Are you curious about viruses, particularly their characteristics, morphology, and life cycle? In this piece, we’ll be studying viruses under the microscope.
Viruses are unique to unicellular organisms as they are generally referred to as particles and not cells.
These particles, or virions, also differ from other unicellular organisms as they are not technically alive and as a result of this they can’t grow or multiply on their own.
Viruses can be difficult to study as due to their size a number of viruses are impossible to view with a light microscope.
If you were to compare viruses to bacteria, bacteria would be on average between 100 to 500 times larger than viruses.
It is suggested by researchers that both unicellular organisms, such as bacteria and viruses, and living organisms host at least one, single virus particle at all times.
Looking At Viruses Under The Microscope
The size of virions, virus particles, range in size but they all tend to be extremely small. They are measured in nanometers.
To put this measurement into context, one centimeter is equal to 10,000,000 nanometers.
The largest virions that have been discovered are the poxviruses and the mimivirus, which both measure 400 nm.
One of the smallest virions is the poliovirus, which can make people extremely ill, it measures 25 nm.
Most conventional and compound microscopes have limited resolution and are best used for particles measuring more than 200 nm and so they can only be used for virions of 200 nm and above, which are referred to as giant viruses.
As a result of this, certain techniques have been developed to aid scientists in discovering particles that are smaller than 200 nm.
Below we look at four such techniques that are often used in laboratories around the globe.
Fluorescence Microscopy Technique
It was in the early 1900, that the Fluorescence Microscopy technique was first developed, making it quite a recently developed technique in the vast history of the field of science.
This technique is used to monitor viral DNA that can be found in the host’s infected cells.
In this technique, Ethyl-modified nucleosides are used to label the DNA of viruses such as herpes virus, vaccinia virus, and adenovirus that have worked their way into the host’s cells, infecting them.
Samples that are taken of the infected cells are studied under a confocal microscope, which uses light from a laser to excite the virions in the sample so that they are easier to identify.
Fluorochromes are used to allow the organisms in the sample to fluoresce when being viewed under the confocal microscope.
Fluorochromes can only fluoresce when they become attached to the target virion, and so if there is a virus present in the sample the fluorochromes will attach themselves, indicating a virus is present in the sample.
If the fluorochromes do not fluoresce then there are no virion particles present in the sample.
This technique is particularly useful when scientists are looking to make an estimate as to the quantity of the virus present in the host.
This process highlights how concentrated virion particles in the sample are which gives a good indication of how many virions may be in the host.
Total Internal Reflection Dark-Field Microscopy (TIRDFM)
TIRDFM is used for viewing large virus particles and is a label-less technique. Using only a perforated mirror rather than a dichroic mirror you can achieve this process.
Dichroic mirrors are typically used in traditional objective-type TIRDFM.
The process of preparing a sample for this process is extremely complicated and has seven main steps that you can expect to be carried out in all circumstances. These steps are explained in brief below:
- Taking a sample of Influenza A/Puerto Rico/8/34 (H1N1) this is inoculated into an allantoic fluid which has been taken from an embryonated egg at 37 degrees for three days
- The allantoic fluid solution is then centrifuged at 2,500g for approximately 20 minutes at 4 degrees
- The supernatant is then passed through a membrane filter, with a pore size of 0.45um
- The resulting solution from the above steps is then subjected to 10 to 60 percent sucrose density gradient centrifugation. This lasts for two hours at 120,000g and at 4 degrees in a swing bucket rotor
- 2ml aliquots of the gradient are next obtained
- The fractions are analyzed next. To do so an SDS polyacrylamide gel is used and then this is centrifuged for 2 hours at 120,000g at 4 degrees
- Lastly, the pellet is suspended in phosphate-buffered saline
When viewing the pellet-shaped samples under a TIRDFM you can expect to see a dark background with bright spots.
Transmission Electron Microscopy
To prepare a sample that you intend to view through the transmission electron microscope you need to follow certain steps carefully. These steps are outlined below:
- 10um of suspension is applied to a hydrophobic surface of a parafilm square which has already been placed in a petri dish
- A formvar-coated grid is then floated on the drop of the suspension for around 60 seconds. When completing this step it is important to ensure that the formvar side comes in contact with the liquid sample
- The grid is next floated in a angle drop of 1.5% phosphotungstic acid, these are then used as stains
The above image shows a transmission electron micrograph of a tissue section containing variola viruses.
Virions viewed under this microscope can often be viewed as small particles inside the host cells that they have infected. An example of what you may observe using this technique is below.
The above image shows a colorized transmission electron micrograph that is showing the H1N1 influenza virus particles. You can clearly see the smaller virion particles inside the infected host cells.
When scientists want to get both a better view and understanding of the structure of viruses cryo-electron microscopy is often a technique that is used.
This technique helps scientists get a deeper look into the viruses’ structure while also giving insight into how the virion particles attach themselves to host cells, and how they assimilate as they replicate.
And also the associations the particles may have with multiple cellular mechanisms during the replication stage of the virus’ life cycle.
A sample is frozen using liquid nitrogen prior to being viewed under a transmission electron microscope that has a cryo-stage.
This process is used when using the cryo-electron microscopy technique.
The images obtained are then put through software that creates three-dimensional versions of the viruses, giving the researchers a unique 360-degree view of the structure of a virus.
Below is an example of such an image.
Here you can clearly see all of the many proteins and particles that make up a virus.
The virus has an appearance similar to a head of broccoli, made of multiple spores and then a core that holds all of the important information regarding the makeup of the virus.
The above techniques are only a small collection of the many ways that researchers and scientists study viruses.
As science continues to progress, new techniques and equipment will be developed too so that viruses can continue to be studied at a similar rate to their production rate.
This will help to prevent outbreaks and also enable scientists to quickly research sudden outbreaks of viruses to create treatments and vaccines to continue to keep living organisms, such as animals and humans safe.
General Virus Characteristics
As viruses continue to evolve there are a few characteristics that continue to be shared, however, researchers often find one of the below three in all found and studied viruses.
- Viruses are dependent on the host that they infiltrate to reproduce, they cannot do so alone
- A receptor-binding protein can be found which allows them to become attached to host cell surfaces, before embedding themselves into the cell and causing infection.
- Most viruses are very small, in most cases, they are 100 times smaller in diameter than bacteria
The Morphology Of Viruses
When looking at a virus’ structure under a strong microscope often they are observed to be a helical or icosahedral shape. A protein coat contains the genetic material which is often called the nucleocapsid.
The capsid size and shape often vary and are influenced by the virus family the virus has evolved from. Capsomeres are subunits of proteins and how they are arranged will determine the shape of the capsid.
Over time, more giant viruses continue to evolve and be discovered, an example of which being the mimivirus which can be up to 700 nm, making it bigger than some bacteria.
The virus’s genome is carried inside the capsid. The genome is either DNA or RNA, depending on the virus. Both help to produce new morphing viruses.
Small viruses carry between 2 to 100 genes in the capsid and larger viruses can carry up to 1,000 genes in their capsid.
Viruses differ from other unicellular organisms as they do not have other types of critical organelles, such as mitochondria and ribosomes which you would typically find.
It is the lack of these organelles that means that they cannot reproduce and grow independently, hence their reliance on a host cell to do so.
Once viruses have infected a host cell they can then begin to reproduce, develop and grow.
As they do so they destroy the host’s cells which results in new viral particles that go and infect other cells in the host, causing the infection to spread and the virus to slowly take over.
Science has been able to keep up with new emerging viruses up to this point and researchers claim that only ten viruses can exist at any given moment.
A Viruses Life Cycle
The life cycle of a virus depends on the host that the virion particles have infected. There are however certain stages that all viruses go through and these are explained below.
This is the first stage of the life cycle of a virus. Attachment proteins are used by the virus to build receptors on the surface of the host’s cells.
These attachment proteins are extremely important as they will only mesh with certain cells, depending on the type of virus.
An example of this is the rabies virus. This particular virus uses glycoprotein as the attachment protein and this enables the virus to attach to cells on neurons.
The genetic material of a virus is injected into the host cells during this stage of the virus life cycle. There are two ways that this can happen, through either direct membrane fusion or endocytosis.
In the event of endocytosis, a virus will attach to the receptors at coated pits. The pits are then pinched off and this causes a coated vesicle to form.
These then become fused with endocytic vesicles and then also fuse with lysosomes.
If fusion proteins are present, the DNA or RNA of the virus will fuse with the membrane and this helps to prevent the fusion protein from being ruined by the lysosome enzymes.
Penetration is important as the RNA of the virus needs to reach the cytoplasm of the cell without being ruined in order for the virus’s life cycle to continue.
If the virus does not reach the cytoplasm it can attempt to use the plasma membrane as an enveloped virus that has been induced by the binding of the protein receptor.
This process differs from endocytosis, as endocytosis is triggered by how the pH levels change.
Targeting & Uncoating
Uncoating refers to the release of the virus’s genome, which begins in the penetration stage.
The replication cycle begins in the capsid for a number of viruses, such as the retrovirus. This is a result of the usual changes of the capsid.
Gene Expression And Replication
This phrase is when new viral genomes and the other virion components are created. This leads to the genome of the virus embedding itself into the DNA of the host.
The severity of this stage on the host depends on the virus.
Virus Assembly And Release
The final stage of the life cycle of viruses is when all of the new virion particles come together.
These newly created genomes are injected into the capsid, which forms a new nucleocapsid and the cycle is then repeated.
It is the morphing of viruses that can make some viruses extremely difficult to treat, however as the last century has shown, many viruses mutate a number of times, and often the last mutations are weaker versions of the initial virus.
Deadly viruses have now become treatable as they continue to mutate into weaker versions they too become less contagious, which benefits everyone.
While certain characteristics may differ between different viruses, the above information highlights the most commonly shared traits amongst virus particles within their characteristics, morphology, and life cycle.
Viruses are extremely complex organisms and as the world continues to evolve so too do more viruses and older viruses continue to adapt and become smarter.
This puts pressure on science to keep up to date so that new viruses can be studied and fought against at a fast rate.
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