Microbial GrowthCharacterizing Microbes By Optimal Growth TemperatureTemperature Growth Ranges and Food SafetyPhysical Requirements: pHPhysical Requirements: Osmotic PressureSlide 6The Requirements for Growth: Chemical RequirementsSlide 8Slide 9How Toxic Forms of Oxygen Are HandledGrowth in thioglycollate broth reveals oxygen preferencesSlide 12Culture Media: Chemically Defined or ComplexAnaerobic and Low O2 Culture MethodsUnusual Culture MethodsSlide 16Selective MediaDifferential MediaEnrichment MediaSlide 20Slide 21Slide 22Slide 23Estimating Bacterial Numbers by Indirect methodsPlate Assays: Spread Plate or Pour Plate MethodsDirect Measurements of Microbial GrowthSlide 27Slide 28Slide 29Estimating Bacterial Numbers by Indirect MethodsMetabolic Conversion/Enzyme AssayDetermining dry mass of a fixed volumeSlide 33Microbial Growth•Physical Requirements of Microbes•Temperature (optimal enzyme operation)•Psychrophiles, mesophiles, thermophiles•pH (optimal enzyme operation)•Using buffers in media•Molds & yeasts versus bacteria•Chemical Requirements•Carbon source in medium •Nitrogen, sulfur, phosphorous, trace elements•Oxygen requirements•Obligate aerobes, anaerobes, facultative anaerobes•Free radical oxygen (O2-) and H2O2 dangers; superoxide dismutase and catalase = aerobes•Culture Media for Microbes•Chemically defined vs. complex media•Anaerobes: reducing media/Brewer jar•Other: animals, eggs, tissue culture, CO2•Media types•Selective, Differential, Enrichment•Bacterial Population Growth•Growth Curve: Lag, Log, Stationary, Death•Quantifying GrowthCharacterizing Microbes By Optimal Growth TemperatureFigure 6.1Temperature Growth Ranges and Food SafetyFigure 6.2“2-40-140”If > 2 hrs at 40-140oF, don’t eat it!Physical Requirements: pH•Most bacteria grow between pH 6.5 and 7.5•Molds and yeasts grow between pH 5 and 6Physical Requirements: Osmotic Pressure•Hypertonic environments, increase salt or sugar, cause plasmolysis•Extreme or obligate halophiles require high osmotic pressure•Facultative halophiles tolerate high osmotic pressureMicrobial Growth•Physical Requirements of Microbes•Temperature (optimal enzyme operation)•Psychrophiles, mesophiles, thermophiles•pH (optimal enzyme operation)•Using buffers in media•Molds & yeasts versus bacteria•Chemical Requirements•Carbon source in medium •Nitrogen, sulfur, phosphorous, trace elements•Oxygen requirements•Obligate aerobes, anaerobes, facultative anaerobes•Free radical oxygen (O2-) and H2O2 dangers; superoxide dismutase and catalase = aerobes•Culture Media for Microbes•Chemically defined vs. complex media•Anaerobes: reducing media/Brewer jar•Other: animals, eggs, tissue culture, CO2•Media types•Selective, Differential, Enrichment•Bacterial Population Growth•Growth Curve: Lag, Log, Stationary, Death•Quantifying Growth•Carbon•Structural organic molecules, energy source•Chemoheterotrophs use organic carbon sources•Autotrophs use CO2The Requirements for Growth: Chemical Requirements•Nitrogen•In amino acids, proteins•Most bacteria decompose proteins•Some bacteria use NH4+ or NO3•A few bacteria use N2 in nitrogen fixation•Sulfur•In amino acids, thiamine, biotin•Most bacteria decompose proteins•Some bacteria use SO42 or H2S•Phosphorus •In DNA, RNA, ATP, and membranes•PO43 is a source of phosphorus•Trace Elements•Inorganic elements required in small amounts•Usually as enzyme cofactorsThe Requirements for Growth: Chemical RequirementsMicrobial Growth•Physical Requirements of Microbes•Temperature (optimal enzyme operation)•Psychrophiles, mesophiles, thermophiles•pH (optimal enzyme operation)•Using buffers in media•Molds & yeasts versus bacteria•Chemical Requirements•Carbon source in medium •Nitrogen, sulfur, phosphorous, trace elements•Oxygen requirements•Obligate aerobes, anaerobes, facultative anaerobes•Free radical oxygen (O2-) and H2O2 dangers; superoxide dismutase and catalase = aerobes•Culture Media for Microbes•Chemically defined vs. complex media•Anaerobes: reducing media/Brewer jar•Other: animals, eggs, tissue culture, CO2•Media types•Selective, Differential, Enrichment•Bacterial Population Growth•Growth Curve: Lag, Log, Stationary, Death•Quantifying Growth•Singlet oxygen: O2 boosted to a higher-energy state•Handling superoxide free radicals: O22O2- + 2O2- + 8H+  4H2O2 oxygen radicals hydrogen peroxide•Superoxide Dismutase (SODS)•Handling peroxide anion: O22 2H2O2  2H2O + O2 hydrogen peroxide water oxygen gasCatalase (Peroxidase)Catalase Test: Bacteria + H2O2 bubblesHow Toxic Forms of Oxygen Are HandledObligate aerobesFaultative anaerobesObligate anaerobesAerotolerant anaerobesMicroaerophiles Thyoglycollate binds molecular oxygen, reducing it and removing it: R-SH + O2  R-SO2Microbial Growth•Physical Requirements of Microbes•Temperature (optimal enzyme operation)•Psychrophiles, mesophiles, thermophiles•pH (optimal enzyme operation)•Using buffers in media•Molds & yeasts versus bacteria•Chemical Requirements•Carbon source in medium •Nitrogen, sulfur, phosphorous, trace elements•Oxygen requirements•Obligate aerobes, anaerobes, facultative anaerobes•Free radical oxygen (O2-) and H2O2 dangers; superoxide dismutase and catalase = aerobes•Culture Media for Microbes•Chemically defined vs. complex media•Anaerobes: reducing media/Brewer jar•Other: animals, eggs, tissue culture, CO2•Media types•Selective, Differential, Enrichment•Bacterial Population Growth•Growth Curve: Lag, Log, Stationary, Death•Quantifying GrowthCulture Media: Chemically Defined or ComplexTable 6.2 & 6.4Anaerobic and Low O2 Culture MethodsBrewer or anaerobic jarCO2 packetCandle jarUnusual Culture MethodsGrows only in certain cell types: using armadillos to culture M. lepraeGrows only inside live cells: eggs as culture vessels for influenza virusGrows only in certain cell types: using tissue culture with low O2, enriched CO2 incubatorsMicrobial Growth•Physical Requirements of Microbes•Temperature (optimal enzyme operation)•Psychrophiles, mesophiles, thermophiles•pH (optimal enzyme operation)•Using buffers in media•Molds & yeasts versus bacteria•Chemical Requirements•Carbon source in medium •Nitrogen, sulfur, phosphorous, trace elements•Oxygen