Unit 1 MIP 300

Compare and contrast prokaryotic cells and eukaryotic cells.

Prokaryotic cells lack a nucleus and membrane bound organelles. Eukaryotic cells have membrane bound organelles such as the golgi, er, mitochondria, and lysosomes.

List 4 types of cellular microbes and give a common example of each of these types of microbes.

Fungi (yeast and mold), protozoa (giardia lamblia), bacteria (mycobacterium tuberculosis), archaea (Thermopolis aquaticus)

List 2 types of acellular microbes and describe what each of these acellular microbes is made of.

Acellular microbes are not composed of cells and include viruses and prions. Viruses consist of both a protein shell and nucleic acid (either DNA or RNA). Prions are composed of protein only.

Explain what a “transitional organism” is and name a type of microbe that can be transitional.

one that is eukaryotic but lacks one or more standard membrane bound organelles. Protoza can be transitional like giardia lamblia lacking a golgi and er

Explain how all living things are classified.

using the three domain system that categorizes organisms based on their ribosomal RNA sequences. the three domains are bacteria, archaea, and eukarya

Describe how bacteria are scientifically named and be able write a bacterial name correctly.

Bacteria are named by their genus, species, and strain. The genus must be capitalized while the species is lowercase and the entire name should be italicized

Describe how a bacterial genus is further broken down into groups.

A bacterial genus is broken down into species of groups

Describe how a bacterial species are further broken down into groups.

Bacterial species can be broken down into specific strains which all have unique characteristics

Relate resolving power of a microscope to microbe size.

Resolving power is the ability of a microscope lens to distinguish between two objects that are positioned close together. For a microbe to be visible it must be larger than the microscopes resolving power.

Explain which microbes can be seen using a light microscope and which ones cannot and why.

Bacteria and eukaryotic cells are generally visible while viruses are not. Light microscopes cannot resolve objects smaller than 0.2 micrometers which bacteria is bigger than but viruses are smaller than.

Put microbes in order based on their size from smallest to largest.

prions, viruses, bacteria, eukaryotic microbes and human cells

Which type of microscopy could you use to visualize organelles inside a eukaryotic microbe and why?

phase contrast microscopy as it converts differences in refractive indexes into light intensity variations allowing for visualization of internal structures

Which type of microscopy could you use to visualize bacteria arrangement, cell shape and size and why?

Bright field microscopy and staining

Which type of microscopy could you use to visualize small internal structures of bacteria?

Electron microscopy because its resolution is 1,000 times better than a light microscope

Which type of microscopy could you use to visualize viruses in infected cells and why?

Electron microscopy because viruses are very small and require a huge resolving power

Which type of microscopy could you use to visualize live bacteria in a water sample and why?

Dark-field or phase-contrast microscopy, because these methods do not require heat-fixing or staining, which kills the cells

Which type of microscopy could you use to visualize yeast cells in the brewing process and why?

Bright-field or phase-contrast microscopy, as yeast are eukaryotes and sufficiently large to be viewed with light microscopes

Describe bright field microscopy

Bright-field: Uses light; the background is bright white, and specimens must be stained and heat-fixed/killed to be seen

Describe dark field microscopy

Background is dark while the organism appears light; it visualizes live cells because staining is not required

Describe phase contrast microscopy

Enhances refractive index differences to show internal organelles and structures in live cells without staining

Describe fluorescent microscopy

Uses UV light to excite fluorescent dyes or labeled antibodies, causing the specimen to glow

Describe electron microscopy

Uses electrons instead of light; has a much higher resolution (.0005 µm) and is used to see viruses and detailed internal anatomy

Explain the differences between a simple stain and a differential stain.

A simple stain uses only one dye to increase visibility of shape, size, and arrangement of microbes. A differential stain uses two dyes to categorize microbes into different groups based on structural characteristics

Identify the three primary shapes and arrangements of bacteria.

Cocci are spherical, bacilli are rod-like, and spirals

What are the different arrangements of cocci?

Diplococci (pairs), Streptococci (chains), Staphylococci (clusters), Tetrads (groups of 4), and Sarcina (groups of 8)

What are the different classifications of bacilli?

Coccobacillus (short/wide), Diplobacillus (pairs), and Streptobacillus (chains)

What are the different arrangements of spirals?

Vibrios (comma-shaped), Spirilla (corkscrew with external flagella), and Spirochetes (corkscrew with internal flagella)

Explain the process of a differential stain for gram positive and gram negative bacteria. What appearance does this give them?

Before staining bacteria are heat fixed to a glass slide. A primary stain is applied to microbes. A decolorizing agent is then applied which washed the primary stain out of some of the cells but not others. A counterstain is then applied and taken up by some cells. In gram staining positive bacteria retain the primary stain and are purple while negative bacteria appear pink.

Compare and contrast bacterial and eukaryotic cells.

Both cells have a plasma membrane, ribosomes, and DNA. Bacterial cells lack membrane bound organelles, are smaller, and have hopanoids. Eukaryotic cells have mitochondria, a golgi, and are stabilized by cholesterol.

Compare and contrast the Gram positive and Gram negative envelope.

Gram positive bacteria, the envelope is characterized by a very thick layer of peptidoglycan (PPG). This layer contains teichoic and lipoteichoic acids, which are unique to Gram-positive cells. Gram negative bacteria have a far more complex envelope with an outer membrane containing lipopolysaccharides (LPS) and porin proteins. Their peptidoglycan layer is significantly thinner

What are the differences between the Gram positive and Gram negative envelope.

The peptidoglycan layer is thick in g+ bacteria and thin in g- bacteria. The periplasmic space is small in g+ bacteria and large in g- bacteria and abundant in enzymes. Only g- bacteria have an outer membrane and g+ bacteria rely on exoenzymes due to the size of their periplasmic space.

Describe the function of the plasma membrane

a semi-permeable barrier and the site for metabolic processes like oxidative phosphorylation

Describe the function of the peptidoglycan layer of the bacterial envelope

Provides the cell with its shape and protects it from osmotic pressure, preventing the cell from bursting (lysis) in hypotonic environments

Describe the function of the periplasmic space and enzymes

The space houses periplasmic enzymes (such as permeases and hydrolytic enzymes) that are crucial for nutrient acquisition

Describe the function of the outer membrane (in g- bacteria only)

Functions as a barrier to toxic substances (like antibiotics), helps the cell avoid immune defenses, and contains porins to allow small molecules to pass

Explain the function of LPS/endotoxin and its clinical relevance

Released when the cell dies; the Lipid A component is highly toxic and can cause septic shock in humans

There is variability in the thicknesses of the peptidoglycan layer of bacteria. Please give a structural explanation for why the thickness may vary between two Gram positive bacteria. How would this answer differ for Gram negative bacteria?

The thickness of the peptidoglycan layer varies between species because it is composed of varying amounts of peptides and sugars. In g+ bacteria, this thickness can range widely, which provides a substantial protective layer. For g- bacteria, the layer is consistently thin because it is sandwiched between the inner plasma membrane and the protective outer membrane, which provides additional structural complexity

What are the typical components of a bacterial plasma membrane?

contains a phospholipid bilayer with stabilizing structures like hopanoids/cholesterol and integral, membrane, and glycoproteins

describe characteristics and functions of integral proteins

intramembrane proteins that function as pores to transport nutrients and waste and are involved in metabolic processes

describe characteristics and functions of peripheral proteins

involved in transport and metabolic processes and reside outside of the membrane

describe the characteristic of glycoproteins

consist of oligosaccharides covalently attached to either integral or peripheral proteins and vary between bacterial species

List the steps of the Gram stain procedure

Heat fix the microbes, apply crystal violet (purple), apply grams iodine which binds crystal violet into aggregates, wash with acetone, apply safranin to counterstain cells

List the steps of the acid-fast stain procedure

Heat fix, apply carbol fuchsin (pink) using heat to drive the stain into the cell, wash with acid alcohol, apply methylene blue

List the steps of the endospore staining procedure

Heat fix, apply malachite green while using heat to penetrate the spore, wash with water, apply safranin

Compare and contrast the three staining procedures

All three procedures are differential stains that use heat fixation. The gram stain uses iodine as a mordant to trap dye while the acid-fast and endospore stains rely on physical heat. Also acid fast staining requires a stronger decolorizer than endospore staining

What do each of the staining procedures target?

The gram stain targets the peptidoglycan layer, the acid fast stain targets the thick/waxy layer of mycolic acid, and endospore stains target the endospore

What are the positive and negative results for a gram stain?

A positive result is purple as it retained crystal violet while a negative stain is pink

What are the positive and negative results for an acid fast stain?

A positive result is hot pink indicating acid fast cells while a negative result is blue

What are the positive and negative results for an endospore stain?

a positive result is green a negative result shows pink vegetative cells

You isolate a bacterium with an LPS layer and a 3 nm peptidoglycan layer. What color would you expect this cell to be after properly performing a Gram stain? What if the bacterium lacked an LPS layer and had teichoic acid?

a bacterium with an LPS layer and a 3 nm peptidoglycan layer will appear pink. a bacterium lacking LPS and containing teichoic acid will be purple

What will be the result of a gram stain if you forget grams iodine?

Iodine acts as a mordant that binds with crystal violet to create aggregates too large to escape the thick peptidoglycan of g+ cells. Without it, the crystal violet remains small and is easily washed out of both types of bacteria by the alcohol so all cell types will appear pink

What will be the result of a gram stain if you forget safranin?

Safranin is the secondary stain used to visualize cells that lost the primary stain; g+ cells will be purple and g- cells will be clear

What will be the result of a gram stain if you forget crystal violet?

both cell types will be pink

What will be the result of a gram stain if you forget alcohol?

The alcohol is the decolorizer that specifically removes the crystal violet-iodine complex from the thin peptidoglycan of g- cells. If this step is skipped, the purple stain is never removed from any cells.

What is the importance of N-acetyl-glucosamine (NAG) and N-acetyl muramic acid (NAM)?

NAG and NAM are the two primary carbohydrates that serve as the fundamental building blocks of peptidoglycan. Their functions include formation of the structural backbone, anchoring tetrapeptide chains through cross linking or peptide interbridges (g+), protection against osmotic pressure, and are a target for antibacterial agents

Describe the internal structural features of prokaryotic cells, and how these differ from eukaryotic cells.

Prokaryotic cells are structurally simplistic and lack a membrane bound nucleus and other lipid bilayer-bound organelles. Instead of a nucleus, they have a nucleoid region, 70S ribosomes, plasmids, and inclusion bodies. Eukaryotic cells contain a membrane bound nucleus and various specialized organelles, and primarily use larger 80S ribosomes.

Identify which internal bacterial structures would make good antibiotic targets and explain why.

70S ribosomes because antibiotics can be targeted without harming the host and ppg because drugs can target cross links

What is the composition and function of inclusion bodies? Give an example of an organic vs inorganic inclusion body.

amorphous aggregates of organic or inorganic substances, such as granules, crystals, or globules that serve as storage sites for essential nutrients like carbon, energy, phosphate, and iron. An organic example is glycogen granules storing carbon. An inorganic example is polyphosphate granules storing phosphate for building DNA

Explain the endosymbiont theory. How does this theory relate to antibiotic targeting?

The endosymbiont theory suggests that an ancestral eukaryotic cell engulfed a bacterium that eventually became the mitochondria. Because of this bacterial ancestry, human mitochondria still contain 70S ribosomes and circular DNA. This relates to antibiotic targeting because drugs designed to attack bacterial 70S ribosomes can sometimes cause side effects in human cells

Discuss selective toxicity.

Selective toxicity is the principle of using antibacterial substances to target specific characteristics of a bacterial cell that are absent in the host

What events would cause a spore forming bacteria to undergo sporulation?

Sporulation is triggered by a lack of food or nutrients

What are the seven steps of sporulation?

Axial filament formation, septum formation, engulfment of forespore, cortex formation, coat synthesis, completion of synthesis and increase in heat resistance, and lysis of sporangium

Explain the function of dipicolinic acid in spores.

Calcium-dipicolinic acid comprising about 15 percent of the spore core serves to stabilize the DNA within the endospore

Describe why spores are so heat resistant.

Endospores are highly heat resistant due to two primary factors: the presence of calcium-dipicolinic acid to protect the DNA and the fact that the spore core is dehydrated (dry)

Discuss the stages of germination.

Activation triggered by temperature change, gemination triggered by the presence of food, and outgrowth where the protoplast makes new components and a vegatative cell emerges from the spore

Without using complex membranes, how do bacteria store essential nutrients?

Bacteria store nutrients by forming aggregates of substances (inclusion bodies) that are either free floating or enclosed in a single layer membrane made of protein or a single layer of phospholipids and proteins, rather than a complex double lipid bilayer

Which of the following would you expect to find within a bacterial cell? Nucleus, Golgi apparatus, Endoplasmic reticulum, peptidoglycan, lysosome, ribosome (70S, 80S), cytoskeleton, DNA.

Peptidoglycan, 70S ribosomes, cytoskeleton, and DNA

Can bacteria reproduce via sporulation?

No, there is no increase in the number of cells in the population when sporulation occurs

Name a famous spore-forming bacterium that causes disease in animals and humans. Describe the signs of this disease

Clostridium tetani is a famous spore-former that causes tetanus. Signs of the disease include constant muscle contractions, which often begin as lockjaw

Identify the 4 phases of a microbial growth curve in a closed system.

First is the lag phase where cells are making copies of things and increasing in volume, then the log phase where bacteria divide at their maximal and constant rate, in the stationary phase the growth curve plateaus because the number of dividing cells equals the number of dying cells (as waste accumulates and nutrients are depleted), last the death phase where cells die at an exponential rated because waste is becoming toxic

Identify the phase or phases on the microbial growth curve during which penicillin would be effective and why.

Penicillin would be most effective during the exponential phase because it inhibits synthesis of the peptidoglycan cells wall and only targets bacteria that are actively dividing

Identify the phase or phases on the microbial growth curve during which lysozyme would be effective and why.

Lysozyme is an enzyme that destroys the integrity of the cell wall by targeting the glycosidic bonds between NAG and NAM and would be most effective during active cell division

Explain what bacterial generation time means.

Bacterial generation time (or doubling time) is the interval required for a single cell to divide into two cells

Predict what will happen to an organism’s growth rate when the generation time increases or decreases.

Growth rate is inversely proportional to generation time

If given different doubling times or numbers of generations for bacteria grown in different environments, be able to determine the environments in which these organisms grow best.

The best environment will show the shortest doubling time, highest number of generations in a fixed amount of time, and they will also show higher growth rates in the environments they prefer

Define the optimal conditions of psychrophiles

grow at 0-20 degrees with an optimal temp at 15

Define the optimal conditions of psyhotrophs

Grow at 0 degrees but grow optimally between 20-30 degrees

Define the optimal conditions of mesophiles

have optimum growth between 20-40 degrees

Define the optimal conditions of thermophiles

grow at 45-100 degrees

Define the optimal conditions of acidophiles

grow at pH 1-5.5

Define the optimal conditions of neutrophiles

grow at pH 5.5-8

Define the optimal conditions of growth for halophiles and barophiles

halophiles require high salt concentrations and barophiles require high pressure

Explain what a colony is.

A colony is a visible cluster of bacteria growing on a solid surface. Each colony originates from a single cell, known as a Colony Forming Unit which has divided through binary fission millions of times until it becomes visible

Explain how Helicobacter pylori can grow in the stomach even though it is a neutrophile

it survives the highly acidic stomach by producing the enzyme urease which breaks down urea in the stomach into carbon dioxide and ammonia. Because ammonia is very basic, it neutralizes the stomach acid immediately surrounding the bacterium

Describe and give examples of the ways in which microbes can be controlled with chemical or physical agents.

Chemical agents like disinfectants, antiseptics, and sanitizers reduce microbial populations. Physical agents like temperature can kill microbes or slow growth and radiation can also affect them.

Describe and give examples of the ways in which microbes can be controlled with antibiotics.

biological or synthetic agents that exhibit selective toxicity, targeting specific bacterial functions while minimizing harm to the host

Describe the 6 conditions that effect microbial killing by a disinfectant.

population size and composition, concentration of the agent, length of exposure, temperature, and local environment

Name the 5 bacterial functions that antibiotics can inhibit

protein synthesis (binding 70S ribosomes), cell wall synthesis (inhibiting ppg formation), cell membrane integrity, nucleic acid synthesis, specific metabolic pathway inhibition

What are the two most common mistakes people make when taking antibiotics? Why are these problems?

Failure to follow dosage directions which prevents the drug from maintaining the necessary concentration in the blood. Failure to finish the course which allows resistant bacteria to survive and grow

Please explain why viral diseases are harder to treat than bacterial diseases.

Viral diseases are harder to treat because viruses are obligate intracellular parasites that use the hosts own cellular machinery to replicate, making it difficult to find a drug that kills the virus without also harming host cells

Please explain why fungal and protozoal diseases are harder to treat than bacterial diseases.

Because fungi and protozoa are eukaryotic cells, they share many structural and functional similarities with human cells. This makes achieving selective toxicity difficult, as drugs often have toxic side effects for the host

Explain the 5 ways that bacteria can be resistant to antibiotics.

Changing the RNA sequence of ribosomes or amino acids so the drug cannot bind, making enzymes to chemically change the antibiotic, using efflux pumps. bypassing blocked metabolic steps

Describe 3 ways bacteria may be inherently resistant to antibiotics.

In g- bacteria the outer membrane is a barrier to large molecules, in mycobacterium the mycolic acid layer is nearly impenetrable, and if the cells have a lack of target like their cell wall

Describe 2 ways that bacteria may acquire antibiotic resistance.

through horizontal gene transfer allowing them to receive resistance genes from other bacteria or though spontaneous mutations

Describe what selective toxicity means.

Selective toxicity means a drug is more toxic to the infecting microbe than it is to the host. This is achieved by targeting unique microbial characteristics that the host lacks.

Explain to your roommate how soap works and why you shouldn’t purchase soaps/products that say “antibacterial” on the label.

Soap forms structures called micelles, which have hydrophilic heads and hydrophobic tails. The tails trap microbes while the heads allow the entire complex to be washed away with water making it inherently antimicrobial

What unique bacterial cell characteristics can antibiotics target?

70S ribosomes, peptidoglycan, and folic acid which cannot be synthesized in animal cells

Please list and briefly describe some differences between bacterial and animal cells that have not been targeted with antibiotics. For each one you list indicate whether you think this could be a target and why.

Bacteria use condensin and have circular DNA, while animals use histones and have linear DNA. This is a potential target because the proteins managing DNA are distinct. Bacteria use specific machinery to create a septum during binary fission. This is a potential target to stop bacterial replication specifically.

Antibiotics work differently on Gram positive and Gram negative bacteria. Please explain why this might be. For the 5 antibiotic targets that are currently utilized, which ones will work differently on Gram positive vs. Gram negative bacteria and why?

Penicillin is much more effective against Gram-positives because it can easily reach their thick, exposed peptidoglycan. In Gram-negatives, the outer membrane prevents access. Nucleic acid targets are broad spectrum and would work on both bacterial cell types

Explain quorum sensing and how it relates to biofilms

Quorum sensing triggers increased polysaccharide production (to build the film), increased antibiotic resistance, and decreased flagella production (because they are stationary)