Microbial Control

Terminology for Microbial Control0 Sterilization- removal or destruction of all forms of

microbial life

0 Commercial sterilization- subjects canned food to only enough heat to destroy the endospores of Clostridium botulinum

0 Disinfection- is the destruction of vegetative pathogens on a surface, usually with chemicals0 Spores and viruses are not necessarily destroyed

0 Antisepsis- is the chemical disinfection of living tissue, such as skin or mucous membranes

Terminology for Microbial Control

0 Asepsis- is the absence of pathogens on an object or area, as in antiseptic surgery

0 Degerming (degermation)- is the removal of transient microbes from the skin by mechanical cleansing or by antiseptic

0 Sanitation- is the reduction of microbial populations on objects to safe public health levels

0 A biocide or germicide- kills microorganisms0 Fungicides kill fungi, virucides kill viruses 0 Suffix – cide means the killer of a specified microorganism0 Suffix- stat used in this way indicates only that the substance

inhibits – for example bacteriostasis

Rate of Microbial Death0 Bacterial populations killed by heat or chemicals tend

to die at constant rates—for example, 90% every 10 minutes. Plotted logarithmically, these figures form straight descending lines.

Factors that influence effectiveness of anantimicrobial treatment:

01. Number of microbes0 -more cells, more time needed to kill all

02. Environmental influences0 -organics often inhibit chemical agents (blood,

feces, vomit)0 -temperature (disinfectants work better in

warm temperatures)0 -pH (heat is more effective in an acid pH)

03. Time of exposure0 -Chemical antimicrobials require a certain

amount of exposure; same agent may need longer on resistant organisms or spores

0 -with heat, lower temps require longer to kill

04. Microbial characteristics0 -resistance genes, protective structures0 (e.g.capsules) etc. can inhibit action0 -bioflims prevent penetration

Factors that influence effectiveness of anantimicrobial treatment:

Type and Age of Microbe

0 Bacteria- susceptible to protein denaturing BUT mycobacteria is not because of its hydrophobic coat

0 Growth cycles- physiologically young bacteria(early in growth cycle) susceptible to heat

0 Endospores more resistant to heat the older they get

Properties of Materials:

0 Plastic and rubber- not heat tolerant

0 Cutting edges of surgical instruments- no moisture or corrosive chemicals

0 Edges can become pitted due to rust and corrosion, microbes can hide there

0 Fabrics- no chemical disinfection; some too fragile

Actions of Microbial Control Agents

0 1. Alteration of membrane permeability0 The plasma membrane controls the passage of nutrients

and wastes into and out of the cell.

0 Damage to the plasma membrane causes:0 leakage of cellular contents 0 interferes with cell growth.

0= Leak lysis, death

Actions of Microbial Control Agents

0 2. Damage to proteins and Nucleic Acids0 Enzymes and other proteins are essential for cell function

0Denatures proteins0Enzymes (no reactions)

0 Proteins necessary for bacteria metabolism0 Hydrogen bonds are broken0 Covalent bonds are also broken

0 3. Damage to nucleic acids0 Prevent replication, transcription, or translation

Physical Methods of Microbial Control

0 -to disinfect objects, food, and solutions common methods:0 -Temperature: kill or inhibit growth

0Heat0Low Temps.

0 -Filtration: physical removal0 -Desiccation: inhibit growth0 -Osmotic pressure0 -Radiation: kill

Methods involving Heat:0 Heat: Common food preservation

0 Denatures protein (changes shape)

0 Thermal Death Point (TDP) = lowest temp at which all microbes in liquid suspension will be killed in 10 min

0 Thermal Death Time (TDT)= minimal length of time for all microbes in liquid suspension to be killed at given temp

0 *Both are different for different species due to microbial variation in heat tolerance

0 Decimal Reduction Time (DRT) = Time in minutes in which 90% of bacteria at a given temperature will be killed

Methods involving Heat:

0 1. Moist heat will always kill faster than dry heat at the same temperature

0 Kills microbes by coagulation of cell proteins

0 A. Boiling (100°C) kills vegetative forms of bacterial pathogens, many viruses, and fungi within 10 minutes

0Some mo’s and viruses are resistant0 Endospores (up to 20 hrs.) and some viruses (30 minutes) survive

boiling for longer times.

Methods involving Heat:

0 B. Autoclaves0 Moist Heat (steam) and pressure for sterilization0 Achieves higher temps. than boiling0 Preferred method (sealed chamber, air is exhausted, and

steam under pressure is injected) for all materials that can withstand it

0 Kills all organisms and their endospores in about 15 - 20 minutes

0 An autoclave is shown in Figure 1.1 and 1.2

Autoclave

Figure 1.1

Figure 1.2

Autoclave tape

Methods involving Heat:

0 C. Pasteurization0 Louis Pasteur0 Mild Heating

0Kills most pathogens0Kills bacteria that cause spoilage0Preserves taste in product0Lowers bacterial numbers

Methods involving Heat:

0 2. Dry Heat0 Kills by burning to ashes or by oxidation0 Flaming- we use this on loops in labs0 Incineration- burning of contaminated paper0 Hot air sterilization

0Hot ovens0 Mainly used for items not suitable for autoclaving

0 Oily substances, powders0 Large amounts of glassware

Low Temperature

0 Lower temp inhibits growth, rapid freezing limits moisture (bacteriostatic)

0 Refrigerator temperatures (0° to 7°C) slow the metabolic rate of microbes; however0 Psychrotrophic species still

grow slowly.

0 Some organisms grow at temperatures slightly below freezing, but microbes at the usual temperatures of freezer compartments are completely dormant.

Filtration0 Liquids

0 Heat- sensitive materials 0 Small pores prevent passage of bacteria

0 High-efficiency particulate air filters (HEPA)0Operating rooms0Special clean rooms 0Masks

0 Used with solutions for: renal dialysis, heart bypass machines, I.V.’s

0Catch particles autoclaves miss

Desiccation0 Microbes require water for growth, and adequately

dried (desiccated) foods will not support their growth, therefore inhibiting growth

0 Absence of Water0 Lyophilization- rapid freeze drying

0Used for blood products , serum products, enzymes, cultures

0Avoids ice crystal formation; cells burst when water expands

Osmotic Pressure0 High salt or sugar concentrations cause water to leave

the cell; this is an example of osmosis. 0 Generally, molds and yeasts resist osmotic pressures

better than bacteria.

Radiation0 Ionization Radiation

0 includes X rays, gamma rays, and high-energy electron beams

0very short wavelengths and high levels of energy0Penetrate deeply0 Ionizes water to form hydroxyl radicals

0 These destroy cell components, especially DNA

0 Kills: vegetative cells, viruses, most endospores with adequate exposure

0 Applications: food preservation, sterilization of pharmaceuticals, medical supplies, mail

Radiation0 Nonionizing radiation

0 Have a longer wavelength and less energy0 Ultraviolet (UV) light is the common example

0 Causes the formation of thymine dimers, which interferes with DNA replication and formation of mRNA.

0 UV lamps are used in hospitals and in food service0 This method does not sterilize, but it does reduce bacterial growth0 Penetrating power is very low, so any type of covering protects microbes.

0 Sunlight has some weak antimicrobial effects (biocidal), but the wavelengths of sunlight are too long to work well.

Types of Chemical Agents

0 1. Phenols and Phenolics0 A. Phenol (carbolic acid)

0 Irritating to skin and mucous membranes0Bad odor

0 Rarely used today

0 B. Phenolics 0Chemicals derived from phenol

0 Chemically altered to make it less irritating and more effective

0Damages plama mbs., inactivates enzymes, denature proteins0Often used as disinfectants as they remain active in the presence

of organic matter

Types of Chemical Agents

0 C. Bisphenols0 contain 2 phenolic groups connected by a bridge0 Hexachlorophene

0pHisoHex is an example0 Prescription antibacterial lotion0 Used in nurseries to control gram + bacteria

0 Skin bacteria: Staphylococcus and Streptococcus

0 Triclosan0Found in antibacterial soap0Effective against G+ and G- bacteria

Types of Chemical Agents

0 2. Biguanides0 Chlorhexidine is an example

0Hibiclens soap0Used on skin and mucous membranes

0 Similar to phenolics but less toxic0 Disrupt plasma mb. 0 Broad spectrum

0Effective against most vegetative bacteria and fungi, but not against endospores and many viruses.

0 Damaging to eyes

Types of Chemical Agents

0 3. Halogens- Effective alone or in compounds0 A. Iodine (I2)

0One of the oldest and most effective 0Very effective on: all bacteria, many endospores, fungi and some

viruses 0Combines with amino acids in proteins and denatures proteins0Alters plasma mb.0Negative Aspects

0 Staining0 Sometimes irritating to the skin0 May trigger allergies

0Applications0 Skin disinfection, wound treatment, water treatment

Types of Chemical Agents0 B. Chlorine (Cl2)

0 Action: forms hypochlorus acid with water → oxidizing agent, denatures proteins

0 Broad spectrum: bacteria, fungi, some endospores, some viruses0 Positive aspects: -effective against all vegetative cells including

Mycobacterium0 -cost effective

0 Negative aspects: -action inhibited by organics0 -can form carcinogenic compounds

0 Applications: water and sewage treatment, surface and instrument disinfection

0 Sodium hypochlorite (NaClO): Is active ingredient of bleach

Types of Chemical Agents

0 C. Alcohols- frequently used for skin degerming0 Kill bacteria, fungi, but not endospores or naked viruses.0 Act by denaturing proteins and disrupting cell membranes.0 Used to mechanically wipe microbes off skin before injections or blood

drawing, instrument disinfection0 Not good for open wounds, because cause proteins to coagulate and leave bacteria

unharmed

0 Ethanol (ethyl alcohol): Optimum concentration is 70%.0 Isopropanol: Rubbing alcohol

0 Better disinfectant than ethanol0 Usual concentration is 90%0 Also cheaper and less volatile.

Types of Chemical Agents0 5. Heavy metals and their compounds

0 Oligodynamic action- Very tiny amounts are effective0 Includes- silver, mercury, copper, zinc0 Bind sulfur groups causing inactivation or denaturing of proteins0 A. Silver:

0 1% silver nitrate used to protect infants against gonorrheal eye infections until recently.

0 B. Mercury0 Organic mercury compounds like merthiolate and0 mercurochrome are used to disinfect skin wounds.

0 C. Copper0 Copper sulfate is used to kill algae in pools and fish tanks (fungicidal, algicidal)

0 D. Zinc0 Used in mouthwashes0 Superficial fungal and bacterial infections

Types of Chemical Agents0 6. Surface-Acting Agents

0 Disrupt plasma membrane & denature proteins0 Decrease surface tension0 Include soaps and detergents

0 Washing with soap breaks up the oily film that covers skin and allows microbes and dirt to be washed away

0 7. Quaternary Ammonium Compounds0 Widely used surface active agents0 Denature proteins & disrupt cell membranes0 Cationic (positively charge) detergents0 Effective against gram positive bacteria, less effective against gram-negative bacteria.0 Also destroy fungi, amoebas, and enveloped viruses.

Types of Chemical Agents0 8. Aldehydes

0 These can act very effectively against microbes

0 Inactivate proteins

0 Action: cross-link (thus inactivate) nucleic acids and proteins0 High activity (sterilization)0 biocidal including endospores

0 Positive aspects0 achieves sterilization

0 Negative aspects0 unstable0 -toxic0 -volatile with noxious fumes

0 Applications: specimen preservation (embalming), vaccine sterilization

Types of Chemical Agents

0 9. Gaseous Sterilizers

0 Chemicals that sterilize in a chamber similar to an autoclave.0 Denature proteins, by replacing functional groups with alkyl

groups0 A. Ethylene Oxide

0Kills all microbes and endospores , but requires exposure of 4 to 18 hours.

0Toxic and explosive in pure form.0Highly penetrating.0Most hospitals have ethylene oxide chambers to sterilize

mattresses and large equipment.

Types of Chemical Agents

0 10. Peroxygens

0 Oxidize cellular components0 A. Hydrogen Peroxide

0 Common household antiseptic0 Not good for open wounds because quickly broken down by catalase present in

human cells0 used in deep wounds because it releases oxygen as it breaks down, which makes

conditions unfavorable for anaerobic bacteria

0Effective in disinfection of inanimate objects (kills endospores)

0Sporicidal at higher temperatures

0Used by food industry and to disinfect contact lenses

Types of Chemical Agents0 B. Benzoyl Peroxide

0 Main ingredient in many acne treatments 0 May be used in treating wound infections caused by

anaerobes

0 C. Peracetic Acid0 One of the most effective liquid sporicides available0 Sterilant :

0 Kills bacteria and fungi in less than 5 minutes0 Kills endosporesand viruses within 30 minutes

0 Used widely in disinfection of food and medical instrument because it does not leave toxic residues