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When I first started watching the Burning Zone I was student. I was studying Biomedical Sciences. What drove me on was the dream that one day I would be able to help diagnose people’s illnesses and assist in the research for a cure to some of the terrible diseases that exist out there. I still hold on to that dream and I am now one step closer. In this piece I hope to be able to share with you the experiences I have as I work alongside real life Virologists and Microbiologists and give some insight into the techniques used and the knowledge I have gained as result of my training.
To start with, a bit of background. There are three main testing areas in Virology. There is Serology, Molecular Biology and Virus Isolation. I started my learning experience in Serology.
Serology is the study of serum essentially. In this section we look for Antigens (bits of virus) and Antibodies (our response to having had a virus). We look at some bacteria and parasites too such as Trepanema pallidum, which causes Syphilis, Legionella, Brucella and Toxoplasma. The viruses we look at in this department include The Herpes family (Epstein Barr Virus, Herpes Simplex, Cytomegalovirus, Varicella Zoster), The Hepatitis group and also HIV.
The tests are quite simple no real hi-tech. equipment and the only microscope we have is an Immunofluorescent one (which I have to say is my favourite bit, it’s very exciting!) These are the type of tests Marcase and Shiroma would run if they had some idea of which virus was causing the problem and they would take blood samples to test. They would also use these methods to check if someone was immune to a certain virus. Here are some pictures that I found on the net of exactly what I look at down the immunofluorescent microscope.

Figure 1	Figure 2

These pictures are showing what you see down an immunofluorescent microscope when you are looking for Epstein Barr Virus (EBV), which is the cause of Infectious Mononucleosis (Glandular Fever). Figure 1 shows a serum sample that has not been infected with EBV. Figure 2 shows a sample that has been infected with EBV. Antibodies to the virus glow bright apple green and cover the infected blood cells.

All the blood samples have to be separated. It’s very simple, you put the blood tube into a centrifuge and spin the sample for 5 minutes. All the blood cells go to the bottom and we just take off the plasma/serum at the top. That’s all we want.
The thing that struck me most and I would have loved to see Jeff Morgan try this one, is the language. For my first week I felt like I had walked into a different land. I was clueless as to what people were talking about. Virologists seem to have a knack of being able to abbreviate everything. The first day our chief MLSO (Medical Laboratory Scientific Officer) was dishing out the days work “Ok” he said. “ Today we have to do the EBVM’s and the VZM’s. Oh, and we’d better print out the EBVG’s. If any of the IgM’s come out positive we’ll have to RF’s on those. Can someone get the IFA kits out? How are the CFT’s going and has anyone FU’d the checklist. Can everyone keep their eyes open for this urgent VZG please and there’s a CMV to run on the VIDAS.” I felt like crying. I’m NEVER going to understand this. I’m happy to report two years down the line I’m talking like a native!!
The next section I worked in was Molecular Biology. This would definitely been Dr. Shiroma’s domain. The whole thing is based on a technique called PCR, which stands for Polymerase Chain Reaction. The idea is that if you have a virus, you can extract its genetic material and then by heating it and cooling it in certain way you can makes lots of copies of this DNA, which you can then detect. This is quite useful, for example, with viral meningitis. One of the viruses that can cause this is a Herpes virus. If you have some cerebral spinal fluid from a patient with meningitis, one of the tests you can perform is PCR. It is very quick, highly specific and you don’t need buckets and buckets of virus present to be able to give a positive diagnosis. You’ve all seen genetic fingerprinting on TV, well that is done by PCR. The bits of DNA along with a dye are put into holes in a gel (imagine a layer of jelly/Jell-O about 5mm thick). An electric current is applied and this drives the DNA along through the gel. The results are detected as bands. How far the bands have traveled over a period of time tells us what size of DNA fragment we have. They are specific genes to the virus, which are of a specific molecular weight. The heavier/bigger ones don’t travel as far down the gel as the smaller/lighter ones.

A picture to represent the visualisation of PCR product. This is done using gel electrophoresis which uses a chemical called Ethidium Bromide which binds with the DNA and then this glows when exposed to Ultraviolet light.

This is the future of virology it will replace Virus Isolation because of how quick, specific and sensitive it is. It’s a real shame because Virus Isolation is the best bit!!
So, onto Virus Isolation. This was my favourite section during my training. This is very Dr Marcase. The whole idea is in the name, you isolate virus in the specimen (which are very varied). We use, blood, faeces, urine, vomitus, spit, skin scrapings, pieces of flesh and swabs from anywhere you care to imagine. I have quite often come over all nauseous :O). I loved this section, there is a lot of microscope work involved. Viruses particles are extremely small, far too small to visualise using a conventional microscope. The only way you can see them is using a high-powered electron microscope, which isn’t always practical as they are about the size of a room! The main test in Virus Isolation is called culturing. We have test tubes in which we grow a layer of living cells. We inoculate these with some of the patient’s specimen, whatever it might be. We then incubate this at body temperature. If there are any viruses present they will cause the same damage or disease to these cells as they do in the body. When you look at the cells down the microscope you can see the damage. Different viruses cause different types of damage. It’s very exciting when you see something. Sometimes the change is dramatic (it’s called Cytopathic Effect or CPE). In our lab we are lucky enough to have an electron microscope, it is so fascinating to look at an actual virus. I had to try and remember what they looked like for my exam so just for your information Herpes looks like a fried egg!

A picture to represent Herpes CPE in Monkey Kidney Cells. Note how the normal cells are pale, in a complete sheet and quite angular. The CPE presents itself as a rounding up (ballooning) of the cells that tears them away from the cell sheet.

And some electron micrographs: - (Clare’s interpretations)

Herpes Virus: Fried Egg!	Molluscum Contagiosum: Thumbprints

Finally, I think I may have found my Shiroma. One of my colleagues wouldn’t let on when her birthday was. Another colleague said, “Ok tell us what your star sign is?”
“I don’t believe in things like that” she replied, “Therefore I don’t have a star sign”
“Dr. Shiroma, is that you in there????.....”

For Virology on the web please visit http://www.tulane.edu/~dmsander/garryfavweb.html

Thanks to Gramps for the much-needed light relief from my studies and his pearls of wisdom…It is probably best not to make-out with a person who may have Ebola... and the genius of…Never share a band-aid with someone who has any kind of flesh-eating virus. The examiners were impressed ;o)