Microbiology: A Systems Approach, 2nd ed. Chapter 7: Microbial Nutrition, Ecology, and Growth

7.1 Microbial Nutrition Nutrition: a process by which chemical substances (nutrients) are acquired from the environment and used in cellular activities All living things require a source of elements such as C, H, O, P, K, N, S, Ca, Fe, Na, Cl, Mg- but the relative amounts vary depending on the microbe Essential Nutrient: any substances that must be provided to an organism Macronutrients: Required in relatively large quantities, play principal roles in cell structure and metabolism (ex. C, H, O) Micronutrients: aka trace elements, present in smaller amounts and involved in enzyme function and maintenance of protein structure (ex. Mn, Zn, Ni) Nutrients are processed and transformed into the chemicals of the cell after absorption Can also categorize nutrients according to C content Inorganic nutrients: A combination of atoms other than C and H Organic nutrients: Contain C and H, usually the products of living things

Chemical Analysis of Microbial Cytoplasm

Sources of Essential Nutrients Carbon sources Nitrogen sources Oxygen sources Hydrogen sources Phosphorus sources Sulfur sources Others

Carbon Sources The majority of C compounds involved in normal structure and metabolism of all cells are organic Heterotroph: Must obtain C in organic form (nutritionally dependent on other living things) Autotroph: Uses inorganic CO2 as its carbon source (not nutritionally dependent on other living things)

Nitrogen Sources Main reservoir- N2 Primary nitrogen source for heterotrophs- proteins, DNA, RNA Some bacteria and algae utilize inorganic nitrogenous nutrients Small number can transform N2 into usable compounds through nitrogen fixation Regardless of the initial form, must be converted to NH3 (the only form that can be directly combined with C to synthesize amino acids and other compounds)

Oxygen Sources O is a major component of organic compounds Also a common component of inorganic salts O2 makes up 20% of the atmosphere

Hydrogen Sources H is a major element in all organic and several inorganic compounds Performs overlapping roles in the biochemistry of cells: Maintaining pH Forming hydrogen bonds between molecules Serving as the source of free energy in oxidation-reduction reactions of respiration

Phosphorus (Phosphate) Sources Main inorganic source of phosphorus is phosphate (PO43-) Derived from phosphoric acid Found in rocks and oceanic mineral deposits Key component of all nucleotides Phospholipids in cell membranes Coenzymes

Sulfur Sources Widely distributed throughout the environment in mineral form Essential component of some vitamins Amino acids- methionine and cysteine

Other Nutrients Important in microbial Metabolism Potassium- protein synthesis and membrane function Sodium- certain types of cell transport Calcium- stabilizer of cell walls and endospores Magnesium- component of chlorophyll and stabilizer of membranes and ribosomes Iron- important component of cytochrome proteins Zinc- essential regulatory element for eukaryotic genetics, and binding factors for enzymes Copper, cobalt, nickel, molybdenum, manganese, silicon, iodine, and boron- needed in small amounts by some microbes but not others

Growth Factors: Essential Organic Nutrients Growth factor: An organic compound such as an amino acid, nitrogenous base, or vitamin that cannot be synthesized by an organism and must be provided as a nutrient For example, many cells cannot synthesize all 20 amino acids so they must obtain them from food (essential amino acids)

How Microbes Feed: Nutritional Types

Main Determinants of Nutritional Type Sources of carbon and energy Phototrophs- Microbes that photosynthesize Chemotrophs- Microbes that gain energy from chemical compounds

Autotrophs and Their Energy Sources Photoautotrophs Photosynthetic Form the basis for most food webs Chemoautotrophs Chemoorganic autotrophs- use organic compounds for energy and inorganic compounds as a carbon source Lithoautotrophs- rely totally on inorganic minerals Methanogens- produce methane from hydrogen gas and carbon dioxide Archaea Some live in extreme habitats

Figure 7.1

Heterotrophs and Their Energy Sources Majority are chemoheterotrophs that derive both carbon and energy from organic compounds Saprobes Free-living microorganisms Feed primarily on organic detritus from dead organisms Decomposers of plant litter, animal matter, and dead microbes Most have rigid cell wall, so they release enzymes to the extracellular environment and digest food particles into smaller molecules Obligate saprobes- exist strictly on dead organic matter in soil and water Facultative parasite- when a saprobe infects a host, usually when the host is compromised (opportunistic pathogen)

Figure 7.2

Other Chemoheterotrophs Parasites Derive nutrients from the cells or tissues of a host Also called pathogens because they cause damage to tissues or even death Ectoparasites- live on the body Endoparasites- live in organs and tissues Intracellular parasites- live within cells Obligate parasites- unable to grow outside of a living host

Transport Mechanisms for Nutrient Absorption Cells must take nutrients in and transport waste out Transport occurs across the cell membrane, even in organisms with cell walls

The Movement of Water: Osmosis Osmosis: Diffusion of water through a selectively permeable membrane The membrane is selectively permeable- having passageways that allow free diffusion of water but can block certain other dissolved molecules When the membrane is between solutions of differing concentrations and the solute is not diffusible, water will diffuse at a fast rate from the side that has more water to the side that has less water

Figure 7.3

Osmotic Relationships The osmotic relationship between cells and their environment is determined by the relative concentrations of the solutions on either side of the cell membrane Isotonic: The environment is equal in solute concentration to the cell’s internal environment No net change in cell volume Generally the most stable environment for cells Hypotonic: The solute concentration of the external environment is lower than that of the cell’s internal environment Net direction of osmosis is from the hypotonic solution into the cell Cells without cell walls swell and can burst Hypertonic: The environment has a higher solute concentration than the cytoplasm Will force water to diffuse out of a cell Said to have high osmotic pressure

Figure 7.4

Adaptations to Osmotic Variations in the Environment Example: fresh pond water- hypotonic conditions Bacteria- cell wall protects them from bursting Amoeba- a water (or contractile) vacuole that moves excess water out of the cell Example: high-salt environment- hypertonic conditions Halobacteria living in the Great Salt Lake- absorb salt to make their cells isotonic with the environment

The Movement of Molecules: Diffusion and Transport Diffusion: When atoms or molecules move in a gradient from an area of higher density or concentration to an area of lower density or concentration Random thermal movement of molecules will eventually distribute the molecules from an area of higher concentration to an area of lower concentration Evenly distributes the molecules Diffusion of molecules across the cell membrane is largely determined by the concentration gradient and permeability of the substance Simple or passive diffusion is limited to small nonpolar molecules or lipid soluble molecules

Figure 7.5

Facilitated Diffusion Utilizes a carrier protein that binds a specific substance, changes the conformation of the carrier protein, and the substance is moved across the membrane Once the substance is transported, the carrier protein resumes its original shape Carrier proteins exhibit specificity Saturation: The rate of facilitated diffusion of a substance is limited by the number of binding sites on the transport proteins Competition: When two molecules of similar shape can bind to the same binding site on a carrier protein

Figure 7.6

Active Transport Nutrients are transported against the diffusion gradient or in the same direction as the natural gradient but at a rate faster than by diffusion alone Requires the presence of specific membrane proteins (permeases and pumps) Requires the expenditure of energy Items that require active transport: monosaccharides, amino acids, organic acids, phosphates, and metal ions Specialized pumps- an important type of active transport Group translocation: couples the transport of a nutrient with its conversion to a substance that is immediately useful inside the cell

Figure 7.7

Endocytosis: Eating and Drinking by Cells A form of active transport Transporting large molecules, particles, lipids, or other cells Occurs in some eukaryotic cells The cell encloses the substance in its cell membrane, simultaneously forming a vacuole and engulfing it Phagocytosis- amoebas and certain white blood cells; ingesting whole cells or large solid matter Pinocytosis- Transport of liquids such as oils or molecules in solution

7.2 Environmental Factors that Influence Microbes Temperature Adaptations Microbial cells cannot control their temperature, so they assume the ambient temperature of their natural habitat The range of temperatures for the growth of a given microbial species can be expressed as three cardinal temperatures: Minimum temperature: the lowest temperature that permits a microbe’s continued growth and metabolism Maximum temperature: The highest temperature at which growth and metabolism can proceed Optimum temperature: A small range, intermediate between the minimum and maximum, which promotes the fast rate of growth and metabolism Some microbes have a narrow cardinal range while others have a broad one Another way to express temperature adaptation- to describe whether an organism grows optimally in a cold, moderate, or hot temperature range

Psychrophile A microorganism that has an optimum temperature below 15°C and is capable of growth at 0°C. True psychrophiles are obligate with respect to cold and cannot grow above 20°C. Psychrotrophs or facultative psychrophiles- grow slowly in cold but have an optimum temperature above 20°C.

Figure 7.8

Figure 7.9

Mesophile An organism that grows at intermediate temperatures Optimum growth temperature of most: 20°C to 40°C Temperate, subtropical, and tropical regions Most human pathogens have optima between 30°C and 40°C

Thermophile A microbe that grows optimally at temperatures greater than 45°C Vary in heat requirements General range of growth of 45°C to 80°C Hyperthermophiles- grow between 80°C and 120°C

Gas Requirements Atmospheric gases that most influence microbial growth- O2 and CO2 Oxygen gas has the greatest impact on microbial growth As oxygen enters into cellular reactions, it is transformed into several toxic products Most cells have developed enzymes that go about scavenging and neutralizing these chemicals Superoxide dismutase Catalase Essential for aerobic organisms

Several General Categories of Oxygen Requirements Aerobe: can use gaseous oxygen in its metabolism and possesses the enzymes needed to process toxic oxygen products Obligate aerobe: cannot grow without oxygen Facultative anaerobe: an aerobe that does not require oxygen for its metabolism and is capable of growth in the absence of it Microaerophile: does not grow at normal atmospheric concentrations of oxygen but requires a small amount of it in metabolism Anaerobe: lacks the metabolic enzyme systems for using oxygen in respiration Strict, or obligate, anaerobes: also lack the enzymes for processing toxic oxygen and cannot tolerate any free oxygen in the immediate environment and will die if exposed to it. Aerotolerant anaerobes: do not utilize oxygen but can survive and grow to a limited extent in its presence

Figure 7.10

Figure 7.11

Carbon Dioxide All microbes require some carbon dioxide in their metabolism Capnophiles grow best at a higher CO2 tension than is normally present in the atmosphere

Effects of pH Majority of organisms live or grow in habitats between pH 6 and 8 Obligate acidophiles Euglena mutabilis- alga that grows between 0 and 1.0 pH Thermoplasma- archaea that lives in hot coal piles at a pH of 1 to 2, and would lyse if exposed to pH 7

Osmotic Pressure Most microbes live either under hypotonic or isotonic conditions Osmophiles- live in habitats with a high solute concentration Halophiles- prefer high concentrations of salt Obligate halophiles- grow optimally in solutions of 25% NaCl but require at least 9% NaCl for growth

Miscellaneous Environmental Factors Nonphotosynthetic microbes tend to be damaged by the toxic oxygen products produced by contact with light Some produce yellow carotenoid pigments to protect against the damaging effects of light by absorbing and dismantling toxic oxygen Other types of radiation that can damage microbes are ultraviolet and ionizing rays Barophiles: deep-sea microbes that exist under hydrostatic pressures ranging from a few times to over 1,000 times the pressure of the atmosphere All cells require water- only dormant, dehydrated cell stages tolerate extreme drying

Ecological Associations Among Microorganisms Most microbes live in shared habitats Interactions can have beneficial, harmful, or no particular effects on the organisms involved They can be obligatory or nonobligatory to the members They often involve nutritional interactions

Symbiosis A general term used to denote a situation in which two organisms live together in a close partnership Members are termed symbionts Three main types of symbionts Mutualism: when organisms live in an obligatory but mutually beneficial relationship Commensalism: the member called the commensal receives benefits, while its coinhabitant is neither harmed nor benefited Satellitism: when one member provides nutritional or protective factors needed by the other Parasitism: a relationship in which the host organism provides the parasitic microbe with nutrients and a habitat

Figure 7.12

Nonsymbiotic Relationship Organisms are free-living and relationships are not required for survival Synergism: an interrelationship between two or more free-living organisms that benefits them but is not necessary for their survival Antagonism: an association between free-living species that arises when members of a community compete One microbe secretes chemical substances into the surrounding environment that inhibit or destroy another microbe in the same habitat

Interrelationships Between microbes and Humans Normal microbiotia: microbes that normally live on the skin, in the alimentary tract, and in other sites in humans Can be commensal, parasitic, and synergistic relationships

7.3 The Study of Microbial Growth Growth takes place on two levels Cell synthesizes new cell components and increases in size The numer of cells in the population increases The Basis of Population Growth: Binary Fission

Figure 7.13

Figure 7.14

The Rate of Population Growth Generation or doubling time: The time required for a complete fission cycle Each new fission cycle or generation increases the population by a factor of 2 As long as the environment is favorable, the doubling effect continues at a constant rate The length of the generation time- a measure of the growth rate of an organism Average generation time- 30 to 60 minutes under optimum conditions Can be as short as 10 to 12 minutes This growth pattern is termed exponential

Graphing Bacterial Growth The data from growing bacterial populations are graphed by plotting the number of cells as a function of time If plotted logarithmically- a straight line If plotted arithmetically- a constantly curved slope To calculate the size of a population over time: Nf = (Ni)2n Nf is the total number of cells in the population at some point in the growth phase Ni is the starting number N denotes the generation number

The Population Growth Curve A population of bacteria does not maintain its potential growth rate and double endlessly A population displays a predictable pattern called a growth curve The method to observe the population growth pattern: Place a tiny number of cells in a sterile liquid medium Incubate this culture over a period of several hours Sampling the growth at regular intervals during incubation Plating each sample onto solid media Counting the number of colonies present after incubation

Stages in the Normal Growth Curve Data from an entire growth period typically produce a curve with a series of phases Lag Phase Exponential Growth Phase Stationary Growth Phase Death Phase

Lag Phase Relatively “flat” period Newly inoculated cells require a period of adjustment, enlargement, and synthesis The cells are not yet multiplying at their maximum rate The population of cells is so sparse that the sampling misses them Length of lag period varies from one population to another

Exponential Growth (Logarithmic or log) Phase When the growth curve increases geometrically Cells reach the maximum rate of cell division Will continue as long as cells have adequate nutrients and the environment is favorable

Stationary Growth Phase The population enters a survival mode in which cells stop growing or grow slowly The rate of cell inhibition or death balances out the rate of multiplication Depleted nutrients and oxygen Excretion of organic acids and other biochemical pollutants into the growth medium

Death Phase The curve dips downward Cells begin to die at an exponential rate

Figure 7.15

Potential Importance of the Growth Curve Implications in microbial control, infection, food microbiology, and culture technology Growth patterns in microorganisms can account for the stages of infection Understanding the stages of cell growth is crucial for working with cultures In some applications, closed batch culturing is inefficient, and instead, must use a chemostat or continuous culture system

Other Methods of Analyzing Population Growth Turbidometry- a tube of clear nutrient solution becomes turbid as microbes grow in it

Figure 7.16

Enumeration of Bacteria Direct or total cell count- counting the number of cells in a sample microscopically Uses a special microscope slide (cytometer) Used to estimate the total number of cells in a larger sample

Figure 7.17

Automated Counting Coulter counter- electronically scans a culture as it passes through a tiny pipette Flow cytometer also measures cell size and differentiates between live and dead cells Real-time PCR allows scientists to quantify bacteria and other microorganisms that are present in environmental or tissue samples without isolating or culturing them

Figure 7.18