Bio 220 chapter 6

Biofilms: Multicellular microbes?

A biofilm is a mass of bacteria that stick to and multiply on a solid surface, such as a stone in a lake or a lung in a cystic fibrosis patient. Can be constructed of single species or by multiple collaborating species and can form on a range of organic and inorganic surfaces Ex: Plaque on teeth is a mixed-species biofilm. P. aeruginosa in lungs of CF patient Staphylococci on medical implants

Lag phase

A lag period after cells are transferred from an old culture to fresh growth media. Bacteria do not divide during this time. Factors that affect the duration of lag phase: Aged cells from the old culture will need time to repair before they can replicate again Changes in carbon, nitrogen, energy source must be sensed Type of medium to which the cells are being introduced

Bacterial Growth Curve

A plot of culture growth as the logarithm of the cell number versus incubation time 1. lag phase 2. log or exponential phase 3. stationary phase 4. death phase

Pure culture

A single, genetically homogeneous strain of a single species. can be grown in liquid or solid media used to examine growth rates & microbial biochemistry useful for separating mixtures of different organisms

Aerobes versus anaerobes

Aerobes, but not anaerobes, destroy ROS with the aid of enzymes such as: superoxide dismutase (to remove superoxide) peroxidase (to remove hydrogen peroxide) catalase (to remove hydrogen peroxide) Also contain resourceful enzyme systems that detect and repair macromolecules damaged by oxidation

Rood nodules (arrow

After rhizobia invade the plant root, symbiosis between plant and microbe produces nodules/

Bacteria growth in a tube

As oxygen diffuses and equilibrates with the air, oxygen levels in different areas of a simple nutrient broth tube will diminish the farther down the tube you look. Inoculating a series of tubes like this with bacteria having different oxygen requirements will result in anaerobes (not strict anaerobes) growing at the bottom of the tubes. Microaerophilic bacteria will grow in the middle and strict aerobes will grow only at the top. Facultative organisms can grow throughout.

Budding

Caulobacter, Hyphomicrobium reproduce by budding

Endospores in Suspended Animation

Certain Gram-positive genera have the remarkable ability to develop dormant spores that are heat and desiccation resistant. Clostridia, such as Clostridium botulinum and Clostridium tetani Bacillus, such as Bacillus anthracis

How do microbes

Concentration of hydrogen ions [H+] also have a direct effect on the cell's macromolecular structures. Extreme concentrations of either hydrogen (H+) or hydroxide (OH-) ions in solution will limit growth. pH influences growth by altering protein shape, which in turn changes protein activity. Microbes have adapted to inhabit diverse pH environments from 0-11.5.

Variations in pressure

Creatures living at sea level are subjected to a pressure of 1 atmosphere (1 atm). High pressure can kill these organisms. At the bottom of the ocean, hydrostatic pressure reaches 400 atm and can go as high as 1000 atm in ocean trenches.

Nitrogen cycle

Dinitrogen gas (N2) is fixed in ammonium ions (NH4+) by species of bacteria (nitrogen fixers) that possess the enzyme nitrogenase. Other bacteria (nitrifiers) oxidize (NH4+) to generate energy. Still others (denitrifiers) use oxidized forms of nitrogen such as NO3- , as an alternative electron acceptor in place of O2.

Quorum sensing

During the formation of a biofilm, more and more cells bind to the surface and begin to communicate with each other by sending and receiving chemical signals in a process known as quorum sensing. Individual cells continually make these chemical signal molecules, but once the population reaches a certain number, the chemical signal reaches a concentration that the cells can sense. Reception of this signal triggers genetically regulated changes that cause the bacteria to bind to the surface substrate and each other.

Generations hour (growth rate graph)

Each curve represents the temperature growth range for different classes of bacteria. Growth rate increases linearly with temperature until an optimum and then decreases. These curves are only an approximation. Growth rates and optimal ranges for individual species within a class will vary.

mitosis

Eukaryotic microbes divide this way

Optimum Temperature

Every organism has an optimum temperature at which it grows most quickly. Minimum and maximum temperatures define the limits of growth. Growth stops when: rising temperatures cause critical enzymes or cell structures to fail. at cold temperatures, enzymatic processes become too sluggish and the cell membrane is less fluid.

Effect of enviroment

Every protein and macromolecular structure within a cell is affected by changes in the environment.

Phases of Growth

Exponential growth is not indefinite because nutrient consumption and toxic by-products eventually slow the growth rate until it halts. As medium conditions deteriorate, alterations occur in membrane composition, cell size, and metabolic pathways, all of which impact generation time.

Log phase

Exponential growth phase, balanced growth where all cell components are synthesized at constant rates in relation to each other Maximum rate, largest in size at this time

Exponential Growth

Growth in which the population size doubles at a fixed rate (such as every 20 minutes) Plotting the increase in the number of cells over time generates an exponential curve.

Continuous Culture

In open systems where fresh medium is continually added to a culture and an equal amount of culture is constantly siphoned off, bacterial populations can be kept in exponential phase at a constant cell mass for long periods of time.

chemically defined minimal medium

In the laboratory, many organisms can be grown with

Spread plates

Isolation of pure cultures Allows counting the number of viable colonies viable = successfully replicates to form a colony One cell = 1 colony does not always hold. Streptococcus & Staphylococcus usually do not exist as single cells, but as chains or clusters. One colony = 1 CFU (colony forming unit)

Limits to bacterial growth

Limiting nutrients helps control bacterial overgrowth. Escherichia coli can divide every 20-30 minutes. One cell can potentially multiply to over 1 × 1014 cells in 24 hours.

Can microbes assemble their own compounds?

Many microbes cannot assemble all the compounds they need for growth on their own.

Can bacteria cooperate with each other?

Many, if not most, bacteria form specialized, surface-attached communities called biofilms. Under environmental stress, many organisms also may undergo complex differentiation programs.

pH Homeostasis Mechanisms

Microbes can prevent an unwanted influx of protons in several ways. E. coli can reverse proton influx by importing a variety of cations such as K+ and Na+. Under extremely alkaline conditions the cells can use Na+/H+ anti-porters to recruit protons into the cell in exchange for expelling Na+. Some organisms can change the pH of the medium by using various amino acid decarboxylases and deaminases, producing alkaline and acidic products, respectively.

What are most microbes?

Most microbes in the world are anaerobic, growing buried in the soil, within our anaerobic digestive tract, or within biofilms on our teeth. some respire using electron transport systems without oxygen as the terminal electron acceptor (such as nitrate) to conduct anaerobic respiration. Organisms without electron transport chains cannot respire and must rely on carbohydrate fermentation for energy.

Obtaining Nitrogen

Nitrogen is a critical component of proteins, nucleic acids, and other cellular constituents and is required in large amounts by living organisms. N2 makes up nearly 79% of Earth's atmosphere, but the nitrogen in N2 is unavailable for use by most organisms. Nitrogen from N2 must be "fixed," or converted to ammonium ions (NH4+) through the nitrogen cycle

barophiles or piezophiles.

Organisms that have adapted to grow at high pressures are called

Symbionts

Organisms that live in intimate association with a second organism. Rhizobium, Sinorhizobium, Bradyrhizobium

Phases of Growth diagram

Phases of bacterial growth that occur in a typical batch culture are represented by different shadings under the curve. (note that in this graph, the y-axis is a logarithmic scale that reflects order of magnitude differences between cell numbers, not simple increases in cell number. A one order of magnitude difference equals a tenfold increase in cell number.

Rickettsias

Some bacteria, such as Rickettsias, have such complex nutrient requirements in their natural habitat that we still do not know how to grow them using artificial media.

halophiles

Species that have evolved to require high salt (NaCl) concentrations are known a

Stages of Endospores

Stage I: Sporulation begins when the bacterial chromosome replicates and stretches into a long axial filament spanning the length of the cell. Stage II: The cell makes a decision to divide at one of the two cell poles instead of the middle, as normally occurs during growth. Stage III: Septation divides the cell into two unequal compartments: the forespore, which will ultimately become the spore, and the larger mother cell. Stage IV: The mother cell chromosome is destroyed and the mother cell membrane engulfs the forespore. Stage V: A thick peptidoglycan layer (cortex) is placed between the two membranes surrounding the forespore. Layers of coat proteins are deposited. Stage VI: Spore formation is completed with resistance to heat and chemical insults. Stage VII: The mother cell, now called a sporangium, dies and releases the mature spore.

Exponential Growth (example)

Starting with any number of organisms at time zero (N0), the number of organisms after n generations will be N0 × 2n. For example: a single cell after 3 generations (n = 3) will produce: 1 cell × 23 = 8 cells

What are the environmental limits beyond which microbes will not grow?

The answer depends on the microbe. Ecological niches outside the norm for humans are considered extreme.

Human Gastrointestinal tract

The human gastrointestinal tract is engineered much like a chemostat in that new nutrients are always arriving from the throat while equal amounts of bacterial culture exit in fecal waste. The basic chemostat ensures exponential growth by constantly adding and removing equal amounts of culture media.

Bio film creation

The stages of biofilm development in Pseudomonas generally apply to the formation of many kinds of biofilms.

What determines the rate of microbial growth?

The ultimate goal of any species is to make more of its own kind.

Temperature Extremes

Unlike other organisms, microbes cannot control their temperature. Bacterial temperature matches that of the immediate environment. Temperature affects the rate of motion, membrane fluidity, nutrient transport, DNA/RNA stability, & enzyme structure and function.

Water Availability and Salt Concentration

Water availability is measured as water activity, a quantity approximated by concentration. Interactions with solutes (such as NaCl) lower water activity. The more solutes in a solution, the less water is available for microbes to use for growth.

Death phase

When the rate at which cells die is exponential. Nutrients are limited and toxic by-products are increasing. Can be prolonged.

Stationary phase

When viable cell numbers stop rising owing to a lack of key nutrients or buildup of waste products. Growth of individual cells slows and the number of cells dividing approximately equals the number of cells dying. Resistance to antibiotics and host defenses develops during this time.

generation time or doubling time.

With unlimited resources, bacteria divide at a constant interval called the

Vibrio vulnificus

a Gram-negative species that causes fatal septicemia and Vibrio cholerae, a cause of extreme diarrhea, are considered marine microorganisms that prefer growth at an elevated salt concentration.

Acidophiles

are bacteria and archaea that live in acidic environments. They are often chemoheterotrophs that oxidize reduced metals and generate strong acids such as sulfuric acid pH 1-4

Neutralophiles

are bacteria that generally grow between pH 5 and 8 and include most human pathogens.

Extremophiles

are bacteria, archaea, and some eukaryotes that can grow in extreme environments.

Strict aerobes

are organisms that not only exist in oxygen but also use oxygen as a terminal electron acceptor. They grow only when oxygen is present and consume oxygen during metabolism.

Spores

are resistant to many environmental stresses that kill vegetative cells. dessication of the spore packing with small acid-soluble proteins (SASPs) that bind to and protect DNA The SASP coat protects from damage due to UV light and various toxic chemicals. Once proper conditions are restored, germination is triggered that wakes up the dormant cell, dissolves the spore coat, and releases a viable vegetative cell.

Essential nutrients

are those compounds that a microbe cannot make itself but must gather from its immediate environment. carbon (C) magnesium (Mg2+ ) nitrogen (N) ferrous iron (Fe2+ ) phosphorus (P) potassium (K+) hydrogen (H) oxygen (O) sulfur (S) Trace elements: cobalt, copper, manganese, molybdenum, nickel and zinc

Escherichia coli and Salmonella (Growth)

at much lower salt concentrations. They inhabit freshwater contaminated by fecal matter from humans and other animals. Most bacteria prefer salt concentrations at 0.05-1M (0.2 -5%). Moderate halophiles grow optimally at 0.85-3.4M (5-20%). Seawater is about 3.5% NaCl. Extremely halophilic organisms require 3.4-5.1 M (20-30%) salt to grow. Halotolerant organisms can grow in high or low salt (0-30% NaCl).

Hyperthermophiles

can grow at temperatures as high as 121°C.

Psychrophiles

can grow at temperatures as low as 0° C but their optimum growth temperature is around 14° C. Cold LOVINGINGINGING

Symbiotic Rhizobium cell

cells clustered on a clover root tip. The rhizobia shown here are clustered on the surface of the root. Soon they will start to invade the root and begin a symbiotic partnership that will benefit both organisms

Strict anaerobes

die in the least bit of oxygen. They do not use oxygen as an electron acceptor and die because they are vulnerable to the highly toxic, chemically reactive oxygen species produced by their own metabolism when exposed to oxygen. Reactive Oxygen Species (ROS) are oxygen molecules or ions with one too few or two many electrons.

Differential media

expose biochemical differences between 2 species that grow equally well.

Phototrophs

extract energy from absorption of light.

Chemotrophs

extract energy from oxidation-reduction reactions that remove electrons from high energy compounds to produce lower energy compounds. lithotrophy organotrophy

Selective media

favor the growth of one organism over another.

Heterotrophs (energy)

gain energy by degrading complex organic compounds, such as polysaccharides, to smaller compounds, such as glucose and pyruvate. The carbon from pyruvate moves through the tricarboxylic acid (TCA) cycle, or Krebs cycle, and CO2 is released. In the absence of a TCA cycle, the carbon can end up as fermentation products, such as ethanol or acetic acid.

Thermophiles

have adapted to growth at high temperatures, typically at 55°C and higher.

Mesophiles

have optimal growth between 20-40°C, with a minimum of 15°C and a maximum of 45°C. Bacteria, Things that cause disease.

chemostat

is a continuous culture system in which the diluting medium contains a growth-limiting amount of an essential nutrient. Increasing the flow rate increases the amount of nutrient available to the microbe. The more nutrient available, the faster a cell's mass will increase. The faster the cell's mass increases, the more quickly it will divide.

Osmolarity

is a measure of the number of solute molecules in a solution. The more particles in a solution, the greater the osmolarity and the lower the water activity.

Nostoc

is an autotroph. These cyanobacteria live in a gelatinous sphere. The spheres shown are colonies (approx. 1 mm diameter) that are just beginning to divide. The individual cyanobacteria are the small cells forming the long strands inside the spheres.

Binary fission

is one parent cell splitting into two equal daughter cells. symmetrical or asymmetrical

Nitrogen-fixing bacteria

may be free-living in soil or water may form symbiotic associations with plants or other organisms

Alkaliphiles

occupy the opposite end of the pH spectrum, growing best at values ranging from pH 9 to pH 11.

Organotrophy

oxidizes organic compounds such as sugars to obtain energy.

Kidney Stones

proteus urease increase pH of urine, which therefore cause kidney stones

Heterotrophs

rely on other organisms to make the organic compounds that they use as carbon sources.

Exopolysaccharides (EPSs)

such as alginate produced by P. aeruginosa and colonic acid produced by E. coli, increase antibiotic resistance by limiting antibiotic access to the center of the biofim. Hallmark of chronic infectious disease

Facultative anaerobes

that possess the enzymes to detoxify oxygen radicals and also the machinery for both fermentation and aerobic respiration. Both energy production

H. influenzae

the causative agent of meningitis cannot synthesize NAD (V factor) or heme (X factor), and thus cannot grow on typical laboratory media unless they are added

Autotrophs (energy)

use light energy or energy derived from the oxidation of minerals to capture CO2 and convert it to complex organic molecules

Aerotolerant anaerobes

use only fermentation to provide energy but contain superoxide dismutase and catalase or peroxidase to protect them from ROS. Doesnt use oxygen; can tolerate but not use it.

Autotrophs

use the CO2 discarded by heterotrophs to make complex cell constituents made up of C, H, and O, such as carbohydrates.

Lithotrophy

uses inorganic chemicals such as hydrogen (H2), hydrogen sulfide (H2S), ammonium (NH4), nitrate (NO2-) and ferrous iron (Fe2+) for energy.

Microaerophiles

will grow only at low oxygen concentrations. They possess a decreased level of superoxide dismutase and/or catalase Grow where there is a little bit of oxygen