Conclusion

This post marks the end of our submission for our blog on virology. It has been fun learning so much about viruses in just one semester. I hope to have more virus topics for the coming semesters.

What have we covered throughout the semester on the blog:
1. Our first insight into Virology --> The two classification of viruses.
2. DNA viruses --> Hepadnaviridae and Herpesviridae
3. RNA viruses --> Orthomyxoviridae and Picornaviridae
4. Flavivirade --> A deadly viruses with major diseases: Dengue fever, Japanese encephalitis
5. Retroviridae --> The HIV virus -- Resulting in AIDS, and also Human T-lymphotropic virus (HTLV-1)
6. Poxviridae --> The story of small pox eradication, and the creation of vaccination
7. Methods of study of viruses --> Tissue culture, serological methods & molecular virology study methods.
8. Virus Host Interaction --> The stages of virus interaction with host: Attachment, penetration, uncoating, replication, assembly, and release.
9. Emerging viruses --> Virus that are becoming more prevalent
10. Prions, Viriods and virusoids --> Unqiue infectious agents that are similar in nature to virus

The End..... That's the conclusion of our virology knowledge acquired for this Semester (2008/2009).

Prions, Viroids and Virusoids

In today's lecture, an interesting virus-like deadly protein was introudced to us. It was amazing! I was astounded by the fact that proteins can be like a virus, and causes such deadly infection.

What are prions?

Two independent research groups have established conclusively that prions are proteins, and that they do not depend on genes or other factors for transmission of their traits. According to the scientists, the studies answer a nagging question that had raised doubts among some researchers about the validity of the so-called "protein-only" hypothesis of prion infectivity. They are rogue protein that transform other cellular protein (PrPC) to the prion form PrP^SC.More and more PrP^SC gets transformed until they completely clogged brain cells. So Cells misfire, work poorly or don’t work at all and when Cells die, prions are release into the blood stream to re-infect other cells.

Prion Diseases

Prion diseases belong to group of progressive conditions that affect the nervous system in humans and animals. In people, prion diseases impair brain function, causing memory changes, personality changes, a decline in intellectual function (dementia), and problems with movement that worsen over time. The signs and symptoms of these conditions typically begin in adulthood, and these disorders lead to death within a few months to several years.

History of prions

Now let’s look at the history of prions and understand more about it. Conventional View of Infectious Disease: Caused by either viruses, bacteria, fungi or other parasites. However in 1986 - Mad Cow disease in England challenged the convention. Scrapie is a fatal, degenerative disease affecting the central nervous system of sheep and goats .This is a very old disease, documented over 200 years ago. The disease classified as transmissible spongiform encephalopathy (TSE).

Pathogenesis

Spongiform encephalitis and it looks like “Sick” brains, riddled with holes. It also looks like Swiss cheese.

What genes are related to prion disease?

Mutations in the PRNP gene cause prion disease.

Only a small percentage of prion disease cases run in families. Most cases are sporadic, which means they occur in people without any known risk factors or gene mutations. Rarely, prion diseases can be transmitted by accidental exposure to prion-contaminated tissues during a medical procedure. This type of prion disease is called iatrogenic.

One type of prion disease in humans, variant Creutzfeldt-Jakob disease (vCJD), is acquired by eating beef products obtained from cattle with prion disease. In cows, this form of the disease is known as bovine spongiform encephalopathy (BSE) or, more commonly, "mad cow" disease. Another example of an acquired human prion disease is kuru, which was identified in the South Fore tribe in Papua New Guinea. The disorder was transmitted when tribe members ate the tissue of affected people during cannibalistic funeral rituals.

Familial forms of prion disease are caused by inherited mutations in the PRNP gene. This gene provides instructions for making a protein called prion protein (PrP). Normally, this protein is likely involved in transporting copper into cells. It may also play a role in protecting brain cells and helping them communicate. In familial cases of prion disease, mutations in the PRNP gene cause cells to produce an abnormal form of the prion protein known as PrP^Sc. In iatrogenic and acquired cases, an affected person develops prion disease from exposure to this abnormal protein.

How is the transmission of prions caused?

It is caused by eating contaminated beef or contaminated mutton and prion may be acquired through blood transfusion

Virusoid

Virusoid can be found in plants, an infectious agent consisting of a single circular strand of RNA, containing some 220 to 350 of the basic structural units known as nucleotides. A virusoid is like a viroid and unlike a virus, in that its genome does not code for any protein. Unlike viroids and viruses, virusoids cannot self-replicate inside a cell. Instead, the cell must also be infected with a certain “helper” virus, which serves as a host for the virusoid and provides RNA that the latter needs in order to reproduce; the virusoid is thus regarded as a type of so-called satellite RNA. Among plant diseases associated with virusoids are lucerne transient streak, subterranean clover mottle, and velvet tobacco mottle.

The sizes of virusoids are similar to viroids in size, structure and means of replication (rolling-circle replication)

Emerging Viruses

This lecture was taught by Mr William How, our lecture. What are emerging viruses? They are known viruses that has newly appeared in a human population and is rapidly increasing in disease incidence. Any examples come to mind? Examples are SARS in Nov 2002, and something more current is Influenza virus and dengue virus. So why are emerging viruses appearing? It is because of Virus and Human factors.

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 natural habitat of animal

Virus-Host Interaction

Today we finally completed Virus-Host interaction. This topic is a killer!!! It took so many hours to sum it up. We learnt that there are 6 phase of interaction. In short, they are APURAR, Attachment, Penetration, Uncoating, Replication, Assembly and Release. Below is the summarized version from an A student who went through the topic 10 times flat!!!

Today we finally completed Virus-Host interaction. This topic is a killer!!! It took so many hours to sum it up. We learnt that there are 6 phase of interaction. In short, they are APURAR, Attachment, Penetration, Uncoating, Replication, Assembly and Release. Below is the summarized version from an A student who went through the topic 10 times flat!!!

Methods of study of Viruses

Week 7 Lecture was taught by Mr Jay, our lecturer. The topic was methods of study of Viruses. It’s quite boring. We learned about the different way of isolating and cultivating virus. Methods include using animal and eggs as host usually by injection. This method is the first method for virus cultivation but in very inconvenient.

Obviously, plants were used to study viruses that affect plants. Now, tissue culture can be use. Cell tissue culture are grown in vitro. The are 2 types of cell culture. The primary cell line and continuous cell line. A primary cell line in made by taking cells from a animal part and growing it in vitro.Continuous cell lines are usually derived from primary cell lines. They are transformed/cancerous cell which can be sub-cultured indefinitely. After learning how to cultivate and isolate viruses, we learned about how to detect, identify and diagnose them. To detect and identify viruses using cell culture methods, we learned about cytopathic effects (CPE) and plaque assay.

Physical methods also can be used. They are:

• X-ray crystallography
• Electron microscopy
  • Transmission EM
  • Scanning EM
  • STEM
• Ultracentrifugation
  • Purification

Serological and Immunological methods such as:

• Haemagglutionation (HA)
• Haemagglutionation Inhibition (HI)
• Virus neutralisation
• Complement fixation
• Immunostaining
  • Immunofluorescence
  • Immuno-gold EM
• Immunoprecipitation/ Immunoblot
• Enzyme-linked immunosorbant assay (ELISA)

Molecular Virology

After learning how to detect and identify them. He talked about how to analyse viral protein by methods of:

• PAGE / SDS PAGE
• Western blot
• Protein Sequencing
• X-ray crystallography

We also learnt how to analyse viral genomes using the methods he mentioned:

• Agarose gels
• Restriction analysis
• Sequencing
• Southern blot
• Northern blot
• PCR / RT-PCR

That was all for the 2 lectures. All about methods to study viruses.

Poxviridae

In today’s lecture, I learnt about this very deadly virus in the past that killed lots of people, which is extinct already. It is called, “small pox”. It comes from a family called, poxviridate, which is the largest family of viruses, and can be seen under a microscope. But be careful, it is deadly as it can remain stable for hours in air allowing it to spread easily!

http://www.stanford.edu/group/virus/pox/2000/IMAGES/poxtoon.gif

Its genome is double stranded DNA, covalently cross-linked at ends.

Small pox infects human only. It causes respiratory secretions, and is associated with skin lesions.

http://www.nlm.nih.gov/medlineplus/ency/images/ency/fullsize/19059.jpg

It symptoms is initially influenza-like. It will result in neurological damage, blindness, and even death.

Variola major and Variola minor are variants of small pox.

Some statistics of these variants were gathered in the lecture:

Variola Major – 25 to 30% fatalities

Variola Minor – less than 1% death

Now, let me tell you about vaccination that helped in smallpox eradication.

http://www.ebinrushed.com/history/images/history_7.jpg

It was found by Edward Jenner who observed that milkmaids got cowpox virus, also known as vaccinia, from cow, and didn't seem to get very sick when infected with small pox and always recovered. So, he decided to test out his theory, he vaccinated an 8 year old boy, by the name of James Phipps. There after, he was challenged with small pox, and the similar effects were observed - the symptoms of small pox were milder and recovery was quick.

From this virus, the term VACCINATION was coined. The derivative of vaccinia, the cowpox virus.

With this new weapon called vaccination, and people obtaining lifelong immunity after recovering from small pox, and human being its only reservoir to infect, the smallpox virus is finally eradicated.

It was announced to be eradicated by WHO (World Health Organisatio) in 1980! What an exciteful lecture on VIROLOGY!

Retroviridae

http://www.stanford.edu/group/virus/retro/2005gongishmail/hiv1.jpg

Week 5 Lecture on Retroviruses was not bad. What is retrovirus? A retrovirus is a virus with an RNA genome that replicates by using a viral reverse transcriptase enzyme to transcribe its RNA into DNA in the host cell. The lecture was mostly about Human T-lymphotropic virus (HTLV-1) and Human Immunodeficiency virus (HIV). We also learned about the history and characteristics of retroviruses. Did you know that the first human retrovirus was discovered in 1981? It was the Human T-lymphotropic virus. Retroviruses have received much attention even before 1980. It had a long history that dates back
to 1908. That is almost 100 years ago.

• 1908: Ellerman and Bang, searching for an infectious cause for leukaemia, studied leukaemia in chickens and succeed in transferring the disease from one to another by cell-free tissue filtrates
• 1911: Peyton Rous discovered chicken sarcoma caused by a virus
• 1960’s: Howard Temin knew that retrovirus genomes were composed of RNA and observed that replication was inhibited by actinomycin D which inhibits DNA synthesis therefore he proposed the concept of reverse transcription.
• 1970:Michael Bishop and Harold Varmus discovered oncogenes and role of oncogenic viruses and cancer
• 1980: Human T-cell leukaemia virus discovered. The first pathogenic human retrovirus.
• 1983: Human immunodeficiency virus discovered.

Retroviridae are broken down further into the genus as show in the table below:

Genus	Examples
Alpharetrovirus	Avian leukosis virus
Betaretrovirus	Mouse mammary tumor virus
Gammaretrovirus	Murine leukernia virus
Deltaretrovirus	Human T-lymphotropic virus
Epsilonretrovirus	Walley dermal sarcoma virus
Lentivirus	Human immunodeficiency virus
Spumavirus	Human foamy virus

The life cycle of retroviruses:

http://www.gene.com/AE/AB/GG/examples_of_viruses.html

Main steps are Attachment, Entry, Uncoating, Transcription, Translation, Assembly and Release.

HTLV-1

What is HTLV-1? It’s the virus that causes Adult T-cell leukaemia (ATLL) which is a rare cancer to the T-cells. Acute aggressive leukaemia can kill in 12 months. Unfortunately, there is no known cure yet.

HIV

HIV is the virus that cause acquired immunodeficiency syndrome (AIDS), a condition in humans in which the immune system begins to fail, leading to life-threatening opportunistic infections.The picture below show the life cycle and structure of HIV:

http://commons.wikimedia.org/wiki/File:Hiv_gross.png

HIV can be transmitted through sexual contact, blood or from mother to child. Symptom of a HIV infection are:

During Primary infection:

• acute stage
• flu-like symptoms
• fever
• skin rash
• swollen lymph nodes

During Asymptomatic stage:

• fatigue
• depression
• weight loss
• memory disorders

Another deadly virus, yet no cure is available. However a lot of research is in progress. Possible methods is vaccines or to inhibit any of the stages of the HIV life cycle.

Flaviridae

During week five, the topic on Flaviviridae was taught during lecture time. Let me give you more details about this particular virus.

Baltimore Classification

Viruses; ssRNA positive-strand viruses, no DNA stage; Flaviviridae

Description and Significance

• Flaviviridae contains a myriad of viruses that cause disease in humans. Flaviviridae has a total of 69 pathogens in its rank. The family gets its name from Yellow Fever virus, a type virus of Flaviviridae; flavus means yellow in Latin.
• The Flavivirus genus contains several nasty critters that include yellow fever virus, dengue fever virus, and Japanese encephalitis (JE) virus.

Morphology

• Virions uniform in shape; spheroidal; enveloped; 40-60 nm in diameter. Surface projections of envelope small (surface appears rough), or distinct (and are obvious fringes in negative stains). Nucleocapsids isometric; 25-30 nm in diameter. Symmetry polyhedral.

Major diseases caused by the Flaviviridae family include:

• Dengue fever
• Japanese encephalitis
• Kyasanur Forest disease

Dengue Fever

http://www.stanford.edu/group/virus/flavi/2000/mosquito_mov.gif

• Dengue Fever and Dengue Haemorrhagic Fever (a more severe form) are the most common mosquito-borne viral diseases in the world.
• The diagnosis of dengue is usually made clinically. The classic picture is high fever with no localising source of infection, a petechial rash with thrombocytopenia and relative leukopenia - low platelet and white blood cell count.
• Signs and symptoms include a sudden onset of severe headache, muscle and joint pains, fever, and rash.
• The classic dengue fever lasts about six to seven days, with a smaller peak of fever at the trailing end of the disease (the so-called biphasic pattern). Dengue Vaccine Initiative which was set up in 2003 with the aim of accelerating the development and introduction of dengue vaccine(s) that are affordable and accessible to poor children in endemic countries.

Japanese encephalitis

• Japanese encephalitis is a disease caused by the mosquito-borne Japanese encephalitis virus. The Japanese encephalitis virus is a virus from the family Flaviviridae.
• Domestic pigs and wild birds are reservoirs of the virus; transmission to humans may cause severe symptoms. One of the most important vectors of this disease is the mosquito Culex tritaeniorhynchus. This disease is most prevalent in Southeast Asia and the Far East.
• Japanese encephalitis has an incubation period of 5 to 15 days and the vast majority of infections are asymptomatic: only 1 in 250 infections develop into encephalitis. There is no specific treatment for Japanese encephalitis and treatment is supportive.

The use of arctigenin has been shown to be effective in a mouse model of Japanese encephalitis, but there is as yet no clinical evidence to support its use.

RNA viruses -Orthomyxoviridae & Picornaviridae

All living things have DNA as genetic material. Only viruses have RNA as its genetic material! RNA virus’s genome is highly infectious(+RNA viruses) as it can be read as mRNA directly.

All RNA virus’s have unusual structural characteristics (leader sequence) and can be differentiated between viral mRNA and host mRNA.

Two types of RNA viruses

Orthomyxoviridae and Picornaviridae

Picornaviridae also known as common cold

http://education.expasy.org/images/Picornaviridae_virion.jpg

Morphology(shape)

Virions consist of a capsid. Virus capsid is not enveloped, round with icosahedral symmetry. The capsid is isometricand and capsids appear round. The capsid consists of 12 capsomers.

Incomplete particles are common.

Genome

The genomic RNA of picornaviridae is infectious. It is long untranslated region end of 5’ end “clover leaf” structure (IRES). It is one linear (+) RNA.

Single polyprotein of 2100 –2400 aa

Both of the ends are modified.

Pathogenesis

Upper Respiratory Tract infection

Short Incubation ( 2 –3 days)

Endogenous IFN helps!

Locally synthesized IgA but titer decline with time

So what are the symptoms?

• Water nasal discharge
• Sneezing
• Little/No fever
• Congestion

When do we get Picornaviridae?

All throughout the year, virus abundant in nasal discharge.

How can we control it?

• Wash hands
• Keep hands away from eyes and nose
• Sneeze into tissue & throw away
• Maintain personal hygiene
• Stay away from people who have colds.

Is vaccination possible for Picornaviridae?

Yes. It is possible. However vaccination is not so useful because we get common cold throughout the whole year and so it is not much use. Another reason is that vaccination is very expensive. So vaccination is not that useful.

Orthomyxoviridae also known as Influenza

http://www.pasteur.ac.ir/researchDepartment/flu/images/flu_structure.gif

Morphology(shape)

Typically Spherical (100nm diameter)

• Enveloped ,Pleomorphic (many forms)
• Spikes on envelope
• Groups of HA (hemagglutinin) or NA (neuraminidase)
• Ratio of HA to NA
  • 5:1

Genome

ss (-) RNA in 8 segments

• 3 polymerase polypeptides with each segment
• 5’ and 3’ end of all segments highly conserved

Pathogenesis

• Respiratory Tract in human
• Affinity of the HA for receptors in the epithelium of the tract
• Innate resistance
• Mucus blanket/cilia
• mmunocompromised

Lab Diagnosis

• Throat swab/gargle/nasal wash/
• Virus culture (MDCK cell line or chick embryo)

Epidemiology

• All throughout the year!
• Incubation (1 –4 days)
• A few serotypes circulating simultaneously

Differences between Flu and Cold

Symptoms	Cold	Flu
Fever	Rare	Characteristic, high (100-102 degrees F); lasts 3-4 days
Headache	Rare	Prominent
General Aches, Pains	Slight	Usual; often severe
Fatigue, Weakness	Quite mild	Can last up to 2-3 weeks
Extreme Exhaustion	Never	Early and prominent
Stuffy Nose	Common	Sometimes
Sneezing	Usual	Sometimes
Sore Throat	Common	Sometimes
Chest Discomfort, Cough	Mild to moderate; hacking cough	Common; can become severe

Complications	Sinus congestion or earache	Bronchitis, pneumonia; can be life-threatening
Prevention	Good hygiene	Annual flu shot or FluMist
Treatment	Only temporary relief of symptoms	Antiviral drugs (oseltamivir or zanamavir) within 24-48 hours of onset

DNA viruses -- Hepadnaviridae & Herpesviridae

$
The lecturer went thorough hepadnaviridae and herpesviridae. Learnt something new on tuesday! Hepatitis B belongs to hepadna family and it has the smallest genome (super small!!!) and has a partial double-stranded DNA. Also, hepatitis patient can either be acutely or chronically infected but only acute infections show symptoms. Yet both kind of infected carrier is characterized by the presence of HBsAG, hepatitis B surface antigen, HBeAG, hepatitis B core antigen and antibodies to the two antigens. Detection of Hepatitis core antigen has been detected by immunofluorescence in both cytoplasm and nucleus in cells of acute and chronic patients. Here’s something shocking, it can be transmitted via INSECTS!!!

Hepadnaviridae

http://www.hon.ch/Library/Theme/HepB/hbvirus.GIF

Unique features of hepadna genome

Hepadna virus family has the smallest genome of all replication competent of animal DNA viruses. It is a partial double strand DNA.

All viruses in the hepadna family have common behavior such as:

– They are enveloped

– They contain polymerases that can repair the viral DNA genome during replication

– They produce lipoproteins containing envelope proteins

– They infects species that are closely related to that of their natural host

– They produce chronic infections in the liver of the host

HepatitisB

- smallest genome

- A partial double-stranded DNA: A longer negative strand with a piece missing at a nicked site

An incomplete positive strand

Pathogenesis

ACUTE and CHRONIC liver infection depending on age group

90% of neonates and 50% of young children become chronic infected

5% to 10% of Immuno-competent adults infected with HBV develop chronic hepatitis B

ACUTE

From subclinical to fulminant(full-blown)

More than 90% of acutely infected adults recover

5%-10% of acutely infected adults become chronically infected

Symptoms: loss of appetite, fever, vomiting, nausea and jaundice

CHRONIC

No symptoms and no abnormalities on laboratory testing

Some have clinically apparent chronic hepatitis and some develop cirrhosis (a degeneration of liver with tissue fibrosis and scarring)

Lab diagnosis

Carrier with HBV infection is characterized by the presence of HBsAG, hepatitis B surface antigen, HBeAG, hepatitis B core antigen and antibodies to the two antigens.

Detection of Hepatitis core antigen can be detected by immunofluorescence in both cytoplasm and nucleus in cells of acute and chronic patients.

Ways of transmission

-Breast feeding

-Unprotected sex

-Contact with contaminated blood

-Insect transmission

Control

-Post 1987-baby vaccinated against HBV

-Pre-1987- screening prenatal infants and mothers

-Avoid unprotected sex

-Avoid sharing needles

Reference-http://virus.stanford.edu/hepadna/index.html

HERPESVIRIDAE

http://stdgen.northwestern.edu/stdgen/bacteria/hhv2/herpes.diagram.jpg

Unique feature

It is a primary infection-virus, which will remain latent until reactivation. It hides in nerve tissues and re-emerges as secondary infection when host undergoes stress.

Herpes is a greek word, which means to creep upon.

GENOME

It has a linear double stranded DNA with 3 origin of replication, and is a concentric virion.

PATHOGENESIS

Herpes simplex virus:

HSV1(cold sore)-oral cavity

HSV2 (genital herpes)-genital

Clinical features

HSV1 OR HSV2?

Cold sores -1 week

Genital herpes- 1 to 3 weeks

Epidemiology of herpes simplex viruses

-HSV1: almost 100% of adult population

-HSV2: up to 20% of the population

Control of herpes simplex virus

-Avoid over kissing

-Avoid risky sex

Varicella zoster virus:

Varicella (chicken pox) - Respiratory tract, pharynx primary infection

Herpes (zoster shingles)-secondary infection

CLINICAL FEATURES- VARICELLA ZOSTER

-fever

-Lesion- scratching leads to secondary infection

-Scarring

-Dangerous in pregnant women

-May affect nervous system (Guillain Barre Syndrome)

Lab diagnosis

- virus culture

- antigen test-EIA

- blood test

Control of varicella zoster

Availability of vaccines

Avoid infected people

Cytomegalovirus-virus found in saliva, urine, semen

EBV-nasopharynx, salivary gland

Epidemiology of EBV

Almost 100% of adult population

Control of EBV

Avoid over kissing

Avoid risky sex

NO vaccine available

Our first Insight into VIROLOGY!

It began with our lecturer teaching us the basic of virology, how viruses are classified. There’s two classification used, Lwoff’s and Baltimore’s.

Let me tell you more about these classifications:

http://www.nlv.ch/images/Lwoff.jpg

Lwoff’s Scheme for classification is based on shared properties of viruses, nucleic acid of the virus, symmetry of capsid, presence or absence of envelope, and dimensions of virion and capsid.

In the picture below, it shows the Lwoff Scheme for classification of virus:

http://www.nlv.ch/Virologytutorials/graphics/classificationtotal.jpg

Baltimore’s System for classification is based on viral genome and its relationship to mRNA.

In total, there is 7 group of classification of viruses.

Group 1 - double stranded DNA viruses

Group 2 - Single stranded DNA viruses

Group 3 - double stranded RNA viruses

Group 4 - positive sense single stranded RNA viruses

Group 5 - negative sense single stranded RNA viruses

Group 6 - RNA reverse transcribing viruses

Group 7 - DNA reverse transcribing viruses

http://education.expasy.org/images/Baltimore.jpg