Ch. 7 -- Control of Microbial Growth

Resistant Bacteria

*Gram -* bc of their external lipopolysaccharide layer & *porins* that are extremely selective of what crosses the membrane *Mycobacteria* bc they have a waxy, lipid-rich component Nonenveloped viruses (made up only of protein) are more resistant than enveloped viruses To destroy prions, animal carcasses are incinerated

Evaluating a Disinfectant

*Use-dilution Test* -- current standard; cultures placed in solution of disinfectant & effectiveness of disinfectant determined by # of cultures grown *Disk-diffusion Method* -- used in teaching labs; a disk of filter paper is soaked with a chemical & placed on an agar plate previously inoculated with a test organism; if effective, a clear zone representing inhibition of growth can be seen

Physical [*Radiation*]

2 types: ionizing + nonionizing *Ionizing* -- gamma rays, x rays, & high-energy electron beams; shorter wavelengths than nonionizing = more energy; the ionization of water forms highly reactive hydroxyl radicals that kill organisms by reacting with organic cellular components, esp. DNA, & damaging them; "hits" can cause enough mutations to kill the microbe *Nonionizing Radiation* -- longer wavelength greater than 1 nm; UV light damages the DNA of exposed cells by causing bonds to form b/w adj. pyramidine bases, usually thymines, in DNA chains; thymine dimers inhibit correct replication of DNA

Peroxygens

A group of oxidizing agents that includes hydrogen peroxide + peracetic acid Hydrogen peroxide not good for open wounds but its good against inanimate objects Peracetic acid is one of the most effective liquid chemical sporcides available & can be used as a sterilant; used on endospores + viruses

Plasmas

A state of matter in which a gas is excited by an electromagnetic field to make a mix of nuclei with assorted electrical charges + free elections; used in sterilization for plastic/metal surgical instruments; they have free radicals that quickly destroy endospores; requires low temps.

Sterilant

A sterilizing agent; liquids + gases can be sterilized by filtration

Aldehydes

Among the most effective antimicrobials Ex: formaldehyde + glutaraldehyde They inactivate proteins by forming covalent cross-links with several organic functional groups on proteins; glutaraldehyde works better & used to disinfect hospital equipment; it could also be called a sterilizing agent; both are used by morticians for embalming

Physical [*High Pressure*]

Applied to liquid suspension is transferred instantly & evenly throughout the sample; if pressure is high enough, it alters molecular structures of proteins + carbohydrates resulting in rapid inactivation of vegetative bacterial cells Endospores resistant to high pressure

Equivalent Treatments

As the temp. is increased, much less time is needed to kill the same number of microbes

Surface-active Agents/Surfactants

Can decrease surface tension among molecules of a liquid Ex: soap + detergents Soap used as mechanical removal of microbes through scrubbing

Phenol + Phenolics

Carbolic acid to control surgical infections in the operating room by Lister; usually used in cough drops; but above 1%, it has significant antibacterial effect Phenolics are derivatives of phenol & contain a molecule of phenol that has been chemically altered to reduce its irritating qualities or increase its antibacterial activity in combination with soap/detergent; they injure *lipid-containing plasma membranes* which makes cell contents leak out; remain active in presence of organic compounds, are stable, & persist for long periods after application; disinfect pus, saliva, & feces; most popular is *cresols*

Supercritical Fluid

Combines physical + chemical methods; when CO2 is compressed into a supercritical state, it has properties of both a liquid (with increase solubility) & a gas (with lowered surface tension)

Membrane Filters

Composed of cellulose esters or plastic polymers & are popular for industrial + lab use; effective for bacteria + viruses

Disinfection

Control directed at destroying harmful microorganisms; refers to destruction of vegetative (non-endospore-forming) pathogens, which is not the same as complete sterility; physical or chemical methods Ex: chemicals, UV radiation, boiling water, steam A disinfectant is used to treat an inert surface or substance

Bisphenols

Derivatives of phenol that contain 2 phenolic groups connected by a bridge Hexachlorophene used in prescription lotion & Triclosan used in antibacterial soaps; but Triclosan inhibits an enzyme needed for the biosynthesis of fatty acids which affects the integrity of the plasma membrane; effective against gram +

Physical [*Sterilization*]

Done by ultra-high-temperature (UHT) treatments; can be done to sterilize milk for several months without refrigeration; liquid milk is sprayed through a nozzle into a chamber filled with high-temp. steam under pressure

Physical [*Dry Heat Sterilization*]

Dry heat kills by oxidation effects Direct flaming is a simple method (flaming a loop) as well as incineration Hot air sterilization is when items are placed in an oven; hot temp. & longer period of time needed

Biguanides

Effective against gram +; affect bacterial cell membranes; best known is Chlorhexidine which is used on skin + mucous membranes

Alcohols

Effectively kill bacteria + fungi but not endospores & nonenveloped viruses; denatures protein & disrupt membranes + dissolves many lipids; evaporate rapidly & leave no residue; not good for skin wounds Ethanol + isopropanol are most commonly used; pure ethanol less effective than aqueous solutions b/c water denatures proteins; isopropanol works better than ethanol

Physical [*Pasteurization*]

Eliminates pathogenic microbes without damaging the taste of a product; it lowers microbial numbers Thermoduric (heat-resistant) bacteria survive this process but don't cause disease or spoilage

Sepsis

From the Greek word for decay/putrid; indicates bacterial contamination, as in septic tanks

Chemical Sterilization

Gaseous chemosterilants used as substitutes for physical sterilization processes; requires a closed chamber similar to an autoclave Ex: ethylene oxide; activity depends on alkylation (replacing the proteins' labile hydrogen atoms with a chemical radical which leads to cross-linking of nucleic acids + proteins & inhibits vital cellular functions); kills all microbes + endospores but it takes a long time to do so

Physical [*Heat*]

Heat denatures enzymes which leads to inactivation

Physical [*Desiccation*]

In the absence of water, microorganisms can't grow/reproduce but can remain viable for years; but when water is reintroduced they resume their growth Lyophilization is known as freeze-drying

Bacteriostasis

Inhibits the growth + multiplication of bacteria

Biocide/Germicide

Kills microorganisms (not endospores)

Physical [*Moist Heat Sterilization*]

Kills microorganisms primarily by coagulating proteins (denaturation) caused by the breakage of hydrogen bonds that give the proteins their 3D structure - Boiling kills vegetative forms within 10 min - Higher temp. achieved when using an *autoclave* (steam under pressure); used to sterilize culture media, instruments, equipment, syringes, etc.

Commercial Sterilization

Limited heat treatment that subjects canned food to enough heat to destroy the endospores of Clostridium botulinum

Sanitization

Lowers microbial counts to safe public health levels & minimize the chances of disease transmission b/w people Ex: restaurant stuff; done by high temp. washing or by dipping in a chemical disinfectant

Quaternary Ammonium Compounds (Quats)

Most widely used surface-active agent; a cationic detergent; modifications of the 4 valence ammonium ion; strongly bactericidal against gram +; fungicidal, amebicidal, & virucidal against enveloped viruses; don't kill endospores or mycobacteria; they change the cell's permeability & cause loss of essential cytoplasmic constituents

Halogens

Particularly iodine + chlorine, are effective both alone & as constituents of inorganic/organic compounds Iodine is the most oldest + effective antiseptic against many bacteria, endospores, fungi, & some viruses; it impairs protein synthesis & alters cell membranes by forming complexes with a.a & unsaturated fatty acids; available as a *tincture* (solution in aqueous alcohol) + as an *iodophor* (combo for Iodine + an organic molecule); Iodine mainly used in skin disinfection + wound treatment Chlorine as a gas or combo with other chemicals; germicidal action when added to water; hypochlorous acid is the most effective form; liquid form of compressed gas used for water; bleach is a household disinfectant; chloramines used for clarifying water & sanitizing glassware

Physical [*Low Temperatures*]

Reduces metabolic rate of most microbes that they can't produce toxins or reproduce; has a bacteriostatic effect

High-efficiency Particulate Air (HEPA) Filters

Remove almost all microorganisms larger than 0.3 micrometers in diameter

Thermal Death Point (TDP)

The *lowest* temperature at which all the microorganisms in a particular liquid suspension will be killed in *10 minutes*

Thermal Death Time (TDT)

The *minimal* length of time for all bacteria in a particular liquid culture to be killed at a given temperature

Oligodynamic Action

The ability of very small amounts of heavy metals, esp. silver + copper, to exert antimicrobial activity; zinc is also another one used When metal ions combine with sulfhydryl groups on cellular proteins, denaturation results

Asepsis

The absence of significant contamination

Degerming

The mechanical removal, rather than the killing, of most of the microbes in a limited area Ex: swabbing skin with alcohol

Physical [*Filtration*]

The passage of liquid/gas through a screenlike material with pores small enough to retain microorganisms; a vacuum is created in the receiving flask & air pressure then forces the liquid through the filter Used to sterilize heat-sensitive materials like culture media, enzymes, vaccines, & antibiotic solutions

Target

The plasma membrane is the target of many microbial control agents; damage to lipids/proteins of the membrane causes cellular components to leak into the surrounding medium & interferes with the cell's growth

Nitrosamines

The reaction of nitrates with a.a can form certain carcinogenic products; prevent botulism in processed meats

Sterilization

The removal/destruction of *ALL* living microorganisms; heating is the most common method used Done by steam under pressure or a sterilizing gas such as ethylene oxide

Decimal Reduction Time (DRT - D value)

The time, in *minutes*, in which *90%* of a population of bacteria at a given temp. will be killed

Physical [*Osmotic Pressure*]

The use of high concentrations of salts + sugars to preserve food; causes a *hypertonic* environment that causes water to leave the cell; like preservation by desiccation Molds + yeasts grow more easily than bacteria with low moisture & high osmotic pressures

High-temperature Short-time (HTST) Pasteurization

Treatment applied as the milk flows continuously past a heat exchanger; uses temp. of at least 72C for 15 sec.; lowers bacterial count

Antisepsis

Treatment directed at living tissue; chemical called an antiseptic; treatment always by chemical antimicrobials

Chemical Agents

Used to control the growth of microbes on both living tissue + inanimate objects; few achieve sterility; most reduce microbial populations to safe levels or remove vegetative forms of pathogens from objects

Acid-Anionic Sanitizers

Very imp. in cleaning food-processing facilities, esp. dairy utensils + equipment; usually combos of phosphoric acid + a surface-active agent Negatively charged portion (anion) of the molecule reacts with the cell membrane; they are odorless, nontoxic, noncorrosive, & fast acting; used on a wide array of microbes like thermoduric