Microbiology Textbook - Ch.6 Microbial Growth PDF

CHAPTER 6 Microbial Nutrition and Growth  Growth of Microbial Organisms  Microbial ‘growth’  Increase in a population of microbes  Due to reproduction of individual microbes  Results of microbial growth  Discrete colony – an aggregation of cells arising from singl...

CHAPTER 6 Microbial Nutrition and Growth  Growth of Microbial Organisms  Microbial ‘growth’  Increase in a population of microbes  Due to reproduction of individual microbes  Results of microbial growth  Discrete colony – an aggregation of cells arising from single parent cell  Biofilm – collection of microbes living on a surface in a complex community Microbial Growth Requirements  Oxygen Requirements  Oxygen in O2 and H2O is non-toxic  Highly reactive oxygen affects protein/enzyme/lipids  Oxygen will steal electrons from compounds  Oxidizing agents  Four toxic forms of oxygen  Singlet oxygen (1O2)  Superoxide radicals (O2-)  Peroxide anion (O22-)  Hydroxyl radical (OH.) Growth Requirements  Oxygen Requirements  Many organisms can live in various O2 concentrations  Obligate Aerobes * oxygen is essential; as final e- acceptor  Obligate Anaerobes * oxygen is deadly; highly reactive O2 is toxic  Facultative anaerobes or facultative aerobes * with or without oxygen  Aerotolerant anaerobes * do not use oxygen, but tolerate it  Microaerophiles * low % of oxygen required Growth Requirements  Figure 6.2 Using a liquid thioglycolate growth medium to identify the oxygen requirements of organisms Growth Requirements Growth Requirements  Other Chemical Requirements  Carbon (autotrophs, heterotrophs)  Nitrogen  Required for amino acids, nucleotide bases  Phosphorus (nucleic acids)  Sulfur (protein folding)  Trace elements  Only required in small amounts  Growth factors  Necessary organic chemicals that cannot be synthesized by certain organisms Growth Requirements Growth Requirements  Temperature  Affects three-dimensional structure of proteins  Lipid-containing membranes of cells and organelles are temperature sensitive  If too low (cold), membranes become rigid and fragile  If too high (warmer), membranes become too fluid  Temperature and metabolic activity:  Minimum growth temp  Maximum growth temp  Optimum growth temp Physical Growth Requirements  Figure 6.3 The effects of temperature on microbial growth for E. coli (its temperature range), incubated for 18 hrs Physical Growth Requirements  Figure 6.4 FIVE CATEGORIES OF MICROBES based on temperature ranges for growth Physical Growth Requirements  pH  Organisms are sensitive to changes in acidity  Neutrophiles * Grow best in a narrow range around neutral pH [6.5 - 7.5]  Acidophiles * Grow best in acidic habitats * Also obligate acidophiles and acid-tolerant microbes  Alkalinophiles * Live in alkaline soils and water (i.e. Vibrio cholerae) Physical Growth Requirements  Water Requirements & Growth  Microbes require water to dissolve enzymes & nutrients  Important reactant in many metabolic reactions  Most cells die in absence of water  Endospores and cysts cease most metabolic activity  Two effects of water:  Osmotic pressure  Hydrostatic pressure Physical Growth Requirements  Osmotic Pressure  The pressure exerted on a semipermeable membrane by a solution containing solutes that cannot freely cross the membrane  Hypotonic solutions have lower solute concentrations  Hypertonic solutions have greater solute concentrations  Restricts organisms to certain environments  Obligate halophiles (require high salt conc)  Facultative halophiles (tolerate high salt concentration) * i.e Staphylococcus aureus Effects of Water – Physical Growth Requirements  Hydrostatic Pressure  Water exerts pressure in proportion to its depth  Barophiles live under extreme hydrostatic pressure  Membranes and enzymes depend on pressure to maintain their three-dimensional, functional shape  Unable to cause disease in organisms that do not live at similar depths Effects of Water – Physical Growth Requirements  Microbial Associations  Organisms live in association with different species and *do* harm:  Antagonistic relationships  Organisms live in association with different species and *do not* harm:  Synergistic relationships  Symbiotic relationships Physical Growth Requirements  Biofilms  Primary residence of MOs in nature  Form on surfaces, medical devices, rocks  Form as a result of quorum sensing  Complex relationships among numerous MOs  Many MOs are more harmful as part of a biofilm  Scientists seeking ways to prevent biofilm formation Physical Growth Requirements  Figure 6.6 Biofilm development Physical Growth Requirements  Figure 6.7 Quorum sensing  Prevention:  Block signal receptors  Use enzymes  Artificially amplify quorum sensing Physical Growth Requirements  Culturing Microorganisms  Inoculum (sample) is introduced into medium  The growth that results is a culture  Collected samples:  Environmental specimens  Stored specimens  Clinical specimens * Suspected pathogen is taken from patient, cultured, and ID’d * Standard precautions must be followed o Not to contaminate the sample o Not to infect themselves Culturing MOs Culturing MOs  Obtaining Pure Cultures  Pure cultures are composed of cells arising from a single progenitor (aka - axenic)  Progenitor is termed a colony-forming unit (CFU).  Aseptic technique prevents contamination of sterile substances or objects  Two common isolation techniques:  Streak plates  Pour plates Culturing MOs  Figure 6.9 The streak-plate method of isolation: A sterile inoculating loop is used to spread inoculum across the solid surface of growth media CFUs are isolated E.coli – on blood agar Culturing MOs  Figure 6.10 The pour-plate method of isolation Culturing MOs  Figure 6.8 Characteristics of bacterial colonies  Serratia marcescens * Circular, entire, convex, large, smooth, shiny, red, and opaque Culturing MOs  Obtaining Pure Cultures  Other isolation techniques:  Some fungi can be isolated with streak and pour plates  Protozoa and motile unicellular algae * isolated through dilution of broth cultures  Large microorganisms * Can individually pick a single cell and use that cell to establish a culture Acanthamoeba spp. Trophozoites https://www.cdc.gov/dpdx/freelivingamebic/index.html Culturing MOs  Culture Media  Variety of liquid and solid media are used to culture microbes  Nutrient broth * A common liquid medium * Beef extract and peptones  Agar * A common addition to make media solid * Used to make Petri plates and slant tubes Figure 6.11 Slant tubes containing solid media (citrate agar medium) Culture Media  Culture Media  Six types of general culture media:  Defined (synthetic)  Complex  Selective  Differential  Anaerobic  Transport https://bit.ly/2MKJvJe Culture Media Culture Media  Culture Media  Complex media  Exact chemical composition is unknown  Nutrients commonly derived from breakdown of yeast, beef, soy, and proteins  Supports growth of a wide variety of microorganisms  Useful when nutritional needs of an organism are unknown  Selective media  Contain substances that favor or inhibit growth of particular microorganisms Culture Media  Figure 6.12 An example of the use of a selective medium with a soil sample: media on right is selective for fungi (acidic pH) Culture Media  Differential Media  Visible changes in the media or visible changes in the colonies  Microbes can utilize nutrients in different ways Figure 6.13: The use of blood agar as a differential medium Culture Media – Differential  Figure 6.14 The use of carbohydrate utilization tubes as differential media Culture Media Culture Media  Figure 6.15 The use of MacConkey agar as a selective and differential medium Culture Media Proteus vulgaris Salmonella Staphylococcus aureus MacConkey’s Agar – explain these results MacConkey’s Agar – explain these results  Mannitol Salt Agar (MSA): Selective and Differential Medium What does this media tell us? How is it Selective? How is it Differential?  Figure 6.16 An anaerobic culture system MO must be able to survive brief oxygen exposure Methylene blue turns colourless in the absence of oxygen Culture Media  Transport Media  Used by health care personnel to ensure clinical specimens are not contaminated and to protect people from infection  Rapid transport of samples is important  Blood, saliva, feces, urine  In buffered media  If anaerobic specimen, in a syringe for less than an hour, or use an anaerobic transport chamber Culture Media  Preserving Cultures  Refrigeration  Stores for short periods of time  Deep-freezing (-50oC to -95oC)  Stores for years  Lyophilization  Stores for decades https://bit.ly/2wIBkDB Preserving Cultures  Growth of Microbial Populations  Most microorganisms reproduce by binary fission  One cell divides in half to produce two daughter cells  Involves four steps Step 1: Step 4: Step 2: Step 3: Growth of Microbial Populations  Generation Time  Time required for a bacterial cell to grow and divide  Time required for a population of cells to double in number  Generation time varies among populations  Dependent on chemical and physical conditions  Some bacteria have a generation time of 20 minutes, for others it may be 10 days Growth of Microbial Populations  Figure 6.20 A typical microbial growth curve Growth of Microbial Populations  Bacteria Have Four Distinct Growth Phases Growth of Microbial Populations  Continuous Culture in a Chemostat  A chemostat is used to maintain a microbial population in a particular phase of growth  Open system  Requires addition of fresh medium and removal of old medium  Used in several industrial settings Figure 6.21 Schematic of chemostat Growth of Microbial Populations  Estimating Microbial Numbers  Estimating the number of microorganisms is useful  Determine severity of certain infections  Determine effectiveness of food preservation techniques  Measure the degree of contamination of water supplies  Evaluate disinfectants and antibiotics Measuring Population Growth