Microbio exam 2

Types of motility

flagellar, twitching, gliding (movement is not the same as adhesion)

Flagella

• - Long helical structures made of polymerized flagellin protein

• - Flagellin is very similar across flagellated bacteria. Vertebrate innate immunity exploits this": TLR-5 (a pattern recognition receptor) specifically senses flagellin

• - Invasive pathogens often tightly regulate flagellin genes so they are not expressed once inside the host, which helps them avoid TLR-5 detection

Vibrio cholerae

A single polar flagellum (at one end). It lets the bacterium move through low-viscosity water and high-viscosity mucus. Non-motile mutants can’t reach the intestinal epithelium, so they are less virulent than wildtype

Spirochetes (Treponema pallidum)

A single flagellum inside the periplasm, which helps avoid immune detection

Taxis: Directional movement in response to stimulus (toward or away from the signla sensed)

chemicals (chemotaxis), oxygen (aerotaxis), light (phototaxis), magnetic fields (magnetotaxis), salinity/ osmotic strength (osmotaxis), hydrotaxis (water)

Chemotaxis

• - Movement towards attractants and away from “repellents”

• - Key role in the virulence of many pathogens (ie H. pylori uses chemotaxis to penetrate deep into the gastric pits of the stomach mucosa, and Pseudomonas aeruginosa also relies on it)

• - Measured by inserting a capillary tube containing an attractant or repellent culture medium of motile bacteria

• - Can also be seen under a microscope

E. coli “Random Walk” Model

E. coli has 4-10 flagella that rotate in two ways:

• - CCW is the default. The flagella form one helical bundle, so the cell makes a straight run

• - CW breaks the bundle apart, so the cell tumbles in place and picks a new random direction

Key Logic: if the cell senses it’s going the wrong way (away from the attractant or toward a repellent), it tumbles sooner and more often. If it’s going the right way, it runs longer; the net result is a biased random walk toward the attractant

How to visualize and measure motility

Soft agar stab tubes:

• Motile bacteria swim away from the stab line, so growth is diffuse and cloudy throughout the tube

• Non-motile bacteria grow only along the stab line (Proteus was the example organism)

Actin-Based Motility: Listeria monocytogenes

The disease (listeriosis, severe human and animal disease caused by the Gram-pos pathogen L. mono and L. ivanov)

• - Food-borne pathogen, responsible for ~28% of food-borne deaths in the USA

• - Can cause spontaneous abortion, transit the placenta

• - Hits immunocompromised people hardest (fetuses, neonates, immunosuppressed and elderly)

• - Untreated, it can progress to CNS infections, menigitis, and fatal bacteremia (sepsis)

• - MAJOR problem": Listeria is a pyschrophile, it grows well at 2-3 C so refrigeration doesn’t stop it, that’s why pregnant patients are told to avoid deli meats, cold cuts, and soft cheeses

Listeria monocytogenes cell-to-cell spread

Entry into the host cell (Internalins InIA) and InIB

Escape from the phagosome (Lysis of the vacuole)

Intracellular movement (ActA builds actin comet tails that propel the bacterium)

Cell-to-cell spread (Pusehs into the neighboring cell, creating 2 membrane vacuole)

Escape from the two-membrane vacuole

Why Listeria monocytogenes spread matters

Spreading cell to cell means Listeria is never exposed to humoral (antibody) immunity. It lives intracellularly to evade the immune system

• LLO mutant: stuck in the phagosome and killed

• ActA mutant: can’t make comet tails, so no cell to cell spread and lower virulence

• In1A/In1B mutant: reduced cell entry

Adhesion/Attachment

Bacterial adhesion/attachment to other cells or surface is an important step for colonization of a new environment or host

Adhesion is an essential first step for colonization and contributes to bacterial pathogenesis (virulence)

Adhesins are bacterial surface components that binds to surfaces, proteins, or sugars on other cells

Attachment can be nonspecific (hydrophobic or charge interactions) or hihgly specific (ligand-receptor binding to its cognate receptor)

Specific binding leads to cell or tissue tropism, a preference for growing at a particular location in the body)

Types of Adhesins

Pili/fimbriae (called this when short): Gram-neg, structure: long and rod-like, helical arrays of pilin, a tip protein is the actual adhesin, binds: host cell sugar groups

Fibrillar adhesins: Gram-pos, structure: short, thin, rod-like surface structures, binds: connective tissue and ECM proteins (fibronectin, collagen, fibrinogen, laminin)

Afimbrial adhesins, both, structure: proteins embedded in the surface, not rod-like, close to the cell surface, binds: mostly host surface proteins

Colonization in the host: 2-Step attachment of pathogenic E. coli

1. 1. Loose attachment is mediated by pili/fimbriae or fibrillar adhesins

2. 2. Tight (intimate) attachment is mediated by afimbrial adhesins (surface proteins)

Some pathogens go further by injecting additional adhesin proteins and effector proteins into the host cell to promote tight binding and modulate the host cell

Adhesion examples

Staphylococcus aureus: adhering to a surface

Staphylococcus epidermidis: adhering to each other (biofilm-like)

Caulobacter crescentus: a holdfast binds to surfaces, divides asymmetrically into two kinds of daughter cells - stalked cell (stationary, attached by holdfast at the end of its stalk) and swarmer cell (motile, with a flagellum, swims away)

Intracellular Bacterial Pathogens

Intracellular

• Enter and reside inside host cells

• Evade host immune killing from inside the cell

• Food-borne (Listeria monocytogenes, Brucella abortus, Salmonella entrica, Shigella flexneri), respiratory (M. tuberculosis, Coxiella), STD (Chlamydia)

Extracellular Bacterial Pathogens

Extracellular

• Outside host cells, on their surface, or in the medium

• Produce toxins and enzymes to fight the immune system

• Acne (Propionibacterium acnes, Staphylococcus epidermidis), Gastritis/Ulcers/Stomach cancer (Heliobacter pylori), skin toxic-shock syndrome (MRSA = Staphylococcus aureus), water-borne diseases (Pseudomononas aeruginosa, Vibrio Cholerae), air-borne diseases (Haemophilus influenzae)

Extracellular Bacterial Pathogens

Acne

• Skin infection from normal microbiota, mainly Gram-pos S. epidermidis and Propionibacterium acnes

• P. acnes: Gram-pos, anaerobic bacillus that colonizes sebaceous glands and eats the oils in sebum. It helps create low pH that protects against other microbes

• How acne forms: dired sebum plus dead skin cells plug the follicle and form a comedone. The plug creates an anaerobic environment that boosts growth of trapped bacteria, causing infection, inflammation, and a pimple

H. pylori (Gastritis, Ulcers, Stomach Cancer)

• Helical, motile (4-6 flagella), Gram-neg

• The stomach lumen is pH 1-2, H. pylori survives by:

• 1. Flagella to swim through mucus layer to the epithelum, where pH is closer to neutral (7-8)

• 2. Urease, which breaks urea into CO2 and ammonia (a base) to neutralize acid

• 3. Adhesins (BapA, SabA) to attach

• 4. Colonization factors and immune modulators in the gastric pit

Survival in the Host (Tissue Penetration and Dissemination - “spreading factors”)

DNases: Degrade DNA in pus from lysed host cells, which thins the viscous pus so bacteria can move

Hyaluronidases (glycosidases): Degrade hyaluronic acid in connective tissue, so bacteria spread along tissue planes

Collagenases/elastases/proteases (“meat tenderizers”): Degrade connective tissue matrix proteins; some also degrade antibodies and complement proteins

Plasminogen activator-like proteases: Degrade fibrin clots so bacteria can escape from blood clots

Streptokinase: activates host serine protease plasminogen, which converts plasminogen to plasmin, which dissolves clots

Staphylokinase (S. aureus): dissolves clots

Staphylococcus aureus (MRSA, ‘superbug’)

• Causes toxic shock syndrome and flesh-eating skin infections (necrotizing fasciitis)

• Staphylokinase: dissolving clots

• Hyaluronidase

• V8 protease: cleaves human IgG

• Exfoliative toxins: causes scalded-skin syndrome (blistering/peeling)

• Also know impetigo as a Staph skin infection

Nutrient Acquisition

Pathogens secrete enzymes that break down host macromolecules into nutrients that bacteria absorb or transport inside

Nutrient Acquisition examples

Glycosidases (neuraminidases/sialidases/hyaluronidases): sugars and polysaccharides (sialic acid on glycolipids & glycoprotein; hyaluronic acid)

Proteases: proteins

Nucleases (DNases, RNases): DNA, RNA

Phospholipases: membrane lipids, which lyse host cells and release contents

Toxins (pore-forming hemolysins): disrupt cells to release cytosolic contents (nutrients and iron)

Brucella abortus (special case)

Gram-neg, causes brucellosis (zoonotic), which leads to spontaneous abortion in cows, sheep, and horses, sometimes humans

Grows slowly in most tissues but very rapidly in the placenta because of high erythritol, a sugar it easily metabolizes

Human risk: raw milk (pasteurization & fermentation break the chain)

Iron Acquisition: The “Tug of War”

The problem: Free iron in the body is extremely low (10^-18 to 10^-9 M) because host proteins sequester it:

• Lactoferrin (mucosal surfaces and milk)

• Transferrin (made in liver, found in serum)

• Ferritin (intracellular iron storage)

• Heme / hemoglobin (~70% of total iron in humans)

• Siderphores - primary molecular components prokaryotes utilize during the 'tug of war' for iron acquisition against eukaryotic hosts

Iron Acquisition: Bacterial counter-strategies

1. 1. Siderophore (catechols or hydroxamates): secreted, chelate Fe with very high affinity, stealing it from host proteins. The iron-siderophore complex is taken back up via specific receptors (Gram-neg through outer membrane and inner membrane transporter powered by TonB/ExbD and ATP) and the iron is released inside

2. 2. Surface receptors that bind transferrin, lactoferrin, ferritin, hemin directly

3. 3. Proteases that degrade host iron-binding proteins to release iron

4. 4. Cytolytic/pore-forming toxins that lyse host cells and release iron, usually regulated and only made under low-iron conditions

Special case: Borrelia burgdorferi (Lyme disease) uses Mn instead of Fe so it bypasses the iron problem

Special case: H. influenzae requires heme for growth because it needs it to bind iron for transport and acquisition

Invasion & Intracellular Survival

Normal clearance: most bacteria taken up by host cells are killed by phagosomal killing, cell death, xenophagy, or immune mediators (few if any, escape). Intracellular pathogens have evolved ways to avoid this

How specific intracellular bacteria avoid killing

Strategy: Escape the phagosome into the cytosol, Organism: Listeria, Shigella, Rickettsia,

Strategy: Block phagosome-lysosome fusion, Organism: Salmonella, Mycobacterium tuberculosis, Brucella

Strategy: Specialized replicative compartments, Organism: Salmonella, Brucella, Legionella, Chlamydia, Coxiella

Obligate intracellular two-form life cycle, Organism: Chlamydia, Coxiella

Obligate intracellular bacteria (2 forms)

Infectious EBs - Metabolically inactive, infectious form that enters the cell (

Noninfectious RBs - Metabolically active and reproduces inside an inclusion, later converts back to EBs and are released

Chlamydia, Coxiella

Advantage of living inside host cells: bacteria can hide from antibodies and immune cells

Samonella enterica

Gram-neg, food-borne, causes ~30% of US food-borne deaths

Induces inflammatory diarrhea, enters host immune cells called macrophages and prevents phagolysosomal fusion and then survives and replicates within specialized Salmonella-containing vacuoles

Bacterial toxins (endo or exo)

Toxins can damage host cells at the site of infection or travel away from the infection site and cause damage to tissues at distant sites

Small molecule toxins: (bacterial membrane components), endotoxin= LPS lipid A, released upon cell lysis)

Small peptide toxins: enterotoxins (toxic shock syndrome toxin), superantigens (released into the medium, cause massive inflammatory immune response)

Large protein toxins (exotoxins): pore-forming/membrane-disrupting cytolytic toxins, hemolysins (released into the medium)

A-B type protein toxins: A = Activity: usually an enzyme with toxic activity, responsible for the cellular outcome, B = Binding: binds host cell surface receptors and mediates entry, trafficking, and translocation of the A part into the cytosol

Steps: Toxin binding → Entry → Translocation → Modifying target → Toxic effects (toxin-mediated diseases are irreversible)

Logic: B deleted→ can’t bind or enter → no toxicity, A deleted → can still bind and enter, but has no catalytic activity → no toxic effect

Bacterial Secretion Systems

Deliver toxic effectors/ proteins (also nontoxic proteins) into the medium and on the surface of the cell

Type I, II, V: Inner membrane components are shared with Gram-pos and Gram-neg (toxic and nontoxic proteins into the medium or onto cell surface)

Type III, IV (4), VI (6): Gram-neg bacteria only T6SS effectors can be delivered to euk and prok

Type VII (7): Mycobacteria only

The Tree of Life

• Carl Woese used the 16S rRNA gene to build the phylogenetic tree. Before the late 1980s, life was split into plants, animals, fungi, protists, and bacteria (simple to complex)

• The 16S tree showed three domains: Bacteria, Archaea, Eucarya. Archaea are genetically distinct from bacteria, even though both are prokaryotes

• All three domains trace back to LUCA (Last Universal Common Ancestor)

Two main Phyla of archaea

Crenarchaeota: mostly thermophiles

Euryarchaeota: methanogens and halophiles

Archaeal Cell Structure (S, R, S-layer, G, F, F, H, CW, M, R, DNA, P)

Shapes: Cocci, rods, spirals, pleomorphic, even square/cuboidal

Reproduction: binary fission, budding, or fragmentation. No spores

S-layer: protein/glycoprotein surface layer, present in almost all archaea (& many bacteria)

Glycocalyces: form biofilms and stick cells to each other and to surfaces

Flagella: same structure and function as bacterial flagella (basal body, hook, filament), but made of different proteins (evolved separately)

Fimbriae: present in many

Hami: Unique to archaea. Fimbria-like, with grappling hooks for attaching to surfaces

Cell wall: No peptidoglycan (all archaea and all eukaryotes lack it; some bacteria lack it, specialized polysaccharides and proteins instead)

Membrane lipids: Ether-linked, branched/cyclized (phytanyl side chains). Some form a monolayer, mechanically stronger (allows archaea to survive extremes)

DNA/genetics: usually a single circular chromosome, but the genetic code, amino acid usage, and cell division proteins are more eukaryote-like

Pathogenicity: no archaea are currently known to cause infectious disease

Extremophiles and Metabolism

Thermophiles (mostly Crenarchaeota) love heat

Halophiles (Euryarchaeota) love salt

Methanogens (Euryarchaeota) are the largest group of archaea

• Many are chemolithoautotrophs, obligate anaerobes (no oxy), convert CO2, H2, and organic acids into methane

• Live in pond, lake, ocean sediments and in animal colons, and are a major source of environmental methane, all methanogens are archaea

Types of microbial growth in different levels of oxygen

Obligate aerobe - only at the top (needs O2)

Obligate anaerobe - only at the bottom (O2 is toxic)

Facultative anaerobe - throughout, densest at top (prefers O2, but can live w/o)

Aerotolerant anaerobe - evenly throughout (doesn’t use O2, but isn’t harmed by it)

Microaerophile - a band just below the top (needs a little O2, but too much is harmful)

Archaea vs. Bacteria vs. Eukaryotes

Big picture Algae & Fungi

Microbes run the biosphere. More than 50% of Earth’s oxygen comes from photosynthesis by cyanobacteria (prokaryotes) and chlorophyll-containing algae (euk)

Algae

Eukaryotic photoautotrophs

They make their own energy and carbon from light and CO2

Aquatic, can be unicellular or simple muticellular, and some form communities (like kelp)

Marine algae have holdfasts to anchor to surfaces

Blue-green algae

Not algae! They are cyanobacteria (prokaryotes)

What products do we get from algae?

Oxygen: from photosynthesis of chlorophyll-containing algae

Agar: red algae, solidifying agent for lab culture media

Carrageenan: red algae, a commercial thickening agent

Alginate: brown algae, used medicinally, linked to wound dressess and textiles in the study guide

Food: seaweeds

Chalk: geological

Types of Algae

Chrysophyta (golden, yellow-green, carotene), includes diatoms, water molds(marine phytoplankton), major source of the world’s O2

Chloraphyta (green, carotene), many have flagella

Rhodophyta (red), cell walls contain agar and carrageenan (thickening agents), only one that just has chlorophyll a (no b)

Phaeophyta (brown, kelp), cell walls contain cellulose and alginate (medicinal)

Fungi

Chemoheterotrophs, get energy and carbon from organic chemicals, lack chlorophyll and do not photosynthesize

Reproduce by budding and producing spores, not the same as bacterial endospores

Mycology is the study of fungi

Most fungi are aerobes (need oxygen), Many yeasts (Saccharomyces) are facultative anaerobes that ferment

Cell walls are made of chitin

Most fungi are saprobes, absorbing nutrients from dead organisms. Soe use modified hyphae called haustoria to penetrate living plant or animal hosts

Fungal infection is called mycosis. Severity ranges from mild to severe, and more fungal pathogens are dimorphic (change to yeast at body temp and mold form)

Fungal Forms

Yeasts - Unicellular. They reproduce asexually by budding (forming a blastopore). They form hyphae only during mating, Saccharomyces, Candida

Molds - Multicellular, with long tangled filaments called hyphae

Dimorphic fungi - 2 forms and switch with the environment. At ~25C (ambient) they are filamentous molds (vegetative). At 37C (body temp), they become unicellular pathogenic yeast (infectious)

Basidiomycetes (mushrooms) lifecycle

1. Most of the organism is a vegetative mycelium underground, replicating asexually by budding

2. During mating, a fruiting body (reproductive structure) forms above ground

3. The fruiting body has a cap, gills, and a stipe (stalk)

4. The gills contain basidia, which make basidiospores at the ends of the hyphae (single/many filaments)

5. Mature spores are released and carried by wind

6. Spores either fuse with other hyphae to mate or germinate into new mycelia (compact tuft/mass of hyphae). The cycle repeats

Basidia generate basidiospores at the ends of the hyphae before releasing the mature spores → airborne, attach to other mushroom hyphae and fuse to mate or germinate to form new mycelia

Characteristics of Lichens

A community of ascomycete + basidiomycete fungi + photosynthetic cyanobacterium or alga

- The meshwork of fungal hyphae makes up most of the lichen body

- Soredia trap the algal or cyanobacterial cells in the meshwork

- The algae/cyanobacteria form the phototrophic layer, which makes food through photosynthesis

- The fungus provides structure and attachment to the substrate

Fungal Pathogens

Fungi are eukaryotes like us, so their cells are metabolically similar to us, so drugs that kill fungi tend to harm human cells too which is why systemic (whole body) fungal infections are hard to treat safely. Bacteria are prokaryotes, so antibiotics can target things we don’t have

Penicillium: Molds (Ascomycetes)

Species are rapidly growing molds with highly branched network of mutlinucleated, septate hyphae that have many branching conidiophore which chains of conidia sprouting from the ends, cause some harmful diseases

• Important in our lives - making cheese, producing antibiotics, antifungals, only causing some harmful diseases

• Griseofulvin from P. griseofulvum & chemo drug

Aspergillus: Molds (Ascomycetes)

Species produce several important substances that can affect us - aflatoxins = carcinogens; ergot alkaloids = psychotics; major allergens, mildew, and cause some harmful diseases, mostly the troublemaker (toxins, allergens, invasive disease)

Yeasts

Candida albicans

Histoplasma capsulatum

Blastomyces dermatitdis

Pneumocystis jirovecii

Coccidioides immitis

Trichophyton rubrum

Cryptococcus neoformans

Candida albicans

Yeast Ascomycetes

Diploid that grow as yeast and filamentous (w/ hyphae) forms

Reversible process of phenotypic switching (environ regulation of genes and spontaneous gene switching generates different morphologies)

Part of our normal microbiota ~80% of human population, but can cause opportunistic infections oral thrush, genital vaginitis, systemic candidiasis (esp. immunocompromised)

Histoplasma capsulatum

Yeast Ascomycetes

Yeast form at 37C in body, histoplasmosis resembles TB and associated with AIDS and lesions of lungs and other organs, common in Midwest (dry, dusty soil)

Bird and bat droppings

Mostly immunocompromised people

Pneumocystis jiroveci

Yeast-like fungi taking place in the lungs, cyst-formation during sexual cycle

Trophozoite → sporozoites → cyst

Causes lethal pneumonia in AIDS patients

Cannot be grown in culture, spreads person to person by air droplets

Blastomyces dermatidis

Common in Midwest USA

Yeast form at 37C (in lungs)

Causes skin and bone lesions, ARDS, and lung disease

Coccidioides immitis

Outside body (saprobic cycle) septate mycelium that fragments into airborne arthrospores

Endemic in desert areas of SW USA

In lungs (coccidiodomycosis - associated with AIDS, presents as difficult to treat meningitis, lethal pneumonia, ARDS)

The spores swell into spherule (cyst-like) form that fragments into endospores that are released into the environment

Trichophyton rubrum

Athlete’s foot, jock itch, ringworm - common sports or gym ailments

Cryptococcus neoformans

Encapsulated yeast (characteristic anti-phagocytic capsular polysaccharide) Urban soil

Pigeon droppings

Spore airborne

Immunocompromised patients have invasive lung and brain infections (meningitis)

Protozoans

Single-celled eukaryotic protists. Parasitic ones alternative between two forms:

• Trophozoite: inside the host, what it does: mature, feeding, reproducing form, often motile, vulnerable, active form cannot survive changes

• Cyst: outside the host, in the environment, what it does: dormant, infectious, survives harsh conditions (pH, nutrient, temp changes)

General cycle of protozoans

1. A cyst is ingested (or transmitted by a vector such as an insect)

2. Inside the host it becomes a trophozoite, which feeds and reproduces

3. Some trophozoites form cysts, which are released (in stool or by insect bite)

4. The cyst is stable and infectious in the environment, ready for the next host

Eukaryotic flagella & motility

1. Whipping motion, not spinning

2. Made of tubulin, not flagella

3. Microtubules in a 9+2 arrangement

4. Covered by plasma membrane

5. Powered by ATP and dynein, not proton motive force

Free-living protozoan models (movement & structure)

Paramecium & Tetrahymena: cilia, contractile vacuole, undulating movement, P. also has a cytostome (mouth) and cytoproct (anus)

Amoeba: shape-changing pseudopods, contractile vacuole, ecto/endplasm

Euglena: flagellum, contractile vacuole, ecto/endoplasm

Dinoflagellates cause “red-tides”

Amoebas (parasitic protozoan infections)

Amoebas - Entamoeba histolytica (dysentery, diarrhea, liver/lung/brain abscesses), Acanthamoeba (eye infections), N. fowleri (soil & stagnant water, brain eating, encephalitis)

Flagellates (parasitic protozoan infections) Localized, anaerobic

Shared features: 2 nuclei, each with 4 flagella, no mitochondria, no Golgi

• Giardia intestinalis: phylum Diplomonada, colonizes small intestine, causing localized diarrhea. Trophozoite swims, cyst is infective and environmentally resistant. EX. hikers drinking untreated stream water

• Trichomonas vaginalis: phylum Metamonada. Infects the urogenital tract of men and women, causing vaginitis. It is an STD and has no cyst

Flagellates (parasitic protozoan infections) Invasive

Insect vector-borne

Membrane-sheathed flagella:

• Trypanosoma brucei - African sleeping sickness, tsetse fly

• Trypanosoma cruzi - Chaga’s disease, found in Americas, triatomine (“bedbug”), parasite enters through bite wound contaminated with insect feces

  • Forms: trypomastigote (in blood, infective, non-replicating) & amastigote (intracellular)

Unsheathed flagella:

• Leishmania species (leischmaniasis) - transmitted by sandflies, SW and Central Asia, L. major - skin lesions, problem with soldiers returning, L. donovani - visceral (organ) leish

Apicomplexans (invasive)

• Cryptosporidium parvum (diarrhea), localized, life cycle within single host including infective cysts (oocysts) — problem for outdoor water sports

• Toxoplasma gondii (flu-like illness, tissue cysts/oocysts, congenital defects), ingested, sexual cycle of cats, transmitted via feces (oocysts in cat feces) - problem for pregnant women with cats as pets

• Plasmodium falciparum (malaria - a global health problem), cycling fever occurs every 48-72 hrs due to lysis of red blood cell (~300-500 million child deaths/yr). Bite of female mosquito

• Cyclospora cayetanensis: explosive diarrhea, iceberg lettuce, contaminated fresh produce or water

Plasmodium’s 3 life stages

1. Sporozoite: injected by the mosquito, infects liver

2. Merozoite: released from the liver, infects, bursts red blood cells (cause of fever cycle)

3. Gametocyte: formed in RBCs, taken up by a mosquito, where sexual reproduction occurs

Minimal components of a virus

A protein capsid and a genome of DNA or RNA

A membrane is not required

Virion

An individual, infectious virus outside a host cell

Capsid

Protein forms the capsid

The protein coat that protects the genome. Euk viruses sometimes also have a membrane envelope made of lipids taken from the host cell

Genome

DNA or RNA, never both. Either type can be single-stranded (ss) or double-stranded (ds)

• DNA genomes can be linear, circular, or partially circular

• RNA genomes are linear, never circular, and are sometimes segmented

Genome size

About 3,000 bp up to 2.8 million bp (E. coli is about 4.6 million bp)

Size and how to see viruses

Generally small, around 100 nm (0.1 µm). Bacteria are about 1 µm, and human cells are about 10 µm

Need an electron microscope to see most viruses; TEM is best for identifying them

Lytic cycle, bacteriophage replication

1. Attachment: virus binds the host cell

2. Entry: virus genome is transferred into the host

3. Synthesis: host machinery makes all the virus parts

4. Assembly: parts are organized into new virions

5. Release: host cell is lysed, freeing new virion

Lysogenic cycle, bacteriophage replication

Attachment→ Entry → Integration into the host chromosome (prophage)→ Lysogeny (replication with host cell) → Induction (excise from chromosome and enter lytic cycle) → Synthesis → Assembly → Release

Replicates using host DNA machinery

Viral DNA is integrated into the host genome

Integration in the lysogenic cycle

The phage genome inserts into the host chromosome. It is now called a prophage

Only in lysogenic phages

Lysogeny in the lysogenic cycle

Normal cell division replicates the host chromosome and the prophage, so every daughter cell inherits it

Only in lysogenic phages

Induction in the lysogenic cycle

Genome is excised from the host chromosome, and the phage enters the lytic cycle (synthesis, assembly, release)

Only in lysogenic phages

Burst time and burst size

Attachment (0 min): starting virions are present

Entry (~0 to 5 min): count drops, bc the virion is gone and only the genome is inside the cell

Synthesis and assembly (~5 to 25 min): count stays flat near zero, bc no complete virions exist yet

Burst time: time at which the cell lyses and virions suddenly appear, ~ 25 min here. It is the time required to complete the lytic cycle

Burst size: the number of virions released per cell, which is the plateau height after the jump (~200)

Bacteriophage (phage)

A virus that infects and replicates within bacteria

Animal virus replication cycle

Attachment - diff from phage (tail fibers), animal viruses do not have tails, so they use glycoprotein spikes to bind host receptors

Entry - can be similar or different (three routes)

Synthesis - similar; both hijack host replication, transcription & translation. Animal DNA viruses usually do this in the nucleus

Assembly - diff from phage (assemble only in cytoplasm); animal viruses can assemble in nucleus, cytoplasm, plasma membrane, or organelles

Release - can be similar or different (two routes)

Why a difference in Animal viruses with phage

Animal cells are euk (nucleus, organelles, glycosylation) and have no cell wall, but they do have a membrane that can wrap a virion. Most bacteria can’t add sugar to proteins, which is why glycoprotein spikes are an animal-virus feature

3 entry routes for animal virus during cell replication

1. Direct penetration (injection): This is the most like phage, viruses inject genome into host cells

2. Membrane fusion: envelope fuses with the host membrane, capsid enters the cytoplasm and uncoats (falls apart) and releases the genome

3. Endocytosis: host membrane engulfs the virion into an endosome. The virion escapes into the cytoplasm then uncoats

Uncoating is the new idea. Phage don’t do it because they inject only the genome

2 release routes for animal virus during cell replication

1. Lysis (non-enveloped viruses): the host cell bursts and releases “naked” virions. This is like phage, and it kills the cell (dsDNA viruses like herpes and papilloma)

2. Budding (enveloped viruses): the capsid gets coated in host membrane that already has viral proteins embedded in it; the virion pinches off. The cells isn’t necessarily killed. Most budding viruses hijack host proteins normal used for cell division

Enveloped

Host cell membrane with viral proteins in it. Envelopes can add complex surface receptors, help hide the virus from the immune system, and let the host cell survive release, more fragile, can be targets for antibodies/drugs

Non-enveloped

“Naked” may persist longer in the environment

Aiding fecal-oral or surface transmission

More resistant to environmental stress and disinfectant, surviving longer

DsDNA viruses and symptoms (disease)

Pox viruses - enveloped - smallpox, cowpox, etc

Herpes viruses - enveloped - Oral (HSV-1), genital (HSV-2), chicken pox/shingles (varicella zoster), Epstein-Barr virus

Papillomaviruses - nonenveloped - warts

Adenoviruses - nonenveloped - common cold, pinkeye, GI illnesses

Hepadnaviruses - enveloped - Hepatitis B (partly double and single stranded)

Parvoviruses - nonenveloped

Infection outcomes

Lytic infections - virus released and cleared

Latent infection - dormant, then recurs

Persistent/chronic infection - slow active infection, mild or no symptoms

Transformation to malignancy - cancer

RNA viruses

• Categorized by genomic structure, presence or absence of an envelope, and capsid size and shape

• Flimsy, mutate fast, change often, spread easily, can jump between species (bird flu→humans), change enough that you need new vaccines (flu shots), evolve into new variants (like COVID-19 variants)

• Four types: +ssRNA, -ssRNA, +ssRNA retroviruses (convert to DNA), dsRNA → + can be used directly by ribosome like mRNA, - must first be transcribed into mRNA

• The genome structure shapes how the virus replicates. The biggest differences between virus types are in the synthesis step

+ssRNA viruses

Can directly be used by ribosome just like mRNA to translate protein

Examples & impact on human & animal health:

• Coronaviruses, rhinoviruses, polioviruses, norovirus, Zika virus, West Nile, Dengue Fever, Yellow fever, hep C virus

• Mosquitoes are vectors for many of these viruses. Most are zoonotic (transferred to humans from animals)

• Coronavirus lifecycle - SARS-CoV2

Coronavirus Life-cycle

1. Attachment (spikes bind)

2. Entry (uncoating, endocytosis (cell engulfs external materials))

3. Synthesis (replication and transcription of viral RNA, then translation using host machinery)

4. Assembly (at the ER membrane)

5. Release (virus released through the Golgi apparatus, leaves by exocytosis - fuse to plasma membrane then release)

-ssRNA viruses

Must first be transcribed into mRNA before being used by the ribosome to translate into protein

Examples & impact on human health:

• Ebola virus, influenza A virus (acquires mutations rapidly, swapping genome segments.) Antigenic drift vs. shift

Antigenic drift (type of genetic variation in viruses)

Drift: a new strain that differs slightly from the original; it happens constantly, which is why seasonal flu strains vary slightly from year to year

Surface proteins HA (hemagglutinin) and NA (neuraminidase) are responsible for the production of new strains of influenza A & B

Antigenic shift (type of genetic variation in viruses)

Shift: a new virus that can be very different from both parents, roughly once a decade, two different strains, influenza A only (B does not undergo shift)

+ssRNA retroviruses

Use reverse transcriptase to convert +ssRNA into dsDNA that gets integrated into the host chromosome

The +ssRNA is also used directly by ribosome to translate viral protein

Examples & impact on human health: HIV lifecycle

HIV lifecycle

1. Attachment: binding to the host cell. GP 120 binds CD4 receptor, then coreceptor (CCR5 or CXCR4)

2. Entry: fusion and transfer of the genome into the cell

3. Integration: reverse transcriptase makes dsDNA from RNA, permanently inserting the genome into the host genome. Allows ‘latency’ to occur

4. Synthesis: using the host machinery to make all of the parts of virus (transcription, then translation)

5. Assembly: organization of all the virus parts to make new viruses

6. Release: budding through the envelope, so new virions leave to repeat the cycle

Latency of animal viruses

When animal viruses integrate into host chromosomes and remain dormant (can be for many years) in host cells (like HIV), the viruses are latent viruses or proviruses (like prophages). This integration is permanent (unlike prophage)

Disease course

Primary infection → acute infection→ clinical latency → immune dysfunction → AIDS (death from being immunocompromised and unable to fight other microbes)

having'/living with HIV does not mean having AIDS

Bacteriophage vs. animal viruses

Bacteriophage

• Tail proteins bind proteins on the cell wall

• Genome injected or diffuses in

• No coating

• Site of synthesis & assembly: cytoplasm

• Release - lysis

• Chronic infection: lysogeny: always incorporated and may leave chromosome

Animal virus

• Spikes, capsid, or envelope proteins bind proteins/glycoproteins on the cell membrane

• Capsid enters by direct penetration, fusion, or endocytosis

• capsid removed by cell enzymes (coating)

• Site of synthesis & assembly: RNA viruses - cytoplasm, Most DNA virus: nucleus

• Release - naked: exocytosis or lysis, enveloped: budding

• Chronic infection: latency: with or without incorporation, incorporation is permanent