CALS Farm and Industry Short Course Program: Farm Microbiology: Notes

SECTION 2.
Basic Structure, Genetics, Habitats, Physiology, Nutrition and Growth of Bacteria

  1. Basic Morphology.

    1. Microscopic.  As an example, the average size of an E. coli cell is 1.3 micrometers (or 0.000052 inch) wide by 4.0 micrometers (or 0.00016 inch) long. It would take 6250 strung end-to-end to make an inch. Note: 1000 micrometers = 1 mm (millimeter) = 0.04 inch.

    2. Some rare exceptions.

      1. Epulopiscium.  Long rod-shaped organism, visible to the naked eye at half a millimeter in length; found in fish in the Red Sea and Australia.

      2. Thiomargarita.  Thiomargarita (which means "sulfur jewel"), a spherical organism, was found in the ocean off Africa.

    3. Shapes.  Bacteria come in a variety of shapes, but the most common are shown in the figure below. Sometimes terms dealing with cellular shape and arrangement can be found in the official scientific names given to some bacteria – such as Bacillus, Staphylococcus (original meaning: spherical cells arranged in clusters), Streptococcus (original meaning: spherical cells arranged in chains), Aquaspirillum (spiral-shaped cell found in water), etc.

      1. Coccus.  Spherical, berry-shaped.

      2. Bacillus.  Cylindrical, rod-shaped.

      3. Spirillum.  Spiral, helical-shaped.


    Shapes of Bacterial Cells

  2. Types of Cells.

    1. Vegetative cell.  Bacteria actively metabolize as "vegetative cells," and "growth" of bacteria refers to an increasing population of living vegetative cells – generally by binary fission. For some genera of bacteria, vegetative cells can give rise to "resting cells" (following) which have various degrees of survival capability in the environment.

    2. Endospore.

      1. Formation.  Formed mainly by vegetative cells of the genera Bacillus and Clostridium. As a population of cells grows by binary fission, a signal triggered by decreasing nutrients can cause a cell to divide into two cells – but this time, one of the cells (i.e. the endospore) is formed inside of the other. Eventually the outer cell breaks apart and the endospore is "freed."

      2. Features.  These are the most durable and long-lived of all types of cells, possessing many resistant outer layers and very little (if any) water. When endospore-forming bacteria cause diseases in animals, such as anthrax, tetanus and botulism, the spores are usually involved in transmission and survival of the organism.

    3. Reproductive spore.  Formed by branched bacteria in the soil such as Streptomyces. Not as hardy as endospore. Used mainly for reproduction.

    4. Cyst.  Highly resistant cell formed by certain soil bacteria.

  3. Some Structural Components.  Compared to eukaryotic cells, procaryotes have a relatively simple internal architecture and organization. Unicellular organisms without a true nucleus or nuclear membrane.

    1. Cell wall and cell membrane = cell envelope.

      1. Cell wall.  The cell wall prevents osmotic rupture or lysis of the cells. Most bacteria have a "gram-positive" or a "gram-negative" type of cell wall. The former is made up of one thick rigid layer made of peptidoglycan, and the latter is made up of two relatively thin layers – a peptidoglycan layer and a cell membrane on the outside. This is a topic gone over more thoroughly in lab during the gram-staining exercise. Bergey's Manual starts classifying bacteria with shape and gram reaction

      2. Cell membrane.  Composed of phospholipids and proteins, it completely surrounds and encloses the protoplasm of the cell. In procaryotes the membrane is a permeability barrier that is used to take up nutrients and to expel waste products. It is also a site for various types of energy generation including respiration.

    2. Capsule.  A coherent slime layer which forms on the outside of the cell. Composed of protein or polysaccharide and is not part of the cell wall or envelope. Produced by some species of bacteria. Used mainly for protection against engulfment by other organisms and for attachment to surfaces.

    3. Ribosomes.  These are the sites of protein synthesis. Ribosomes are composed of ribosomal RNA (rRNA) and protein. The ribosomes of procaryotes are smaller in size and slightly different than eukaryotic ribosomes.

    4. Storage granules.  These are distinct granules or inclusions in the cells. Usually they are reserve materials of some sort, analogous to starch or fat bodies in higher organisms.

    5. Flagella.  Whip-like filaments that are responsible for swimming movement.

    6. Fimbriae (also called Pili).  Hair-like structures on the covering the cell surface. Fimbriae have similar functions to capsules in that they are involved in attachment to surfaces or protection against engulfment by higher organisms.
  4. Genetic Material and Associated Activities.

    See diagrams and discussion on special handout, reproduced here.

    1. Nucleic acids – DNA and RNA.  DNA=deoxyribonucleic acid. The procaryotic chromosome is usually a single circular molecule of DNA. RNA=ribonucleic acid.

    2. Structure of DNA.

      1. Basic unit = nucleotide – composed of:

        1. Base molecule: adenine, guanine, cytosine or thymine.  This is hooked to a sugar and phosphate.

        2. Sugar molecule = deoxyribose.

        3. Phosphate.

      2. Linkage of nucleotides = strand of DNA.  The nucleotides are linked together by the phosphate molecules to form a strand of the DNA molecule.

    3. Replication of DNA.

    4. The genetic code.

      1. Bases in groups of three.  The bases (in groups of 3) each code for a specific amino acid.

      2. Genes.  About two thousand different genes code for various structures and enzymes. Each gene is composed of many nucleotides that (in groups of 3) code for the various amino acids that eventually get strung together to produce a certain protein.

    5. RNA.

      1. Synthesis = transcription.  Strands of messenger RNA (mRNA) are synthesized according to the DNA code.

      2. Differences from DNA.

        1. Base molecule: adenine, guanine, cytosine or uracil.

        2. Sugar molecule = ribose.

    6. Protein synthesis = translation.  The code on the mRNA orders up the various amino acids which are ultimately strung together in the precise order to produce a specific protein. This happens at the site of the ribosome.

    7. Activities (DNA replication, transcription, translation) accomplished in any order?  These activities occur simultaneously and constantly in actively-metabolizing cells.

  5. Genetic Differentiation and Identification of Bacteria – Base-Sequencing.  (Note: Traditional methods of differentiating and identifying bacteria will be discussed in lab as well as how bacteria and other forms of life are named. Serotyping will be discussed in Section 3.)

    1. Differentiating between species and establishing relationships.  See diagram below.

    2. Identification of unknown organisms and establishing new species.


    Basis for Constructing a Phylogenetic Tree
    (Series of Diagrams from Bacteriology 102)

  6. Habitats.  Anywhere that you can see that life exists, procaryotes are always there. In addition, procaryotes may be the only form of life in many environments.

  7. Unique Processes Not Found in Eukaryotes.

    1. Special types of photosynthesis.  Photosynthetic bacteria other than cyanobacteria produce no O2, and in many cases organic material (instead of CO2) is used as the carbon source. Often such organisms assume a variety of red-based colors.

    2. Special types of fermentation.  End products include lactic acid, butyric acid, acetic acid (used in production of vinegar), alcohols, etc.

    3. Use of inorganic compounds as energy sources.  This is called lithotrophy.

    4. Respiration without oxygen.  An "oxygen substitute" such as nitrate or sulfate can be used by some organisms. This is anaerobic respiration.

    5. Nitrogen-fixation.  This is where molecular nitrogen (N2) from the atmosphere is used as the source of cellular nitrogen compounds.

    6. Degradation of complex organic substances.  Many bacteria decompose materials that other microorganisms cannot.

  8. General Overview of Microbial Metabolism and Nutritional Requirements.  Under the right physical conditions, every microorganism has to find (1) an energy source and also (2) compounds containing certain chemical elements in order to grow and reproduce. Exactly how the cells go about doing this is called metabolism.

    1. Metabolism.  The term metabolism refers to the sum of the biochemical reactions required for energy generation and the use of energy to synthesize cell material from small molecules in the environment. Hence, metabolism has an energy-generating component, called catabolism, and an energy-consuming, biosynthetic component, called anabolism. Catabolic reactions or sequences produce energy as ATP, which can be utilized in anabolic reactions to build cell material. The relationship between catabolism and anabolism is illustrated in the figure below.


      A Very generalized Overview of Metabolism.
      Note: "Energy source" is better called "source of electrons." Also, the lateral lines (directed to the right) represent the transfer of energy (1) given off when electrons are released and (2) given off when ATP goes back to ADP.

    2. Nutritional requirements.  Chemicals (elements or molecules) that cells need to grow and produce energy.

      1. Overview of major elements.

        Elements of Prime Importance and Their Sources and Functions in Bacterial Cells.

        Element % of dry weight Source Major Functions
        Carbon (C) 50 organic compounds or CO2 Essential elemental constituent of cellular material.
        Oxygen (O) 20 H2O, organic compounds, CO2, and O2 Constituent of cell material and cell water; O2 is electron acceptor in aerobic respiration.
        Nitrogen (N) 14 NH3+, NO3–, organic compounds, N2 Constituent of amino acids, the bases in nucleic acids and nucleotides, coenzymes, vitamins.
        Hydrogen (H) 8 H2O, organic compounds, H2 Most numerous elemental constituent of organic compounds and cell water.
        Phosphorus (P) 3 inorganic phosphates (PO43–) Constituent of ATP and nucleic acids (the phosphate component); also phospholipids, lipopolysaccharide, teichoic acids, vitamins.
        Sulfur (S) 1 SO42–, H2S, S0, organic sulfur compounds Constituent of two amino acids (cysteine, methionine), some coenzymes, vitamins.
        Potassium (K) 1 Potassium salts Main cellular inorganic cation and cofactor for certain enzymes
        Magnesium (Mg) 0.5 Magnesium salts Inorganic cellular cation, cofactor for certain enzymatic reactions
        Calcium (Ca) 0.5 Calcium salts Inorganic cellular cation, cofactor for certain enzymes and a component of endospores
        Iron (Fe) 0.2 Iron salts Component of cytochromes and certain nonheme iron-proteins and a cofactor for some enzymatic reactions
      2. Trace elements.  These are elements whose compounds are present in "trace" amounts. Sometimes elements such as cobalt (Co), zinc (Zn), manganese (Mn), and molybdenum (Mo) are also needed, although the requirement cannot always be demonstrated.

      3. Growth factors.  These are essential organic compounds which an organism cannot synthesize from the materials at hand; they then have to be provided pre-made to the organism. Examples include vitamins, amino acids, nucleic acid bases, and fatty acids. Various organisms can synthesize these compounds to a greater or lesser degree. E. coli has no special growth factor requirement; it can synthesize all it needs from a medium in which glucose is the only organic compound. Some microbes need a wide variety of vitamins and other growth factors in their nutrition as do animals; those that require a wide variety of growth factors are termed "fastidious."

      4. Importance of water.  Cells are 95% H2O. All enzymatic activities take place in H2O. All nutrients must be dissolved in H2O for the cell to take them in. The more solute tied up in water, the less likely there will be growth although there are some, as an example, that require a high requirement of NaCl in their medium which would be deleterious to other organisms.

  9. Sources of Energy and Reducing Power for Catabolic Reactions.


    An Attempt to Produce an Ultra-general "Universal Diagram" to Summarize Catabolism.

    1. Purposes of catabolism.

      1. Generate "reducing power."  Electrons released can be used in various cellular metabolic reactions.

      2. Generate energy.  Energy generation is associated with the release of electrons and is used to generate ATP (adenosine triphosphate) from ADP and phosphate. ATP provides energy in a variety of cellular metabolic reactions, most essentially in anabolism.

      3. Provide some of the "building blocks" for anabolism.

    2. Overview of terms and reactions.

      1. Chemotrophs vs. phototrophs.  Chemical compounds are oxidized by chemotrophs as the sole means of obtaining energy. (This is accomplished in any such organism by respiration or fermentation.) Chemical compounds are also oxidized by phototrophs, but the ultimate energy source is light.

      2. Organotrophs vs. lithotrophs.  Organotrophs oxidize organic compounds. Lithotrophs oxidize inorganic compounds.

        Examples of electron donors that are oxidized include:

        • H2 (hydrogen gas), oxidized to H2O
        • S– (sulfide), oxidized to SO42– (sulfate)
        • Fe2+ (ferrous ion), oxidized to Fe3+ (ferric ion)
        • NH4+ (ammonium), oxidized to NO2– (nitrite)
        • NO2– (nitrite), oxidized to NO3– (nitrate)
      3. Combining/compounding terms.  A way in which organisms can be described or classified: chemoorganotrophs, chemolithotrophs, photoorganotrophs or photolithotrophs.

    3. Further details and applications (see figure below).

      1. Fermentation.

        1. End products.  Result from the reduction reactions which are required to balance oxidation reactions (the reactions that produce energy). End products are waste products to the organisms that produce them, but are necessary to maintain the energy- producing reactions.

        2. Practical applications.  The fermentation industry is usually in it for the end products which may be used (1) as foods, (2) to prepare, flavor and preserve foods (e.g., lactic acid), (3) to produce alcoholic beverages, (e.g., ethanol) or (4) to produce various fuels or solvents (e.g., acetate, butanol, acetone, gasohol).

      2. Respiration.  Metabolism in which energy is derived from the complete oxidation of a substrate using an outside electron acceptor. Compared to fermentation as a means of oxidizing organic compounds, respiration is a lot more complicated. Complete oxidation of an organic compound results in CO2 production.

        1. Aerobic respiration.  Uses O2 as the final electron acceptor which is reduced to H2O. Animals, plants, fungi, protozoa and many bacteria are aerobic respirers. Overall pathway for aerobic respiration by chemoorganotrophic organisms is:
          CH2O + O2 --------> CO2 + H2O

        2. Anaerobic respiration.  Use of some chemical other than O2 as an electron acceptor. Anaerobic respiration is a metabolic feature that is unique to procaryotes and it means that they can respire in the absence of oxygen, i.e., in anaerobic environments. Compounds like NO3– (nitrate) or SO42– (sulfate) may be used as electron acceptors by anaerobic respirers.

      3. Oxygenic (plant) photosynthesis.  Production of O2. Conducted by plants, algae and cyanobacteria.

      4. Anoxygenic (bacterial) photosynthesis.  No production of O2. Conducted by the purple and green bacteria under anaerobic conditions.

    " FERMENTATION:

    Example: On the left side, glucose is oxidized to pyruvate. On the right side, pyruvate can be reduced to acids, alcohols and gases.

    " RESPIRATION:

    Above shows AEROBIC RESPIRATION with the use of oxygen. ANAEROBIC RESPIRATION uses an "oxygen substitute" such as nitrate, sulfate, etc.

    " PHOTOTROPHY:


    ("chl" represents chlorophyll.)

    Phototrophy can be OXYGENIC (evolving O2 when H2O serves as the electron donor) or ANOXYGENIC (non-O2-evolving).

    Simplified Representations of Fermentation, Respiration and Phototrophy.

  10. Sources of Carbon.

    1. Importance of carbon.  Carbon is the backbone of all organic molecules that make up cell material. Oxidation of the C to H bond is the basis for the energy release from organic molecules.

    2. Heterotrophy vs. autotrophy.  Organisms are also identified on the basis of their carbon source for growth. Basically, autotrophs use CO2 as a source of carbon and heterotrophs use organic carbon. In macroscopic life, plants are autotrophs and animals are heterotrophs.

  11. Nutritional Types of Microorganisms – Based on Carbon and Energy Sources.  There is a wide range of physical conditions over which the microbes will grow. In the bacteria there are species that will grow at freezing and others that will grow at boiling temperatures. Some grow in 20% NaCl or in 1 N sulfuric acid. The physical restrictions placed on procaryotic growth define the limits of the existence of life on earth. The response of microbes to three environmental criteria – temperature, pH and oxygen – is described below in association with the terms that are applied to microorganisms depending on their behavior under certain environmental conditions.

    Major Nutritional Types of Microorganisms.

    Nutritional Type Carbon Source Energy Source Examples
    Photoautotrophs CO2 Light (These organisms are generally "photolithotrophic" in that electron transfer usually involves the oxidation of inorganic compounds.) Algae; cyanobacteria; some purple and green bacteria.
    Photoheterotrophs Organic compounds Light (These organisms are generally "photoorganotrophic" in that electron transfer usually involves the oxidation of organic compounds.) Some purple and green bacteria; a few algae.
    Chemoautotrophs CO2 Usually inorganic compounds (chemolithotrophy) – e.g., H2, NH4+, NO2–, H2S Relatively few bacteria and many archaea.
    Chemoheterotrophs Organic compounds Usually organic compounds (chemoorganotrophy) Protozoa, fungi, most bacteria, some archaea.

  12. Physical Requirements and Restrictions.

    1. Temperature.  The procaryotes can grow over a range of temperature from freezing to boiling. The only temperature restriction is that water must be in a liquid form, not ice or steam. No eukaryote grows above 73°C.

      1. Range (cardinal points): minimum, maximum, optimum.  Every organism has a range of temperature over which it will grow: a minimum temperature below which growth does not occur, an optimum temperature where growth is fastest and most efficient, and a maximum temperature above which growth does not occur, These three temps are called the cardinal points. For most bacteria there are about 30°C between the minimum and the maximum. Bacteria are classified according to their range of temperature for growth.

        1. Psychrophiles.  Able to grow at freezing, 0°C. Most psychrophiles are free living organisms that grow below 10°C.

        2. Psychrotrophs.  Grow at 0°C, but they grow much better at room temperature (20-25°C). They are the organisms most likely to grow at refrigerator temperatures (0-2°C) and consequently spoil food.

        3. Mesophiles.  Optimum temperature near 37°C, the body temp of humans. Mesophiles may be free living in soil and water or live in associations with plants and animals.

        4. Thermophiles.  "Heat loving" bacteria that grow above 45°C, as in compost piles.

        5. Extreme thermophiles.  Usually archaea that are able to grow at boiling temperatures of 100°C and above. See Figure below.


        Effect of Temperature on Growth of Bacteria.

      2. Effect on rate of cell division.  An increase in T from the minimum to the optimum is directly related to an increase in growth and cell division. Beyond the optimum some critical thermal event takes place, and the growth rate falls off sharply (see figure above).

      3. Thermal death.  All organisms can be killed by a certain amount of heat (above their growth maximum) in a certain amount of time. Thermal death is brought about by inactivation of cellular enzymes, nucleic acids and membrane. Time and temperature are always related in regards to thermal death. Generally, as the temperature is increased, the time (at that temp) of death is decreased.

    2. pH (acidity and alkalinity).

      1. Range.  Bacteria grow over a range of about 3 pH units and are classified accordingly. (See figure below.)

      2. Effect on growth rate.


      Effect of pH on Growth Rates of Different Types of Bacteria.

    3. Oxygen (O2).

      1. Classification of organisms as to use and tolerance of O2 – terms used in the most general sense.

        1. Aerobes.  Require O2 for growth using it as a final electron acceptor in aerobic respiration.

        2. Anaerobes.  Do not need or use O2 as a nutrient. In fact, O2 is a toxic substance, which either kills or inhibits their growth. Obligate anaerobic procaryotes may live by fermentation, anaerobic respiration, anoxygenic photosynthesis or the novel process of methanogenesis (which we often lump under anaerobic respiration for convenience).

        3. Facultative anaerobes.  Organisms that can switch between aerobic and anaerobic types of metabolism. In the presence of O2 they switch to aerobic respiration.

      2. Terms applied in the "oxygen relationship" laboratory test.  This test involves a standard medium in which most common chemoheterotrophic bacteria can grow. Any anaerobic growth is attributed to fermentation of the glucose in the medium; test does not allow for anaerobic growth due to anaerobic respiration or anoxygenic photosynthesis.

    Oxygen Relationships among Chemotrophic Microorganisms

    Group Aerobic
    growth    		 Anaerobic
    growth    		 Ability to
    respire
    (with O2)    		 Ability to
    ferment    		 Representative Organisms
    Obligate
    Aerobe    		 + – + – Animals, plants, algae, molds, protozoa, many prokaryotes.
    Facultative
    Anaerobe    		 + + + + Many bacteria, yeasts, some protozoa, few animal cells.
    Aerotolerant
    Anaerobe    		 + + – + The "lactic acid bacteria." (Aerotolerant anaerobes would be included in facultative anaerobes in the more general use of these terms as listed above.)
    Obligate
    Anaerobe    		 – + – + or – Many prokaryotes (e.g., Clostridium).

  13. Growth of Microorganisms.  Growth is defined as an orderly increase in cell mass and cell numbers.

    1. Binary fission and exponential growth.  Most bacteria grow by the process of binary fission, which is a simple splitting process. The cell grows in size and mass, replicates its chromosome, forms a cross-wall (septum) and splits into two cells. For this reason the cell population doubles every time the cells divide and the population grows in a geometric progression of 1, 2, 4, 8, 16, etc. This pattern of growth is called exponential growth.

    2. Generation time.  This is the time it takes for a population of bacterial cells to double. For many bacteria this is between 15 minutes and one hour under the right conditions. Bacterial populations can grow fast and reach very high populations of cells (greater than 109 cells per ml)

    3. Growth curves.  Bacteria growing in the laboratory, and sometimes in nature, exhibit a typical pattern of growth in a closed system called the typical bacterial growth curve which is shown below.


      Typical Bacterial Growth Curve.

    4. Growth relations with one another.  Most bacteria grow in associations with other organisms, including plants, animals, and other microorganisms. These associations are not necessarily mutually beneficial.

      1. Mutualistic associations.  These are associations between two organisms where both organisms benefit. This is what is typically thought of as" symbiosis."

        1. Normal flora.  The normal flora present – in/on such sites as the intestinal tract or skin – prevent growth of pathogenic bacteria by competing for nutrients or colonization sites or by producing specific chemicals that prevent growth of the harmful bacteria. These bacteria are living in mutualistic associations with their animal hosts. Bacteria that live on the skin produce acids that prevent the growth of fungi that cause athletes foot or ringworm.

        2. Root nodule bacteria (bacterial association with plant roots).  Main examples are Rhizobium and Bradyrhizobium. Organisms in these genera form nodules on the roots of leguminous plants such as peas, clover, alfalfa and soy beans. The bacteria that live in the nodule fix nitrogen by converting N2 to ammonia which can be used by the plant as a nitrogen source.

        3. Mycorrhiza (fungal association with plant roots – mentioned here for comparison).  This refers to an association between fungi in plant roots. The fungi live on the root and sometimes penetrate it. The fungi obtain nutrients from the plant without harming it and in return they produce growth-stimulating substances for the plant.

      2. Parasitic associations.  These are situations where one organism (a parasite) lives at the expense of another organism (the host). Often the result is damage or harm to the host so that parasitic microorganisms may be the agents of disease.

      3. Commensal associations.  Commensal associations between two organisms are situations wherein two organisms live together but have no obvious effect on one another. Commensalism is really a provisional category between mutualism and parasitism. If you study a commensal situation long enough, you will probably discover that one organism has some effect on the other, for better or worse.

    5. Some obvious changes produced by microbial growth.  There are many changes in the environments that microorganisms grow in. Below are some familiar changes that are caused in foods and other substrates as a result of microbial growth. Some are beneficial to a consumer while some are not.

      1. Acid formation.  Examples: silage, buttermilk, cheese, sausage.

      2. Alcohol production.  Examples: beer, wines, etc.

      3. Gas production.  Examples: cheese, bread, silo gas.

      4. Protein decomposition.  Examples: cheese flavors, putrefaction.

      5. Fat decomposition.  Examples: blue cheese, rancidity.

      6. Ropiness and slime.  Examples: milk, meats.

  14. Control of Microbial Growth.  The control of microbial growth is necessary in many practical situations, and significant advances in agriculture, medicine, and food science have been made through study of this area of microbiology.

    1. Practicality.

      1. In medicine.  Example: prevention or cure for infectious disease.

      2. In agriculture.  Example: prevention or treatment of microbial diseases of plants and animals; increasing soil fertility; silage fermentations.

      3. In food science.  Examples: prevention of food spoilage; use of bacteria for useful fermentations of foods.

    2. Principles of prevention.

      1. Killing vs. inhibition of growth – use of cidal vs. static agents.  Control of growth usually involves the use of physical or chemical agents which either kill or prevent growth (inhibit) microorganisms. Agents which kill cells are called cidal agents; agents which inhibit the growth of cells (without killing them) are referred to as static agents. The term "bactericidal" refers to killing bacteria and "bacteriostatic" refers to inhibiting the growth of bacterial cells. A bactericide kills bacteria, a fungicide kills fungi, etc.

      2. Sterilization.  This is the complete removal of living things. Various physical or chemical cidal agents can be used, and sterilization procedures often involve the use of heat. Used for sterile products, supplies, medicines, etc. An object which is sterilized and remains out of contact with environmental microorganisms will remain sterile indefinitely, as the killing of a microorganism obviously prevents its growth.

    3. Methods of control.  (For food, must keep it "too hot" or "too cold" for growth of spoilage and pathogenic organisms.)

      1. Low temperatures (refrigeration) – lower rate of growth.  Most organisms grow very little or not at all at 0°C. Store perishable foods at low temperatures to slow rate of growth and consequent spoilage (e.g. milk). Low temperatures are not bactericidal. Psychrotrophs, rather than true psychrophiles, are the usual cause of food spoilage in refrigerated foods.

      2. High temperatures – destroy microorganisms.  Heat is the most important and widely used method to destroy microorganisms. For any heat treatment, one must always consider the appropriate (1) type of heat, (2) time of application and (3) temperature to ensure success, as these factors are related to the type of product being heated and the concentration of microorganisms in the product. Endospores of bacteria are considered the most thermoduric of all cells, so any mechanism of heat sterilization must be aimed at them. Other types of spores may show some degree of heat resistance.

        1. Simple boiling.  100°C for 30 minutes. Kills everything except some endospores (Actually, for the purposes of purifying drinking water 100°C for 5 minutes is probably adequate though there have been some reports that Giardia cysts can survive this process). To kill endospores, and therefore sterilize the solution, very long or intermittent boiling is required.

        2. Boiling under pressure.  Autoclaving (steam under pressure or pressure cooker): 121°C for 15 minutes (15#/in2 pressure). This kills endospores. Good for sterilizing almost anything, but heat-labile substances will be denatured or destroyed. The basis of the canning industry is to preserve foods indefinitely by pressure cooking (autoclaving) them in a sealed environment.

        3. Mild heat (pasteurization).  This is a relatively mild heat treatment which is intended to kill disease-causing microorganisms. Although pasteurization uses heat to reduce the total number of microorganisms in milk, its main function as a public health measure is to prevent the transmission of bacteria in milk that may cause disease in humans. The time and temperature depend on killing potential pathogens that are transmitted in milk, i.e., staphylococci, streptococci, Brucella abortus and Mycobacterium tuberculosis. Last-named organism was considered to be the most heat-resistant pathogen associated with milk until Coxiella burnetii (causative agent for Q fever) was found to be more so. Therefore, based on the heat-resistance of C. burnetii, the pasteurization temperatures and times were adjusted accordingly. So, for pasteurization of milk by the batch method: 63°C for 30 minutes; by the flash method: 72°C for 15 seconds.

        4. Heating in microwave.  High-frequency radio waves cause food particles to vibrate and heat is generated. Temperature obtained depends on type and consistency of food.

        5. Dry heat (hot air oven).  160°C for 2 hours or 170°C for 1 hour. Used for glassware, metal, and objects that won't melt.

        6. Incineration.  Incineration burns organisms and physically destroys them. Used for needles , inoculating wires, glassware, etc. and objects not destroyed in the incineration process.

      3. Increased acidity (lower pH).  Acid foods (e.g., yogurt, buttermilk, pickles, sauerkraut) are much less susceptible to spoilage by the microbial acidity (lactic) acid present.

      4. Reduced water activity.  Drying (removal of water by dehydration) or addition of salt or sugar or other solute. Microorganisms in food are associated with the moisture in the food where their nutrients are available and enzymatic activities can take place. Solutes such as salt and sugar, when added to a food, dissolve in the water and can inhibit most microorganisms if the solute concentration in solution is 10% or more. Example: If a food contains 50% water and 5% NaCl, the actual "brine strength" where the microorganisms are growing – i.e., the water – is 10%. Drying increases the brine strength in that the solutes are dissolved in less water. A few spoilage organisms – particularly molds – need very little moisture to be able to grow on aerobic surfaces, and some spoilage organisms may require a high solute concentration in order to grow.

      5. Irradiation.  Irradiation usually destroys or distorts nucleic acids. Ultraviolet light is usually used (commonly used to sterilize the surfaces of objects), although x-rays and microwaves are possibly useful.

      6. Gas.  Formaldehyde, glutaraldehyde, and ethylene oxide are examples of toxic chemicals which are used to kill all forms of life in a specialized gas chamber.

      7. Filtration.  Filtration involves the physical removal (exclusion) of all cells in a liquid or gas, especially important to sterilize solutions which would be denatured by heat (e.g. antibiotics, injectable drugs, amino acids, vitamins, etc.).

      8. Chemical agents (antimicrobial agents).

        1. Antiseptics.  Antiseptics are microbiocidal agents harmless enough to be applied to the skin and mucous membrane; should not be taken internally. Examples: mercurials, silver nitrate, iodine solution, alcohols, detergents.

        2. Disinfectants.  Disinfectants are agents that kill microorganisms, but not necessarily their spores, not safe for application to living tissues; they are used on inanimate objects such as tables, floors, utensils, etc. Examples: chlorine, hypochlorites, chlorine compounds, lye, copper sulfate, quaternary ammonium compounds.

        3. Preservatives.  Preservatives are static agents used to inhibit the growth of microorganisms, most often in foods. If eaten they should be nontoxic. Examples: calcium propionate, sodium benzoate, formaldehyde, nitrate, sulfur dioxide.

        4. Antibiotics.  These are antimicrobial agents produced by microorganisms that kill or inhibit certain other microorganisms. Among the molds, the notable antibiotic producers are Penicillium and Cephalosporium which are the main source of the beta-lactam antibiotics (penicillin and its relatives). In the Bacteria, the Actinomycetes, notably Streptomyces species, produce a variety of types of antibiotics including the aminoglycosides (e.g. streptomycin), macrolides (e.g. erythromycin), and the tetracyclines. Endospore-forming Bacillus species produce polypeptide antibiotics such as polymyxin and bacitracin.

        5. Chemotherapeutic agents.  These are antimicrobial agents of synthetic origin useful in the treatment of microbial or viral disease and used like antibiotics. Examples: sulfonilamides, isoniazid, AZT, chloramphenicol.

GO 
TO:		 Outline for this section.
Farm Microbiology Home Page.
CALS Farm and Industry Short Course Home Page.
Bacteriology Department Web Site.

Page last modified on
3/23/03 at 5:30 PM, CST.
John Lindquist, Dept. of Bacteriology,
University of Wisconsin – Madison