VIRUSES

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Are Viruses Living or NonNon-living?

Viruses are both and neither They have some properties of life but not others For example, viruses can be killed, even crystallized like table salt However, they cant maintain a constant internal state (homeostasis).
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What a Virus Isnt

Not a bacterium... Not independent... Cannot survive in absence of a living cell within which to replicate... Antibiotics generally dont work on them...

What are Viruses?

A virus is a noncellular particle made up of genetic material and protein that can invade living cells.

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What Viruses Are...

Infectious agents composed mainly of nucleic acid with a protein coat (capsid) Visible with electron microscope (10-200 nM) Carry on normal cell-like function (unless free, then infectious) In infectious form: no growth; no respiration??? Can enter living plant, animal or bacterial cell

Discovery of Viruses
Beijerinck (1897) coined the Latin name virus meaning poison He studied filtered plant juices & found they caused healthy plants to become sick

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Tobacco Mosaic Virus

Wendell Stanley (1935) crystallized sap from sick tobacco plants He discovered viruses were made of nucleic acid and protein
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Smallpox
Edward Jenner (1796) developed a smallpox vaccine using milder cowpox viruses Deadly viruses are said to be virulent Smallpox has been eradicated in the world today
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Viewing Viruses
Viruses are smaller than the smallest cell Measured in nanometers Viruses couldnt be seen until the electron microscope was invented in the 20th century
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Electron Microscopy

Mitra, K. & Frank, J., 2006. Ribosome dynamics: insights from atomic structure modeling into cryo-electron microscopy maps. Annual review of biophysics and biomolecular structure, 35, 299-317.

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X-ray Crystallography of Viruses

Symmetry of protein shells makes them uniquely well-suited to crystallographic methods Viruses are the largest assemblies of biological macromolecules whose structures have been determined at high resolution

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Viral Structure
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Virus Appearence?
1. Capsid 2. Core and genetic material (DNA/RNA) Capsid: outer shell of the virus which encloses genetic material (link: chemical structure of capsid helps determine immune response to virus) capsid is made of many identical individual proteins protein core under capsid protecting genetic material sometimes an additional covering (lipid bilayer w/embedded proteins) on outside known as an envelope ( like a baseball) various forms: rods, filaments, spheres, cubes, crystals
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Characteristics
Non living structures Noncellular Contain a protein coat called the capsid Have a nucleic acid core containing DNA or RNA Capable of reproducing only when inside a HOST cell
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Characteristics
Some viruses are DNA enclosed in an protective envelope Some viruses may have spikes to help attach to the host cell Most viruses infect only SPECIFIC host ENVELOPE cells
CAPSID

SPIKES

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Characteristics
Viral capsids (coats) are made of individual protein subunits Individual subunits are called capsomeres

CAPSOMERES

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Capsid

capsomere: unit/molecule associated with capsid structure

Characteristics
Outside of host cells, viruses are inactive Lack ribosomes and enzymes needed for metabolism Use the raw materials and enzymes of the host cell to be able to reproduce
HIV VIRUS
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EBOLA VIRUS

Size of Viruses

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Viral Shapes
Viruses come in a variety of shapes Some may be helical shape like the Ebola virus Some may be polyhedral shapes like the influenza virus Others have more complex shapes like bacteriophages
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Typical Virus Shapes

RODS

SPHERES

CUBES

The Structure of Viruses

Tobacco Mosaic Virus

HIV

Bacteriophage

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Helical Viruses

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Polyhedral Viruses

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More Virus Shapes

Complex Viruses

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Taxonomy of Viruses

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Viral Taxonomy
Family names end in -viridae Genus names end in -virus Viral species: A group of viruses sharing the same genetic information and ecological niche (host). Common names are used for species Subspecies are designated by a number
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Viral Taxonomy Examples

Herpesviridae Herpesvirus Human herpes virus 1, HHV 2, HHV 3 Retroviridae Lentivirus Human Immunodeficiency Virus 1, HIV 2

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Herpes Virus

SIMPLEX I and II

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Adenovirus

COMMON COLD
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Influenza Virus

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Papillomavirus Warts!

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Used for Virus Identification

RNA or DNA Virus Do or do NOT have an envelope Capsid shape HOST they infect

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Bacteriophages

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Phages
Viruses that attack bacteria are called bacteriophage or just phage T-phages are a specific class of bacteriophages with icosahedral heads, double-stranded DNA, and tails
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T-phages
The most commonly studied T-phages are T4 and T7 They infect E. coli , an intestinal bacteria Six small spikes at the base of a contractile tail are used to attach to the host cell Inject viral DNA into cell

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Escherichia Coli Bacterium

T - EVEN PHAGES ATTACK THIS BACTERIUM

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E. Coli and the Bacteriophage

What it looks like in real life

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T-Even Bacteriophages

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Diagram of T-4 TBacteriophage

Head with 20 triangular surfaces Capsid contains DNA Head & tail fibers made of protein
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Composition of T-Even Bacteriophage

Capsid: brains of virus, tightly-wound protein protecting nucleic acids Body: attached to capsid head, rod-like structure w/retractible sheath, hollow core Tail: at end of core is a spiked plate carrying 6 slender tail fibers, anchor virus to its host
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Retroviruses

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Characteristics of Retroviruses
Contain RNA, not DNA Family Retroviridae Contain enzyme called Reverse Transcriptase When a retrovirus infects a cell, it injects its RNA and reverse transcriptase enzyme into the cytoplasm of that cell
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 Change DNA into RNA.

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ENZYME

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Retroviruses
The enzyme reverse transcriptase (or RTase), which causes synthesis of a complementary DNA molecule (cDNA) using virus RNA as a template
RTase
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Retroviruses
HIV, the AIDS virus, is a retrovirus Feline Leukemia Virus is also a retrovirus

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Viroids & Prions

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Viroids
Small, circular RNA molecules without a protein coat Infect plants Potato famine in Ireland Resemble introns cut out of eukaryotic
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Prions
Prions are infectious proteins They are normal body proteins that get converted into an alternate configuration by contact with other prion proteins They have no DNA or RNA The main protein involved in human and mammalian prion diseases is called PrP
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Prion Diseases
Prions form insoluble deposits in the brain Causes neurons to rapidly degeneration. Mad cow disease (bovine spongiform encephalitis: BSE) is an example People in New Guinea used to suffer from kuru, kuru, which they got from eating the brains of their enemies
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Viral Replication

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How do viruses work?

Viruses make use of the host cells chemical energy, protein and nucleic acid synthesizing ability to replicate themselves... each virus attacks a specific type of cell
cold viruses attack cells of the lung the AIDS virus attacks T4 cells of the immune system fish viruses are just as specific

Bottom Line...
All viruses only exist to make more viruses Most are harmful Replication = host cell death.

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Virusal Mechanism
Viruses contain single- or double- stranded DNA or RNA Often, the virus alters the intracellular environment enough to damage or kill the cell (oops!!) If enough cells are destroyed, disease results!

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Role of RNA/DNA
Supplies the codes for building the protein coat (capsid) and for producing enzymes needed to replicate more viruses Information given so newly-built viruses can lyse cells (e.g., bacteriophage) Result: cell destroyed.

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Viral Attack
Viruses are very specific as to which species they attack HOST specific Humans rarely share viral diseases with other animals Eukaryotic viruses usually have protective envelopes made from the host cell membrane
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5 Steps of Lytic Cycle

1. Attachment to the cell 2. Penetration (injection) of viral DNA or RNA 3. Replication (Biosynthesis) of new viral proteins and nucleic acids 4. Assembly (Maturation) of the new viruses 5. Release of the new viruses into the environment (cell lyses) : DNA incorporated (10 minutes) hundreds of virions appear causing the cell to rupture, releasing hundreds of small viral replicates
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Bacteriophage Replication Bacteriophage inject their nucleic acid They lyse (break open) the bacterial cell when replication is finished
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Bacteriophage Attack

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Lytic Cycle Review

Attachment Penetration Phage attaches by tail fibers to host cell Phage lysozyme opens cell wall, tail sheath contracts to force tail core and DNA into cell Production of phage DNA and proteins Assembly of phage particles Phage lysozyme breaks cell wall
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Biosynthesis Maturation Release

Common steps in the assembly of all dsDNA viruses

Unique portal ring at one Vertex Scaffolding proteins Procapsid assembled empty of DNA DNA pumped into procapsid through portal ring DNA moves back through portal to enter cell

Pathway

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Bacterial cell wall

Bacterial chromosome

Capsid

DNA Capsid

Sheath

Tail fiber Attachment: Phage attaches to host cell. Base plate Pin Cell wall

Tail

Plasma membrane

Penetration: Phage pnetrates host cell and injects its DNA.

Sheath contracted

Tail core

3 Merozoites released into bloodsteam from

liver may infect new red blood cells UY - 2011

Tail DNA

4 Maturation: are Viral components

assembled into virions. Capsid

5 Release: lyses and Host cell

new virions are released. Tail fibers

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OneOne-step Growth Curve

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Viral Latency
Some viruses have the ability to become dormant inside the cell Called latent viruses They may remain inactive for long periods of time (years) Later, they activate to produce new viruses in response to some external signal HIV and Herpes viruses are examples UY - 2011

Lysogenic Cycle
Phage DNA injected into host cell Viral DNA joins host DNA forming a

prophage
When an activation signal occurs, the phage DNA starts replicating

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Lysogenic Cycle
Viral DNA (part of prophage) may stay inactive in host cell for long periods of time Replicated during each binary fission Over time, many cells form containing the prophages

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Viral Latency
Once a prophage cell is activated, host cell enters the lytic cell New viruses form a & the cell lyses (bursts) Virus said to be virulent (deadly)

ACTIVE STAGE

INACTIVE STAGE UY - 2011

Virulent Viruses

HOST CELL LYSES & DIES

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The Lysogenic Cycle

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Latency in Eukaryotes
Some eukaryotic viruses remain dormant for many years in the nervous system tissues Chickenpox (caused by the virus Varicella zoster) is a childhood infection It can reappear later in life as shingles, a painful itching rash limited to small areas of the body

SHINGLES

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Latency in Eukaryotes
Herpes viruses also become latent in the nervous system A herpes infection lasts for a persons lifetime Genital herpes (Herpes Simplex 2) Cold sores or fever blisters (Herpes Simplex1)
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SKIN TO SKIN CONTACT

PASSED AT BIRTH TO BABY

The Virus Invasion

Whats Infected by a Virus?

All living things have some susceptibility to a particular virus Virus is specific for the organism Within a species, there may be a 100 or more different viruses which can affect that species alone Specific: for example, a virus that only affects one organism (humans and smallpox) Influenza can infect humans and two animals

Different Types of Viruses

Major classifications: animal, plant, bacterial Sub-classified by arrangement and type of nucleic acid Animal virus group: double-stranded DNA, singlestranded DNA, double-stranded RNA, singlestranded RNA, retrovirus Influenza: SS-RNA

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Do Viruses ever Change?

Mutations do occur. If the mutation is harmful, the new virus particle might no longer be functional (infectious) However, because a given virus can generate many, many copies, a small number of non-functional viruses is not important Mutation is not necessarily damaging to the virus -it can lead to a functional but new strain of virus

Virulence

VIRUS DESTROYING HOST CELL

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Lytic and Lysogenic Cycles

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Defense Against Viruses

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Defense Against Viruses

First Line: skin and mucous membrane, which also lines the gastrointestinal and respiratory passageways skin is tough and stomach acidity acts as a disinfectant Second Line: after the virus enters the blood and other tissues, white blood cells and related cells (phagocytes) consume them accumulation of phagocytes in area of infection is known as puss

Defense Against Viruses

Antibodies attacking chickenpox virus

Defense Against Viruses

Antibodies are the best defense against viruses unfortunately, they are specific in their action chickenpox antibody will only attack a chickenpox virus a particular virus stimulates the production of a particular antibody

Defense Against Viral Infection

Animals are protected in several ways: 1) intracellular: if a particular virus attacks cells, our bodies produce interferons interferons (alpha, beta or gamma) are proteins which interact with adjacent cells and cause them to become more resistant to infection by the virus if the resistance is not quite good enough, we become sick

Defense Against Viral Infection

2) immune system (extracellular): kills the virus outside the cell also kills the infected cells virus cannot spread eventually the virus is completely removed and we get better exception: HIV because it infects cells of the immune system, itself chemicals/drugs: acyclovir, AZT, HIV protease inhibitor, several fish vaccines available.

Major Fish Viruses

Major Viral Infections in Fish

Infectious pancreatic necrosis (IPN) Viral hemorrhagic septicemia (VHS) Infectious hematopoetic necrosis (IHN) Channel catfish virus disease (CCVD)

(1) Infectious Pancreatic Necrosis (IPN)

What?: viral infection of salmonids (trout and char) Time: Acute Result: high mortality (fry and fingerlings) Rare in larger fish (good thing!) History: Isolated in Pacific NW in 1960s, wiped out brook trout in Oregon in 1971-73 Size: Only 65 nM in diam., smallest of fish viruses

IPN: general notes

Single capsid shell, icosohedral symmetry, no envelope Contains two segments of DS-RNA Fairly stable and resistant to chemicals (acid, ether, etc.), variable resistance to freezing Remains infectious for 3 months in water (uh oh!) Targets pancreas and hematopoietic tissues of kidney and spleen

IPN: epizootiology (disease process)

Who?: All salmonids, brook trout most susceptible, marine fish (flounder?) Reservoirs (where)?: carriers, once a carrier always a carrier, virus particles shed in feces/urine Transmission (how?): horizontal, by waters via carriers or infected fry; vertical from adults to progeny; experimentally by feeding infected material, IP injection Pathogenesis: entry via gills, digestive tract Environmental factors: mortality reduced at lower temps (why?); however, carriers not reduced

IPN: pathology (what do we see?)

IPN: detection, diagnosis and control

Isolation: whole fry, kidney, spleen, pyloric cecae, sex fluids are all good sources, .i.e. check these!!! Presumptive tests: epizootiological evidence and/or typical PCR in infected cells Definitive tests: serology (fluorescent antibody test (FAT)) Control: avoid virus in water, virus-free stock, destruction of infected stock, vaccine exists now!

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How Bad Can It Be??

Fish severely affected by IPNV:

Atlantic salmon* (Salmo salar) brook trout* (Salvelinus fontinalis) brown trout* (Salmo trutta) danio zebrafish* (Brachydanio rerio) rainbow trout* (Oncorhynchus mykiss) yellowtail* (Seriola lalandi)

Other species known to be susceptible

amago salmon (Oncorhynchus rhodurus) Arctic char (Salvelinus alpinus) Atlantic menhadden (Brevoortia tyrannus) carangids (Carangidae) chinook salmon (Oncorhynchus tshawytscha) chum salmon (Oncorhynchus keta) cichlids (Cichlidae) coho salmon (Oncorhynchus kisutch) common scallop (Pecten maximus) cutthroat trout (Salmo clarki) cyprinids (Cyprinidae) Danube salmon (Salmo hucho) drums/croakers (Sciaenidae) eels (Anguilla spp) grayling (Thymallus thymallus)

More
halibut (Hippoglossus stenolepis) Jap. amberjack (Seriola quinqueradiata) lampreys (Petromyzontyidae) loach (Misgurnus anguillicaudatus) masou salmon (Oncorhynchus masou) perches (Percidae) silversides (Atherinidae) soles (Soleidae) striped snakehead (Channa striatus) summer flounder (Paralichthys dentatus) white seabass (Moronidae) carp (Cyprinus carpio) redfin perch (Perca fluviatilis) yellowfin bream (Acanthopagrus australis) herrings/sardines (Clupidae) lake trout (Salvelinus namaycush) left-eye flounders (Bothidae) loaches (Cobitidae) Pacific salmon (Oncorhynchus spp) pikes (Esocidae) sockeye salmon (Oncorhynchus nerka) Southwest European nase (C. toxostoma) suckers (Cotostomidae) turbot (Psetta maxima) whitefish (Coregonidae) goldfish (Carassius auratus) southern flounder (P. lethostigma)

Asymptomatic carriers...
coalfish (Pollachius virens) common carp (Cyprinus carpio) discus fish (Symphysodon discus) goldfish (Carrasius auratus) heron (Ardea cinerea) loach (Cobitidae) minnow (Phoxinus phoxinus) noble crayfish (Astacus astacus) Infectious pancreatic necrosis in Atlantic salmon. pike (Esox lucius) Note swollen stomach and 'pop eye' river lamprey (Lampetra fluviatalis) Source: Australian Animal Health Laboratory shore crab (Carcinus maenas) Spanish barbel (Barbus graellsi) white suckers (Catostomas commersoni) ...what now???

(2) Viral Hemorrhagic Septicemia (VHS)

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(2) Viral Hemorrhagic Septicemia (VHS)

What?: Viral disease of European salmonids When?: Recognized in Denmark in 1949, isolated from Pacific Coast in 1989 Size: rhabdovirus, bullet-shaped (one rounded end), 185 x 65 nM, lipoprotein envelope non-segmented SS-RNA Constitution: sensitive to ether and chloroform, heat, acid, resistant to freeze-drying

Viral Hemorrhagic Septicemia

Produces a general viremia, tissue and organ damage, liver necrosis, spleen, kidney Epizootiology: cultured rainbow trout, also brown trout, steelhead, chinook, coho (most cases in WA state) Reservoirs: again...survivors are life-long carriers, usually rainbow trout, brown in Europe Transmission: horizontal through water, virus can occur on eggs spawned by carriers, IP injection, birds, hatchery equipment

Viral Hemorrhagic Septicemia (VHS)

Pathogenesis: infection results in viremia, disrupts many organ systems, 200-300g fish most affected Environmental factors: low temp (< 8oC, 46oF) External pathology: lethargy, hanging downward in water (dropsy), swimming in circles, exopthalmia, dark discoloration, hemorrhages in roof of mouth, pale gills w/focal hemorrhages

Viral Hemorrhagic Septicemia (VHS)

Internal pathology: gut devoid of food, liver pale, hemorrhages in connective tissue, kidney gray and swollen (chronic), red and thin (acute) Histopathology: necrosis of liver, kidney nephrons, spleen, pancreas, melanin in kidneys and spleen (OUCH!) Isolation/tests: isolated from kidney/spleen, epizootiological evidence, definitive test is serum neutralization, or FAT.

Viral Hemorrhagic Septicemia (VHS) External hemorrhages

Liver red in acute stage

Viral haemorrhagic septicaemia in rainbow trout. Note pale color of stomach region, pinpoint haemorrhages in fatty tissue, and pale gills

Source: T Hstein

Viral haemorrhagic septicaemia in rainbow trout. Note swollen stomach and pop eye

Type I-a I-b I-c I-d I-e II

Prevalent host type and location Farmed rainbow trout and a few other freshwater fish in continental Europe[10] Marine fish of the Baltic Sea, Skagerrak, Kattegat, North Sea, Japan[1] Farmed rainbow trout Denmark Farmed rainbow trout in Norway, Finland, Gulf of Bothnia Rainbow trout in Georgia, farmed and wild turbot in the Black Sea[11] Marine fish of the Baltic Sea Marine fish of the British Isles and northern France, farmed turbot in the UK and Ireland, and Greenland halibut (Reinhardtius hippoglossoides) in Greenland[12]

III

IV-a IV-b

Marine fish of the Northwest Pacific (North America), North American north Atlantic coast,[13] Japan, and Korea[1][14] Freshwater fish in North American Great Lakes region[14]

Virus presence spread through much of the Great Lakes from 2003-2007.

Viral Hemorrhagic Septicemia

Prevention: clean broodstock and water = clean fish, avoid infected broodstock, test and slaughter Can spread very quickly from farm to farm: avoid close proximity to other farms Vaccines are under development. One EPA-approved disinfectant: Virkon AQUATIC (made by Dupont). Bleach kills the VHS virus.

(3 ) Infectious Hematopoietic Necrosis (IHN)

Who: sockeye, chinook, rainbows; cohos resistant When?: 1950s in Oregon hatcheries. 100 million mortalities between 1970-1980, if infected, 70% mortality likely, esp. in young fish (fry: 90-95% mort. possible) What?: bullet shaped rhabdovirus, non- segmented SS-RNA, sensitive to heat and pH, glycoprotein is spiked on surface of virus

Infectious Hematopoietic Necrosis (IHN)

Reservoirs: survivors life-long carriers, adults shed virus at spawning Transmission: horizontal, primary mode is vertical via ovarian fluid (virus hitches ride on sperm into egg); however, feces, urine, and external mucus possible. Also, feeding and inoculation have worked experimentally Pathogenesis: gills suspected; incubation period depends on temp, route, dose, age; extensive hemorrhaging, necrosis of many tissues; death usually due to kidney failure

Infectious Hematopoietic Necrosis (IHN)

Environmental factors: temp. very important, slows below 10 C, holding in tanks/handling increase severity (doesnt occur naturally >15 C) External pathology: lethargy, whirling, dropsy, exopthalmia, anemia, hemorrhaging of musculature/fins, scoliosis Internal pathology: liver, kidney, spleen pale; stomach/intestines filled with milky fluid; petechial hemorrhaging Histopathology: extensive necrosis of hematopoetic tissue of kidney/spleen

Infectious Hematopoietic Necrosis (IHN)

Definitive diagnosis: serum neutralization, FAT, ELISA Prevention: avoidance, quarantine, clean water with UV, ozone, virus-free stock; test, slaughter, disinfect; disinfect eggs; vaccines under development; elevated water temp No vaccines as of June 2007.

(4) Channel Catfish Virus Disease (CCVD)

Contagious herpes virus affecting only channel catfish less than four months old Occurs in SE United States, California, Honduras Acute hemorrhagia, high mortality, discovered in 1968 Agent: enveloped capsid, icosohedral nucleocapsid with 162 capsomeres Physio/chemical properties: easy to kill, sensitive to freeze-thaw, acid, ether, etc.

Channel Catfish Virus Disease (CCVD)

Environmental factors: optimal temperature 28-30 C, common during warmer months, cooler water = big difference epizootiology: horizontal, vertical suspected external pathology: spiral swimming; float with head at surface; hemorrhagic fins, abdomen; ascites; pale or hemorrhagic gills; exophthalmia

Channel Catfish Virus Disease (CCVD)

Internal pathology: hemorrhages of liver, kidney, spleen, gut, musculature; congestion of mesenteries and adipose Histopathology: necrosis of kidney, other organs; macrophages in sinusoids of liver, etc.; degeneration of brain Presumptive diagnosis: clinical signs, epizootiological evidence Definitive diagnosis: SN or FAT.

Channel Catfish Virus Disease (CCVD)

Prevention: avoid potential carriers (survivors) or infected fry, keep temperature below 27oC (will still produce carriers), attenuated vaccine shows some promise Therapy: none available...

Channel Catfish Virus Disease

Channel Catfish Virus Disease

However, you can always take precautions!

Treatment for Viral Disease

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Vaccines
An attenuated virus is a weakened, less vigorous virus Attenuate" refers to procedures that weaken an agent of disease (heating) A vaccine against a viral disease can be made from an attenuated, less virulent strain of the virus Attenuated virus is capable of stimulating an immune response and creating immunity, but not causing illness
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Perbandingan beberapa tipe vaksin ikan

Jenis Vaksin
Inactivated vaccine

Kelebihan
Resiko infeksi (-)

Kekurangan
Mahal, Kasus perkasus tdk ada efeknya, Imunitas jangka pendek -> perlu injeksi booster Mahal, Potensi resiko terinfeksi

Jenis Virus
VHSV, RSIV, GCHV, ISAV, IPNV, IHNV, SVCV, CCV IPNV, IHNV, VHSV, CCV

Generasi I
Live attenuated vaccine Imunitas jangka panjang, Menginduksi respon imun cellular dan humoral Tidak mahal, Memungkinkan produksi skala besar

Generasi II Generasi III

Recombinant protein vaccine

Sulit dalam purifikasi (biaya produksi tinggi) Tidak mengaktifkan imun respon seluler

IHNV, VHSV, SVCV

Tanpa resiko infeksi; Mudah Terbatas pada protein DNA vaccine dikembangkan & produksi; immunogens Vaksin Stabil; Menginduksi respon imun cellular & humoral

IHNV, VHSV, HIRRV, IHV1

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Other Viral Treatments

Interferon are naturally occurring proteins made by cells to fight viruses Genetic altering of viruses (attenuated viruses) Antiviral drugs (AZT) Protease inhibitors prevent capsid formation
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