BIO221 Chapter 7 Microbial Control

Degerming

(or degermation) the mechanical removal and reduction of microbes from animate surfaces. Removal of microbes from a limited area, such as the skin around an injection site

Physical Methods of Microbial Control

* Heat - Moist heat sterilization - Pasteurization - Dry heat sterilization * Filtration * Low temperatures * High Pressure * Desiccation * Osmotic Pressure * Radiation - Ionizing - Nonionizing - Microwaves

A suffix meaning "to kill"?

-cide

Decimal reduction time is the time in minutes in which _____ of the population at a given temperature will be killed.

90%

According to the figure, at what rate are the cells dying?

90% each minute

Disinfectant

A chemical that removes pathogens from inanimate surfaces

Use-dilution Tests

A method of determining the effectiveness of a disinfectant using serial dilutions. Metal or glass cylinders (8 mm × 10 mm) are dipped into standardized cultures of the test bacteria grown in liquid media, removed, and dried at 37°C for a short time. The dried cultures are then placed into a solution of the disinfectant at the concentration recommended by the manufacturer and left there for 10 minutes at 20°C. Following this exposure, the cylinders are transferred to a medium that permits the growth of any surviving bacteria. The effectiveness of the disinfectant can then be determined by the number of cultures that grow. Variations of this method are used for testing the effectiveness of antimicrobial agents against endospores, mycobacteria that cause tuberculosis, viruses, and fungi, because they are difficult to control with chemicals. Also, tests of antimicrobials intended for special purposes, such as dairy utensil disinfection, may substitute other test bacteria

Ultra-high-temperature (UHT) Treatments

A method of treating food with high temperatures (140-150°C) for very short times to make the food sterile so that it can be stored at room temperature

High-efficiency Particulate Air (HEPA) Filters

A screenlike material that removes particles larger than 0.3 μm from air

Membrane Filters

A screenlike material with pores small enough to retain microorganisms; a 0.45-μm filter retains most bacteria

Sterilant

A sterilizing agent

Biocide

A substance capable of killing microorganisms

Germicide

A substance capable of killing microorganisms

Bacteriostasis

A treatment capable of inhibiting bacterial growth

Actions of Microbial Control Agents

Alteration of Membrane Permeability Damage to Proteins and Nucleic Acids

Disk-Diffusion Method

An agar-diffusion test to determine microbial susceptibility to chemotherapeutic agents; also called Kirby-Bauer test. used in teaching laboratories to evaluate the efficacy of a chemical agent. A disk of filter paper is soaked with a chemical and placed on an agar plate that has been previously inoculated and incubated with the test organism. After incubation, if the chemical is effective, a clear zone representing inhibition of growth can be seen around the disk. Disks containing antibiotics are commercially available and used to determine microbial susceptibility to antibiotics

Incineration

An effective way to sterilize and dispose of contaminated paper cups, bags, and dressings

Aseptic

An object or area is free of pathogens

The chemical disinfection of living tissue, such as skin or a mucus membrane?

Antisepsis

Disinfection

Any treatment used on inanimate objects to kill or inhibit the growth of microorganisms; a chemical used is called a disinfectant. Destruction of vegetative pathogens

The absence of pathogens on an object or area?

Aspepsis

What physical method of control would be most effective to sterilize microbiological media?

Autoclaved unless media is heat sensitive; otherwise, filteration

Steam under pressure obtained in restorts, pressure cookers, and?

Autoclaves

What physical method of control would be most effective to eliminate endospore-forming pathogens

Autoclaving at 121°C, 15 psi for 15 minutes

Damage to Proteins and Nucleic Acids

Bacteria are sometimes thought of as "little bags of enzymes." Enzymes, which are primarily protein, are vital to all cellular activities. Recall that the functional properties of proteins are the result of their three-dimensional shape. This shape is maintained by chemical bonds that link adjoining portions of the amino acid chain as it folds back and forth upon itself. Some of those bonds are hydrogen bonds, which are susceptible to breakage by heat or certain chemicals; breakage results in denaturation of the protein. Covalent bonds are stronger but are also subject to attack. For example, disulfide bridges, which play an important role in protein structure by joining amino acids with exposed sulfhydryl (—SH) groups, can be broken by certain chemicals or sufficient heat The nucleic acids DNA and RNA are the carriers of the cell's genetic information. Damage to these nucleic acids by heat, radiation, or chemicals is frequently lethal to the cell; the cell can no longer replicate, nor can it carry out normal metabolic functions such as the synthesis of enzymes

Some antimicrobial treatments kill microbes; some inhibit growth. Which term refers to an agent that inhibits bacterial growth?

Bacteriostatic.

Principles of Effective Disinfection

By reading the label, we can learn a great deal about a disinfectant's properties. Usually the label indicates what groups of organisms the disinfectant is effective against. Remember that the concentration of a disinfectant affects its action, so it should always be diluted exactly as specified by the manufacturer. Also consider the nature of the material being disinfected. For example, are organic materials present that might interfere with the action of the disinfectant? Similarly, the pH of the medium often has a great effect on a disinfectant's activity. Another very important consideration is whether the disinfectant will easily make contact with the microbes. An area might need to be scrubbed and rinsed before the disinfectant is applied. In general, disinfection is a gradual process. Thus, to be effective, a disinfectant might need to be left on a surface for several hours

Chemical Methods of Microbial Control

Chemical agents are used to control the growth of microbes on both living tissue and inanimate objects. Unfortunately, few chemical agents achieve sterility; most of them merely reduce microbial populations to safe levels or remove vegetative forms of pathogens from objects. A common problem in disinfection is the selection of an agent. No single disinfectant is appropriate for all circumstances

Antiseptic

Chemical for disinfection of the skin or mucous membranes

Heat sufficient only to kill endospores on the botulism bacterium?

Commercial sterilization

Alteration of Membrane Permeability

Damage to the lipids or proteins of the plasma membrane by antimicrobial agents causes cellular contents to leak into the surrounding medium and interferes with the growth of the cell.

The time in minutes required to kill 90% of a bacterial population?

Decimal reduction time

The removal of transient microbes from skin by mechanical cleansing or by an antiseptic?

Degerming

The absence of water, resulting in a condition of dryness?

Desiccation

Sterilization is the

Destruction of all life forms

Antisepsis

Destruction of vegetative pathogens on living tissue. A chemical method for disinfection of the skin or mucous membranes; the chemical is called an antiseptic

Sterilization

Destruction or removal of all forms of microbial life, including endospores but with the possible exception of prions

Equivalent treatments

Different methods that have the same effect on controlling microbial growth

Dry Heat Sterilization

Dry heat kills by oxidation effects. A simple analogy is the slow charring of paper in a heated oven, even when the temperature remains below the ignition point of paper

Desiccation

Drying;in the absence of water, microorganisms cannot grow or reproduce but can remain viable for years

Microwaves

Electromagnetic radiation with wavelength between 10^-1and 10^-3m. Do not have much direct effect on microorganisms, and bacteria can readily be isolated from the interior of recently operated microwave ovens. Moisture-containing foods are heated by microwave action, and the heat will kill most vegetative pathogens. Solid foods heat unevenly because of the uneven distribution of moisture. For this reason, pork cooked in a microwave oven has been responsible for outbreaks of trichinellosis

Foods to be canned need not be sterile. However, all _______ of Clostridium botulinum must be killed

Endospores

Autoclave

Equipment for sterilization by steam under pressure, usually operated at 15 psi and 121°C. Sterilization in an autoclave is most effective when the organisms either are contacted by the steam directly or are contained in a small volume of aqueous (primarily water) liquid

What physical method of control would be most effective to sterilize vaccines

Filtration

In the figure shown, what is the method of sterilization used?

Fitration

What part of a microbe is typically not a major target for action of antimicrobials?

Flagella

Generally speaking, the group of organisms that is more resistant to osmotic pressure than bacteria is?

Fungi, such as molds and yeasts

Thermoduric

Heat resistant

High Pressure

High pressure applied to liquid suspensions is transferred instantly and evenly throughout the sample. If the pressure is high enough, it alters the molecular structures of proteins and carbohydrates, resulting in the rapid inactivation of vegetative bacterial cells. Endospores are relatively resistant to high pressure. They can, however, be killed by other techniques, such as combining high pressure with elevated temperatures or by alternating pressure cycles that cause spore germination, followed by pressure caused death of the resulting vegetative cells. Fruit juices preserved by high-pressure treatments have been marketed in Japan and the United States. An advantage is that these treatments preserve the flavors, colors, and nutrient values of the products

Ionizing Radiation

High-energy radiation with a wavelength less than 1nm; causes ionization. X rays and gamma rays are examples

Nosocomial Infections

Hospital-acquired infections

Virucide

Inactivates viruses

Heat

Kill microorganisms by denaturing their enzymes; the resultant changes to the three-dimensional shapes of these proteins inactivate them

Fungicide

Kills fungi

Retorts

Large industrial autoclaves

Low Temperatures

The effect of low temperatures on microorganisms depends on the particular microbe and the intensity of the application. For example, at temperatures of ordinary refrigerators (0-7°C), the metabolic rate of most microbes is so reduced that they cannot reproduce or synthesize toxins. In other words, ordinary refrigeration has a bacteriostatic effect. Yet psychrotrophs do grow slowly at refrigerator temperatures and will alter the appearance and taste of foods after a time. For example, a single microbe reproducing only three times a day would reach a population of more than 2 million within a week. Pathogenic bacteria generally will not grow at refrigerator temperatures

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

Filtration

The passage of a liquid or gas through a screenlike material; a 0.45-μm filter removes most bacteria

Sepsis

The presence of a toxin or pathogenic organism in blood and tissue

Pasteurization

The process of mild heating to kill particular spoilage microorganisms or pathogens

Flaming

The process of sterilizing an inoculating loop by holding it in an open flame

Sanitization

The removal of microbes from eating utensils and food preparation areas. Treatment is intended to lower microbial counts on eating and drinking utensils to safe public health levels

Which is not a characteristic of the autoclave?

The requirement for long (hours) exposure times

Decimal Reduction Time (DRT,or D value)

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

Osmotic Pressure

The use of high concentrations of salts and sugars to preserve food is based on the effects of osmotic pressure. High concentrations of these substances create a hypertonic environment that causes water to leave the microbial cell. This process resembles preservation by desiccation, in that both methods deny the cell the moisture it needs for growth. The principle of osmotic pressure is used in the preservation of foods. For example, concentrated salt solutions are used to cure meats, and thick sugar solutions are used to preserve fruits. As a general rule, molds and yeasts are much more capable than bacteria of growing in materials with low moisture or high osmotic pressures. This property of molds, sometimes combined with their ability to grow under acidic conditions, is the reason that molds, rather than bacteria, cause spoilage of fruits and grains. It is also part of the reason molds are able to form mildew on a damp wall or a shower curtain

The lowest temperature required to kill a liquid culture of a certain species of bacteria in 10 minutes?

Thermal death point

What physical method of control would be most effective to sterilize milk for storage at room temperatures

Ultra-high-temperature (UHT) treatment

Steam ____________ allows temperatures above boiling to be reached

Under pressure (as in an autoclave)

Evaluating a Disinfectant

Use-dilution Tests Disk-Diffusion Method

A good example of ionizing radiation is?

X rays, gamma rays, high-energy electrons

An antiseptic is used when one needs to remove microbes from

skin, prior to injection

Microbial Characteristics and Microbial Control

Many biocides tend to be more effective against gram-positive bacteria, as a group, than against gram-negative bacteria. A principal factor in this relative resistance to biocides is the external lipopolysaccharide layer of gram-negative bacteria. Within gram-negative bacteria, members of the genera Pseudomonasand Burkholderiaare of special interest. These closely related bacteria are unusually resistant to biocides and will even grow actively in some disinfectants and antiseptics, most notably the quaternary ammonium compounds. This resistance to chemical antimicrobials is related mostly to the characteristics of their porins (structural openings in the wall of gram-negative bacteria). Porins are highly selective of molecules that they permit to enter the cell. The mycobacteria are another group of non-endosporeforming bacteria that exhibit greater than normal resistance to chemical biocides. This group includes Mycobacterium tuberculosis,the pathogen that causes tuberculosis. The cell wall of this organism and other members of this genus have a waxy, lipid-rich component. Instruction labels on disinfectants often state whether they are tuberculocidal, indicating that they are effective against mycobacteria. Special tuberculocidal tests have been developed to evaluate the effectiveness of biocides against this bacterial group. Bacterial endospores are affected by relatively few biocides. The cysts and oocysts of protozoa are also relatively resistant to chemical disinfection.The resistance of viruses to biocides largely depends on the presence or absence of an envelope. Antimicrobials that are lipid-soluble are more likely to be effective against enveloped viruses. The label of such an agent will indicate that it is effective against lipophilic viruses. Nonenveloped viruses, which have only a protein coat, are more resistant—fewer biocides are active against them. A special problem, not yet completely solved, is the reliable killing of prions. Prions are infectious proteins that are the cause of neurological diseases known as spongiform encephalopathies, such as the popularly named mad cow disease. To destroy prions, infected animal carcasses are incinerated. A major problem is the disinfection of surgical instruments exposed to prion contamination. Normal autoclaving has proven to be inadequate. The World Health Organization (WHO) and the Centers for Disease Controland Prevention (CDC) have recommended the combined use of a solution of sodium hydroxide and autoclaving at 134°C. Recent reports indicate that surgical instruments have been successfully treated to inactivate prions, which are proteins, by addition of protease enzymes to the cleaning solution. Surgeons sometimes resort to using disposable instruments

-cide

Meaning kill

-stasis

Meaning to stop or to steady

-stat

Meaning to stop or to steady

Moist Heat Sterilization

Moist heat kills microorganisms primarily by coagulating proteins (denaturation), which is caused by breakage of the hydrogen bonds that hold the proteins in their three-dimensional structure. This coagulation process is familiar to anyone who has watched an egg white frying

Ultraviolet light is an example of what type of radiation?

Noniionizing

Boiling

One type of moist heat "sterilization" which kills vegetative forms of bacterial pathogens, almost all viruses, and fungi and their spores within about 10 minutes, usually much faster. Free-flowing (unpressurized) steam is essentially thesame temperature as boiling water. Endospores and some viruses, however, are not destroyed this quickly. Some hepatitis viruses, for example, can survive up to 30 minutes of boiling, and some bacterial endospores can resist boiling for more than 20 hours

Mild heating to destroy particular spoilage organisms or disease organisms in milk or similar products?

Pasteurization

Hightemperature Short-time (HTST) Pasteurization

Pasteurizing at 72°C for 15 seconds

A test for the effectiveness of a chemical disinfectant?

Phenol coefficient

Phosphatase Test

Phosphatase is an enzyme naturally present in milk. If the product has been pasteurized, phosphatase will have been inactivated

Radiation

Radiation has various effects on cells, depending on its wavelength, intensity, and duration. Radiation that kills microorganisms (sterilizing radiation) is of two types: ionizing and nonionizing

The reduction of microbial populations to safe public health levels?

Sanitation

The rate of Microbial Death

Several factors influence the effectiveness of antimicrobial treatments: ● The number of microbes. The more microbes there are to begin with, the longer it takes to eliminate the entire population. ● Environmental influences. The presence of organic matter often inhibits the action of chemical antimicrobials. In hospitals, the presence of organic matter in blood, vomitus, or feces influences the selection of disinfectants. Microbes in surface biofilms are difficult for biocides to reach effectively. Because their activity is due to temperature-dependent chemical reactions, disinfectants work somewhat better under warm conditions. The nature of the suspending medium is also a factor in heat treatment. Fats and proteins are especially protective, and a medium rich in these substances protects microbes, which will then have a higher survival rate. Heat is also measurably more effective under acidic conditions. ● Time of exposure. Chemical antimicrobials often require extended exposure to affect more-resistant microbes or endospores. ● Microbial characteristics.

Nonionizing Radiation

Short-wavelength radiation that does not cause ionization; ultraviolet (UV) radiation is an example. UV light damages the DNA of exposed cells by causing bonds to form between adjacent pyrimidine bases, usually thymines, in DNA chains. These thymine dimers inhibit correct replication of the DNA during reproduction of the cell. The UV wavelengths most effective for killing microorganisms are about 260 nm; these wavelengths are specifically absorbed by cellular DNA

Sunlight owes its biocidal activity mainly to the formation of what type of oxygen?

Signlet

Hot-air sterilization

Sterilization by the use of an oven at 170°C for approximately 2 hours

Destroying or removing all forms of microbial life?

Sterlization

Commercial Sterilization

Sufficient heat treatment to kill endospores of Clostridium botulinumin canned food

Aseptic Surgery

Techniques used in surgery to prevent microbial contamination of the patient

Asepsis

The absence of contamination by unwanted organisms