05_Bacterial_Culture_Growth_2.pdf

CHAPTER 4 Bacterial Culture, Growth, and Development Copyright © 2024 by W. W. Norton & Company, Inc. Learning Objectives Know the basic techniques to culture and count microbes Know the different types of media Analyze a bacterial growth curve to conclude its stages of growth 2 Agar plate culture m...

CHAPTER 4 Bacterial Culture, Growth, and Development Copyright © 2024 by W. W. Norton & Company, Inc. Learning Objectives Know the basic techniques to culture and count microbes Know the different types of media Analyze a bacterial growth curve to conclude its stages of growth 2 Agar plate culture made from swabbing patient elevator buttons and escalator rails at a university hospital. 3 4.3 Culturing and Counting Bacteria § Microbes in nature exist in complex, multispecies communities, but for detailed studies they must be grown separately in pure culture. § After almost 140 years of trying, we have succeeded in culturing less than 1% of the microorganisms around us. § The Great Plate Count Anomaly 4 The vast majority has yet to be tamed, why? Very complex, narrow growth requirements Nutrients Environmental conditions Mandatory symbioses Intracellular growth Extremely long growth times 5 Bacteria Are Grown in Culture Media § Bacteria are grown in culture media, which are of two main types: 1. Liquid or broth – Useful for studying the growth characteristics of a pure culture 6 Bacteria Are Grown in Culture Media § Bacteria are grown in culture media, which are of two main types: 1. Liquid or broth – Useful for studying the growth characteristics of a pure culture 2. Solid (usually gelled with agar) – Useful for trying to separate mixed cultures from clinical specimens or natural environments 7 Isolation/Dilution Streaking Serial Dilutions and Spread Plates Each colony on an agar plate represents one viable organism (or colony-forming unit; CFU) present in the original liquid culture. Dilution Streaking and Spread Plates Types of Media § Complex media are nutrient rich but poorly defined. § Grows most chemoheterotrophic organisms § Contains components with variable nutrient content § For example, animal products 11 Types of Media § Minimal defined (synthetic) media contain only those nutrients that are essential for growth of a given microbe. § Used to stringently control growth conditions § Can be difficult to determine § Ability to know EXACT chemical make-up 12 When might you use complex vs. synthetic media? 13 Types of Media § Enriched media are complex media to which specific high nutrient components are added. § For example, egg yolks or blood serum 14 Types of Media § Selective media favor the growth of one organism over another to isolate types of microbes. § E.g., Eosin Methylene Blue 15 Types of Media § Differential media exploit differences between two species that grow equally well to aid in identification. § E.g., Mannitol Salt Agar 16 Types of Media § Several media used in clinical microbiology are both selective and differential. E.g., MacConkey medium, Mannitol salt agar 17 Growth Factors and Uncultured Microbes § Growth factors are specific nutrients not required by other species. A microbe needs them in order to grow in laboratory media. e.g., Streptococcus pyogenes requires glutamic acid and alanine because it can no longer synthesize them. Why does S. pyogenes no longer synthesize these necessary nutrients? 18 Growth Factors and Uncultured Microbes § Microbe growth factors depend on the nutrient profile of their natural ecological niche. § The specific growth factors needed can change over time as organisms evolve. 19 Some species have adapted so well to their natural habitats that we still do not know how to grow them in the lab. § But, some ingenious ways can help us isolate new organisms! 20 Techniques for Counting Bacteria Viable plate counts 21 Techniques for Counting Bacteria Live/Dead Staining 22 Techniques for Monitoring Growth Dynamics Optical Density What is a limitation to this method? 23 4.4 The Growth Cycle § Most bacteria divide by binary fission, where one parent cell splits into two equal daughter cells. § Generation time, or doubling time, is the time it takes for a population to double. 24 Exponential Growth § The growth rate, or rate of increase in cell numbers, is proportional to the population size at a given time. Population: 100 -> 200 -> 400 -> 800 -> 1600 -> 3200... 25 Exponential Growth § The growth rate, or rate of increase in cell numbers, is proportional to the population size at a given time. “Exponential” because it generates an exponential curve, a curve whose slope increases continually. Exponential growth doesn’t typically last forever, what can stop this growth rate? 26 Stages of Growth in a Batch Culture § The simplest way to model the effects of a changing environment is to culture bacteria in a batch culture. A liquid medium within a closed system § The changing conditions in this system greatly affect bacterial physiology and growth. 27 Bacterial Growth Curves 28 Bacterial Growth Curves 29 Bacterial Growth Curves 30 Bacterial Growth Curves 31 Continuous Culture § In a continuous culture, all cells in a population achieve a steady state § Chemostats ensures logarithmic growth by constantly adding and removing equal amounts of culture medium 32