Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Lecture prepared by Mindy Miller-Kittrell, University of Tennessee, Knoxville M I C R O B I O L O G Y WITH DISEASES BY BODY SYSTEM SECOND EDITION Chapter 6 Microbial Nutrition and Growth

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Microbial growth – an increase in a population of microbes rather than an increase in size of an individual Result of microbial growth is discrete colony – an aggregation of cells arising from single parent cell Reproduction results in growth

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Organisms use a variety of nutrients for their energy needs and to build organic molecules and cellular structures Most common nutrients – those containing necessary elements such as carbon, oxygen, nitrogen, and hydrogen Microbes obtain nutrients from variety of sources

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT TABLE 6.1]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Sources of carbon, energy, and electrons – Two groups of organisms based on source of carbon – Autotrophs use an inorganic source of carbon (carbon dioxide) – Heterotrophs catabolize reduced organic molecules (proteins, carbohydrates, amino acids, and fatty acids) – Two groups of organisms based on use of chemicals or light as source of energy – Chemotrophs acquire energy from redox reactions involving inorganic and organic chemicals – Phototrophs use light as their energy source

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT FIGURE 6.1]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Oxygen requirements – Oxygen is essential for obligate aerobes (final electron acceptor in ETC) – Oxygen is deadly for obligate anaerobes – How can this be true? – Neither gaseous O 2 nor oxygen covalently bound in compounds is poisonous – Rather the forms of oxygen that are toxic are highly reactive and also excellent oxidizing agents – Resulting chain of oxidations causes irreparable damage to cells by oxidizing compounds such as proteins and lipids

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Oxygen requirements – Four toxic forms of oxygen 1. Singlet oxygen – molecular oxygen with electrons boosted to higher energy state – Occurs during photosynthesis so phototropic organisms have carotenoids that remove the excess energy of singlet oxygen 2. Superoxide radicals – some form during incomplete reduction of oxygen in aerobic and anaerobic respiration – So reactive that aerobes produce superoxide dismutases to detoxify them – Anaerobes lack superoxide dismutase and die as a result of oxidizing reactions of superoxide radicals formed in presence of oxygen

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Oxygen requirements – Four toxic forms of oxygen 3. Peroxide anion – formed during reactions catalyzed by superoxide dismutase and other reactions – Aerobes contain either catalase or peroxidase to detoxify peroxide anion – Obligate anaerobes either lack both enzymes or have only a small amount of each 4. Hydroxyl radical – results from ionizing radiation and from incomplete reduction of hydrogen peroxide – The most reactive of the four toxic forms of oxygen – Not a threat to aerobes due to action of catalase and peroxidase

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT FIGURE 6.2]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Oxygen requirements – Aerobes – undergo aerobic respiration – Anaerobes – do not use aerobic metabolism – Facultative anaerobes – can maintain life via fermentation or anaerobic respiration or by aerobic respiration – Aerotolerant anaerobes – do not use aerobic metabolism but have some enzymes that detoxify oxygen’s poisonous forms – Microaerophiles – aerobes that require oxygen levels from 2 to 10% and have a limited ability to detoxify hydrogen peroxide and superoxide radicals

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT FIGURE 6.3]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Nitrogen requirements – Anabolism often ceases due to insufficient nitrogen needed for proteins and nucleotides – Nitrogen acquired from organic and inorganic nutrients, plus all cells recycle nitrogen from amino acids and nucleotides – The reduction of nitrogen gas to ammonia (nitrogen fixation) by certain bacteria is essential to life on Earth because nitrogen is made available in a usable form

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Nutrients: Chemical and Energy Requirements – Other chemical requirements – Phosphorus is a component of phospholipid membranes, DNA, RNA, ATP, and some proteins – Sulfur is a component of sulfur-containing amino acids, disulfide bonds critical to tertiary structure of proteins, and in vitamins (thiamin and biotin) – Trace elements – only required in small amounts; usually found in sufficient quantities in tap water – Growth factors – necessary organic chemicals that cannot be synthesized by certain organisms (vitamins, certain amino acids, purines, pyrimidines, cholesterol, NADH, and heme)

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT TABLE 6.2]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Physical Requirements – Temperature – Effect of temperature on proteins – Effect of temperature on lipid-containing membranes of cells and organelles – If too low, membranes become rigid and fragile – If too high, membranes become too fluid and cannot contain the cell or organelle

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT FIGURE 6.4]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT FIGURE 6.5]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements [INSERT FIGURE 6.6]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Physical Requirements – pH – Organisms sensitive to changes in acidity because H + and OH - interfere with H bonding in proteins and nucleic acids – Neutrophiles are bacteria and protozoa that grow best in a narrow range around neutral pH ( ) – Acidophiles are bacteria and fungi that grow best in acidic habitats – Acidic waste products can help preserve foods by preventing further microbial growth – Alkalinophiles live in alkaline soils and water up to pH 11.5

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Physical Requirements – Physical effects of water – Microbes require water to dissolve enzymes and nutrients required in metabolism – 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 in a dry environment for years – Two physical effects of water – Osmotic pressure – Hydrostatic pressure

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Physical Requirements – Physical effects of water – Osmotic pressure – The pressure exerted on a semipermeable membrane by a solution containing solutes that cannot freely cross membrane; related to concentration of dissolved molecules and ions in a solution – Hypotonic solutions have lower solute concentrations; cells placed in these solutions will swell and burst – Hypertonic solutions have greater solute concentrations; cells placed in these solutions will undergo crenation – This effect helps preserve some foods – Restricts organisms to certain environments – Obligate halophiles – grow in up to 30% salt – Facultative halophiles – can tolerate high salt concentrations

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Physical Requirements – Physical effects of water – Hydrostatic pressure – Water exerts pressure in proportion to its depth - For every additional 10 m of depth, water pressure increases 1 atm – Organisms that live under extreme pressure are barophiles - Their membranes and enzymes depend on this pressure to maintain their three-dimensional, functional shape

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Associations and Biofilms – Organisms live in association with different species – Antagonistic relationships – Synergistic relationships – Symbiotic relationships

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth Requirements Associations and Biofilms – Biofilms – Complex relationships among numerous microorganisms – Develop an extracellular matrix that adheres cells to one another, allows attachment to a substrate, sequesters nutrients, and may protect individuals in the biofilm – Form on surfaces often as a result of quorum sensing – Many microorganisms become more harmful when part of a biofilm

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms Inoculum introduced into medium (broth or solid) – Environmental specimens – Clinical specimens – Stored specimens Culture – refers to act of cultivating microorganisms or the microorganisms that are cultivated

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.7]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT TABLE 6.3]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms Obtaining Pure Cultures – Cultures composed of cells arising from a single progenitor – Progenitor is termed a CFU – Aseptic technique is used to prevent contamination of sterile substances or objects – Two common isolation techniques – Streak plates – Pour plates

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.8]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.9]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms Culture Media – Majority of prokaryotes have never been grown in culture medium – Six types of general culture media – Defined media – Complex media – Selective media – Differential media – Anaerobic media – Transport media

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.10]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT TABLE 6.4]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.11]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.12]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.13]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT TABLE 6.5]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.14]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms [INSERT FIGURE 6.15]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms Special Culture Techniques – Techniques developed for culturing microorganisms – Animal and cell culture – Low-oxygen culture – Enrichment culture

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Culturing Microorganisms Preserving Cultures – Refrigeration – stores for short periods of time – Deep-freezing – stores for years – Lyophilization – stores for decades

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.17]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations Generation Time – The time required for a bacterial cell (or population of cells) to grow and divide – Dependent on chemical and physical conditions

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.19]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.20]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations Measuring Microbial Growth – Direct methods – Viable plate counts – Membrane filtration – Microscopic counts – Electronic counters

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.21]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.22]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.24]

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations Measuring Microbial Growth – Indirect methods – Metabolic activity – Dry weight – Turbidity

Copyright © 2009 Pearson Education Inc., publishing as Pearson Benjamin Cummings Growth of Microbial Populations [INSERT FIGURE 6.25]