Micro Exam 1

Outcomes of Citric Acid Cycle.

Citric acid cycle (CAC): pathway through which pyruvate is completely oxidized to CO2; much greater ATP yield than fermentation (38 vs. 2) • >>>decarboxylation of pyruvate to CO2, NADH, and acetyl- CoA • >>>Acetyl-CoA + oxaloacetate forms citric acid. • >>>>>>>2 CO2, 3 NADH, 1 FADH2 • >>>>>>oxaloacetate regenerated per pyruvate, total = 3 CO2, 4 NADH, 1 FADH2 • per glucose molecule, 6 CO2 molecules released and NADH and FADH2 generated • NADH and FADH oxidized in electron transport chain: consumes electrons and produces ATP

dark field microscopy,

Light reaches the specimen from the sides. • Only light reaching the lens has been scattered by specimen. • Image appears light on a dark background. •better resolution than light microscopy • excellent for observing motility (flagella)

thermophilic microbes.

Microorganisms can be classified into groups by their growth temperature optima. high, found in hot environments organisms with growth temperature optima between 45°C and 80°C Thermophiles inhabit moderately hot or intermittently hot environments.

Psychrophilic microbes.

Microorganisms can be classified into groups by their growth temperature optima. low, found in cold environments • organisms with optimal growth temperature ≤ 15°C, maximum ≤ 20°C, minimum ≤ 0°C • inhabit constantly cold environments

mesophilic microbes.

Microorganisms can be classified into groups by their growth temperature optima. midrange, most commonly studied

heperthermophilic microbes.

Microorganisms can be classified into groups by their growth temperature optima. very high, found in extremely hot habitats organisms with optima greater than 80°C • inhabit hot environments, including boiling hot springs and seafloor hydrothermal vents, that can experience temperatures in excess of 100°C Hyperthermophiles in hot springs >>>• chemoorganotrophic and chemolithotrophic species present • >>>generation times (g) as low as one hour common • >>>high prokaryotic diversity (both Archaea and Bacteria represented)

Lipopolysaccharides, (LPS)

Most of cell wall composed of outer membrane or the lipopolysaccharide (LPS) layer. • >>>barrier against antibiotics and other harmful agents • >>>LPS consists of core polysaccharide, O-polysaccharide, and lipid A. •>>> LPS replaces most of phospholipids in outer half of outer membrane. • >>>endotoxin: lipid A, the toxic component of LPS

Molecular adaptation to osmolarity.

Osmosis: Water diffuses from high to low concentrations. Typically, the cytoplasm has a higher solute concentration than the surrounding environment; thus, the tendency is for water to move into the cell (positive water balance). • When a cell is in an environment with a higher external solute concentration, water will flow out unless the cell has a mechanism to prevent this. Osmophiles: organisms that live in environments high in sugar as solute • Compatible solutes: used by cell to maintain positive water balance • >>pumping solutes from environment into cell • >>>synthesizing cytoplasmic solutes • >>>highly water-soluble, (e.g., sugars, alcohols, glycine betaine, KCl)

Molecular adaptation to life at high temperature.

Protein and membrane stability at high temperatures • >Enzymes and proteins function optimally at high temperatures, features that provide thermal stability. • >>>Critical amino acid substitutions in a few locations provide more heat-tolerant folds. • >>>Increased number of ionic bonds between basic and acidic amino acids resists unfolding in the aqueous cytoplasm. • >>>highly hydrophobic interiors • >>>Production of solutes (e.g., di-inositol phosphate, diglycerol phosphate) helps stabilize proteins. • >enzymes commercially useful • >>>prolong shelf life, (e.g., Taq polymerase for polymerase chain reaction) • Protein and membrane stability at high temperatures • >modifications in cytoplasmic membranes to ensure heat stability • >>>Bacteria have lipids rich in long-chain and saturated fatty acids, fewer unsaturated fatty acids. • >>>Archaea have C40 hydrocarbons made of repeating isoprene units bonded to glycerol phosphate, and membrane forms lipid monolayer rather than bilayer.

Sterilization using autoclaving versus pasteurization.

The autoclave is a sealed device that uses steam under pressure. • allows temperature of water to get above 100°C • kills endospores • not the pressure but the high temperature that kills the microbes • Pasteurization is the process of using precisely controlled heat to reduce the microbial load in heat-sensitive liquids. • does not kill all organisms, so it is different from sterilization

Advantages for microbes to be small

The cultivation of microorganisms is foundational to microbiology. A microbial culture is a collection of cells that are grown in/on nutrient mediums which is a liquid or solid mixture that contains all of the nutrients required for a MO to grow. A single microbial cell can grow to form a visible colony. The formation of visible colonies makes it easier to see and grow MO.

Description and significance of Redox tower.

The redox tower represents the range of possible reduction potentials. • Substances toward the top (reduced) prefer to donate electrons. • Substances toward the bottom (oxidized) prefer to accept electrons. • The farther the electrons "drop," the greater the amount of energy released (ΔE0′). • Oxygen (O2): strongest significant natural electron acceptor

Bright field microscopy,

The specimen are visualized because of differences in contrast (density) between specimen and surroundings • pigmented microbes add contrast

Fermentation versus respiration.

Two reaction series are linked to energy conservation in chemoorganotrophs: fermentation and respiration. • fermentation: anaerobic catabolism in which organic compounds donate and accept electrons • respiration: aerobic or anaerobic catabolism in which a donor is oxidized with O2 (aerobic) or another compound (anaerobic) as an electron acceptor

bactericidal antibacterial agents.

agents bind tightly and kill the cell.

Bacteriostatic antibacterial agents.

agents inhibit biochemical processes such as protein synthesis and bind weakly.

and bacteriolytic antibacterial agents.

agents kill by lysis (e.g., detergents).

Enzymes,

biological catalysts • typically proteins (some RNAs) • highly specific • active site: region of enzyme that binds substrate Many enzymes contain small nonprotein, nonsubstrate molecules that participate in catalysis (prosthetic groups and coenzymes)

microaerophiles,

can use oxygen only when it is present at levels reduced from that in air due to limited respiration or oxygen sensitivity

Limitations of microscopic cell counts.

cannot distinguish between live and dead cells without special stains • Precision is difficult to achieve. • Small cells can be overlooked. • phase-contrast microscope required if a stain is not used • cell suspensions of low density (< 106 cells/ml) hard to count • Motile cells need to immobilized. • Debris in sample can be mistaken for cells.

Anaerobes:

cannot respire oxygen

Chemoorganotrophs

conserve energy from organic chemicals.

Louis Pasteur contributions,

discovered that living organisms discriminate between optical isomers • discovered that alcoholic fermentation was a biologically (not just chemically) mediated process • Using the swan necked Pasteur flask, he disproved theory of spontaneous generation: Life arose spontaneously from nonliving material. • led to sterilization methods and food preservation • developed vaccines for anthrax, fowl cholera, and rabies

Simple transport,

driven by proton motive force • either symport: solute and H+ cotransported in one direction • >>>E. coli lac permease, phosphate, sulfate, other organics • or antiport: solute and H+ transported in opposite directions

lipid A,

endotoxin: lipid A, the toxic component of LPS

optimum temp

enzymatic reactions occurring at maximal possible rate

Robert Koch contributions,

experimentally demonstrated the link between microbes and infectious diseases (germ theory of infectious disease) • identified causative agents of anthrax, tuberculosis, and cholera • Koch's postulates • developed solid media for obtaining pure cultures of microbes • observed that masses of cells (called colonies) have different shapes, colors, and sizes • awarded Nobel Prize for Physiology and Medicine in 1905

Fimbriae and pili.

filamentous protein structures ~2-10 nm wide • Fimbriae enable organisms to stick to surfaces or form pellicles (thin sheets of cells on a liquid surface). • Pili are typically longer, and fewer (1 or a few) found per cell than fimbriae. • >>>Conjugative/sex pili facilitate genetic exchange between cells (conjugation). • >>>Type IV pili stick to host tissues and support twitching motility (e.g., Pseudomonas and Moraxella).

Pseudomurine,

found in cell walls of certain methanogenic Archaea • polysaccharide similar to peptidoglycan • composed of N-acetylglucosamine (in peptidoglycan) and N-acetyltalosaminuronic acid (different, replaces N-acetylmuramic acid) • β-1,3 glycosidic bonds instead of β-1,4 • amino acids all L-stereoisomer • cannot be destroyed by lysozyme and penicillin Cell walls of some Archaea lack pseudomurein. • contain other polysaccharide polymers

Difference between prokaryotic cell and eukaryotic cell in size structure and genome structure.

genome: a cell's full complement of genes • eukaryotic DNA • linear chromosomes within nucleus • much larger/more DNA (up to billions of base pairs) • prokaryotic DNA • generally single circular chromosome that aggregates to form the nucleoid region • may also have plasmids (extrachromosomal DNA) that confer special properties (e.g., antibiotic resistance) • small, compact (0.5-10 million base pairs)

Role and molecular structure of gas vesicles.

give buoyancy in planktonic cells • Conical-shaped, gas-filled structures made of protein • Impermeable to water and solutes • Molecular structure • >>>Gas vesicles are composed of two proteins, GvpA and GvpC. • >>>function by decreasing cell density, increasing buoyancy

Difference between Gram positive and Gram negative bacterial cell walls.

gram stain differences because of cell wall structure • Bacteria can be divided into two major groups: • Gram-positive bacteria appear purple-violet, and gram-negative bacteria appear pink.

phase contrast microscopy,

improves image contrast of unstained, live cells • Phase ring amplifies differences in the refractive index of cell and surroundings. • Resulting image—dark cells on a light background

obligate anaerobes,

inhibited or killed by oxygen, (e.g., some Bacteria and Archaea, few fungi, and few protozoa)

coenzymes.

loosely bound • Most are derivatives of vitamins

minimum temp

membrane gelling(solidify/harde/thicken); transport processes so slow that growth cannot occur

S-Layers.

most common cell wall type • consist of protein or glycoprotein • paracrystalline structure • in many organisms, this layer is present in addition to other cell wall components, usually polysaccharides • always outermost layer

transmission electron microscopy,

much greater resolving power (0.2 nm) than light microscope • enables visualization of structures at the molecular level • Specimen must be very thin (20-60 nm) and stained with high atomic weight substances that scatter electrons well and improve contrast. • Negative staining allows direct observation of intact cells/components

Negative and positive impacts of microbes on food.

negative impacts • can cause food spoilage and foodborne disease • harvest, storage, safety, prevention of spoilage influenced by microbes positive impacts • improving food safety, preservation • dairy products (e.g., cheeses, yogurt, buttermilk) • other food products (e.g., sauerkraut, kimchi, pickles, chocolate, coffee, leavened breads, beer)

Roles of capsules and slime layers.

not considered part of cell wall because these do not confer significant structural strength • polysaccharide layers • >>>may be thick or thin, rigid or flexible • capsule: if tightly attached, tight matrix; visible if treated with India ink • slime layer: loosely attached, easily deformed (e.g.,Leuconostoc) They both assist in attachment to surfaces • role in development and maintenance of biofilms • protect against phagocytosis • prevent dehydration/desiccation

psychrotolerant microbes.

organisms that can grow at 0°C but have optima of 20°C to 40°C • More widely distributed in nature than psychrophiles • isolated from soils and water in temperate climates and food at 4°C

facultative,

organisms that can live with or without oxygen

chemolithotrophs.

oxidize inorganic compounds (H2, H2S, NH4+).

Functions of cytoplasmic membranes.

permeability barrier • >>Polar and charged molecules must be transported. • >>>Transport proteins accumulate solutes against the concentration gradient. • protein anchor • >>holds transport proteins in place • energy conservation and consumption • >>generation of proton motive force

Prokaryotic versus eukaryotic cells •

prokaryotes • Bacteria and Archaea • no membrane-enclosed organelles (membrane-enclosed structures), no nucleus • eukaryotes • plants, animals, algae, protozoa, fungi • contain organelles • DNA enclosed in a membrane-bound nucleus .

maximum temp

protein denaturation; collapse of the cytoplasmic membrane; thermal lysis

Carl Woese contributions.

realized rRNA sequences could be used to infer evolutionary relationships. • discovered rRNA from methanogens distinct from Bacteria and Eukarya • named new group Archaea •found relationships can be deduced by comparing genetic information in the different specimens

Aerobes,

require oxygen (respiration) and grow at full oxygen tension (~21 percent)

periplasm,

space located between cytoplasmic and outer membranes • ~15 nm wide • houses many extracellular proteins

prosthetic group,

tightly bound • usually bind covalently and permanently (e.g., heme in cytochromes)

Generation time,

time required for microbial cells to double in number • depends on nutritional and genetic factors and temperature • example: Escherichia coli = 20 minutes

aerotolerant anaerobes.

tolerate oxygen and grow in its presence even though they cannot respire

Porines,

transmembrane protein channels for entrance and exit of solutes

Overview of the gram-positive cell wall •

up to 90 percent peptidoglycan • common to have teichoic acids (acidic substances) covalently bound to peptidoglycan • >>>bind divalent metal ions (e.g., Ca+2 and Mg+2) prior to transport • >>>lipoteichoic acids: teichoic acids covalently bound to membrane lipids

fluorescence microscopy,

used to visualize specimens that fluoresce (emit light after illumination with different wavelength) • Cells appear to glow on black background due to filters. • fluoresce naturally (autofluorescence) or after they have been stained with a fluorescent dye such as DAPI • widely used in microbial ecology to count bacteria in natural samples

confocal scanning laser microscopy,

uses a computerized microscope with a laser source to generate a 3D image • Computer can focus the laser on single layers of the specimen. • Different layers can then be compiled for a three- dimensional image.

differential interference microscopy,

uses a polarizer to create two distinct beams of polarized light (light in single plane) • gives structures such as nuclei, endospores, vacuoles, and inclusions a 3D appearance

Enzymecatalysis

• Catalysis depends on: • >>>substrate binding • >>position of substrate relative to catalytically active amino acids in active site • Endergonic and exergonic reactions coupled • >>>example: ATP hydrolysis or proton motive force

Lower limits of cell size

• Cellular organisms <0.15 μm in diameter are unlikely. • >>>>need volume to house proteins, nucleic acids, ribosomes, and so on • Open oceans tend to contain small cells (0.2-0.4 μm in diameter) known as "ultramicrobacteria." • >>>>Genomes are highly streamlined, missing functions that must be supplied by other microbes or hosts (plants and animals).

Species of Bacteria separated into two groups based on Gram stain

• Gram-positives and gram-negatives have different cell wall structures. • gram-negative cell wall (also called cell envelope) >>>• at least two layers: LPS and peptidoglycan • gram-positive cell wall • >thicker, primarily one layer of peptidoglycan

Stationary phase

• Growth rate of population is zero. • Either an essential nutrient is used up or waste products accumulate. • Metabolism continues at greatly reduced rate. • Some cells grow while others die, balancing each other.

Death phase

• If incubation continues after cells reach stationary phase, the cells will eventually die. • exponential rate • typically much slower than exponential growth • Viable cells remain for months or years.

Chemolithotrophy

• Microorganisms demonstrate a wide range of mechanisms for generating energy one of them being this (others are anaerobic respiration and phototrophy) uses inorganic chemicals as electron donors • >>>examples: (H2S), (H2), (Fe2+), (NH4+) • >>>waste products of chemotrophs • typically aerobic • begins with oxidation of inorganic electron donor • electron transport generates proton motive force • autotrophic; uses CO2 as carbon source

Sergei Winogradsky contributions,

• demonstrated that specific bacteria are linked to specific biogeochemical transformations (e.g., N and S cycles) • proposed concept of chemolithotrophy = oxidation of inorganic compounds to yield energy •demonstrated chemolithotrophs use carbon from CO2 (autotrophy) • first to demonstrate nitrogen fixation (Clostridium pasteurianum) and nitrification

Hydrogenosomes, what are they and they work?

• found in anaerobic, strict fermenters (e.g., Trichomonas and some protists) • similar size to mitochondria • lack TCA cycle enzymes and cristae • Major function is oxidation of pyruvate to H2, CO2, and acetate. • >>>Some methanogenic Archaea live in some anaerobic eukaryotes and consume H2 and CO2, producing CH4; acetate is secreted.

Lag phase

• interval between inoculation of a culture and beginning of growth • time needed for biosynthesis of new enzymes and to produce required metabolites before growth can begin

advantages to being small

• more surface area relative to cell volume than large cells (i.e., higher S/V ratio). • support greater nutrient and waste product exchange per unit cell volume • tend to grow faster than larger cells • Mutations lead to faster evolution. • Eukaryotic cells adapt slower.

The Great Plate Count Anomaly"

=Direct microscopic counts of natural samples reveal far more organisms than those recoverable on plates. • Why is this? • >>>Microscopic methods count dead cells, whereas viable methods do not. • >>>Different organisms may have vastly different requirements for growth.

log phase

(exponential phase) Cells in this phase are typically in the healthiest state.

Koch Postulates,

1. The suspected pathogen must be present in all cases of the disease and absent from healthy animals.- We could use Microscopy/ staining in order to look at the blood/ tissue of the diseased animals and the healthy animals. We would see that S.pyogenes was only in the diseased animals 2. The suspected pathogen must be grown in pure culture. - We would then streak one agar plate with sample from a diseased animal and another plate from a healthy animal. We would see that there would be colonies of the suspected pathogen in the one from the diseased animal and no organism present from the one of the healthy animal. 3. Cells from a pure culture of the suspected pathogen must cause disease in a healthy animal. - We would then put inoculate a healthy animal with cells of suspected pathogen. and then the animal would die. 4. The suspected pathogen must be preisolated and shown to be the same as the original. - we would then remove blood or tissue sample from the dead animal and observe by microscopy.

Advantages and disadvantages of turbidimetric measures of microbial cell numbers.

Advantages • quick and easy to perform • typically do not require destruction or significant disturbance of sample • Same sample can be checked repeatedly. • Disadvantages • sometimes problematic (e.g., microbes that form clumps or biofilms in liquid medium)

Properties of all microbial cells and properties of some microbial cells.

All Cells: Metabolism, growth, evolution Some cells: Differentiation, Communication , Genetic exchange, Motility

ABC transporters.

Also called ATP-binding cassette) systems • 200+ different systems identified in prokaryotes for organic and inorganic compounds • high substrate affinity • ATP drives uptake. • requires transmembrane and ATP-hydrolyzing proteins plus: • >>>Gram-negatives employ periplasmic binding proteins. • >>>Gram-positives and Archaea employ substrate-binding proteins on external surface of cytoplasmic membrane.

Compare and contrast the chemical composition and structure of the cytoplasmic membranes found in Bacteria and Archaea. What is the advantage of the archaeal membranes in relationship to the types of environments archaea may inhabit?

Answer: Bacterial cytoplasmic membranes contain fatty acids with ester linkages that always form a phospholipid bilayer that is highly fluid. Archaeal cytoplasmic membranes contain glycerol ethers that are either diether or tetraethers. Glycerol diether molecules form a bilayer membrane and glycerol tetraether form a monolayer membrane. Both the ether linkages and the monolayer structure are more rigid than the bilayer ester-linked fatty acids and are more stable at high temperatures and pressures. Archaea tend to inhabit more extreme environments in terms of temperature, pressure, and salt, thus the more stable membrane components allow archaea to survive under these conditions.

Why is 100% ethanol less effective as an antimicrobial compared to a 70-95% concentration of ethanol?

Answer: 70% percent alcohol is ideal as opposed to a stronger solution. Pure alcohol coagulates protein on contact. If 100% alcohol is poured over a single-celled organism, the alcohol goes through the cell wall of the organism in all direction and coagulates(solidify, hardens, thickens) the proteins found in the cell membrane. The coagulated membrane proteins would then stop the alcohol from penetrating further into the cell, preventing denaturation of cytoplasmic proteins. If this happened, the cell would become inactive but not dead. Under favorable conditions, the cell would then begin to function. If 70 percent alcohol is poured over a single celled organism, the diluted alcohol also coagulates the protein, but at a slower rate, so that it is able to penetrate all the way through the cell before protein coagulation can block it. Thus, the protein found in the entire cell is coagulated and the organism dies.

Consider a pizza dough made by vigorously mixing to form gluten and evenly disperse the ingredients such as bakerʹs yeast (Saccharomyces cerevisiae). Predict the metabolic differences yeast would have in a thinly flattened dough and in a spherical dough ball.

Answer: A flattened dough would have higher surface area and more oxygen exposure to support aerobic respiration of S. cerevisiae. The dough ball on the other hand would initially have aerobic metabolism of S. cerevisiae due to the mixing. Once oxygen is depleted from respiration the yeast would begin anaerobic fermentation, especially in the center of the dough ball while the surface of the dough ball could still support aerobic growth if not enclosed in a container.

Explain why being small is advantageous for cells and how it affects growth rates.

Answer: A smaller cell has a greater surface area to volume ratio than a larger cell. This means that smaller cells can more rapidly exchange materials with their surroundings because there is so much surface membrane relative to the internal volume. This increases growth rates, meaning that smaller cells generally grow faster than larger cells.

Knowing the concentration of microorganisms in a sample is often an important consideration for environmental, industrial, and medical microbiologists, yet a microscope is sometimes not used to accomplish this task. Explain five major limitations to using a light microscope to directly count microorganisms.

Answer: All of the following eight are potential issues associated with the direct counting method: 1) live and dead cells are indistinguishable without special staining, 2) low cell concentrations are challenging to enumerate, 3) motile cells must be immobilized, 4) precision can be quite poor with manually counting hundreds or thousands of cells, 5) small cells are difficult to observe and count even at 1,000X total magnification, 6) small particulates can be mistaken for microbial cells, 7) unstained cells require more sophisticated microscopy, and 8) it is very time-consuming.

Construct a chart to show at least five major differences between the cytoplasmic membrane and cell wall of bacteria and archaea. What are the implications of these differences?

Answer: Answers could include the following: the abundance or presence of amino acid stereoisomers, polysaccharides, pseudomurein and S-layers, action of lysozyme and penicillin, and type of glycosidic bonds.

Describe two capabilities of microbes that exemplify their dynamic nature in interacting with their environment.

Answer: Answers could possibly include cell-cell communication, ability to move (motility), ability to differentiate, and exchange of materials (any two).

Explain why the amount of energy released in a redox reaction depends on the nature of both the electron donor and the electron acceptor.

Answer: Answers should emphasize that energy does not come from specific molecules but rather from the difference in reduction potential between two molecules. For example, assigning arbitrary values and subtracting them from one another by comparing two different electron acceptors to one donor would indicate differences in energy for an electron acceptor. In a similar way, this could also be shown to mathematically explain electron donors having an equal role in determining ΔE0ʹ.

Categorize the circumstances under which the same substance (molecule) can be either an electron donor or an electron acceptor.

Answer: Answers should explain that not all molecules are strictly one or the other, and each molecule must be compared to the other in a pair to determine which is the electron acceptor and which is an electron donor.

Explain the differences between symporters, and antiporters.

Answer: Answers should highlight differences in transport direction and energy input. Both are forms of simple transport which is driven by proton motive force. Symport- the solute and H+ cotransported in one direction. Ecoli lac permease, phosphate, sulfate, other organics Antiport- solute and H+ transported in opposite directions

A beer-making microbiologist noticed that no matter how long the brewing process went, 3% alcohol was the maximum produced. Hypothesize what is causing this low level of alcohol in reference to the brewer's recipe and recommend how a higher alcohol yield could be achieved. Ethanol is toxic at high concentrations, but ignore this factor to focus on microbial metabolism.

Answer: Answers will vary but one explanation is a low substrate concentration resulted in low fermentation to produce ethanol. Providing more carbohydrates such as glucose to the yeast in the recipe for the same growth period would increase fermentation activity and ethanol production. Another explanation is that there may be too much oxygen introduced during the brewing process, which would result in the complete oxidation of glucose instead of fermentation to ethanol. The brewer would need to take more precautions to exclude oxygen during brewing.

Explain why infectious diseases are much less lethal in developed countries than in underdeveloped countries.

Answer: Answers will vary but should emphasize ways in which increased knowledge about microbial pathogenesis has influenced preventative care (e.g., sanitation) and treatment (e.g., antimicrobial drugs).

Compare and contrast the functions microbes serve in the digestive systems of both humans and rumens (e.g., cattle).

Answer: Answers will vary but should focus on humans having a high cell localized density in the colon (large intestine), whereas rumens have higher microbial populations in the rumen. Microbes in both systems aid in digestion and improve nutrition/health of the host. Microorganisms in the digestive system of rumen are important, as they help to digest and ferment cellulose. This is significant, as rumens ingest a lot of cellulose-rich food such as grass and hay and need these microorganisms to digest their food. The human colon lacks a large number of cellulose-degrading microorganisms but instead has a large number of microbial cells that assist in digestion by synthesizing vitamins/nutrients and compete with pathogenic microorganisms for space.

In an aquatic microbial community where a photoautotroph, chemoorganoheterotroph, and nitrogen fixing bacterium are present, predict an environmental perturbation that would cause only one to be outcompeted by the other two groups and explain how each group would respond.

Answer: Answers will vary but should highlight a unique feature of one of the groups such as: photoautotrophs are sensitive to photon (light) availability, chemoorganoheterotrophs require organic molecules for carbon, and nitrogen fixing bacteria use N2 gas.

Explain why microbial cells are excellent models for understanding cell function in higher organisms.

Answer: Answers will vary but should include commonality of function, biochemical and genetic similarities, and ease and speed with which they can be grown in large quantities.

Explain why prokaryotes tend to survive and adapt more rapidly to extreme and dynamic environmental conditions than eukaryotes.

Answer: Answers will vary, but should reflect an understanding of how the higher surface-to- volume ratio influences the growth rate and total accumulation of mutations in prokaryotes. Another feature that increases mutation rate is the haploid nature of prokaryotes. Lastly, answers could mention that the rigid cell walls and various changes in the cytoplasmic membrane make it easier for prokaryotes to survive in unusual and extreme environments.

Explain what an enzyme must accomplish to catalyze a specific reaction.

Answer: Answers will vary, but the focus of the answer should be on overcoming the required activation energy.

Explain how you would use Robert Koch's postulates to determine that Streptococcus pyogenes is the causative agent of streptococcal pharyngitis ("strep throat").

Answer: Answers will vary but will need to detail how S. pyogenes will be subjected to all four postulates. 1. The suspected pathogen must be present in all cases of the disease and absent from healthy animals.- We could use Microscopy/ staining in order to look at the blood/ tissue of the diseased animals and the healthy animals. We would see that S.pyogenes was only in the diseased animals 2. The suspected pathogen must be grown in pure culture. - We would then streak one agar plate with sample from a diseased animal and another plate from a healthy animal. We would see that there would be colonies of the suspected pathogen in the one from the diseased animal and no organism present from the one of the healthy animal. 3. Cells from a pure culture of the suspected pathogen must cause disease in a healthy animal. - We would then put inoculate a healthy animal with cells of suspected pathogen. and then the animal would die. 4. The suspected pathogen must be preisolated and shown to be the same as the original. - we would then remove blood or tissue sample from the dead animal and observe by microscopy.

Compare and contrast the leading causes of death in 1900 with the leading causes of death today. What roles have microbiologists played in the dramatic changes that are evident?

Answer: Answers will vary, but a focus should be that pathogens that killed people in the early 1900s are now treatable due to knowledge learned from microbiologists. ---------------During this century we have better control of infectious diseases, that are caused by bacterial and viral pathogens, then we did in the last century. In the 1900s the leading causes of death were influenza and pneumonia, TB, and gastroenteritis which are all from infectious diseases, but since microbiologist have know learned more knowledge about treatments for these disease caused by pathogens we now in todays words have different leading causes of death and the top ones are all non microbial diseases (they are Hear disease, cancer, stroke etc)

Microbes were first formally observed during the mid-1600s, but the cell theory was not enunciated until 1839. Write a brief essay explaining why microbiology did not become a formally recognized science until Louis Pasteur's and Robert Koch's time.

Answer: Answers will vary, but a theme should be the lack of powerful microscopy tools. Without sufficient microscopes individual cells could not be seen, but the activities of microorganisms could be observed, such as the production of ethanol in Louis Pasteur's experiments on fermentation.

Enumerating the viable and total cell concentration of a population of a microbial isolate can be a laborious task for a microbiologist, especially when studying the same isolate for several years. It is often more practical to determine the relationship between optical density (OD) and cell concentration. Once this relationship (determined by a standard curve) is determined, the OD of an isolate in a broth can be used to determine the populationʹs concentration. Why must a standard curve be prepared for each isolate when using OD measurements to determine cell concentration? Also describe an experiment that would generate this type of standard curve.

Answer: Answers will vary, but it should be noted that a particular OD value will correspond to a specific cell concentration due to different shapes and sizes of microorganisms. One such example could be a culture of Citricella sp. SE45 which was determined to have an OD540 of 0.485. This OD fit into the already-constructed standard curve to indicate that 5 × 105 viable cells are in each milliliter of broth, which can be helpful in experiments where the final cell concentration or number is critical. To initially create the standard curve, a traditional growth curve experiment could be performed over time where both OD and cell concentration (viable with viability plating or total with direct counts) would be measured. The values of OD and cell concentration for each time point would then be used to create a standard curve, and a linear regression curve would show the relationship of OD to cell concentration such that subsequent experiments OD could be measured as a proxy for cell concentration.

Compare and contrast the spread plate method and the pour plate method of doing plate counts. Also describe which group of organisms would not be quantifiable in the pour plate method but would still be observed in the spread plate method.

Answer: Answers will vary, but one benefit to the pour plate method is that larger volumes of cells can be dispensed, whereas large liquid volumes dispensed on the surface of agar plates is often impractical. Many cells, especially psychrophiles and some mesophiles, cannot withstand the warm temperature of molten agar (~50°C) and therefore must be spread on top of an agar plate. Colonies are easier to enumerate on a two- rather than three-dimensional surface that favors the spread plate technique over the pour plate method.

Provide evidence supporting the statement that an "ecosystem is controlled by microbial activities."

Answer: Answers will vary, but one example could be oxygen depletion, where a loss of oxygen would then favor anaerobic microorganisms. the metabolic activities of MO can change the habitat in which they live, both chemically and physically, and these changes can affect other organisms. For example, excess nutrients added to a habitat can cause aerobic MO (o2 consuming) to grow rapidly and consume o2, leaving the habitat anoxic (O2 free).

Using specific examples, explain why it is sometimes impossible to satisfy Robert Koch's postulates.

Answer: Answers will vary, but one issue is the consideration for a model animal host that will react to the (human) pathogen in the same manner as in a human host. For example, a chicken would not show flu-like symptoms when infected with the influenza virus. Another issue is the inability to cultivate some microorganisms outside of the host. Some pathogens can cause several different diseases meaning the diseased symptoms created could vary in the exp, some pathogens are caused in humans only such as HIV, some disease may be caused by multiple microbes, the animal may have different symptoms with the disease than a human would

Compare and contrast the works of Louis Pasteur and Robert Koch in terms of both applied techniques and basic science.

Answer: Answers will vary, but should highlight the differences between basic scientific research in which fundamental ideas are discovered opposed to the usage of microbiological principles to solve larger questions. Examples of Pasteur's basic science contributions are his work showing that fermentation was mediated by microorganisms and discovered that living organisms discriminate between optical isomers. Pasteur also applied his ideas to develop sterilization techniques. Robert Koch focused more on the application of microbiology to identify the cause of tuberculosis by developing pure culturing techniques and the four postulates to link microbes to a disease.

How would the presence of endospores in Louis Pasteur's nutrient solutions have affected his conclusions about spontaneous generation?

Answer: Answers will vary, but ultimately this could have confounded Pasteur if the endospores sometimes went into a vegetative growth phase and other times no growth was observed. If the nutrient solution in Pasteur's experiment was not properly sterilized and still some endospores were present in it, they could have grown in the media. Since there was no information about endospores was available that time, it could have been assumed that the growth have happened from the nonliving content of the media only and Theory of spontaneous generation could have become stronger.

Antibiotics such as penicillin interfere with the ability of bacteria to synthesize cell walls. Explain why cell walls are a good target for a useful antibiotic and whether you think that penicillin would be effective against a bacterial cell in an isotonic environment.

Answer: Because human cells do not use peptidoglycan, an antibiotic that affects peptidoglycan can harm bacterial cells with less risk of damage to human cells even though all medications may have some adverse effects (such as causing an allergic reaction). In an isotonic environment, a bacterial cell can survive without a cell wall and therefore penicillin would be less effective than in a hypotonic environment.

Describe beneficial and harmful ways in which microorganisms interact with agricultural crops.

Answer: Certain microbes are beneficial to crops when they produce nutrients (e.g., NH 4+, SO42-) usable by a crop from a substrate that was unusable. Other microbes can cause diseases in plants, much like pathogens cause disease in humans. For example, nitrogen fixing bacteria will convert nitrogen into NH3 through nitrogen fixation. NH3 is the major nutrient in fertilizer and is used as a nitrogen source for plant growth. Another example of a beneficial microbe is cellulose-degrading microbes in rumen.

Elaborate on why discovering endospores was important to microbiology.

Answer: Endospores allow bacteria to survive boiling water, UV light, and extreme desiccation, as well as allowing them to be in a dormant state for very long periods of time. Two possible themes could be how the discovery of endospores changed sterilization procedures for food and surgical instruments, as well as a change in our view of how bacteria can survive, such as surviving in space and surviving 1,000s of years in a dormant state.

Why is energy required for nutrient transport? Give an example of a system that transports nutrients and describe what source of energy is used to move the nutrients into the cell.

Answer: Energy is required for nutrient transport because nutrient concentration outside of the cell is lower than the nutrient concentrations inside the cell, thus nutrient transport moves solutes against a concentration gradient and requires energy. There are three examples in the text: (i) Simple transporter(proton motor force): such as lac permease. Each nutrient molecule is cotransported into the cell with a H+ ion, thus the proton motive force provides the energy to transport nutrients. (driven by the energy in the proton motor force) (ii) Group translocation (another energy rich compound) : such as sugar phosphotransferases. Each nutrient molecule is modified during the transport process. modification, in this case, phosphorylation, releases energy, thus the energy source is an energy- rich compound such as phosphoenol pyruvate or some other phosphorylated compound. (chemical modification of the transported substance driven by phosphoenolpyruvate) (iii) ABC transporters (ATP). In this example specific binding proteins bind to nutrient molecules with high affinity. Movement of the nutrient into the cell is coupled to ATP hydrolysis, thus ATP is the source of energy for transporting nutrients. (periplasmic binding proteins are involved and energy comes from ATP)

Explain the difference between the division rate and the generation time.

Answer: Generation time (g) and division rate (v) are reciprocal values of each other; both are determined by monitoring exponential growth over time. Division rate is reported as the number of generations per time and is calculated by taking 1/g. The units for g are in time per generation, which is the doubling time during exponential growth.

Describe the autoclave, and explain the roles of time, temperature, and pressure in the functioning of the autoclave.

Answer: High pressure is used in an autoclave so that liquids do not boil at temperatures beyond 100°C. The heat (not pressure) is used to kill living cells and is usually used for 15 minutes at 121°C. The time must be increased for bulky samples to ensure the objects or liquids are heated to 121°C for at least 15 minutes.

Explain the present understanding of molecular adaptations to the cytoplasmic membrane that are present in thermophiles.

Answer: In thermophiles and most hyperthermophilic Bacteria, the cytoplasmic membrane has a higher content of long-chain and saturated fatty acids and a lower content of unsaturated fatty acids than are found in the cytoplasmic membranes of mesophiles. Saturated fatty acids form a stronger hydrophobic environment than do unsaturated fatty acids, and longer-chain fatty acids have a higher melting point than shorter-chain fatty acids; collectively these properties increase membrane stability.

Describe the mechanisms by which certain prokaryotes glide. What are the ecological advantages of gliding motility?

Answer: Mechanisms will vary depending on the organism described, some of which include the involvement of proteins in the cytoplasmic membrane, slime extrusion, and type IV pili. Examples of advantages could include biofilm formation on a surface, increased pathogenesis or movement towards a different habitat with new resources.

Explain the present understanding of molecular adaptations to the cytoplasmic membrane as found in psychrophiles.

Answer: Non-psychrophiles do not thrive in the cold environment, in part due to predominantly having large or saturated fatty acid chains in their cytoplasmic membrane, which become increasingly rigid and wax-like with colder temperatures. The psychrophiles have higher relative concentrations of short and unsaturated fatty acids in their membrane and therefore maintain a semi-permeable membrane when exposed to less heat.

Discuss why energy yield in an organism undergoing anaerobic respiration is less than that of an organism undergoing aerobic respiration.

Answer: One possible explanation could point to the substrate-level phosphorylation process itself as being less energy yielding than (aerobic) oxidative phosphorylation. Another reason is the fate of pyruvate itself, where fermentation is unable to take it through the higher energy yielding process, which requires O2 as a terminal electron acceptor. Other answers could discuss the E0ʹ being greatest with the O2/H2O redox couple in aerobic metabolism compared to anaerobic redox couples.

You have discovered a new bacterial strain that causes urinary tract infections. Closely related bacterial species cannot cause infections. You compare the strains and find that your new strain has structures composed of protein external to its cell wall. What structures might your new strain have that the other strains do not? Why?

Answer: Pili or fimbrae are the most likely structures that are found in the new pathogenic strain but missing in the nonpathogenic strains. Both pili and fimbrae aid in attachment of cells to surfaces and tissues. Attachment is important for pathogenesis. In addition, some pili are involved in twitching motility, which can help cells invade the body. (Other answers such as the capsule would only be partly correct, since the capsule is not made of protein. Flagella could also be an acceptable answer if they logically connect swimming motility to pathogenesis.)

Contrast fermentation and respiration in terms of electron donor, electron acceptor, type of ATP production, and relative number of ATP produced.

Answer: Respiration should be distinguished as using separate electron donors and acceptors (such as organic carbon as the electron donor and oxygen as the electron acceptor), while fermentation splits organic molecules in order to oxidize one part of the molecule and reduce the other part in order to regenerate NAD+. Fermentation uses substrate level phosphorylation to generate relatively few ATP, while respiration uses oxidative phosphorylation to generate more ATP.

Explain the biosynthetic and bioenergetic roles of the citric acid cycle.

Answer: Some of the molecules generated during the CAC, such as alpha- ketoglutarate, oxalacetate, and succinyl-CoA, can serve as precursors for the biosynthesis of critical cellular components such as amino acids, chlorophyll, and cytochromes. The bioenergetic component of the cycle should be described in the context of FADH2 and NADH electron donors storing energy potential, usable in electron transport where O2 is reduced to water.

Describe the makeup of the phospholipid bilayer. Include molecular orientation and proteins as well.

Answer: The fatty acid components of the lipids in the cytoplasmic membrane are oriented toward each other, whereas the glycerol-phosphates point out toward the cytoplasm and external environment. Proteins can span the entire membrane (both layers) or be embedded in the phospholipids on either side of the membrane.

You are studying swimming motility in a pathogenic bacillus. You create mutations in random genes and then test which mutations effect swimming motility by looking at the mutant cells under the microscope. One of the mutant bacteria cannot swim anymore, but still rotates around in one spot when you watch them. Using electron microscopy you discover that some parts of the flagella are still present in the cell wall, but no long flagella are visible. Which gene do you think is mutated (i.e., missing) and which motility-related parts are still present in this mutant?

Answer: The flagellar apparatus is put together in a particular order first the MS ring, anchoring proteins, and the hook extend off of the cytoplasmic membrane. Flagellin proteins then pass through the narrow filament channel and cap proteins finally are put onto the end when roughly 20,000 flagellin have been assembled. Flagellar growth thus occurs after the assembly of the basal body, rings, and hook. The basal body provides the rotation and uses the proton motive force to rotate the basal body. From the information given above, one can infer that the basal body and hook can still rotate, but the flagellin proteins are missing or defective so that no long flagella can form. This would explain why the cells might still rotate in place, but not be able to move forward rapidly.

Predict what would happen to a motile bacterium undergoing chemotaxis if the Mot proteins suddenly ceased to function.

Answer: The function of Mot proteins should be described producing the rotation of the flagella. This would probably allow flagella to form, but they would not be able to rotate. The bacterium would not be able to move, but the flagella would still look normal.

What is the function of an endospore and how is an endospore formed?

Answer: The function of an endospore is to allow the cell to survive harsh conditions by going into a dormant state. An environmental trigger, such as nutrient depletion or dehydration triggers the formation of compounds such as SASP and dipicolinic acid to protect the DNA from damage and reduce the water content inside the endospore. These changes protect the DNA and other critical chemical components from heat, desiccation, and UV exposure. The function of an endospore is not for reproduction because every cell forms only one endospore. (The amount of detail expected in this answer should commensurate with the amount of detail discussed in class.)

Why are microbial doubling times in nature typically longer than those obtained in the laboratory?

Answer: The general theme should be on how lab conditions are designed to optimally grow a microorganism (e.g., pH, substrate(s), temperature, vitamins) and minimize pressure. In the environment this microbe often is in conditions that are less than ideal, where they likely will encounter competition for growth substrates and other essential chemicals.

Explain why a hyperthermophile is unlikely to be a human pathogen.

Answer: The human bodyʹs temperature is approximately 37°C, which is not a favorable environment for a hyperthermophile, which has an optimal growth temperature of 80°C or higher.

Use the formulas N = N02n and g = t/n. N = final cell number, N0 = initial cell number, n = number of generations, t = time, g = generation time. Find the generation time if N = 2 × 108, N0 = 3 × 106 and t = 3 hours.

Answer: The number of generations (n) must first be determined through the rearranged formula: n = [(log N)-(log N0)]/(log 2) to determine n = 6.059 generations. Finally, the generation time (g) = t/n, which equates to nearly 0.5 hours per generation or almost 30 minutes per generation.

Summarize the roles the proton motive force has in microbial metabolism.

Answer: The proton motive force uses an energized cell membrane for ATP synthesis via ATPase, transporting some ions and molecules into and out of the cell, and flagellar rotation.

Explain why a eukaryotic cell needs membrane-enclosed lysosomes and peroxisomes.

Answer: The structures both contain high concentrations of enzymes that serve as a localized region to perform specific reactions. Enclosed membranes provide a barrier to maintain conditions within the lysosomes and peroxisomes that are unlike those in the cytoplasm, which is important because environmental conditions for these processes to be optimal are often different than the conditions in the cytoplasm.

What type of microscope would you use to visualize the internal structures of a chloroplast? Support your answer with evidence based on the size of the structures you want to see and the resolution and magnification power of different types of microscopes.

Answer: Transmission electron microscopy would be necessary to visualize the internal structures of a chloroplast. Chloroplasts are less than 5 μm in diameter and the internal membranes are only 10 nm thick. Light microscopes only have a resolution of 200 nm, thus any structure less than 200 nm will not be visible. Individual chloroplasts could be seen with a light microscope, but not the structures inside. Scanning electron microscopy can only see external features because electrons cannot penetrate the cell, thus the cell must be sectioned and prepared for transmission electron microscopy to see the inside of the chloroplasts.

Discuss the pros and cons of using radiation to sterilize items such as foods, drugs, and surgical supplies.

Answer: UV radiation sterilizes only the surfaces of objects which can be problematic for foods that are not fully cooked on the inside. Ionizing radiation can penetrate so they are ideal for foods such as grain cereals and ground beef. The high costs associated with the specialized equipment make it feasible only for large industrial applications. Due to the high energy of some of these radiation procedures, there are many concerns of potential radioactive contamination, production of carcinogenic or toxic products, and nutritional value alteration.

Would you expect a xerophilic organism to be halotolerant? Why or why not?

Answer: Yes, xerophiles are typically halotolerant. For example, the xerophilic halotolerant fungus Xeromyces bisporus inhabits cereal, candy, and dried fruit, all of which are very dry environments that also contain salt. With all else being equal, a decrease in solvent (such as water) translates to an increase in solute (salt) concentration, and therefore a microbe capable of tolerating a low water activity likely can also tolerate higher salt concentrations.

Difference between Bacterial and Archaeal membranes composition.

Bacterial cytoplasmic membrane • General structure is phospholipid bilayer containing embedded proteins. • contain both hydrophobic (water-repelling) and hydrophilic (water-attracting) components • hydrophobic = fatty acids • hydrophilic = glycerol + phosphate and another functional group (e.g., sugars, ethanolamine, choline) • Fatty acids point inward to form hydrophobic environment; hydrophilic portions remain exposed to external environment or the cytoplasm. 8-10 nm wide • some species strengthened by hopanoids (sterol-like molecules) • embedded proteins including integral membrane proteins (significantly embedded) or peripheral membrane proteins (loosely attached) Archaeal membranes • ether linkages in phospholipids of Archaea in contrast to Bacteria and Eukarya that have ester linkages in phospholipids • Archaeal lipids have isoprenes instead of fatty acids. • Major lipids are phosphoglycerol diethers with phytanyl C20 side chains and diphosphoglycerol tetraethers with biphytanyl C40 side chains, which can form lipid monolayers.

Batch culture versus a Chemostat, .

Batch culture: a closed-system microbial culture of fixed volume • Typical growth curve for population of cells grown in a closed system is characterized by four phases. • lag phase • exponential phase • stationary phase • death phase Continuous culture: an open system microbial culture of fixed volume • Chemostat: most common type of continuous culture device • Both growth rate and population density of culture can be controlled independently and simultaneously, depending on: • >>>Dilution rate: F/V (F is flow rate of adding fresh medium and removing spent medium, V is culture volume). • >>>concentration of a limiting nutrient. Experimental Uses • can maintain exponential growth phase for weeks/months • used to study physiology, microbial ecology and evolution, enrichment and isolation of bacteria from nature • growth rate controlled by dilution rate

Biofilm benefits to microbes.

Biofilms prevent harmful chemicals (e.g., antibiotics) from penetrating, prevent protists from grazing, and prevent washing away of cells. • Biofilms affect human health, water distribution systems, and fuel storage.

Minimum, optimum, and maximum temperatures.

Cardinal temperatures: the minimum, optimum, and maximum temperatures at which an organism grows Range is typically < 40°C

Proton Motive Force, Describe briefly how it works?

During electron transfer, protons are released on outside of the membrane. • >>>Protons originate from 1) NADH and 2) dissociation of water. • Results in generation of pH gradient and an electrochemical potential across the membrane (the proton motive force) • >>>The inside becomes electrically negative and alkaline (OH-). • >>>>The outside becomes electrically positive and acidic (H+). • Complex I (NADH: quinone oxidoreductase) • >>>NADH oxidized, quinone reduced, four H+ released • Complex II (succinate dehydrogenase complex) • >>>bypasses Complex I • >>>feeds e- and H+ from FADH2 directly to quinone pool • >>>four fewer H+ pumped per 2e-; reduces ATP yield • Complex III (cytochrome bc1 complex) • transfers e- from ubiquinol (reduced quinone) to cytochrome c • pumps H+ • Cytochrome c shuttles e- to Complex IV. • Complex IV (cytochromes a and a3 ) • >>>terminal oxidase; reduces O2 to H2O • pumps H+ • ATP synthase (ATPase): complex that converts proton motive force into ATP; two components: • >>>F1: multiprotein extramembrane complex extending into cytoplasm • >>>Fo: membrane-integrated proton-translocating multiprotein complex • >>>reversible catalysis of ADP + Pi to ATP • >>>consumes three to four H+ per ATP; three ATP produced per two e- • ATPases in strict fermenters generate proton motive force for flagellar rotation and transport.

Role of catalase and peroxidase in reducing toxicity to reactive oxygen species.

Enzymes are present to neutralize most of these toxic oxygen species. • Catalase and peroxidase convert H2O2 to O2 and H2O. pic

Difference between prokaryotic and eukaryotic flagella.

Flagella and cilia in eukaryotes • organelles of motility that allow cells to move by swimming • Cilia are short flagella. • structurally distinct from prokaryotic flagella and do not rotate; instead whip (flagella) or beat in synchrony (cilia) • bundle of nine pairs of microtubules surrounding a central pair of microtubules • Dynein is attached to the microtubules and uses ATP to drive motility.

Difference of flagella and archealla.

Flagella/archaella: structure that assists in swimming in Bacteria and Archaea, respectively • >>>tiny rotating machines • Flagellar and flagellation • >>>long, thin appendages (15-20 nm wide) • >>>different arrangements: polar, lophotrichous, amphitrichous, peritrichous • >>>increase or decrease rotational speed relative to strength of proton motive force Flagellar structure and activity >>>• helical in shape • >>>consists of several components • >>>Filament composed of flagellin. • >>>reversible rotating machine Flagellar synthesis • Several genes are required. • Filament grows from tip. • MS ring is made first. • Other proteins and hook are made next. Archaella • >>>half the diameter of bacterial flagella (10-13 nm) • >>>move by rotation • >>>composed of several different filament proteins with little homology to bacterial flagellin • >>>speeds vary from 0.1-10x Escherichia coli • >>>structurally similar to type IV pili

Endospore structure and function.

Formed during endosporulation or sporulation • Highly differentiated cells resistant to heat, harsh chemicals, and radiation • Survival structures to endure unfavorable growth conditions • "Dormant" stage of bacterial life cycle • Ideal for dispersal via wind, water, or animal gut • Present only in some gram-positive bacteria, (e.g., Bacillus and Clostridium) Formation and germination • >>>vegetative cell converted to nongrowing, heat-resistant, light-refractive structure • >>>only occurs when growth ceases due to lack of essential nutrient such as carbon or nitrogen >>>can remain dormant for years but converts rapidly back to being vegetative • three steps: activation, germination, and outgrowth Structure and features • many layers: exosporium (outermost), spore coats, cortex, core • contains dipicolinic acid • enriched in Ca2+ • Core contains small acid-soluble spore proteins (SASP), which bind and protect DNA and function as carbon and energy source for outgrowth.

Outcomes of glycolysis

Glycolysis : a common pathway for catabolism of glucose that forms two ATP • >>>Glucose can be fermented or respired. • >>>ATP produced by substrate-level phosphorylation: energy-rich phosphate bond from organic compound is transferred to ADP, making ATP Two ATPs are produced.

Size range for prokaryotic and eukaryotic cells

Size range for prokaryotes: 0.2 μm to >700 μm in diameter Size range for eukaryotic cells: 2 to >600 μm in diameter

scanning electron microscopy.

Specimen is coated with a thin film of heavy metal (e.g., gold). • An electron beam scans the object. • Scattered electrons are collected and projected to produce an image. • Even very large specimens can be observed. • magnification range of 15-100,000x • only surface visualized

Peptidoglycan structure and composition.

Structure of Peptidoglycan (Figure 2.10) • rigid layer that provides strength • typically composed of: • >>>alternating modified glucose (N-acetylglucosamine and N-acetylmuramic acid) in β-1,4 linkages • >>>amino acids L-alanine, D-alanine, D-glutamic acid, and either L-lysine or diaminopimelic acid (DAP) • >>>cross-linked differently in gram-negative bacteria and gram-positive bacteria (often "interbridges") • can be destroyed by lysozyme (enzymes that cleave glycosidic bond between sugars) • >>>found in human secretions, major defense against bacterial infection • 100+ distinct peptidoglycans have been described

group translocation,

Substance transported is chemically modified. • Energy-rich organic compound (not proton-motive force) drives transport. • best-studied system for example energy derived from phosphoenolpyruvate

How Chemotaxis works in E. coli?

Taxis: directed movement in response to chemical or physical gradients chemotaxis: response to chemicals Chemotaxis • best studied in E. coli (peritrichous) • "run and tumble" behavior • >>>run: smooth forward motion, flagellar motor rotates counterclockwise • >>>tumble: stops and jiggles, flagellar motor rotates clockwise, flagellar bundle comes apart • move by rotation • Bacteria respond to temporal, not spatial, difference in chemical concentration. • monitor/sample environment with chemoreceptors that sense attractants and repellents Measuring chemotaxis • measured by inserting a capillary tube containing an attractant or a repellent in a medium of motile bacteria • can also be seen under a microscope

Molecular adaptation to life in the cold.

• production of enzymes that function optimally in the cold • >>>more α-helices than β-sheets → greater flexibility for catalysis at cold temperatures • >>>more polar and fewer hydrophobic amino acids • >>>fewer weak bonds (e.g., hydrogen and ionic bonds) • Cytoplasmic membranes function at low temperatures. •>>> high unsaturated and shorter-chain fatty acid content • >>>some polyunsaturated fatty acids, which remain flexible at very low temperatures • cold shock proteins (chaperones) • Cryoprotectants (e.g., antifreeze proteins, certain solutes) prevent formation of ice crystals. • exopolysaccharide cell surface slime