BIO 326R 1st Edition Lecture 5 Outline of Last Lecture I. Gram Negativea. Periplasmb. Braun’s lipoproteinsc. PorinsII. Growth of Bacterial Cella. Growth mediab. Inoculumc. Binary fissionIII. Measuring Growtha. Absorptionb. Viable countsc. Direct countingOutline of Current Lecture I. Measuring Growth (contd from last lecture)a. Dry WeightII. Growth in a closed systema. Growth curvei. Lag, log, stationary, deathb. Doubling timeThese notes represent a detailed interpretation of the professor’s lecture. GradeBuddy is best used as a supplement to your own notes, not as a substitute.c. Closed system vs. continuous cultured. Persister cellse. VBNCIII. Environmental Factorsa. Temperatureb. Osmotic pressurec. Atmospherei. Aerobic, anaerobic, facultative anaerobes, microanerobesCurrent LectureI. Measuring Growth (continued)- Dry weight= spin out bacteria in centrifuge to dry, weigh to get count- Problem with this:o No differentiation between live and dead and is cumbersomeII. Growth in a Closed System- Growth curve phaseso Lag= metabolically active, but not much cell divisiono Log/exponential phase= lots of cell division Balanced growth= all in about the same state (RNA, growing actively, etc.-- Consistent doubling time, similar cell composition)o Stationary phase= level off of growth due to increase in waste, lack of space, less available nutrients Growth rate= death rate Most bugs in nature are in this phaseo Death phase= toxic waste build-up, no nutrients available Death>growth May not see this phase in OD—not because it isn’t happening, but because OD does not differentiation between dead and live if the dead cells aren’t lysing- Doubling time= time it takes for population to doubleo E. coli in defined medium= 50 minutes and in complex medium= 20 minutes Defined= minimal nutrients (sugar and salt, bacteria need to synthesize the rest) Complex= complete nutrients needed (bacteria do not have to synthesize anything)o Formulas to calculate generation time N=( 2^n)(N0)o N= number of cells, n= number of generations, N0 =number of cells initial N= (logN- logN0 )/0.301o 0.301 is log2 which related to the 2^n in the previous equation G= t/no T= time, n= number of generations, g= generation time- Closed system vs. continuouso Closed system= batch culture No addition of new nutrients during the experimento Continuous culture= chemostat Constantly add nutrients and removing waste and old cells—always in exponential phase Used typically for longterm studies- Death phase almost never goes to zero persister cellso Persister cells= hang on, hard to kill, very resistant to antibioticso VBNC= viable but non culturable living, not dividingIII. Environmental Factors- Temperatureo Psychrophiles= survive at <15 degrees Co Mesophiles= survive at 20-45 degrees Co Thermophiles= survive at >45 degrees C- Osmotic pressure: salt can inhibit cell growth by causing:o Decreased water availability, decreased turgor pressure Water leaves cell, cell shrinkso Reason why people salted meats when the did not have refrigeration Preserved meat by drying it and inhibiting bacteria growth- atmosphereo aerobic= oxygen neededo anaerobic= no oxygen needed, sometimes can’t survive with oxygeno facilitative anaerobes= can switch between aerobic and anaerobic depending on the availability of oxygeno microanaerophiles= too much oxygen is toxic, but some is needed to
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