Microbial Nutrition, Ecology, and Growth

Transcription

1 Chapter 7 Microbial Nutrition, Ecology, and Growth

2 Microbial Nutrition Nutrition process by which chemical substances (nutrients) are acquired from the environment and used in cellular activities Essential nutrients must be provided to an organism Two categories of essential nutrients: Macronutrients required in large quantities; play principal roles in cell structure and metabolism Proteins, carbohydrates Micronutrients or trace elements required in small amounts; involved in enzyme function and maintenance of protein structure Manganese, zinc, nickel 2

3 Gases: the atmosphere is a reservoir for nitrogen, oxygen, and carbon dioxide essential to living processes. Sunlight supplies the basic source of energy on earth for most organisms. Photosynthesizers can use it directly to produce organic nutrients that feed other organisms. Non photosynthetic organisms extract the energy from chemical reactions to power cell processes. Courtesy: Pacific Northwest National Laboratory Plant litter Soil microbes CO2 Nutrients Organic compounds Nutrients are constantly being formed by decomposition and synthesis and released into the environment. Many inorganic nutrients originate from non-living environments such as the air, water, and bedrock. Soil community Kathy Park Talaro Complex communities of microbes exist in nearly every place on earth. Microbes residing in these communities must associate physically and share the habitat, often establishing biofilms and other inter relationships. ph Acidic [H + ] Acid Neutral [OH ] Aquatic microbes Base Basic (alkaline) The acid or base content (ph) can show extreme variations from habitat to habitat. Microbes are the most adaptable organisms with regard to ph. C K The temperature of habitats varies to a significant extent among all places on earth, and microbes exist at most points along this wide temperature scale.

4 Nutrients Organic nutrients contain carbon and hydrogen atoms and are usually the products of living things Methane (CH 4 ), carbohydrates, lipids, proteins, and nucleic acids Inorganic nutrients atom or molecule that contains a combination of atoms other than carbon and hydrogen Metals and their salts (magnesium sulfate, ferric nitrate, sodium phosphate), gases (oxygen, carbon dioxide) and water 4

5 Chemical Analysis of Cell Contents 70% water Proteins 96% of cell is composed of 6 elements: Carbon Hydrogen Oxygen Phosphorous Sulfur Nitrogen 5

6 Essential Biological Nutrients 6

7 Sources of Essential Nutrients Carbon sources Heterotroph must obtain carbon in an organic form made by other living organisms such as proteins, carbohydrates, lipids, and nucleic acids Autotroph an organism that uses CO 2, an inorganic gas as its carbon source Not nutritionally dependent on other living things 7

8 Growth Factors: Essential Organic Nutrients Organic compounds that cannot be synthesized by an organism because they lack the genetic and metabolic mechanisms to synthesize them Growth factors must be provided as a nutrient Essential amino acids, vitamins 8

9 Nutritional Types Main determinants of nutritional type are: Carbon source heterotroph, autotroph Energy source Chemotroph gain energy from chemical compounds Phototrophs gain energy through photosynthesis 9

10 Nutritional Categories 10

11 Autotrophs and Their Energy Sources Photoautotrophs Oxygenic photosynthesis Anoxygenic photosynthesis Chemoautotrophs (lithoautotrophs) survive totally on inorganic substances Methanogens, a kind of chemoautotroph, produce methane gas under anaerobic conditions 11 Kathy Park Talaro

12 Heterotrophs and Their Energy Sources Digestion in Bacteria and Fungi Majority are chemoheterotrophs Aerobic respiration Two categories Saprobes: free-living microorganisms that feed on organic detritus from dead organisms Opportunistic pathogen Facultative parasite Parasites: derive nutrients from host Pathogens (a) (b) (c) Organic debris Walled cell is a barrier. Enzymes Enzymes are transported outside the wall. Enzymes hydrolyze the bonds on nutrients. Some are obligate parasites 12 (d) Smaller molecules are transported across the wall and cell membrane into the cytoplasm.

13 Concept Check: If an organism is degrading large organic molecules to get both carbon and energy, it would be best described as a A. Photoheterotroph B. Photoautotroph C. Chemoheterotroph D. Chemoautotroph

14 Transport: Movement of Chemicals Across the Cell Membrane Passive transport does not require energy; substances exist in a gradient and move from areas of higher concentration toward areas of lower concentration Diffusion Osmosis diffusion of water Facilitated diffusion requires a carrier Active transport requires energy and carrier proteins; gradient independent Active transport Group translocation transported molecule chemically altered Bulk transport endocytosis, exocytosis, pinocytosis 14

15 Diffusion Net Movement of Molecules Down Their Concentration Gradient (Passive Transport) How Molecules Diffuse in Aqueous Solutions 15

16 Osmosis - Diffusion of Water (Passive Transport) Glass tube Membrane sac with solution Solute Water Container with water Pore a. Inset shows a close-up of the osmotic process. b. As the H 2 O diffuses into the sac, the volume c. Even as the solution becomes diluted, there The gradient goes from the outer container (higher concentration of H 2 O) to the sac (lower concentration of H 2 O). Some water will diffuse the opposite direction but the net gradient favors osmosis into the sac. increases and forces the excess solution into the tube, which will rise continually. will still be osmosis into the sac. Equilibrium will not occur because the solutions can never become equal. (Why?) 16

17 Response to solutions of different osmotic content Cells with Cell Wall Isotonic Solution Hypotonic Solution Hypertonic Solution Cell wall Cell membrane Cell membrane Water concentration is equal inside and outside the cell, thus rates of diffusion are equal in both directions. Net diffusion of water is into the cell; this swells the protoplast and pushes it tightly against the wall. Wall usually prevents cell from bursting. Water diffuses out of the cell and shrinks the cell membrane away from the cell wall; process is known as plasmolysis. Cells Lacking Cell Wall Early Early Cell membrane Late (osmolysis) Late Rates of diffusion are equal in both directions. Diffusion of water into the cell causes it to swell, and may burst it if no mechanism exists to remove the water. Water diffusing out of the cell causes it to shrink and become distorted. 17 Direction of net water movement.

18 Facilitated Diffusion (Passive Transport) Outside cell Inside cell Outside cell Inside cell (a) (b) 18

19 Carrier Mediated Active Transport Membrane Membrane Membrane Protein Protein Protein Protein Protein Protein Extracellular Intracellular Extracellular Intracellular Extracellular Intracellular (a) Carrier-mediated active transport. The membrane proteins (permeases) have attachment sites for essential nutrient molecules. As these molecules bind to the permease, energy from ATP pumps them into the cell s interior through special membrane protein channels. Microbes have these systems for transporting various ions (sodium, iron) and small organic molecules. Group Translocation Membrane Membrane Protein Protein Protein Protein 19 Extracellular Intracellular Extracellular Intracellular (b) In group translocation, the molecule is actively captured, but along the route of transport, it is chemically altered. By coupling transport with synthesis, the cell conserves energy.

20 Endocytosis: Eating and Drinking by Cells Endocytosis: bringing substances into the cell through a vesicle or phagosome Phagocytosis ingests substances or cells Pinocytosis ingests liquids Phagocytosis Pinocytosis Pseudopods 3 4 Microvilli Liquid enclosed by microvilli 2 Oil droplet 1 Vacuoles Vesicle with liquid 20

21 Summary of Transport Processes 21

22 Concept Check: If a cell is in a concentrated glucose solution and the glucose is moving into the cell through a carrier protein, this would be an example of A. Diffusion B. Facilitated Diffusion C. Active Transport D. Endocytosis E. Pinocytosis

23 Environmental Factors That Influence Microbes Niche: totality of adaptations organisms make to their habitat Environmental factors affect the function of metabolic enzymes Factors include: Temperature Oxygen requirements ph Osmotic pressure Barometric pressure 23

24 3 Cardinal Temperatures Minimum temperature lowest temperature that permits a microbe s growth and metabolism Maximum temperature highest temperature that permits a microbe s growth and metabolism Optimum temperature promotes the fastest rate of growth and metabolism 24

25 3 Temperature Adaptation Groups Psychrophiles optimum temperature below 15 o C; capable of growth at 0 o C Mesophiles optimum temperature 20 o -40 o C; most human pathogens Thermophiles optimum temperature greater than 45 o C Optimum Psychrophile Mesophile Thermophile Minimum Maximum Temperature C 25

26 Gas Requirements Oxygen As oxygen is utilized it is transformed into several toxic products: Singlet oxygen ( 1 O 2 ), superoxide ion (O 2 - ), peroxide (H 2 O 2 ), and hydroxyl radicals (OH - ) Most cells have developed enzymes that neutralize these chemicals: Superoxide dismutase, catalase If a microbe is not capable of dealing with toxic oxygen, it is forced to live in oxygen free habitats 26

27 Categories of Oxygen Requirement Aerobe utilizes oxygen and can detoxify it Obligate aerobe cannot grow without oxygen Facultative anaerobe utilizes oxygen but can also grow in its absence Microaerophilic requires only a small amount of oxygen Anaerobe does not utilize oxygen Obligate anaerobe lacks the enzymes to detoxify oxygen so cannot survive in an oxygen environment Aerotolerant anaerobes do not utilize oxygen but can survive and grow in its presence 27

28 Culturing by Oxygen Requirement Photo by Keith Weller, USDA/ARS Terese M. Barta, Ph.D.

29 Carbon Dioxide Requirement All microbes require some carbon dioxide in their metabolism Capnophile grows best at higher CO 2 tensions than normally present in the atmosphere 29 Courtesy and Becton, Dickinson and Company

30 Effects of ph Majority of microorganisms grow at a ph between 6 and 8 (neutrophiles) Acidophiles grow at extreme acid ph Alkalinophiles grow at extreme alkaline ph 30

31 Osmotic Pressure Most microbes exist under hypotonic or isotonic conditions Halophiles require a high concentration of salt Osmotolerant do not require high concentration of solute but can tolerate it when it occurs 31

32 Other Environmental Factors Barophiles can survive under extreme pressure and will rupture if exposed to normal atmospheric pressure 32

33 Concept Check: Chlamydomonas nivalis grows on Alaskan glaciers and it s photosynthetic pigments give the snow a red crust. This organism would be best described as a A. Psychrophile B. Alkalinophile C. Microaerophile D. Osmotolerant E. Barophile Image (a) courtesy Nozomu Takeuchi (b) Image courtesy Nozomu Takeuchi

34 Ecological Associations Among Microorganisms Microbial Associations Symbiotic Organisms live in close nutritional relationships; required by one or both members. Nonsymbiotic Organisms are free-living; relationships not required for survival. Mutualism Obligatory, dependent; both members benefit. Commensalism The commensal benefits; other member not harmed. Parasitism Parasite is dependent and benefits; host harmed. Synergism Members cooperate and share nutrients. Antagonism Some members are inhibited or destroyed by others. 34

35 Ecological Associations Symbiotic two organisms live together in a close partnership Mutualism obligatory, dependent; both members benefit Commensalism commensal member benefits, other member neither harmed nor benefited Parasitism parasite is dependent and benefits; host is harmed Staphylococcus aureus growth Science VU/Fred Marsik/Visuals Unlimited Courtesy Arthur Hauck (Germany) Haemophilus satellite colonies 35

36 Ecological Associations Non-symbiotic organisms are free-living; relationships not required for survival Synergism members cooperate to produce a result that none of them could do alone Antagonism actions of one organism affect the success or survival of others in the same community (competition) Antibiosis Nitrogen fixer (Azotobacter) N 2 NH 4 + Glucose Cellulose degrader (Cellulomonas) 36

37 Interrelationships Between Microbes and Humans Human body is a rich habitat for symbiotic bacteria, fungi, and a few protozoa - normal microbial flora Commensal, parasitic, and synergistic relationships 37

38 Microbial Biofilms Biofilms result when organisms attach to a substrate by some form of extracellular matrix that binds them together in complex organized layers Dominate the structure of most natural environments on earth Communicate and cooperate in the formation and function of biofilms quorum sensing 38

39 Biofilm Formation and Quorum Sensing Chromosome Quorum-dependent proteins 1 Inducer molecule Matrix Free-swimming cells settle on a surface and remain there. Cells synthesize a sticky matrix that holds them tightly to the substrate. When biofilm grows to a certain density (quorum), the cells release inducer molecules that can coordinate a response. Enlargement of one cell to show genetic induction. Inducer molecule stimulates expression of a particular gene and synthesis of a protein product, such as an enzyme. Cells secrete their enzymes in unison to digest food particles. 39

40 The Study of Microbial Growth Microbial growth occurs at two levels: growth at a cellular level with increase in size increase in population Division of bacterial cells occurs mainly through binary fission (transverse) Parent cell enlarges, duplicates its chromosome, and forms a central transverse septum dividing the cell into two daughter cells 40

41 Binary Fission 1 A young cell at early phase of cycle 2 A parent cell prepares for division by enlarging its cell wall, cell membrane, and overall volume. Midway in the cell, the wall develops notches that will eventually form the transverse septum, and the duplicated chromosome becomes affixed to a special membrane site. 3 The septum wall grows inward, and the chromosomes are pulled toward opposite cell ends as the membrane enlarges. Other cytoplasmic components are distributed (randomly) to the two developing cells. 4 The septum is synthesized completely through the cell center, and the cell membrane patches itself so that there are two separate cell chambers. 5 At this point, the daughter cells are divided. Some species will separate completely as shown here, while others will remain attached, forming chains or doublets, for example. Cell wall Cell membrane Chromosome 1 Chromosome 2 Ribsomes 41

42 Rate of Population Growth Time required for a complete fission cycle is called the generation, or doubling time Each new fission cycle increases the population by a factor of 2 exponential growth Generation times vary from minutes to days * * Log of 3000 ( ) number Number 2500 ( ) of cells of cells using the power 1500 of Number of cells Number of generations Exponential value (2 1) (2 2) (2 2 2) ( ) ( ) (b) 0 Time 0 42 (a)

43 Rate of Population Growth Equation for calculating population size over time: N ƒ = (N i )2 n N ƒ is total number of cells in the population N i is starting number of cells Exponent n denotes generation time 2 n number of cells in that generation 43

44 Concept Check: Escherichia coli has a doubling time of 20 minutes. If there are 5 cells at the beginning of the experiment, how many will there be in 3 hours?

45 Viable Plate Count Flask inoculated Samples taken at equally spaced intervals (0.1 ml) 500 ml 60 min 0.1 ml 120 min 180 min 240 min 300 min 360 min 420 min 480 min 540 min 600 min Sample is diluted in liquid agar medium and poured or spread over surface of solidified medium Plates are incubated, colonies are counted Number of colonies (CFU) per 0.1 ml Total estimated cell population in flask None <1* <10, , , , , , , , , ,150,000 *Only means that too few cells are present to be assayed.

46 Logrithm (10 n ) of Viable Cells The Population Growth Curve In laboratory studies, populations typically display a predictable pattern over time growth curve Stages in the normal growth curve: 1. Lag phase flat period of adjustment, enlargement; little growth Stationary phase The final outcome varies with the culture Hours T otal cells in population, live and dead, at each phase Few cells Live cells Dead cells (not part of count) 46

47 Logrithm (10 n ) of Viable Cells The Population Growth Curve Stages in the normal growth curve: 1. Lag phase 2. Exponential growth phase a period of maximum growth will continue as long as cells have adequate nutrients and a favorable environment Stationary phase The final outcome varies with the culture Hours T otal cells in population, live and dead, at each phase Few cells Live cells Dead cells (not part of count) 47

48 Logrithm (10 n ) of Viable Cells The Population Growth Curve Stages in the normal growth curve: 1. Lag phase 2. Exponential growth phase 3. Stationary phase rate of cell growth equals rate of cell death caused by depleted nutrients and O 2, excretion of organic acids and pollutants Stationary phase The final outcome varies with the culture Hours T otal cells in population, live and dead, at each phase Few cells Live cells Dead cells (not part of count) 48

49 Logrithm (10 n ) of Viable Cells The Population Growth Curve Stages in the normal growth curve: 1. Lag phase 2. Exponential growth phase 3. Stationary phase 4. Death phase as limiting factors intensify, cells die exponentially Stationary phase The final outcome varies with the culture Hours T otal cells in population, live and dead, at each phase Few cells Live cells Dead cells (not part of count) 49

50 Methods of Analyzing Population Growth Turbidometry most simple Degree of cloudiness, turbidity, reflects the relative population size Percent of light transmitted (1) Kathy Park Talaro/Visuals Unlimited (a) (b) (2) 50

51 Methods of Analyzing Population Growth Enumeration of bacteria: Viable colony count Direct cell count count all cells present; automated or manual Sample in liquid Bacterial cell Tube Automatic counter Counting orifice Electronic detector 51