Microbiology Test 1

What comprises a bacterial cell

Envelope

• Cell wall

• cell membrane

Cytoplasm

• Nucleoid

Cell wall

Rigid outer shell, protects from

osmotic stress

Cell Membrane

Lipid barrier full of proteins that contains the cytoplasm, mediates transport in and out of the cell, and scaffolds cell activity

Cytoplasm

• Gel-like network of proteins, DNA, RNA, cytoskeletal-like elements, etc.

• Contains a cytoskeleton

• Cytoskeletal elements and other proteins localize to specific places and times to coordinate cellular processes

• Proteins orfer the bacterial chromosome in the nucleoid

Nucleoid

DNA, expression machinery, DNA-binding and regulatory

proteins

FtsZ function

mediates proper placement of the septum

Important for placing replication machinery

What is the general function of bacterial cytoskeletal protein filaments?

• They organize cellular processes and help determine cell shape

Caulobacter crescentus

• Rod-shaped bacteria found in wetlands

• Produces different structures at the end of each pole

• Grows stalk, always will know what side to grow on

• 
  

What are bacterial cell membranes primarily composed of

Phospholipids and proteins

Phospholipid Bilayer

1. Hydrophobic tails

2. Hydrophilic head

3. Forms Bi-Layer

4. Gasses, water and uncharged molecules pass through

Facilitated diffusion

• transport across membrane facilitated by membrane protein

Uniport

Transport of a substance in one direction across the membrane

Symport

Transport of substances together in the same direction.

Antiport

Transport of substances in opposite directions.

Besides transport, what other functions can membrane proteins perform?

• receptor

• enzyme

• anchor

Why are membranes sensitive to low temperatures?

Membranes can freeze, and saturated fatty acids tend to "stick" together

What three fatty-acid modifications increase membrane fluidity?

Unsaturation, shorter chains, and branches.

Cells can adjust to thermal damage

Heat speeds up molecular motion; cells compensate with saturated fatty acids

Cold slows down molecular motion; cells compensate with modified fatty acids

What is peptidoglycan composed of?

• Specialized polysaccharides cross-linked by peptides.

What is the major protective function of the sacculus?

Production of osmotic stress

How do bacterial cells grow their cell wall?

By inserting new peptidoglycan into the sacculus.

Shapes of bacteria

Coccus: round

Vibrio- Curved

Helical-spiral

rod- rod

Gram positive

• Has one cell membrane

• Thick peptidoglycan layer outside the cell membrane

• Teichoic acid

• Stain purple

Teichoic acid

• Negatively charged glycerol phosphate polymer

• Protects the cell from chemical damage

• Helps organize the thick peptidoglycan layer.

• Acts as charge barrier in gram positive bacteria

Gram negative bacteria

• 2 membranes

• Peptidoglycan in-between membranes

• LPS found

• Pink

LPS

It protects the cell from chemical damage

It contains the O-antigen, core polysaccharide, and Lipid A.

O-antigen

A repeating pattern of highly variable sugars; each bacterial species has a unique O-antigen.

Lipid A

t anchors LPS to the outer membrane

is very hydrophobic.

Lipoproteins

Lipoproteins attach the outer membrane to peptidoglycan. Structural anchor for the outer membrane.

Porins

Permeability gates in the outer membrane allow small molecules to diffuse into the cell.

Why is the Gram-negative outer membrane considered a diffusion barrier?

It protects against dangerous chemicals but can also prevent nutrients from entering.

Periplasm

• A space, not a structure, between the plasma/inner membrane and the outer membrane.

• A thick gel containing detox enzymes and chemical-binding proteins.

• Chemical defense.

What structures do both Gram-positive and Gram-negative bacteria have?

Peptidoglycan, a cytoplasmic/inner membrane, phospholipids, fatty acids, and membrane proteins.

What is unusual about the cell architecture of Mycobacterium tuberculosis?

thick, waxy envelope/cuticle.

• capsule contains 2 EPS layers with mycolic acid lipid layer between them

Why are mycolic acids important to mycobacteria?

They allow bacteria to grow inside macrophages and are crucial for drug resistance.

S-layer

• A surface layer/array made of proteins.

• They provide additional protection and function for the bacterial cell.

capsule and slime layer

A capsule is anchored to the cell, while a slime layer is loose.

Capsule appears rough

Slime layer appears wet

Capsules and slime layers can produce recognizable differences in colony appearance on agar.

They can change the cell's properties and its ability to withstand certain environments.

Determine structure of given bacterial envelope

Light microscopy,

• Gram stain

Electron Microscopy

• Thin section TEM

• Cryo-EM and tomography

How can bacterial envelope protein content be analyzed?

Cell fractionation, protein gels, and mass spectrometry.

How can bacterial envelope chemical composition be analyzed?

Cell fractionation followed by chromatography and mass spectrometry.

4 types of host-microbe interactions

Mutualism- Both partners benefit

Commensalism- One partner benefits, one is unharmed

Amensalism- One harmed, one unharmed

Parasitism- One benefits, one is harmed

Why is the relationship between humans and their gut flora considered mutualistic?

Humans provide bacteria with a place and nutrients to live, while gut microbes provide benefits to the host

Ruminants

• An animal with a rumen, an organ containing bacteria that degrade cellulose.

• The cow consumes the fermentation acids left over by the bacteria.

• The bacteria obtain cellulose/food and a place to live, while the cow obtains useful fermentation products.

Pea Aphids and Buchnera Aphidicola

• Aphids eat plant sap, which is a poor diet that is mostly sugars (75%) and contains relatively few amino acids (15%).

• Buchnera lives inside host-generated structures called bacteriocytes and provides essential amino acids to the aphid.

• Buchnera can be visualized using FISH (fluorescence in situ hybridization).

What does each partner provide in the aphid–Buchnera mutualism?

• Aphid → Buchnera: energy, carbon, and nitrogen

• Buchnera → aphid: essential amino acids

• Buchnera uses resources such as nitrogen from the host to generate amino acids for the aphid.

How is Buchnera passed from one generation of aphids to the next?

• Vertical transmission:

• passes from mother → offspring.

• Bacteriocytes are near germline cells, and Buchnera escapes into the developing embryo to promote maternal transfer.

How did scientists test whether Buchnera provides essential amino acids to aphids?

• Treating aphids with antibiotics caused rapid reproductive failure.

• When researchers then supplied the essential amino acids encoded by Buchnera, the aphids were rescued.

• This supports the hypothesis that Buchnera supplies essential amino acids.

How does mutualistic Wolbachia benefit bedbugs, and how was this demonstrated?

• Wolbachia lives in a bacteriocyte and supplies B vitamins.

• Eliminating Wolbachia with antibiotics greatly reduced adult emergence, while adding B vitamins to antibiotic-treated bedbugs largely rescued them.

• It's similar to Buchnera, except Buchnera provides amino acids and Wolbachia provides B vitamins.

Can a microbe's relationship with its host change? Give examples from the lecture.

Yes. Relationships can change through evolution or changes in the host/microbe.

Obligate symbionts cannot grow w/out host, maternally inherited

Infected fathers + uninfected mothers = inviable offspring

What is cytoplasmic incompatibility

Wolbachia make CidB toxin and CidA antitoxin in testes

• When an egg is fertilized CidA is degraded, CidB

activated

• Active CidB modifies chromatin & histones

• Unless the egg already has compatible Wolbachia making CidA, chromatin Does not divide properly

Pathogenisis

• Pathogens obtain nutrients by taking them away from other cells because large amounts of carbon and energy are stored in living organisms.

• Pathogens can secrete proteases (proteins), nucleases (nucleic acids), amylases (starches/carbohydrates), and phospholipases (phospholipids) to break down host material.

Koch’s postulates

1. Isolate the organism from an infected individual

2. Culture it in the lab

3. Re-infect a new individual and reproduce the disease

4. Re-isolate the organism.

What are the important characteristics of Helicobacter pylori?

Gram-negative

Helical rod with polar flagella

human pathogen associated with gastric ulcers and cancer.

It was first described in 1890 and rediscovered in gastric biopsies by Barry Marshall and Robin Warren in 1982.

What are ID₅₀ and LD₅₀, and how do they relate to virulence?

• ID₅₀: number of bacteria required to infect 50% of hosts

• LD₅₀: number of bacteria required to kill 50% of hosts

• A lower ID₅₀ or LD₅₀ = greater virulence, because fewer bacteria are needed to cause infection/death.

What is the general disease cycle?

• Reservoir → invasion → colonization → growth → toxicity.

• A reservoir is an environmental source of a pathogen, such as another animal, soil, or water.

What are virulence factors, and what are their two main functions?

• Properties of bacteria that help them overcome host defenses by increasing:

• Invasiveness: ability to enter and survive in the body

• Toxicity: ability to damage host cells and obtain nutrients

What are Koch’s Molecular Postulates for determining whether a gene encodes a virulence factor?

1. Gene is present in disease-causing strains and absent in non-disease strains.

2. Mutating the gene reduces virulence.

3. Introducing the gene increases virulence.

4. An immune response against the gene product should protect against infection.

How can motility and adhesion act as virulence factors?

• Motility: helps bacteria migrate to target tissues (tissue tropism); examples include swarming through mucus, twitching over surfaces, and endoflagella movement.

• Adhesion: allows bacteria to attach at the infection site and resist being flushed away.

What structures help bacteria adhere to a host?

• Pili: protein fibers that stick to surfaces

• Adhesins: bacterial proteins that bind sugar patterns on eukaryotic cell, host decorates its surface proteins with sugars to distinguish itself, adhesins bind to these sugars

• Capsule: sticky extracellular polysaccharide coat that promotes attachment, sticky substance to attach to host

How can a capsule increase bacterial virulence?

Capsules can:

• Help bacteria attach to the host

• Protect against hydrogen peroxide

• Block engulfment by macrophages

• Shield surface antigens from antibodies

• Help form biofilms

What is antigenic variation and how does it promote immune evasion?

Bacteria can have or switch between different versions of their antigens, making them harder for the immune system to recognize.

For example, Salmonella typhi has two flagellin genes and can switch between them.

What do the O, H, and K antigens represent?

• O-antigen: unique sugar pattern on LPS of Gram-negative bacteria

• H-antigen: flagellin

• K-antigen: capsule
  Different combinations produce different serotypes, such as E. coli O157:h7

Streptococcus pneumoniae and its capsule

• Gram-positive diplococcus that infects the lungs.

• It has about 90 different capsule structures, allowing antigenic variation.

• Some capsules also mimic host-cell sugars, helping it evade the immune system.

• Variation: many capsules to evade the immune system!

• Mimicry: Some capsules look like host cell sugars!

What did the Griffith and Avery experiments show about capsules and virulence?

• Griffith: A capsule is required for virulence, and the capsule-producing trait could be transferred to an avirulent strain.

• Avery: DNA/genes for capsule production were transferred to the avirulent strain through natural competence.

What are siderophores and how do they increase virulence?

• High-affinity iron sequestration systems

• allow bacteria to take iron away from host proteins such as lactoferrin and transferrin

difference between exotoxins and endotoxin

• Exotoxins: secreted enzymes/proteins that disrupt host-cell structures or processes

• Endotoxin: LPS that is part of the bacterial cell itself; it is not secreted and is released when bacteria die.

What do hemolysin, phospholipase, and IgA protease exotoxins do?

• Hemolysin: forms holes in host-cell membranes → cells burst

• Phospholipase: allows bacteria to escape endocytic vesicles → intracellular growth hidden from the immune system

• IgA protease: degrades antibodies → prevents antibodies from targeting bacteria

How do A/B toxins work?

• A subunit = toxin

• B subunit = delivery vehicle
  Together, they form a self-injecting, secreted toxin complex that typically overrides host signaling systems.

How do cholera toxin and Shiga toxin damage the host?

• Cholera toxin (Vibrio cholerae): modifies host signaling → disrupts regulation of ion transport → massive water efflux → severe diarrhea

• Shiga toxin (E. coli O157:H7): 28S rRNA ribonuclease → destroys eukaryotic ribosomes → kills host cells

What are the important parts of LPS/endotoxin?

• O-antigen: species-specific sugar pattern that reacts with antibodies but is not toxic

• Lipid A: the extremely toxic portion of LPS

How does Lipid A cause toxic shock?

Bacteria die

LPS released

LBP binds Lipid A

Docks with TOLL receptor on a white blood cell

Massive cytokine release

widespread blood-vessel inflammation/dilation

blood pressure crashes

toxic shock.

Why is the immune system so sensitive to Lipid A?

It allows the body to detect trace amounts of bacteria in the blood and respond quickly. T

The downside is that it can overreact, causing system-wide inflammation and potentially fatal toxic shock.

What makes the human body an inhospitable environment for pathogens?

• Skin: layer of dead cells that is difficult to penetrate + dry

• Lungs: dry, harsh environment

• Stomach: very acidic

• Throughout body: nutrients are locked up/unavailable to pathogens

How does flushing defend the body against pathogens?

• Tears: flush microbes from the eyes

• Mucus: traps bacteria and carries them away in the lungs and GI tract

• Urine: flushes microbes from the urinary tract

How do enzymes protect the body from bacteria?

• Lysozyme in tears: destroys bacterial peptidoglycan

• Saliva and stomach acid: contain suites of digestive enzymes that help destroy microbes.

How does the native microbial flora defend against pathogens?

• Harmless, well-adapted bacteria already cover the body's external surfaces.

• An invading pathogen must outcompete the normal flora before it can successfully establish itself.

How does iron sequestration protect the body from pathogens?

• Iron is a scarce micronutrient, so the body binds/sequesters iron with proteins such as lactoferrin, essentially starving invading bacteria of the iron they need

• Bacteria fight this defense using siderophores, which steal iron away from host proteins like lactoferrin.

How do macrophages defend against bacteria?

• Engulf bacteria

• Secrete hydrogen peroxide (H₂O₂) to help kill bacteria.

• This explains why a capsule is a virulence factor—it can block macrophage engulfment and protect against hydrogen peroxide.

What are antibodies and antigens?

• Antibodies: proteins made by the immune system that bind to antigens

• Antigens: specific patterns found on bacterial surfaces
  Antibody binding helps target bacteria for destruction.

What are the major host defenses against pathogens?

Inhospitable environments, flushing, enzymes, native microbiome, nutrient/iron sequestration, phagocytes, and antibodies.

What is the traditional vs. modern view of bacterial life?

• Traditional: unicellular, planktonic/free-floating, independent individuals

• Modern: bacteria often live in multicellular, surface-associated, cooperative groups and multispecies communities, using quorum sensing, social motility, and biofilms.

What are the four types of bacterial motility and how do they work?

• Swimming: flagella-driven

• Swarming: flagella-driven group movement across a surface

• Gliding: adhesins are pulled from one end of the cell to the other

• Twitching: pilus retracts and pulls the cell

What is a biofilm, and what is its matrix made of?

• A biofilm is a multicellular aggregate of bacteria held together by EPS.

• The matrix contains exopolysaccharides, proteins, and DNA

What are a colony, pellicle, and biofilm?

They are bacterial aggregates at different interfaces:

• Colony: air–surface interface

• Pellicle: air–water interface

• Biofilm: water–surface interface

What are some beneficial vs. harmful examples of biofilms?

Beneficial: rhizosphere, cyanobacterial mats/blooms, marine snow, normal body flora.
Harmful: dental plaque/caries, clogged catheters/pipelines, contaminated medical supplies, biocorrosion, infections, and increased gene transfer/antibiotic resistance

How do biofilms cause biocorrosion and dental caries?

• Biocorrosion: biofilm metabolism produces acids and anaerobic respiration reduces metals → corrosion.

• Dental caries: bacteria stick to teeth and ferment sugars into acids → enamel degradation/tooth destruction.

How are cells in a biofilm different from planktonic cells?

• Biofilm cells are not motile and experience chemical gradients in oxygen, nutrients, bacterial products, and signaling molecules.

• Because cells experience different environments, they express different genes and produce different proteins.

What are the 3 stages of biofilm development?

• Attachment → growth/maturation → detachment/dispersion.

• Cells transition from planktonic → biofilm → planktonic states.

What happens during biofilm formation?

• Cells switch from motility to EPS production.

• EPS helps cells stick to each other and surfaces, allowing them to grow into aggregates

How does c-di-GMP control the switch between planktonic and biofilm states?

• DGC (diguanylate cyclase) makes c-di-GMP, c-di-GMP makesEPS, ↓ motility → biofilm formation

• PDE (phosphodiesterase) degrades c-di-GMP → ↓ c-di-GMP → ↓ EPS, ↑ motility → biofilm dispersion

Why is EPS necessary for biofilm formation?

• EPS allows cells to stick to one another and to surfaces and build structured aggregates.

• The slide comparing wild type with an eps mutant shows that loss of EPS disrupts normal pellicle and colony architecture.

What types of enzymes can promote biofilm dispersion?

Enzymes that break down components of the biofilm matrix can promote dispersion:

• Amylase → polysaccharides

• Nuclease → DNA

• Protease → proteins

Lysozyme would NOT be expected to specifically break down the biofilm matrix because it targets bacterial peptidoglycan rather than the listed EPS matrix components

What are the 5 major advantages of living in a biofilm?

1. Concentrates digestive enzymes → many bacteria essentially eat together

2. Prevents flushing → allows persistence in favorable environments

3. Collaborative metabolism → products from one species can be used by another

4. Maintains high cell density → facilitates quorum sensing and genetic transfer

5. EPS provides protection → diffusion barrier against antibiotics + large groups are difficult for immune cells/microscopic eukaryotes to engulf.

What are antibiotics from an ecological/microbe–microbe perspective?

Chemical warfare. Some organisms synthesize and secrete chemicals that inhibit the growth of competing organisms. These chemicals are called antibiotics

What is selective toxicity and why is it important for antibiotics?

• An antibiotic should prevent the growth of bacteria while permitting human cells to grow.

• To achieve selective toxicity, antibiotics target specialized cellular functions that differ between bacteria and humans.

How was penicillin discovered?

• In 1928, Alexander Fleming observed that the fungus Penicillium inhibited the growth of Staphylococcus bacteria

• An accidental discovery that led to penicillin.

Where do many naturally occurring antibiotics come from?

• Streptomyces species.

• About 75% of known antibiotic-producing species can produce multiple and unique antibiotics.

Why is soil an important place to search for new antibiotics?

• Soil contains about 10¹¹ bacteria per gram.

• Microbes produce antibiotics to defend their territory against competitors, so soil contains diverse organisms that may produce undiscovered antibiotics.

What are the major cellular targets of antibiotics, and what are examples?

• DNA synthesis: nalidixic acid/fluoroquinolones

• RNA synthesis: rifampicin/rifamycins

• Cell-wall synthesis: ampicillin & penicillin/β-lactams

• Protein synthesis: oxytetracycline

• Folate synthesis: trimethoprim

How can different antibiotics inhibit peptidoglycan synthesis?

They interfere at different steps of the same process:

• Penicillin: inhibits cross-linking

• Vancomycin: inhibits polymerization

• Bacitracin: inhibits activation of the membrane carrier.