making jellies/jams create desiccation
• Molds/yeasts are more capable of growing in low-moisture, high-osmotic pressure environments
  • Fruits and grains typically spoil with mold first

Physical Methods - Desiccation

• Water is essential for bacterial growth/reproduction, but bacteria can survive without water
  • Survive in a dormant state and resume growth once rehydrated
• Tuberculosis can withstand drying for months and endospores survive for years
• Some viruses are relatively resistant to desiccation
  • Drying foods combines with changes in oncotic pressure to preserve them

Physical Methods - Heat

• The most common and economical technique
• Destroys by denaturing or coagulating proteins (bacterial enzymes)
• Dry heat sterilization: Organisms killed by oxidation
  • Flaming loops and incinerating biowaste are examples

Physical Methods - Moist Heat

• Coagulation of vital bacterial proteins occurs more quickly with water present
• Boiling at 100°C (212°F) at sea level kills vegetative bacterial cells, viruses, fungi, and most protozoa but not bacterial endospores
• Hepatitis virus can withstand 30 minutes of boiling, some endospores are viable after 20 hours
• Boiling sanitizes, decreasing microbial numbers, but will not sterilize
• Sterilization requires steam under pressure within an autoclave.
  • Pressurizing liquids makes them hotter than boiling
  • Standard autoclave conditions (time, temperature, pressure) eradicate all organisms and spores: 15 psi, 121 °C (250 °F), 15 minutes

Important Autoclave Considerations

• Steam must reach all surfaces by tipping bowls in surgical packs and using only one click to close instruments
• Ensure not to pack autoclaves too tightly
• Tip surgical pans and bowls at 45-degree angles
• Ensure to use perforated pans
• Wrap pack tightly in cloth or paper
• Always include a sterility indicator to know that autoclave is working

Methods of Sterilization

• High-speed/pressure sterilization with a flash autoclave uses 30 psi at 134 °C (273 °F) for 3-5 minutes
  • Used if an instrument is contaminated but dropped

Further Methods of Sterilization

• Some control techniques that work well are not usable sometimes
• Heat sterilizing delicate medical instruments like endoscopes, suture material, or surgical gloves can damage them.
• Pasteurization: Uses mild heating to preserve taste
• Destroys pathogens and decreases the number of other bacteria
• Extends storage time
• Kills pathogens, but non-pathogens are not removed: Milk will eventually sour

Pasteurization Methods

• Classic: Heating to 63°C (145°F) for 30 minutes
• High Temperature, Short Time (HTST): Heating to 72°C (162°F) for 15 seconds is standard today and to sterilizes milk and food products is 140° C (284° F) for 1 second
• Ultra-High Temperature (UHT) sterilization: 140º C (284º F) for 1 second
  • Can be stored without refrigeration.

Further Sterilization/Filtration Methods

• Filtration with High-Efficiency Particulate Air (HEPA) filters removes particles from air in burn units, using vacuum cleaners, etc
• These filters do not kill anything, but just filters it out
• Membrane filters with different pore sizes remove bacteria from medical and backcountry water
• Refrigeration from 0 C to 7C slows bacteria
  • Bacteriostatic to most bacteria with pathogens (except Listeria) unable to reproduce
• Freezing below 0° C - Though some survive, slow freezing is more effective
• Lyophilization is freeze drying, and removes water from a frozen sample allowing preservation of structure/nutrients

Sterilization and Disinfectant types: Radiation

• Ionizing: Destroys microbial DNA through industrial applications with gamma rays and x-rays
• Nonionizing (Including Ultraviolet Light): Damages surface level DNA of exposed cells (steripens) not able to kill all cells The UV light is able to be utilized in germicidal lamps hospitals, water treatments
• Microwaves do not kill bacteria but heat the water to produce steam

Chemical Methods to Control Microbial Growth

• Agents function on living and inanimate objects as bacteriostatic instead of sterilizing to damage hospital equipment

Action on Microbial Controls

• Alteration of Membrane: Plasma of cells damaged by alcohol and chlorhexidine, weaken cell and cause leakage
• Damage of Protein/ Nucleic Acids: Halogens and Peroxides break down protein bonds, denaturing proteins and cause inability to reproduce
• Surfactants: Agents that have poor cleaning properties, but remove debris and bacteria via floating away from the surface

Disinfectant Principles

• Disrupts the phospholipid layer of cell membrane
• It is important to check the label for concentration and contact in order to cause most damage, or it may lead to:
  • Too weak properties, which means they will be ineffective
  • A tool strong can injure other tissues
• Materials must be in physical contact to work in disinfectant
• Instrument must be cleaned, in order to be sanitized
• Tools containing surfactants may need surfactant to work
• Increase Temp with use of warm/hot water

Disinfectants Types: Phenols

• Phenols (Carbolic Acid): Cidal bacteria that removes Lister, but may causes irritation
• Phenolics: Has less irritant with organic , stable for long period with 2 types
  • Cresol: Derived from coal tar
  • Lysol: Has toxic and is a effective disinfectant
  • Hexachlorophene: Is a surgical scrub, harmful in newborns with absorbed through the scan in damages neurological system
• Triclosan: Common antimicrobial FDA ban

Disinfectants Types: Halogens and Chlorhexidrines

• Chlorhexidine:
  • Binds to skin and mucous membranes and has great effects for ~48 hrs for various uses in detergent, disinfect inaminate objects, etc
• Halogens- Iodines:
  • Has high range effects with 2 Forms of iodines
  • Iodophor is know an excellent killer
  • Has forms of chlorine to use in a variety of uses