IV. Viral Pathogenesis

Viral Pathogenesis is the capacity of a virus to cause disease in a target host. It is a sophisticated study about the relationship between viral replication, host defence and viral evasion of the host defence.

I’ll just organise this topic into different parts for easier explaining.

1. Sites of Viral Entry into the host.
2. Viral spreading in the host body.
3. Modes of Transmission.
4. Virus-induced injury (cellular).
5. Sites of Viral entry into the host.

I shall now start explaining each and every one of these points…

1. Sites of Viral Entry into the host.

There are various places that a virus can enter and start causing disease to the host.

A. Animal host (E.g. Humans, lions, fishes, etc.)

• Skin
• Cuts, abrasions, etc.
• Conjunctiva (eyelids)
• Urogenital tract
• Respiratory tract
• Alimentary tract

B. Plant host

• Any part of the plant as long as there is direct penetration of the cell wall.

2. Virus spreading in the host body.

i. Systemic infection

• Many organs are infected

ii. Haematogenous spread

• Spread through the bloodstream
• Viremia
• Active / Passive
• Primary / Secondary

iii. Neural spread

3. Modes of Transmission.

• Spread through germ cells
• Consumption of infected tissue
• Respiratory Secretions
• Aerosols during speaking, sneezing, coughing, breathing, singing
• Faeces
• Blood

4. Virus-induced injury (cellular) and effects.

The cells that are infected with viruses will display Cytopathic Effects (CPE).

Some of these effects consist of:

• Altered shape
• Detachment from substrate
• Lysis
• Membrane fusion; syncytium
• Membrane permeability
• Inclusion bodies
• Apoptosis

Other effects shown would be the Formation of Syncytium, shutting off of cell functions and Immunopathological lesions.

a. Formation of Syncytium

Below is a diagram describing Syncytium formation.

b. Shutting off of cell functions

• E.g. poliovirus shuts off cellular function in neurons resulting in cell death and hence paralysis

c. Immunopathological lesions

• Impairment of immune response due to infection of immune cells. (E.g. HIV on CD4⁺ & CD8⁺ T lymphocytes)

• Enhancement of immune response causing haemorrhagic fever. (E.g. Dengue haemorrhagic fever, Hantaan, Ebola, etc.)

Viruses? A deadly weapon of war?

We all know how dangerous viruses can be. They can even cause cell death. However, ever thought about what humans could do with the control of such technology? Sounds dangerous, doesn't it? Imagine terorists being able to bring the world under their control and believes, by threatening citizens with the mastery of perhaps, cowpox? Cowpox is dangerous enough to cause death, especially since vaccination stopped over 30 years ago, and if a strain of virus kept for research was to be stolen.

Although we may not be sure of being able to prevent such theft, we can learn how to play our part in fighting against it.

We should always make sure to vaccine ourselves and our close family. Also, we should take precautions by keeping our body stystems healthy enough to fight against diseases, by keeping a healthy diet and taking medication whenever needed to.

We can all play a part to fight against viruses, artificial or natural. It is simply a matter of whether we want to. How about you?

Emerging Viruses. How to fight this new foe?

At all times, we must all be ever ready and prepared for new rapidly spreading infectious viruses within the human population. But what are these new threats called? We call them, 'emerging viruses'. This has already been explained by my colleague. But what I'm unterested in, is as to how we can fight this new enemy.

It is good to cure this disease after researching about it, but prevention is better than cure. Therefore, we shall learn about the causes of these emrging viruses, which we must take care to avoid, in order to prevent teh enemy from even having an opportunity to strike, or at least hinder the birth of such problems.

Firstly, we know about virus factors, those that we cannot avoid yet. Spontaneous evolution of a new virus entity and generation of a novel strain due to co-infection of different strains in an individual.

The second part consists of what we can prevent, that is, human factors, such as concentration of people with shared lifestyles, breakdown in public health, climate change, and man invading natural habitats such as cutting of forests to increase their land for development and comfort, which may cause zoonoses.

Men are typically selfish. However, we must understand what must be done, and prevent all of these factors from happening as much as possible, as much as we can survive with.

However, although I believe in doing so, there are cases where certain choices are up to you. Imagine if you are a farmer with sick chickens that are infected with avian flu. With your family starving, would you rather kill the chickens as teh government demands, or sell them illegally for bread and butter. Personally, if we were to die of hunger, I would rather send my children to teh government to take care of, and die of hunger on my own with dignity, since I do know that tehy would want it that way, and that it is better than knowing that all were killed due to my selfishness.

Think about this. What would you do?

Virusoids? What are they?

A long time ago, most people believed that infectious diseases were caused by either viruses, bacteria, fungi or parasites. However, the mad cow's disease caused by something else proved this theory wrong.

In this case, I shall be talking about a certain infectious agent known as virusoids that infect plants. Similarly to viroids, they are infectious agents composed exclusively of a single piece of circular single stranded RNA which has some double-stranded areas. It has no capsid or envelope, making it susceptible to certain environment hazards such as heat.

The key is here. Unlike viroids, they do not require assistant virus, as viroids do, such as hepatitis with hepatitis D.

Whether this is better, or if this is not, largely depends on the purpose of the virus. This is what I believe. How about you?

Flaviviridae -II

Next up is Yellow fever. Now, the symptoms first.

It has an incubation period of 3-6 days

Headaches, nausea strawberry tongue, flushing of the head and neck, conjunctival injection, just to list a few.

Severe yellow fever is when the patient goes into remission after acute yellow fever

Fever, vomiting, abdominal pain, dehydration, prostration, jaundice, black vomit (coffee-ground diathesis), and bleeding from injection puncture sites. Interestingly, the virusis absent from the blood, which suggests that autoimmunity greatly influences it.

Lastly, the West Nile fever.

It originated in Uganda, discovered way back in 1937, with an epidemic in 2002 in the US. Now let's look at the checklist of symptoms:

Mainly mild to no symptoms

Fever

Headache

Body aches

Skin rash

Swollen lymph glands

Note: these are for the not so severe cases, those are listed below-

Crossing blood-brain barrier- as in it passes through the blood-barrier in the brain, residing there.

Encephalitis

Meningitis

There is no vaccination for this west nile virus and dengue, so please follow this for your sake and your loved ones. 

It's amazing what a tiny insect can do to humans. It's scary, but as long as one takes preventive measures, I'm sure everything will be alright. That's all for now. tune in for more on our blog.

Flaviviridae

This is a family of (+) single-strand RNA viruses that have 3 important structural proteins and a host-derived lipid layer. Below is a simplified diagram on Hepatitis C virus, which belongs to the flaviviridae family.

 Generally, it is  genetic material-containing icosahedron in a lipid-containing envelope. Now, onto the members of this family.

First, let's meet the most important arbovirus (insect-borne) virus today - Dengue.

Dengue- 

It has 4 serotypes, DEN-1, DEN-2, DEN-3 AND DEN-4, based on neutralisation test. DEN-2 has the greatest antigenic and genotypic distance from the others. It is spread by the Aedes aegypti, or Aedes mosquito. 

It causes dengue fever, which has asymptomatic infections. Acute infections has the following symptoms: severe frontal headache, nausea and vomiting. Sever muscle and bone pain are also experiecnced.  

Dengue Haemorrahagic Fever or Dengue Shick Syndrome may also occur. It's symptoms are similar to that of dengue fever, HOWEVER, remission occurs, resulting in the sudden deterioriation of the patient's condition. Severe prostration, hypotension, collapse of the circulatory system, bleeding and shock follows. It can be catergorised into 4 grades. 

Now, onward to pathogenesis. Despite intensive studies conducted, pathogenesis of dengue haemorrhagic fever(DHF), is not really understood, However, there are 2 theories, both of which I will present.

(I)Virulent strain theory

Some strains are more virulent than others, with molecular studies showing variance in sequences among different strains withinserotypes, and early evidence points to DEN-2

(II)Antibody enhancement

This is the main theory, as these cells, the monocytes and marcophages are the main target for the DHF viruses. 

Wow, this was a long post, so I'll be covering on Yellow fever in the next one!! But I realised that something so simple as discarding stagnant water can prevent the breeding of these mosquitos, and eventually, the prevention of such disease. So, get rid of stagnant water, cause Prevention is better that Cure.

Emerging Viruses

First off, what is an emerging virus?? Well, its an known virus, that is thought to infect animals only, but is showing an increasing incidence in humans. Sounds familiar?? If you guessed bird flu or SARS, you're right. 

Now, how does a virus that infects animals only start infecting humans?? There are various factors, which can be lumped into 2 groups: Virus and Human factors.

Virus Factors:
(i) Spontaneous Evolution
Alright, now, viral populations are hetergenous and there IS a limit of variation, but that is a really big number. Furthermore, as there is selection pressure present, this results in high mutation rates, and thus, spontaneous evolution, where a new virus entity is formed.

(ii)Random Assortment, or the generation of a new strain due to the co-infection of different strains in an individual. That was long, so I'll be using Random Assortment instead.

As previously stated, in the life cycle of a virus, the genetic material of the virus undergoes replication and expression. However, when two viruses infect one cell at the same time, or co-infection, the host cell gets mixed up during the assembly stage and a "new" vius is produced and released out of the cell to infect others. Thus RANDOM assortment

Human factors:

(I)Man invading the natural habitat of animals. Truly, this shows why humans, at times, should not destroy Mother Nature. Though other times, we humans, too have no other alternative. Anyway, on with the explanation.

Examples of humans invading the animals' natural habitat would be deforestation, which puts man closer to wild animals. 

(II)Climate change is an important factor, especially with global warming nowadays. 

This increase of global temperature will definitely affect the ecosystem on a whole. Insects, especially, are greatly involved as warmer temperatures have been linked to an increase in reproduction of mosquito, a vector of the virus which causes dengue fever. Hence, increase in temperature = increase in viral vectors = more people infected.

(III)Breakdown in Public Health

Though this might not happen in developed countries, it still happens in less developed countries. Unhygenic surroundings increase the likelihood of incidence. 

(IV)Concentration of people with shared lifestyle

As the concentration of peopple increase, so does the chances of contracting a virus from an infected person. So, when one person is infected, that one person can spread to another 2 people who then infect others, continuously infecting other, increasing incidence.

As seen above, some factors are caused by humanity, others by nature. From this, I realised that the cause of increasing incidence has so many factors, and mostly are results of our own actions. 

III. Virus Assembly and Release

In the 3rd stage of the virus life cycle, the new incomplete virus particles start to arrange and assemble themselves in the host cell’s site of viral synthesis – Areas of protein synthesis and processing of virus material in the host cell.


After the assembly process comes to hail when the host cell is exhausted of its nutrients and energy in making the virus proteins and envelopes, the virus particles will begin to leave the host cell via several methods, thus beginning to re-infect other cells again.



A. Virus Assembly

- Depends on the site of Synthesis

- Sites of virus protein synthesis and processing
1. Endoplasmic reticulum
2. Golgi body

- Sites of Assembly
1. Nucleus
2. Endoplasmic reticulum
3. Golgi body

- Viral Envelope sites
1. Nucleus
2. Endoplasmic reticulum
3. Golgi body
4. Plasma membrane
5. Formed via budding off from plasma membrane

A diagram showing a typical budding process.

B. Virus Release

1. Through cell lysis/cell burst/apoptosis.

2. Through budding at the plasma membrane.

3. Through Accumulation of particles in vesicles and release via Exocytosis.

(Exocytosis is the opposite of Endocytosis,. It is the process of purging unwanted materials from the cell. Below are 2 pictures to show what it is.)

The link to the 2 pictures: http://www.linkpublishing.com/video-transport.htm#Paramecium_-_Exocytosis_

Personal opinion and conclusion...


That would be the gist of what the stage of Virus assembly and release would be talking about.
After going through all the information, I realised that viruses are truly terrifying entities.


Viruses are classified as non-living entity by the central dogma, yet they are able to survive in extremely hard conditions with only some genetic materials, manipulate its host’s mechanism to start reproducing the virus and finally resulted in causing most of the deadly diseases known to man.


Now, is Viruses terrifying or not?

The Original Taxonomy of Viruses

The taxonomy of viruses is well known to all and is still in progress, as science is. But do you know what the original taxonomy was? Quite simply, it consisted of infectivity, filterability and their requirement for human hosts.

Infectivity is how well the virus infects a host, which quite determines it's strength.

Filterability represents how well it can be filtered through the chamberland filter (refer to previous post on the birth of virology).

Finally, their requirement for hosts, such as which kind, perhaps humans. This determines their flexibility.

Through these, viruses were first classified. Of course, like I mentioned earlier, the road of change is never ending. Who knows, maybe you may be next to change this taxonomy.

Plaques assay

Plaques are visible structures formed in a cell culture contained within a nutrient medium. (e.g the plaquing medium). These clearings has no cells in it because the virus infect the cell and eventually lysing it.The cells that were initially seeded in the well and allow the virus to infect it then overlay with plaquing medium for the cells to attach to the agarose and grow. Just like how each colony is derived from a single E.coli cell in an agar plate, each clearing is derived from one virus attaching to one cell, and when the virus infects the cell and multiple it continues to lyse the other cells around and forms a clearing around that area. As that clearing, also known as plaque, arise from one virus, we are able to determine the number of viruses (which is also known as titer which is also = no. of plaque forming units/ ml) by counting the number of plaques.

In plaque assay, virus undergoes serial dilution.

http://www.slic2.wsu.edu:82/hurlbert/micro101/images/virus28.gif

I have generally found that plaques assay is not a very good way to find virus concentration. There are many ways for virus to hide, and so this assay is usually an underestimate of the actual number of virus present. For instance, a pfu (plaque forming unit) may not find a cell by the time the attachment period is ended or it may stick to the plate where no cell is present and by the time the monolayer reaches it, it's too far behind to form a visible plaque.

The Birth of Virology

Previously, we had discussed the origins of viruses. But do you know how viruses were discovered? In other words, in this topic, Iwill be talking about the birth of virology.

This story begins in 1884, where a scientist, the french microbiologist Charles Chamberland invented teh chamberland filter with pores made smaller than bacteria.

In 1892, the Russian Biologist Dimitri Ivanovski used it to filter the tobacco mosaic virus from infected plant leaves. It was speculated to have been produced by bacteria, but was not further studied. From that point on, it was thought taht such infectious agents could be filtered teh same way and grown on a nutrient medium. This was known as the germ theory.

However, later on, down history, many scientists have done research on this newly discovered type of infectious agents, and helped to change and improve the virology that we study today. But, that's another story.

Indeed, science is ever changing and requires the help of all curious and innovative minds such as those of the students of today, like you and I. That, is why this blog was made. For us to share our knowledge and change our views on microbiology.

Where do Viruses Come From?

We study so much about viruses, and yet there is one very much unsolved mystery, that is where do these viruses come from? In other words, their origin. There are three very popular theories about this.

Let us start with the regressive theory. It is said that viruses were once parasites, as they were today. Overtime however, genes into required by their parasitism were lost. This is also known as the degeneracy theory. This is very possible, since viruses cannot reproduce without a host.

Next up is the cellular origin theory. This theory states that viruses could have evloved from stray pieces of RNA or DNA, perhaps even plasimds (DNA that moevs between cells, also known as jumping genes). Overtime, they could have developed their capsids and envelopes.

Last but not least, we have the coevolution theory, which states that viruse evolved overtime from pieces of genome and protein.

Personally, I would choose to believe the regressive theory, seeing as how viruses may have been weaken by their activities as parasites. On a side note, this is also an interesting lesson to us. If we depend too much on others, one day, we will be weaken and unable to stand independantly.

This is just my opinion, but how about you? Which theory do you believe in most?

Antibody and Antigen

I'm sure the title has been heard often, or a least a little. Now then, I'll explain.

Antigen - This is the agent that causes your body to produce antibody, as shown by the following picture. It is foreign to the body, i.e. from the external enivoment or and thus, antibodies are formed

. 
Which brings us to.... Antibodies.

Antibodies - A type of immune system-related protein which binds specifically to an antigen. As explain by the following picture:

Each antibody consists of four polypeptides, 2 heavy and 2 light, joined to form a "Y"-shaped molecule, though the amino acid sequence at the tips of both heavy and light chains vary, which determine the different types of antibody that bind to different types of antigen. Perhaps the next picture will make things seem a little clearer.
 

Now, antibodies are classified into 5 major classes, based on the constant reigon and their immune function.

For those would like an even more detailed explanation on this subject may click on the following link. I hope that readers have gained an understanding on this. I know the two terms are confusing, I know I've been confused by it a few times before finally getting it.

Enzyme-Linked Immuno-Sorbent Assay (ELISA)

As the title states, this post will be on Enzyme-Linked Immuno-Sorbent Assay, or ELISA, for short.

Now, exactly what IS ELISA?

It is a immunochemical test which is used to test for the presence of a specific antibody or antigen.

ELISA has 2 methods, direct, and indirect. First up in the spotlight is the direct method of ELISA

Direct method-

This tests for the presence of a specific antigen. The specific antibody is used to coat the walls of the wells on the plastic strip, using a coating solution. The positive sample, which contains the antigen is introduced into the well, where the antigen is bound to the antibody. This bond is strong enough to withstand the removal of the fluid and rinse. 

Afterwich, the secondary enzyme is added in, which binds to the bound antigen. This secondary antigen has been modified, so that it has an enzyme attached onto it. 

The contents are then rinsed once more, removing excess antibodies. The specific substrate for the enzyme is then added. This causes the enzyme to turn colour, which then turns the colour of the whole contents of the well.

Indirect method- This tests for the presence of a specific antibody. Thus, the antigen is used to coat the walls of the wells. The positive sample contains the antibody, which binds with the antigen. This complex formed withstands rinsing. An anitbody-enzyme conjugate, or reporter antibodies, bind to the antibody, froming a complex. 

Rinsing takes place again, before the specific substrate for the enzyme is added, causing a colour change.

Uses:

ELISA can be used to determine the presence of a specific antigen and detect the presence antigenic properties. The following links will lead to an animation describing ELISA and a virtual lab.

I personally find that ELISA effective, as long as one got all the steps right and can save time, despite the time taken to complete the serial dilutions. And so, I final understand what is done with our blood and urine samples, though only a bit of our sample goes through this.

II. Replication and Expression

I shall discuss about the 2nd stage of Virus life cycle, which is the replication of genome, mRNA production, processing and translation. In short, it is known as Viral Replication & Expression.



In actual fact, the way the virus reproduces itself in the host depends on the nature of their respective genome.



For example, Group I of the Baltimore’s Classification - Double stranded DNA viruses (E.G. Smallpox and other viruses from the family Poxviridae), can only replicates in the host cell’s cytoplasm and require host cell polymerases to replicate its genome and therefore is very dependent on the host cell’s cycle and mechanisms. The viral genome contains all factors for genome replication and transcription.

Below is a diagram that describes replication of the genome.

http://www.chem.ox.ac.uk/smallpox/images/smallpoxreplication.jpg

A side note...

Remember Baltimore’s Classification? In studying the subject of viral replication & expression, this classification is important as it is the classification which identifies virus into groups based on their genome & how it is being replicated. This, in turn, gives us a shortcut method of determining how a particular virus replicates as long as we know which class is the virus from.


I realise this after I went back to revise the lecture about viral replication & expression. I also concluded that everything taught to us in the MicroB lectures have significant connection to all the other different aspects in virology. In this case, for example, the Baltimore’s classification can be use to simplify the method which is use to determine how certain type of viruses replicates.

Virus Entry

In the last entry, I mentioned and explained about virus attachment onto the host cell. Now I will go onto the second part of the 1st stage of the virus life cycle - Routes of host entry (Penetration).



Even if the virus particle attaches itself onto the cell surface, it does not gain entry into the host cell. There are several routes for the virus to take in order to penetrate into the host cell. Without further ado, I shall explain the routes of entry to the best of my abilities.



I. Attachment and Entry

After the virus attaches itself onto the host cell, there are 2 main routes a virus particle would take to enter an animal host cell and only 1 route in the case of a plant and bacteria host cell. In an animal cell, a virus particle can use....



1. Envelope Membrane Fusion

In envelope membrane fusion, the virus's envelope blends with the cell membrane, releasing its genome into the host cell and starts its replication.

2. Endocytosis


2a. Receptor-Mediated Endocytosis (Envelope)

In receptor-mediated endocytosis, the enveloped virus particle tricks the cell into thinking that it is nothing more than nutrients or harmless proteins. The host cell will then take in the virus particle along with its membrane. Once inside, the virus particle will then release its genome and starts its replication.


2b. Clathrin endocytosis (Naked)

In clathrin endocytosis, the naked virus binds with the host cell membrane to gain entry into the host along with the newly binded membrane from the host.

In a plant and bacteria cell particle, virus particle can only gain entry when there is a breach in their respective cell walls. From there, the virus particle will start to uncoat and starts to infect the host.

Below is a link to an animation which summarizes what the whole stage of attachment and entry is all about.

http://www.blackwellpublishing.com/wagner/animations/attach/attach.html

Virus Attachment

I. Attachment and Entry


Viruses are non-living entity because they are non-active outside a host cell and do not display actions which define them as living entity by the central dogma but starts to begin motion and actions of living entities once they are inside a host cell.Therefore, infection only starts when the virus is inside of the host cell.








But... how does the Virus get into the cell?










The answer is here!!!












The virus got to attach itself to gain entry to the host cell!!!














Alright... now that you get that... SO, do you know how does the virus attach itself to the cell?














Let me tell you how now!











- Attachment (Virus)

To be able to attach to their respective host cells, every type of viruses has their own specific virus attachment protein (VAP) to the host cell receptor. Therefore, Viruses do not attach itself to all the living cells they come across. They are only able attach themselves to those cells which has specific surface protein receptors that can receive and connect to the viruses’ own attachment proteins.




- Receptors (Host cells)

The receptors can be any surface molecule of the cell. It can be Glycoprotein or Glycolipid. Usually, a cell has multiple receptors and each receptor have their own function and opportunistically, viruses would use it as a mean for attachment. It is also because of that, viruses have a certain host range and tissue tropism.

A typical cell surface (**Note that the glycoprotein and glycolipid are the green structure protruding on the surface)

Now That's all for attachment part. The next post will cover about the routes of penetration by the virus into the host.

Virus-Host Interaction & Virus Life Cycle

http://cimurr.deviantart.com/art/Viruses-89730723

Woosh!!!!! I’m back with a series of updates about the virus-host interaction and virus life cycle. After 4 lectures about the Virus-host interaction and the virus life cycle by Mr William HOW, I’ve receive considerable knowledge about the life cycle of viruses.

As molecular biology students, we know that Viruses are not a living organism defined by the Central Dogma but interestingly, they can still infect living organisms and cause diseases that vary from asymptomatic to death as a result of contracting such viruses. Viruses challenged the boundaries of life science as once doctors and scientists assumed that the only things that cause diseases were only bacteria, fungi and parasites. As Viruses are non-living entity, it would be interesting to look at how they infect the host and therefore cause the disease that they are known to cause.

With this, I’ve decided to share my knowledge as well as giving some of my opinions and comments about lectures on virus life cycle. I hope to get comments for my posts too.

Firstly, I’ve copied the stages of the Virus Life Cycle from the lecture for better understanding of the cycle itself. Below are the stages of the Virus Life Cycle. There are 5 stages all together.

Stages of Virus Life Cycle

I. Attachment and Entry

II. Replication and Expression
o Genome replication
o mRNA production, processing and translation

III. Assembly and Exit

IV. Viral Pathogenesis

Now that the stages of Virus life cycles have been labelled out, I hope that other students out there studying virology like me could get a better idea of the Virus-host interactions and the virus life cycle. This way of classifying the stages helped me in memorising and understanding the whole hoo-haa about Virus-life cycle.

In studying virus life cycle, there is this one thing that one must commit to mind.

1. Attachment
2. Penetration
3. Uncoating
4. Replication & Expression
5. Assembly
6. Release

Diagram showing a typical virus life cycle