Chapter 6 microbial growth
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Chapter 6: Microbial Growth. Microbial Growth. Microbial growth = growth in population Increase in number of cells, not cell size Two main categories of requirements for microbial growth: Physical requirements (environmental conditions) Temperature, pH, osmotic pressure

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Chapter 6: Microbial Growth

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Chapter 6:Microbial Growth


Microbial Growth

  • Microbial growth = growth in population

    • Increase in number of cells, not cell size

  • Two main categories of requirements for microbial growth:

    • Physical requirements (environmental conditions)

      • Temperature, pH, osmotic pressure

    • Chemical requirements


Physical Requirements for Growth: Temperature

  • Temperature

    • Minimum growth temperature

    • Optimum growth temperature

    • Maximum growth temperature

  • Three main classifications

    • Psychrophiles (optimum ~120C)

    • Psychrotrophs (optimum ~230C)

    • Mesophiles (optimum ~370C)

    • Thermophiles (optimum above 500C)


Physical Requirements for Growth: Temperature

Refrigeration

Cause majority of food spoilage

Figure 6.1


Hansen’s Disease(Leprosy)

  • Mycobacterium leprae

  • Optimal growth temperature: 30°C

    • Grows in peripheral nerves, nasal mucosa and skin cells

Figure 22.8


Physical Requirements for Growth: pH

  • pH

    • Most bacteria grow between pH 6.5 and 7.5

    • Molds and yeasts grow optimally between pH 5 and 6

    • Acidophiles grow in acidic environments (pH<5.5)

    • Alkaliphiles grow in basic environments (pH>8.5)

  • Acidic foods (pickles, sauerkraut) preserved by acids from bacterial fermentation

  • Growth media used in the laboratory contain buffers


Physical Requirements for Growth: Osmotic Pressure

  • Osmotic Pressure

    • Hypertonic environments (=high osmotic pressure), increased salt or sugar, cause plasmolysis

      • Obligate halophiles require high osmotic pressure

      • Facultative halophiles tolerate high osmotic pressure(>2% salt)

    • Nutrient agar has a high percentage of water to maintain low osmotic pressure (bacterial cells are 80-90% water)

Low osmotic pressure

High osmotic pressure

Water flow

High solute concentration/

Low water concentration

Low solute concentration/

High water concentration


Physical Requirements for Growth: Osmotic Pressure

  • Plasmolysis: cell growth is inhibited when the plasma membrane pulls away from the cell wall

    • Added salt or sugar is another method of preserving food

Hypertonic solution

(high osmotic pressure)

Isotonic solution

Figure 6.4


Chemical Requirements for Growth

  • Carbon

    • Structural organic molecules, energy source

      • Heterotrophs use organic carbon sources

      • Autotrophs use CO2

  • Nitrogen, Sulfur, Phosphorus

    • For synthesis of amino acids, nucleotides, vitamins, phospholipids

    • Most bacteria decompose proteins to obtain N

    • Inorganic ions are sources for these elements (NH4+, NO3-, PO43-, SO42-)


Chemical Requirements for Growth

  • Trace Elements (Iron, Copper, Zinc)

    • Inorganic elements required in small amounts, usually as enzyme cofactors

    • Often present in tap water

  • Organic Growth Factors

    • Organic compounds obtained from the environment (i.e. the organism cannot synthesize them)

    • Vitamins, amino acids


Chemical Requirements for Growth: Oxygen

  • Oxygen (O2)


Chemical Requirements for Growth: Oxygen

  • Aerotolerance of individual organisms depends on their ability to handle oxygen toxicity

    • Oxygen radical species: O2-, O22-, OH

    • Presence/lack of enzymes that neutralize toxic oxygen species

      • SOD (Superoxide dismutase)

      • Catalase/peroxidase

.


Chemical Requirements for Growth: Oxygen

  • Oxygen (O2)

Express SOD and catalase

Require oxygen, but at lower levels than in the air

Tolerate oxygen (express SOD/catalase) but incapable of using it for growth

Don’t express SOD/catalase


Culture Media

  • Culture Medium: Nutrients prepared for microbial growth

    • Source of energy, carbon, nitrogen, sulfur, phosphorus, trace elements and organic growth factors

  • Sterile: No living microbes

  • Inoculum: Introduction of microbes into medium to initiate growth

  • Culture: Microbes growing in/on culture medium


Culture Media:Agar

  • Complex polysaccharide

  • Used as solidifying agent for culture media in Petri plates, slants, and deeps

  • Generally not metabolized by microbes

    • Agar is not a nutrient

  • Liquefies above 100°C

    • Can incubate at a wide range of temperatures


Culture Media


Anaerobic Culture Media:Broth cultures

  • Reducing broth media

    • Contain chemicals (thioglycollate) that combine with dissolved O2 to deplete it from the media


Anaerobic Culture Methods:Agar Cultures

  • Anaerobic jar

    • Oxygen and H2 combine to form water

Figure 6.5


Culture Media:Selective and Differential Media

  • Selective media: suppress growth of unwanted microbes and encourage growth of desired microbes

  • Differential media: make it easy to distinguish colonies of different microbes

Enterobacter aerogenes on EMB

E. coli on EMB

Figure 6.9b, c


Obtaining Pure Cultures

  • A pure culture contains only one species or strain

  • A colony is a population of cells arising from a single cell or spore or from a group of attached (identical) cells

    • One colony arises from one colony-forming unit (CFU)

  • Specimens (pus, sputum, food) typically contain many different microorganisms

    • Common way to isolate a single species from a mixture of microorganisms: Streak plate method


Streak Plate Method for Isolation of a Pure Species

  • Use loop to pick colony

  • Inoculate broth

  • Pure culture

Figure 6.10a, b


Microbial Growth in Hosts:Biofilms

Microbial communities

3-dimensional “slime”

i.e. dental plaque, soap scum

Share nutrients

Sheltered from harmful factors

Cell-to-cell communication: quorum sensing

Figure 6.5

Bacterial biofilm growing on a micro-fibrous material


Microbial Growth in Hosts:Biofilms & Quorum Sensing

  • Quorum sensing allows a form of bacterial communication

    • Individual cells can sense the accumulation of signaling molecules (autoinducers)

      • Informs individual cells about surrounding cell density

      • May change the behavior (gene expression) of individual cells

        • Results in a coordinated response by the whole population

http://biofilmbook.hypertextbookshop.com/public_version/


Prokaryotic Reproduction:Binary Fission

Figure 6.11


Reproduction in Prokaryotes:Generation Time

Generation time: the time required for one population doubling

  • Varies with species and environmental conditions


Reproduction in Prokaryotes:Generation Number

  • Generation number: the number of times a cell population has doubled in a given time under given conditions

Figure 6.12b


Reproduction in Prokaryotes:Growth Plot

Logarithmic

Arithmetic

Figure 6.13


Bacterial Growth Curve

  • Lag: little/no cell division

    • Adapting to new medium

    • *Metabolically active*

  • Log: exponential growth

    • Most metabolically active

    • Gen. time at constant minimum

  • Stationary: equilibrium phase

    • Growth rate = death rate

    • Nutrients exhausted, waste accumulation, pH changes

  • Death: logarithmic decline

Figure 6.14


Measuring Microbial Growth

  • To determine the size of a bacterial population in a specimen, cell counting techniques are used

    • Often there are too many cells per ml or gram of specimen…

      • A small proportion of the specimen (a dilution) is counted

      • The number of cells in the original specimen can be calculated based on the count in the small dilution


Direct Measurements of Microbial Growth:Viable Cell Count

  • Plate Counts: Perform serial dilutions of a sample

  • How many cells are in 1 mL of original culture?

DF=1

10-5

DF:

10-3

10-4

10-2

10-1

Figure 6.15, top portion


Direct Measurements of Microbial Growth:Viable Count

  • Inoculate one agar plate with each serial dilution

Figure 6.16


Direct Measurements of Microbial Growth:Viable Count

  • After incubation, count colonies on plates that have 30-300 colonies (CFUs)

Figure 6.15


Direct Measurements of Microbial Growth

  • Filtration

    • Ideal when microbial density is low in a sample

Figure 6.17a, b


Direct Measurements of Microbial Growth

Disadvantages:

-Likely to count dead cells

-Motile cells can be difficult to count

Figure 6.19


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