Methods of Study of Virus

There are six different methods of study of viruses.
The six different methods of study of viruses are as follows:
· Embryonated eggs
· Primary cell culture
· Cytopathic Effect (CPE)
· Plaque assay
· Hemagglutination
· Western blot


Embryonated eggs

This picture shows a typical avian egg with the parts labelled.
In the process of embryonating eggs, the influenza virus is studied through isolation and cultivation.

Primary cell culture
This picture shows the cells in culture. They are stained for keratin in red colour and DNA in green colour.
Primary cell cultures derive cell cultures. They typically will have a finite life span in culture. Only cells, which are grown in vitro, undergo primary cell culture.

Cytopathic Effect (CPE)
This picture shows the structure and cytopathic effect of Nelson Bay Virus.
The cytopathic effects are visible changes in the infected host. These changes come about upon having the virus to subvert its host just to replicate itself. If there is no such visible change in the infected host, this means that the cell morphology has changed. These changes vary inter-virus and inter-host.

Plaque assay
This picture shows how a plaque assay is carried out with words to describe each step.
Plaque assay uses principle of one plaque produced by one virus on the monolayer. It requires a lot of time to complete. Its requirement is such that a cell line is infected by the virus. The result of plaque assay is related to observing cell death in infected cell culture. The accuracy varies inversely to the concentration of virus. The surrounding cells are infected as well upon having one virus to infect one cell. Viruses, which infect monolayer cells or cause cell death, can be used in plaque assay. Plaques are seen in visible circular shape. The number of infected cells varies directly to the number of visible circular plaques formed.

Hemagglutination

There are two different types of hemagglutination.
The two different types of hemagglutination are as follows:
· Hemagglutination
· Hemagglutination Inhibition
This picture shows the result of hemagglutination.
In hemagglutination, there are two spike proteins used. They are known as the neuraminidase and haemagglutinin. Only the haemagglutinin binds specifically to red blood cells.
In hemagglutination inhibition, when there is presence of antibody, then the hemagglutination will only be experimented. The virus neutralisation is a process where the antibody neutralisation decreases virus infectivity. The virus neutralisation inhibits agglutination. Agglutination is where the red blood cells glue together to prevent foreign matter from penetration.

Western blot
This picture above shows an immunoblot (western blot) analysis of proteins separated by SDS-PAGE gradientgel electrophoresis.

This picture shows a western blot using radioactive detection system.
In western blot, electrophoresis helps to separate the HIV protein antigens. The nitrocellulose paper strips are used for blotting of these protein antigens upon separation. The strip undergoes incubation with patient antibody. The removal of the unbound antibody is by washing the strip upon incubation. The result of western blot is obtained upon binding serum from an HIV-infected person and identifying the major HIV antigenic proteins.

Classification of viruses

Classification of Viruses
There are two different types of virus classification.

The two different types of virus classification are as follows:
· The Baltimore’s System for Classification
· The Lwoff’s Scheme for Classification

Baltimore’s System for Classification

In the Baltimore’s System for Classification, there are seven groups of viruses.

The seven groups of viruses are as follows:
· Group I: dsDNA Viruses
· Group II: ssDNA Viruses
· Group III: dsRNA Viruses
· Group IV: (+) sense RNA Viruses
· Group V: (-) sense RNA Viruses
· Group VI: RNA Reverse Transcribing Viruses
· Group VII: DNA Reverse Transcribing Viruses

In group I, families of vertebrate, invertebrate, bacterial, algae, and Mycoplasma viruses are involved respectively.

In group II, families of vertebrate, invertebrate, bacterial, and Spiroplasma viruses are involved respectively.

In group III, families of vertebrate, invertebrate, and protozoa viruses are involved respectively.

In group IV, families of vertebrate, invertebrate, and bacterial viruses are involved respectively.

In group V, the order is Mononegavirales. Families of vertebrate viruses are involved.

In group VI, families of vertebrate viruses are involved.

In group VII, families of vertebrate viruses are involved.



Lwoff’s Scheme for Classification

In the Lwoff’s Scheme for Classification, the families of viruses are grouped according to DNA and RNA.

In terms of DNA, families of viruses are dsDNA and enveloped.

Families of viruses are dsDNA and nonenveloped.
Families of viruses are ssDNA and nonenveloped.
Families of viruses are ds/ssDNA and enveloped.

In terms of RNA, families of viruses are positive dsRNA, nonsegmented and enveloped.

Families of viruses are positive dsRNA, segmented and nonenveloped.
Families of viruses are positive ssRNA, nonsegmented and enveloped.
Families of viruses are positive ssRNA, nonsegmented and nonenveloped.
Families of viruses are positive ssRNA, with DNA step in replication and enveloped.
Families of viruses are negative ssRNA, segmented and enveloped.
Families of viruses are negative ssRNA, nonsegmented and enveloped.
Families of viruses are negative and ambisense ssRNA segmented and enveloped.

Prions, Viroids and Virusoids

Prions
Types of prions:
• Sheep version – scrapie (TSE)
• Cow version – mad cow (BSE- a type of TSE)
• Human version – Creutzfeldt-Jakob disease

A disease-causing agent that is neither bacterial nor fungal nor viral and contains no genetic material. A prion is a protein that occurs normally in a harmless form. By folding into an aberrant shape, the normal prion turns into a rogue agent. It then coopts other normal prions to become rogue prions.

• Prions are rogue protein that transform other cellular protein (PrPC) to the prion form PrPsc.
• PrPc gene on chromosome 20
• More and more PrPsc gets transformed until they completely clogg brain cells
• Cells misfire, work poorly or don’t work at all.
• Cells dir, release prions into blood stream to re-inact other cells
PrPC normally does membrane-anchored glycoprotein

Pathogenesis
• Spongiform encephalitis
• “sick” brains, riddled with holes
• Looks like swiss cheese


CLICK PICTURE FOR A LARGER VIEW! :D
http://www.nature.com/jidsp/journal/v7/n1/thumbs/5640076f5th.gif


CLICK PICTURE FOR A LARGER VIEW! :D
http://www.rkm.com.au/BSE/prionimages/PRIONS-600.jpg


CLICK PICTURE FOR A LARGER VIEW! :D
http://www.mol.biol.ethz.ch/groups/wuthrich_group/research/struc_pics/Mammalian_prions/image

Conversion of prions
• PrPsc forms a heterodimer with normal PrPc
• Template for altering the protein fold
• Tightly coiled α-helix converted to loose β-sheets

This picture illustrates the effect of prions on cells

CLICK PICTURE FOR A LARGER VIEW! :D

Transmission
• Eating contaminated beef or contaminated mutton
• Through blood transfusion

Viroids
• Viroids are infectious agents composed exclusively of a single piece of circular single stranded RNA which has some double-stranded regions
• Plants pathogen (25 main sequences identified)
• Hepatitis D virus that infects man has a structure similar to viroids

Mechanism of action
Viroids are suspected to be ribozymes (ribozymes are RNA molecules, hydrolyze their own phosphodiester bonds, or the hydrolysis of bonds in other RNAs, but they have also been found to catalyze the aminotransferase activity of the ribosome)

Disease caused by Viroids
• The only human disease known to be caused by a viroid-like entity is Hepatitis D
• Hepatitis D was previously ascribed to a defective virus called the delta agent
• There is extensive sequence complementarity between the hepatitis D viroid RNA and human liver cell 7S RNA
- The 7S RNA structure involved in the translocation of secretory and membrane (associated particles)
• The hepatitis D viroid causes liver cell death via sequestering this 7S RNA and/or cleaving it.

Transmission
• Co-infection with Hepatitis B virus.
• Bodily fluid
- Unprotected sex
- Sharing contaminated needles
- Close proximity


CLICK PICTURE FOR A LARGER VIEW! :D
http://www.clinical-virology.org/pages/cvn/sp_vr/viral_info/bbv/HTML/hep_d_big.jpg

Virusoid
Virusoid is an infectious agent that infects plants in conjunction with an assistant virus
• Viroids (circular RNA molecules that infect plants but don't require an assistant virus)

Properties
• Not considered a virus but a subviral particle
• The size and structure is similar to viroids
• Its genome is a single molecule of single stranded circular RNA that is several hundred nucleotides long and codes for nothing but its own stucture.

Poxviridae

Poxviridae
FLOWCHART ON POXVIRIDAE!

CLICK PICTURE FOR LARGER VIEW! :D

Properties of poxviridae
• Largest family of viruses
• Unique oval-shape
• Antigenically very complex
• Remains stable in air for hours
Genome
• Linear double stranded DNA covalently cross-linked at ends
• Terminal repeated sequences
• Inverted repeats at both ends
Clinical Features
• Human only
• Respiratory Secretions
• Always associated with skin lesions
• At least 9 poxviruses cause disease in human
• Variola and Vaccinia
• 7–12 days of incubation
• Initially influenza-like symptoms
• Characteristic pustules
• Scarring of skins
• Neurological damage
• Blindness
• Death
Variola
Variola Major
• 25–30% fatalities
Variola Minor
• Less than 1% death
Practice of variolation
• Smallpox from receipients to protect against future infection
Edward Jenner
• Vaccination
- Used cowpox from milkmaids
- Vaccinate an 8-year-old James Phipps
- Then the cowpox strain challenged with smallpox.
Vaccinia
• Evolved from cowpox or smallpox
• Used as a vector
Eradication of smallpox
• no reservior for VV except fro man
• VV causes only acute infection from which the patient dies
• or, obtains lifelong immunity
• VV is an effective immunogen
• certified by WHO in 1980 to be defeated

EMERGING viruses!

What is emerging viruses?

-Emerging virueses are known viruses that has newly appeared in human population and is rapidly increasing in disease incidence.

Reasons for emergence
Virus factors
-Spontaneous evolution of a new virus entity
-Generation of a novel strain due to co-infection of different strains in an individual ( random assortment )

Human factors
-Concentration of people with shared lifestyle
-Breakdown in public health
-Climate change
-Man invading the natural habitat of animal

Spontaneous evolution of viruses
-Viral population are heterogenous
-High mutation rates
-> due to selection pressureRandom assortment

-H3N2 infects man, H5N1 infects birds
-H5 is efficient at attachment
-Co-infection of both
-Virus replicates inside host
-RNA strands transcribed
-Viral protein synthesized
-A new influenza virion formed, H5N2!
-Release of virus particles

Whats possible?
-H-Hemagglutin
-N-Neuraminidase
-16 different H types
- 9 different N types
-H1N1
-H3N2
-Only 4 strains of avian influenza virus are known to cause disease in humans: H5N2, H7N3, H7N7, H9N2

Breakdown in public health
-Not likely in developed countires
-Examples, HIV in Africa
cholera outbreak in Iraq
http://www.reliefweb.int/rw/RWB.NSF/db900SID/LSGZ-76QHDR?OpenDocument

Climate change and Viral disease
1) http://dsc.discovery.com/news/2009/01/27/mosquito-disease-warming.html
2) http://allafrica.com/stories/200901261750.html

Man invading Animal natural habitats
-Deforestation for agriculture needs
http://www.globalchange.umich.edu/globalchange2/current/lectures/deforest/deforest.html

-Closer to wild animals
- Increase possibility of zoonoes

Statistics of emerging viruses over last 6 years
http://www.dh.gov.uk/en/Publicationsandstatistics/Publications/AnnualReports/Browsable/DH_4874410

Virus Host Interaction Video

Here's a video on Virus Host Interaction on HIV Virus.

Virus-Host Interactions

General Virus Replication Cycle

6 phases in viral replication cycle:
1) Attachment
2) Entry
3) Uncoat
4) Replication
5) Assembly
6) Release

Virus Life Cycle- Growth Curve

During attachment , the virus will attach itself to the host cell receptor and gain entry to the cell. During eclipse, there will be a fall in virus titre when no infectious particles present during replication process and the virus will not be be detected in the external medium until released. During the release period, the virus will burst and new progeny virus will be assembled and released.

1) Attachment and Entry
Attachment is a specific binding between the viral capsid proteins and specific receptors (The receptors can be protein, carbonhydrate or lipid. ) on the host cell surface. This specificity also determines the host range of a virus. To infect the host cell, there must be a specific type for the virus to attach to. The virus is able to enter the host cell in 3 different routes:
-Endocytosis
-> Pinocytosis
-> Phagocytosis
-> Receptor-mediated endocytes
-Fusion
-Direct penetration

Endocytosis
Endocytosis is a process whereby the cells absorb material from outsdie the cell by engulfing it with their cell membrane.

Pinocytosis ( non specific ) is a form of endocytosis in which small particles are brought into the cell suspended within small vesicles which subsequently fuse with lysosomes to hydrolyze, or to break down, the particles.

Phagocytosis is the cellular process of phagocytes and protists of engulfing solid particles by the cell membrane to form an internal phagosome, which is a food vacuole.

Receptor-mediated endocytosis (RME), also known as clathrin-dependent endocytosis, is a process by which cells internalize molecules ( endocytosis ) by the inward budding of plasma membrane vesicles containing proteins with receptor site is specific to the molecules being internalized.

Uncoating
Nucleic acid has to be sufficiently uncoated that virus replication can begin at this stage. When the nucleic acid is uncoated, infectious virus particles cannot be recovered from the cell.

After uncoating
Virus genome will be sensitive to external environment:
-Mechanical shearing
-UV irradiation
-pH
-Dehydration
-Enzymes

The capsid and envelope will provide protection.

After uncoating, it ensures that the infection was successful.


Penetration
(A) Entry by fusing with the plasma membrane.

Some enveloped viruses fuse directly with the plasma membrane. Thus, the internal components of the virion are immediately delivered to the cytoplasm of the cell.

(B) Entry via endosomes at the cell surface

Some enveloped viruses require an acid pH for fusion to occur and are unable to fuse directly with the plasma membrane. These viruses are taken up by invagination of the membrane into endosomes. As the endosomes become acidified, the latent fusion activity of the virus proteins becomes activated by the fall in pH and the virion membrane fuses with the endosome membrane. This results in delivery of the internal components of the virus to the cytoplasm of the cell
Non-enveloped viruses Non-enveloped viruses may cross the plasma membrane directly or may be taken up into endosomes. They then cross (or destroy) the endosomal membrane.

Stages in Virus Life Cycle
-Initiation of infection
-> Attachment and entry
-Replication and expression
-> Genome replication
->mRNA production, processing and translation
-Assembly and exit
-Viral Pathogenesis

Replication and expression
-Genome replication and gene expression are very closely linked
-Characteristics depends on nature of the genome

BALTIMORE CLASSIFICATION








Click for bigger view!

Regulation of Expression

Transcriptional control
-Promoters on viral genome
-Early and late activators / enhancers
-Late repressors
-Viral transciptases for RNA viruses not well understood
-Post-transcriptional control
-Splicing of polycistronic mRNA in nucleus
-Differential rate of splicing


Post-transcriptional control
-Control of mRNA from nucleus to cytoplasm
-Regulatory sequences found on introns
-Translational control
-Differential stability of mRNAs
-Secondary structures close to initiation sequence
-Problem due to overlapping reading frames


Translational control – overlapping reading frames
-IRES – internal ribosomal entry sites
-Frameshifting
-Pseudoknots

Translation of protein
Translation is the first stage of protein synthesis (part of the overall process of gene expression). It is also the production of proteins by decoding mRNA produced in transcription.

Translation occurs in the cytoplasm where the ribosomes are located. Ribosomes are made of a small and large subunit which surrounds the mRNA. In translation, mRNA is decoded to produce a specific polypeptide. This uses an mRNA sequence as a template to guide the synthesis of a chain of amino acids that form a protein. Translation proceeds in four phases: activation, initiation, elongation and termination (all describing the growth of the amino acid chain, or polypeptidethat is the product of translation). Amino acids are brought to ribosomes and assembled into proteins.

Virus Assembly and Exit
Assembly depends on site of synthesis. There are different sites of protein synthesis and processing such as the endoplasmic reticulum and golgi body. There are also sites of assembly such as the nucleus, the endoplasmic reticulum and golgi body.
The cell from which the body itself buds from goes to survive, and sheds more viral particles. The lipid bilayer envelope of the viruses, is derived from the host cell itself and the sources of the envelope can be found in the nuclear membrane, endoplasmic reticulum, golgi body and plasma membrane.

Read more: http://knipelab.med.harvard.edu/pdfs/viral_capsids-Lucas.pdf

Budding
Budding is the process by which enveloped viruses acquire their external envelope, which bulges outwards and takes the virion inside. This method helps the virus leave the cell without lysing the cell, thereby allowing the cellular machinery to produce more viruses.

Release
Virus may be released due to cell lysis, or, if enveloped, may bud from the cell. Budding viruses do not necessarily kill the cell. Thus, some budding viruses may be able to set up persistent infections. Not all released viral particles are infectious. The ratio of non-infectious to infectious particles varies with the virus and the growth conditions.

Viral Pathogenesis
To successfully infect a host, there must be sufficient to initiate the infection and a balance between viral replication, host defence and viral evasion of host defence.

To infect a host, the virus must first enter the body surface.
There are sites such as:

-Skins ( cuts, abrasion..etc)
Entry through this organ occurs when its integrity is breached by
breaks or punctures. Replication is usually limited to the site of entry because the epidermis is devoid of blood or lymphatic vessels that could provide pathways for further spread. Other viruses can gain entry to the vascularized dermis through the bites of arthropod vectors such as mosquitoes, mites, ticks, and sandflies. Even deeper inoculation, into the tissue and muscle below the dermis, can occur by hypodermic needle punctures, body piercing or tattooing, animal bites, or sexual contact when body fluids are mingled through skin abrasions or ulcerations.

-Conjuntiva ( eyelids )
The epithelium covering the exposed part of the sclera and the conjunctivae is the route of entry for several viruses. Every few seconds the eyelid passes over the sclera, bathing it in secretions that wash away foreign particles. There is usually little opportunity for viral infection of the eye, unless it is injured by abrasion. Direct inoculation into the eye may occur during ophthalmologic procedures or from environmental contamination (e.g., improperly sanitized swimming pools).

-Urogenital tract
Some viruses enter the urogenital tract as a result of sexual activities. The urogenital tract is well protected by physical barriers, including mucus and low pH (in the case of the vagina). Normal sexual activity can result in minute tears or abrasions in the vaginal epithelium or the urethra, allowing viruses to enter. Some viruses infect the epithelium and produce local lesions (e.g., certain human papillomaviruses, which cause genital warts). Other viruses gain access to cells in the underlying tissues and infect cells of the immune system (e.g., human immunodeficiency virus type 1), or sensory and autonomic neurons (in the case of herpes simplex viruses).

-Respiratory tract
The most common route for a virus to enter.
Viruses may enter the respiratory tract in the form of aerosolized droplets expelled by an infected individual by coughing or sneezing, or through contact with saliva from an infected individual. Larger virus-containing droplets are deposited in the nose, while smaller droplets find their way into the airways or the alveoli. To infect the respiratory tract successfully, viruses must not be swept away by mucus,
neutralized by antibody, or destroyed by alveolar macrophages.

-Alimentary tract

*click on pictures for bigger view.





Eating, drinking, and some social
activities routinely place viruses in the alimentary tract. It is designed to mix, digest, and absorb food,providing a good opportunity for viruses to encounter a susceptible cell and to interact with cells of the circulatory, lymphatic, and immune systems.




Virus shedding
-Absolutely necessary for survival; exceptions
-> spread through germ cells
-> consumption of infected tissue

-Effective transmission depends on
-virus concentration
-route of transmission

Modes of transmission

-Respiratory secretions
-> aerosols during speaking, sneezing and coughing
-> Virus can be inactivated by drying
-> It can be further transmitted by contaminated hands

-Saliva
-> Kissing

- Faeces
-> Resistant to drying

-Blood
-> Direct blood/ body fluids exposure

picorviridae and orthomyxoviridae

I have learnt about RNA viruses and am going to discuss 2 of it today. Orthomyxoviridae and Picornaviridae. In greek myxo means mucus and pico means very small. Just by stating this u would get a rough idea of what the viruses are about.

Let us now take a look at Picornaviridae.
















It is known to be the largest virus family and it belongs to the Rhinovirus. It’s virions are naked and it’s nucleocapsid is isometric. It is an RNA virus which means its genome is infectious. It is made up of a single poly protein of 2100-2400 amino acids. Let me now explain how the virus infection takes place. The upper respiratory tract gets infected and the virus would be incubated for 2-3 days. IgA is locally synthesized but the titre declines with time but the IgG serum persists for years. There are over 100 rhinoviruses.


Symptoms of Picornaviridae are watery nasal discharge, congestion, sneezing and little or no fever. There are a few methods used in the lab to discover if the virus is present. The methods are virus culture, nasal washings


























































,EIA
or PCR. There are alot of virus present in the nasal discharge.


The common cold is Picornaviridae.


The virus would be easy to control of we follow these steps. We should always wash our hands, try as much as possible not to touch our eyes or nose, always sneeze into a tissue and throw it away rather than sneezing in the air and letting the germs to circulate in the air. Try to avoid if possible people who have a cold or if you are the one having it, stay home.




Now let me tell what I know about Orthomyxoviridae. Influenza virus A and B comes from this group. The first report of influenza epidermic was from the Athens. In 1918 there was a flu epidermic which led to about 25 – 40 million deaths. In 1970 there was 11 epidermics. This was how serious the virus was.
The virus was typically spherical (100nm diameter).








It is enveloped and pleomorphic which means it has many forms. It has spikes on envelope and has groups of HA (hemagglutinin) or NA (neuraminidase ) in the ratio of HA to NA 5:1.
The Genome of Influenza A and B is a single stranded negative RNA in 8 segments and 3 polymerase peptides with each segment.


The infection takes place in the respiratory tract of humans and there is an affinity of the HA for receptors in the epithelium of the tract. There would be an innate resistance and a mucus blanket. It is more easier for the elderly to get it. The existing antibodies are anti-HA AB, IgA and IgG.



To find out if the virus is present, a throat wash/ gargle/ nasal wash would be done. Virus culture ( cell line or chick embryo), Direct EIA or PCR
















would also be done. Alot of virus is present in the nasal discharge.


Antigenic drift is a mutation in the genetic code of surface antigens (HA/NA) type A and B, results in new strain and happens all the time. Antigenic shift occurs when genes re-assort from different subtypes ( only for type A ), results in a new subtype and happens occasionally. The methods of control for influenza is a vaccine and antiviral drugs.

Virus in plants????

Do viruses infect plants?

YES! It also infects animal, fungi, protista and bacteria! Basically, the 5 kingdoms : plantae, fungi, animalia, protista and monera.

PLANT VIRUSES
Plants play an important factor in our life. For example, for food. The human nutrition depends on cereals, RICE, wheat and maize. Other food such as vegetable, fruits and herbs. Animals also have to feed on plants. For example, grass.

Plants are also used for non food products such as wood to build furniture, buildings and paper.

Cotton is often extracted from plants to create cloth for daily wear.

Another important factor is medicine is required to be extracted from plants. For example, aspirin and taxol. Aspirin is used to relieve pain and aches and to reduce fever and taxol used to treat people with cancer.

Plants are also a primary source of basic chemicals for the industrial synthesis of a vast array of organic chemicals. These chemicals are used to for studies and experiments.

However, there is a negative effect for the uses of plants. Take for example, plants which produce windblown pollen which cause allergic reactions to people who suffer hay fever. There is also the poison ivy which is poisonous and causes skin irritations. Some plants also contain psychotropic chemicals ( a chemical substance which will act on the central nervous system where it will alter the brain function, resulting to temporary changes in behaviour, mood or perception ) which can be smoked, including tobacco or cocaine. Too much consumed can caused damages to the body.

















TRANSMISSION
Plant viruses can be transmitted by a vector, most often by insects. The chosen insect vector of a plant virus will often be the determining factor in that virus’s host range: it can only infect plants that the insect vector feeds upon.
















Nematodes are able to transmit viruses too. They acquire and transmit viruses by feeding on infected roots.













Viruses can also be transmitted through seeds, the seed will be infected in the generative cells and the virus will be maintained in the germ cells and sometimes in the seed coat. There is still little known about the mechanism involving in the transmission of plant viruses via seeds. The seed transmission occurs because of a direct invasion of the embryo via the ovule or by an indirect route with an attack on the embryo mediated by infected gametes.


BACTERIAL VIRUSES
bacterial viruses are bacterial infected with viruses also known as bacteriophage. It consists of an outer protein hull enclosing the genetic material. This genetic material can be dsRNA, ssRNA, ssDNA or dsDNA . The nucleic acid usually contains modified or unusual bases. These modified bases will protect the bacteriophage nucleic acid from nucleases that will break down the host nucleic acids during phage infection.

The T4 is one of the largest phages and is about the size of 200nm long and 80-100nm wide. Most phages are 20 to 200nm in length. The capsid is composed of proteins and genetic material can found in the capsid. It also acts as a protective covering for the nucleic material. A phage also consist of a tail.
























Bacteriophages may have lytic cycle ( considered the main method for viral replication, as it results to the destruction of a infected cell ) or lysogenic cycle.
Better understanding....
http://www.ncbi.nlm.nih.gov/books/bv.fcgi?rid=mcb.figgrp.1421





MYCOVIRUSES
mycoviruses are viruses that infect fungi. Right now, only a few fungal viruses are studied and it must be done through electron microscopy. An electron microscopy is used because it has greater resolving power and can obtain higher magnification.
The most identified have dsRNA and spherical in shape. One example of mycovirus being the causal agent of La France Disease which affects the mushroom. Some observations of the infected mushroom are reduced yield, rapid deterioration in tissue and change in morphology.

Retrovirus and Flaviviridae

For Microbiology B which is my year 1 semester 2 module, two topics i have learnt about is called Retrovirus and Flaviviridae.
Most of what I have learnt about Retrovirus is mainly:
2 types of Retroviruses:
• Deltaretrovirus
• Lentivirus
The properties of Retroviruses
• Spherical enveloped virion (A complete virus particle)
• 80 – 100nm
• Ribonucleoprotein in central nucleoid (concentric or truncated cone in lentivirus) within icosahedral capsid
• Envelope with glycoprotein peplomers (a glycoprotein structural unit found in the lipoprotein envelope of enveloped viruses)
• Reversed transcriptase
- Genome doesn’t serve as mRNA
• 2 copies of linear positive sense single stranded RNA
- 7-10kb long
• 3’ polyadenylated tail (protects the mRNA from digestion with nuclease and greatly increases the efficiency of translation)
• 5’ cap (specially altered nucleotide end to the 5' end of precursor messenger RNA)
- The process of 5' capping is vital to creating mature messenger RNA which is then able to undergo translation. Capping ensures that the messenger RNA is stable while it undergoes translation in the process of protein synthesis. It is a highly regulated process, which occurs in the nucleus
• Formation of long terminal repeats before provirus DNA inserted into host genome
• gag, pol, env genes some regulatory genes, some oncogenes
- gag
• Group-specific antigen
• Virion core (capsid protein)
- pol
• Polymerase (reverse transcriptase)
• RNase H (part of the viral reverse transcriptase enzyme, it is absolutely necessary for the proliferation)
- env
• envelope
Types of Retroviruses
• Tumour-forming
- Trans-activating (HTLV)
• Non-tumour-forming
- HIV

HTLV-1
The image below shows the infection and life cycle of what a HTLV-1 virus goes through.

CLICK PICTURE FOR LARGER VIEW! :D
http://www.ambion.com/tools/pathway/loadImage.php?pos=br&im=images/Cellular%20Transformation%20by%20HTLV1.jpg

• Adult T-cell leukemia/lymphoma (ATLL)
- Acute aggressive leukemia resulting in death in 12 months
- No known cure yet
• Tropical spastic paraparesis / HTLV-1 associated myelopathy
- Wasting of neurons (lost of myelin sheaf)
- Back pain followed by paralysis

HIV
The image below show the infection and life cycle of what a HIV virus goes through.

CLICK PICTURE FOR LARGER VIEW! :D
http://www.nwabr.org/education/pdfs/hiv_lifecycle.jpg

This is a video illustrating the infection and a life cycle of the HIV virus.



This picture illustrates the pathogenesis of HIV.

CLICK PICTURE FOR LARGER VIEW! :D

Primary Infection
Acute Stage
• Flu-like Symptoms
• Fever
• Skin Rash
• Swollen Lymph nodes
- Virulence factors (degree of pathogenicity of an organism, the relative ability of a pathogen to cause disease)
• Rate of replication
• Propensity to mutate
• Cytopathogenicity (the production of pathological changes in cells)
- Host resistance
• Suppression by CD8 T suppressor cells
• Presence of cytotoxic T-lymphocytes
Asymptomatic Stage
• Fatigue
• Depression
• Weight loss
• Memory disorder
- No apparent disease
- Fall in CD4 T-lymphocytes (primary target cells)
Symptomatic Stage
• AIDS-related complex
- Diseases not considered definitive of AIDS
- May be attributed to HIV infection
- Indicative of defect in cell-mediate immunity
• AIDS
- Opportunistic infections as a result of fall in CD4 T-lymphocytes
AIDS Therapy
• Non-specific therapeutic management
- To boost general health
- Vitamin
- Minerals
- Anti-oxidants
• Specific therapeutic management: antiretroviral therapy
- Nucleoside Reverse Transcriptase Inhibitors
• AZT (azidothymidine)
• 3TC (lamivudine)
- Non-nucleoside Reverse Transcriptase Inhibitors
• Efavirenz
• Nevirapine
- Protease inhibitors
• Indinavir
• Ritonavir
- Rapid mutations due to inefficiency of reverse transcriptase
- Combination therapy to combat resistance

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• Immunomodulation
- Enhancement of immune system through treatment with:
- Interleukin-2 (still under study)
• Vaccines
- Many candidates under development and trials
- None so far proven useful
That's my knowledge of Retroviruses.

Here's a picture of AIDS Awareness just for the fun of it:

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Flaviviridae
Most of what I have learnt about Flaviviridae is mainly:
Two types of viruses:
• Flavivirus
- Yellow Fever
- Dengue
- West Nile
• Hepacivirus
- Hepatitis C
The properties of Flaviviridae
• Spherical enveloped virion (A complete virus particle)
• 40 – 50nm
• Inner core protein C
• Membrane / matrix protein M
• Envelope with glycoprotein peplomers (E)
• Single linear 11kb positive sense single stranded RNA
- Infectious mRNA
• 3’ polyadenylated tail (protects the mRNA from digestion with nuclease and greatly increases the efficiency of translation)
• 5’ cap (specially altered nucleotide end to the 5' end of precursor messenger RNA)
- The process of 5' capping is vital to creating mature messenger RNA which is then able to undergo translation. Capping ensures that the messenger RNA is stable while it undergoes translation in the process of protein synthesis. It is a highly regulated process, which occurs in the nucleus
• Cytoplasmic replication (perinuclear)
• Polyprotein from genomic RNA cleaved
• 3 structural proteins
• Several non-structural proteins
The image below shows how the strain of flaviviridae virus is passed.

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http://www.microbeworld.org/images/scientists/interviews/arbocycle.jpg

Dengue
The image below shows the infection and life cycle of what a Dengue virus goes through.

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http://www.klinikum.uni-heidelberg.de/typo3temp/pics/e102b53902.gif

Dengue
• Most important type of arbovirus presently
• Range which infection is prominent in is, Southeast Asia, America, Pacific regions and Africa.
• Two types of signs in which dengue causes:
- Non-fatal dengue fever (DF)
- Fatal dengue haemorrhagic fever(DHF) / Dengue shock syndrome (DSS)
• 4 distinct serotypes based on neutralization test:
- DEN-1
- DEN-2 (shows greatest antigenic and genotypic distance from the others)
- DEN-3
- DEN-4
• Protective immunity after infection homotypic
Symptoms of dengue fever
• Many infections are asymptomatic
• Acute infection:
- Fever
- Severe headache (frontal)
- Retro-orbital pain
- Nausea
- Vomiting
• Severe muscle and bone pain
• Severe arthralgia (joint swelling, mainly the back) which is also known as the break bone fever
• Maculopapular rash just before recovery
Picture of maculopapular rash:

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http://www.pedrheumonlinejournal.org/nov-dec/Images/figure2.jpg

Dengue Haemorrhagic Fever (DHF) / Dengue Shock Syndrome (DSS)
Key factors of DHF and DSS:
• Prior infection
• Age (seldom occurs in individuals above the age of 15)
Similar to Yellow Fever in biphasic nature (having two phases):
• Initial symptoms similar to DF but then it is followed by remission (a complete or partial disappearance of the signs and symptoms)
• Sudden deterioration of patient condition
Symptoms of DHF / DSS
• Severe prostration
• Hypotension
• Circulatory collapse
• Bleeding
- Petechiae in skin, mucous membranes (mouth) [a small (1-2mm) red or purple spot on the body, caused by a minor haemorrhage]
- Injection and punction sites
- Gastrointestinal bleeding
- Haemorrhagic pneumonia
• Shock

World Health Organisation (WHO) Grading of DHF:
• GRADE 1 (Fever with non-specific, constitutional symptoms and haemorrhagic manifestations being a positive tournique test)
• GRADE 2 (Fever with non-specific, constitutional symptoms and specific haemorrhagic manifestations)
• GRADE 3 (Signs of circulatory failure and hypotension)
• GRADE 4 (Profound shick with pulse and blood pressure undetectable)

This picture illustrates the possible pathogenesis of DHF/DSS.

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http://www.umassmed.edu/cidvr/graphics/rothman.gif

Pathogenesis of DHF / DSS
Not well understood despite intensive studies
2 types of theories:
• Virulent strain theory
- Some strains are more virulent than others
- Molecular studies shows variations in sequences amongst different strains within serotypes
- Early evidence pointed to DEN-2
• Antibody enhancement
- Main theory for DHF / DSS
- Main cell target of dengue: Monocytes or macrophages
- Most cases of DHF / DSS had prior infection of in infants below 1 year had material Ab
- Experiments on monkeys showed similar enhancement.
• Possible causes of DHF / DSS
- Immune system overreacting
- Severe Acute Respiratory Syndrome
Control of dengue:
1. Insecticide
2. Mosquito screen
3. Remove stagnant water

Yellow Fever
The image below shows the infection and life cycle of what a Yellow Fever virus goes through.

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http://jvi.asm.org/content/vol81/issue17/images/large/zjv0170795060001.jpeg

Yellow Fever
Tropical disease in Latin America and Africa
Incubation period: 3–6 days
Symptoms of Yellow Fever:
• Viraemia
• Infectious
• Headache
• Malaise
• Nausea
• Lassitude
• Muscle ache (3days)
• Flushing of head and neck
• Conjunctival injection
• Strawberry tongue
Symptoms of Severe Yellow Fever:
• Remission after acute yellow fever
• Haemorrhagic, hepatic and renal disease
• Fever
• Vomiting
• Abdominal pain
• Dehydration
• Prostration
• Haemorrhagic/coffee-ground diathesis(black vomit)
• Bleeding from puncture site of injections and drip needles
• Jaundice
• Massive haematemesis/haemoptysis/intra-abdominal bleeding
• Renal failure
• Hypotension
• Shock
Virus absent from blood, but antibody titre is high – implying autoimmunity may play a major role
Mortality 20–50%
Survivors suffers from extended chronic jaundice before full recovery, hepatic and renal failure may persist

Control of dengue:
1. Insecticide
2. Mosquito screen
3. Remove stagnant water
4. Attenuated vaccine

West Nile Fever
The picture below shows the west nile virus transmission cycle

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http://dittamore.us/images/west_20nile_20virus_20life_20cycle.gif

West Nile Fever
Common disease in Africa, West Asia, Europe and Middle East
Epidemic in US in 2002
Symptoms of West Nile Fever
• Mainly mild to no symptoms
• Fever
• Headache, body aches
• Skin rash
• Swollen lymph glands
Severe symptoms of West Nile Fever (mainly in people aged 50 years and above)
• Crossing blood-brain barrier
• Encephalitis
• Meningitis
Control of dengue:
1. Insecticide
2. Mosquito screen
3. Remove stagnant water

HERPESSSSSSSSSS

Herpes actually means to creep upon. Below shows how herpes virus actually ‘creep upon’ your body.



The features of the Herpes virus are very special. When herpes virus infects you, the virus will remain present until there is a reactivation. Reactivation such as stress, or during menstrual period. The virus will hide in the nerves tissue which will cause encephalitis ( acute inflammation of the brain ) and if passed from mother to baby, it can cause brain damage in the child.



The herpes virus genome consists of 3 main points. The concentric virion, linear dsDNA and three origin of replication (ORI).Under concentric virion, we have the inner core, iscosahedral capsid, amorphous tegument and envelope.



















http://sg.images.search.yahoo.com/search/images?ei=UTF-8&p=herpes+virus+genome&rd=r1&fr2=tab-web&fr=yfp-t-web




Glycoprotein serve to identify and bind to receptor sites on the host’s membrane and the envelope is to help virus enter host cells. This can happen by that when the glycoprotein binds to the receptor sites, the envelope will fuse with the host’s membrane to allow the capsid and genome to enter and infect the host.



Tegument is a cluster of non-essential and essential proteins between the envelope and capsid. It aids in viral replication and usually released after infection into cytoplasm.



A capsid is the protein shell of a virus and encloses the genetic material of the virus.





There are many types of viruses under the family of herpesviridae.

However, only a few viruses are more focused on such as Herpes simplex 1 & 2 virus, Varicella zoster virus, cytomegalovirus and esptein barr virus (EPV).



Under Herpes simplex viruses, there is HSV1 and HSV2. HSV1, which is oral herpes, involves the symptoms such as cold sores which will infect both the face and mouth. HSV2, genital herpes involves blisters, burning sensation and discharge.



The common pox which will occur mostly when we were young, the chickenpox and shingles are under Varicella Zoster virus. Fever, lesion, scarring are the few clinical features of varicella zoster. Varicella zoster is also very dangerous in pregnant women as it can cause the child and the mother to be in danger of getting infected by the Guillan Barre syndrome.The Guillan Barre syndrome is a disease which affects both the brain and the spinal cord, thus exhibiting as paralysis.



Cytomegalovirus infection is associated with salivary glands, urine, and semen and breast milk. It is also known as Human Herpesvirus 5. It is life threatening for patients who are immunocompromised. Some symptoms of this virus are fever, sorethroat and a mild hepatitis.



EBV infection is often asymptomatic and causes glandular fever. Symptoms of the glandular fever are fever, sorethroat and swollen lymph glands. The virus can be found in the nasopharynx and salivary gland.



To avoid Herpes simplex virus and EBV, practically don’t kiss or any risky behaviour. For Varicella Zoster Virus, avoid infected people and for cytomegalovirus, avoid any sexual behaviour or intimate contact.





The lab diagnosis of these viruses are to do a virus culture, a antigen test- EIA( Enzyme Immuno Assay ) and Blood test. The EIA is to detect antibody or antigen in a sample.

*click on pictures for bigger view