Microbial Nutrition and Growth Chapter 6 Microbial Nutrition and Growth

Result of microbial growth is discrete colony Growth Requirements Microbial growth Increase in a population of microbes Result of microbial growth is discrete colony An aggregation of cells arising from single parent cell Reproduction results in growth © 2012 Pearson Education Inc. 2

Microbes obtain nutrients from variety of sources Growth Requirements Organisms use a variety of nutrients for their energy needs and to build organic molecules and cellular structures Most common nutrients contain necessary elements such as carbon, oxygen, nitrogen, and hydrogen Microbes obtain nutrients from variety of sources © 2012 Pearson Education Inc. 3

Nutrients: Chemical and Energy Requirements Growth Requirements Nutrients: Chemical and Energy Requirements Sources of carbon, energy, and electrons Two groups of organisms based on source of carbon Autotrophs Heterotrophs Two groups of organisms based on source of energy Chemotrophs Phototrophs © 2012 Pearson Education Inc. 4

Figure 6.1 Four basic groups of organisms

Nutrients: Chemical and Energy Requirements Growth Requirements Nutrients: Chemical and Energy Requirements Oxygen requirements Oxygen is essential for obligate aerobes Oxygen is deadly for obligate anaerobes How can this be true? Toxic forms of oxygen are highly reactive and excellent oxidizing agents Resulting oxidation causes irreparable damage to cells © 2012 Pearson Education Inc. 6

Nutrients: Chemical and Energy Requirements Growth Requirements Nutrients: Chemical and Energy Requirements Oxygen requirements Four toxic forms of oxygen Singlet oxygen Superoxide radicals Peroxide anion Hydroxyl radical © 2012 Pearson Education Inc. 7

Figure 6.2 Catalase test

Nutrients: Chemical and Energy Requirements Growth Requirements Nutrients: Chemical and Energy Requirements Oxygen requirements Aerobes Anaerobes Facultative anaerobes Aerotolerant anaerobes Microaerophiles © 2012 Pearson Education Inc. 9

Figure 6.3 Oxygen requirements of organisms-overview

Nutrients: Chemical and Energy Requirements Growth Requirements Nutrients: Chemical and Energy Requirements Nitrogen requirements Anabolism often ceases because of insufficient nitrogen Nitrogen acquired from organic and inorganic nutrients All cells recycle nitrogen from amino acids and nucleotides Nitrogen fixation by certain bacteria is essential to life on Earth © 2012 Pearson Education Inc. 11

Nutrients: Chemical and Energy Requirements Growth Requirements Nutrients: Chemical and Energy Requirements Other chemical requirements Phosphorus Sulfur Trace elements Required only in small amounts Growth factors Necessary organic chemicals that cannot be synthesized by certain organisms © 2012 Pearson Education Inc. 12

Physical Requirements Growth Requirements Physical Requirements Temperature Effect of temperature on proteins Effect of temperature on membranes of cells and organelles If too low, membranes become rigid and fragile If too high, membranes become too fluid © 2012 Pearson Education Inc. 13

Figure 6.4 Microbial growth-overview

Thermophiles Mesophiles Hyperthermophiles Growth rate Psychrophiles Figure 6.5 Four categories of microbes based on temperature ranges for growth Thermophiles Mesophiles Hyperthermophiles Growth rate Psychrophiles Temperature (°C)

Figure 6.6 An example of psychrophile-overview

Physical Requirements Growth Requirements Physical Requirements pH Organisms are sensitive to changes in acidity H+ and OH– interfere with H bonding Neutrophiles grow best in a narrow range around neutral pH Acidophiles grow best in acidic habitats Alkalinophiles live in alkaline soils and water © 2012 Pearson Education Inc. 17

Physical Requirements Growth Requirements Physical Requirements Physical effects of water Microbes require water to dissolve enzymes and nutrients Water is important reactant in many metabolic reactions Most cells die in absence of water Some have cell walls that retain water Endospores and cysts cease most metabolic activity Two physical effects of water Osmotic pressure Hydrostatic pressure © 2012 Pearson Education Inc. 18

Physical Requirements Growth Requirements Physical Requirements Physical effects of water Osmotic pressure Pressure exerted on a semipermeable membrane by a solution containing solutes that cannot freely cross membrane Hypotonic solutions have lower solute concentrations Hypertonic solutions have greater solute concentrations Restricts organisms to certain environments Obligate and facultative halophiles © 2012 Pearson Education Inc. 19

Physical Requirements Growth Requirements Physical Requirements Physical effects of water Hydrostatic pressure Water exerts pressure in proportion to its depth Barophiles live under extreme pressure Their membranes and enzymes depend on pressure to maintain their shape © 2012 Pearson Education Inc. 20

Associations and Biofilms Growth Requirements Associations and Biofilms Organisms live in association with different species Antagonistic relationships Synergistic relationships Symbiotic relationships © 2012 Pearson Education Inc. 21

Associations and Biofilms Growth Requirements Associations and Biofilms Biofilms Complex relationships among numerous microorganisms Develop an extracellular matrix Adheres cells to one another Allows attachment to a substrate Sequesters nutrients May protect individuals in the biofilm Form on surfaces often as a result of quorum sensing Many microorganisms more harmful as part of a biofilm © 2012 Pearson Education Inc. 22

Figure 6.7 Plaque (biofilm) on a human tooth

Culturing Microorganisms Inoculum introduced into medium Environmental specimens Clinical specimens Stored specimens Culture Act of cultivating microorganisms or the microorganisms that are cultivated © 2012 Pearson Education Inc. 24

Figure 6.8 Characteristics of bacterial colonies-overview

Culturing Microorganisms Obtaining Pure Cultures Cultures composed of cells arising from a single progenitor Progenitor is termed a CFU Aseptic technique prevents contamination of sterile substances or objects Two common isolation techniques Streak plates Pour plates © 2012 Pearson Education Inc. 26

Figure 6.9 Streak plate method of isolation-overview

Figure 6.10 Pour plate method of isolation-overview

Culturing Microorganisms Culture Media Majority of prokaryotes have not been grown in culture medium Six types of general culture media Defined media Complex media Selective media Differential media Anaerobic media Transport media © 2012 Pearson Education Inc. 29

Figure 6.11 Slant tube containing solid media Butt

Figure 6.12 An example of the use of a selective medium Bacterial colonies Fungal colonies pH 7.3 pH 5.6

Figure 6.13 The use of blood agar as a differential medium Beta-hemolysis Alpha-hemolysis No hemolysis (gamme-hemolysis)

Acid fermentation with gas Figure 6.14 The use of carbohydrate utilization tubes as differential media Durham tube (inverted tube to trap gas) No fermentation Acid fermentation with gas

Figure 6.15 Use of MacConkey agar as a selective and differential medium-overview

Figure 6.16 An anaerobic culture system Clamp Airtight lid Chamber Palladium pellets to catalyze reaction removing O2 Envelope containing chemicals to release CO2 and H2 Methylene blue (anaerobic indicator) Petri plates

Culturing Microorganisms Special Culture Techniques Techniques developed for culturing microorganisms Animal and cell culture Low-oxygen culture Enrichment culture © 2012 Pearson Education Inc. 36

Culturing Microorganisms Preserving Cultures Refrigeration Stores for short periods of time Deep-freezing Stores for years Lyophilization Stores for decades © 2012 Pearson Education Inc. 37

Growth of Microbial Populations ANIMATION Bacterial Growth: Overview © 2012 Pearson Education Inc. 38

Growth of Microbial Populations ANIMATION Binary Fission © 2012 Pearson Education Inc. 39

Figure 6.17 Binary fission events-overview

Figure 6.18 Comparison of arithmetic and logarithmic growth-overview

Growth of Microbial Populations Generation Time Time required for a bacterial cell to grow and divide Dependent on chemical and physical conditions © 2012 Pearson Education Inc. 42

Figure 6.19 Two growth curves of logarithmic growth-overview

Figure 6.20 Typical microbial growth curve Stationary phase Death (decline) phase Log (exponential) phase Number of live cells (log) Lag phase Time

Growth of Microbial Populations ANIMATION Bacterial Growth Curve © 2012 Pearson Education Inc. 45

Figure 6.21 Schematic of chemostat Fresh medium with a limiting amount of a nutrient Flow-rate regulator Sterile air of other gas Culture vessel Culture Overflow tube

Growth of Microbial Populations Measuring Microbial Reproduction Direct methods Serial dilution and viable plate counts Membrane filtration Most probable number Microscopic counts Electronic counters © 2012 Pearson Education Inc. 47

Figure 6.22 Estimating microbial population size-overview

Figure 6.23 Use of membrane filtration to estimate microbial population-overview

Inoculate 1.0 ml into each of 5 tubes Figure 6.24 The most probable number (MPN) method for estimating microbial numbers 1.0 ml 1.0 ml Undiluted 1:10 1:100 Inoculate 1.0 ml into each of 5 tubes Phenol red, pH color indicator, added Incubate Results 4 tubes positive 2 tubes positive 1 tube positive

Figure 6.25 The use of a cell counter for estimating microbial numbers-overview

Growth of Microbial Populations Measuring Microbial Growth Indirect methods Metabolic activity Dry weight Turbidity © 2012 Pearson Education Inc. 52

Figure 6.26 Spectrophotometry-overview

Growth of Microbial Populations Measuring Microbial Reproduction Genetic methods Isolate DNA sequences of unculturable prokaryotes Used to estimate the number of these microbes © 2012 Pearson Education Inc. 54