May 23, 2020: I see a lot of confusion in the literature, with all the different tests being done. Many times, it is not the appropriate test for what is being studied. The big question right now is actually "Infectivity?" Is an infectious unit, a virion, present and "viable, " to be transmitted to the next host. This requires a viral cell culture, where the virus is inoculated into a cell medium, to see if it can replicate, thus proving it is viable.
PCR shows the presence of the specific viral RNA, but does not prove viability. The RNA made be nonviable, from chemical removal of its envelope (e.g. detergents), from heat exposure, from drying, from IgA or IgE attachment, enzyme degradation, etc. If the virus is inactivated, then this test may be a false positive, or if degraded further might produce a false negative result on PCR.
Serology (blood) tests are the body's response to viral exposure or infection. They may not show active infection, but that there was enough viral exposure previously to mount an immune response. These are antibodies that can take about a week to develop after sufficient exposure. They can last various times; IgM, IGA, IGE, in the range of months; and IgG in the range of years. You can monitor an exposure or infection, starting from no antibodies and increasing in time. With some of the new tests for this virus, there appear to be varying degrees of false positives and negatives, at least for the time being. Again, they do not prove or deny infectivity of the patient.
Infectivity, the test which shows if a virus can infect the host, requires a viral cell culture, where the virus is inoculated into a cell medium, to see if it can replicate, thus proving it is viable. Ideally, as we are treating humans, one would take the virus in its inoculum and expose a human. But, this is obviously unethical. Sometimes, vulnerable animals are used. Usually a cell culture is used. This does take a few days, but many people are impatient, so the other quicker tests may be used. It has been estimated the human infectious dose is 100 virions. A cell culture may only require one viable virion to grow.
As long as one does not forget the other tests (PCR and serology) are not measuring "infectivity."
Thank you. Gary Ordog, MD
May 23, 2020: I spent the day reading JAMA, Lancet, CDC reports, WHO reports, etc, not one measure of infectivity, not one mention of a culture. All of the studies reported PCR or serology, or sometimes both. But with the big question of who can be released from isolation, and when, there was never any concern for actual infectivity. As I said, those other tests cannot prove or disprove infectivity. Something for everyone to think about. Thank you, Gary Ordog.
One reason that's not done as a diagnostic test is specificity -- many viruses lyse cells in culture. Seeing lysis alone does not tell you that it's SARS2 that's doing it.
Dear Gary I read the discussion with interest and found it intriguing. Well I agree with all that has been deliberated upon, yes theoretically the virus may not be viable or the infectious dose may not have been achieved. But we have to keep the entire clinical picture in totality and then treat the patient. So there are clinical indicators which include fever, breathlessness etc along with the diagnostic tests aiding the diagnosis. Because performing a virus culture requires biosafety level 3+ infact 4 along with the complete virology setup which is available at only a few centres it becomes the next best option to detect the viral nucleic acid in various clinical samples. However asymptomatic individuals may be positive for the virus but do not have the disease, are totally symptomless or have had mild symptoms that just passed off. So the entire spectrum is in shades of grey not exactly black and not truly white. A complete picture and clinical correlation is a more pragmatic approach. I hope I have provided another view.
John Schloendorn < many viruses lyse cells in culture. Seeing lysis alone does not tell you that it's SARS2 that's doing it. >
Is any method to avoid other viruses when conduct SARS-CoV-2 alone lysis experiment?
Mahima Lall There are some patients having long time (several months) with PCR positive. We know dead virus RNA can be PCR amplified. In this case, should 'infectivity' of the viruses be tested on the patients?
My point is that I see no mention of testing infectivity out of all of the hundred of articles I reviewed. Someone needs to do this, I would think.
Here is one prime example, as a physician, have to deal with thousands of times. The patient has COVID-19, and recovers, of which there may be a quarter million in the USA right now. I need a test for infectivity to send the patient back. JAMA shows today they can have positive PCR, eg stool, for months. Serology obviously doesn't tell infectivity currently. Only a culture . Obviously, there are lots of provisos for each.
Cultures have a high specificity, but low sensitivity. As a physician, I can increase sensitivity of my infectivity culture technique by doing more samples, altered by timing and location. I can approach 100% because there is no other ready method for determining infectivity. The poor fellow who is isolated for six months because he is shedding dead RNA in his stool, and what if he is the only neurosurgeon around? Cultures may be of help in certain cases.
To one of the questions above, when you see lysis, then you identify it, PCR etc. What I would like to know, when you have killed the virus and have all of this deactivated RNA everywhere, when is it safe to go outside and mingle? You need a viability, infectivity, contagion type test. A cell culture, or even give it to a similar animal? All of these studies that are coming out maybe testing for inactivated virus, because no one is adequately testing infectivity. As an example, lets say I am taking my tonic water (l-quinine) and eating a good diet of leptins, I have no illness, but my nose is shedding inactivated RNA, test positive by PCR, positive by serology IgM at this stage. The IgG is starting to increase. Am I infectious or not. Three negative cultures, I say "not." But the rest of the world right not would probably say "yes."
To answer another of the above questions, how do you isolate only one virus in a culture/ the so-called lysis experiment. I guess that is the cool thing about 40 years of clinical practice, with access to any lab test of the day. People don't seem to get two viruses at once. I cannot think of one case where a patient had two active viral infection at the same time. Now I was told and had assumed over the years that it was because the first virus caused massive interferon production that blocked any other virus from infecting the person. Maybe someone else has a different experience, let me know. So for example, we would usually do three separate cultures, all three would be H1N1, for example and nothing else. If it is Herpes virus causing encephalitis, then there was never CMV as well, and on and on. Perhaps others has studied this anecdotal observation. So to answer that question above, If the patient has a viremia, and you take a culture, and you get disk on your " lysis experiment" that should be your virus, because you only get one at a time. Also we did three separate cultures, which if all three are the same virus, the probability os greater than 95% that is your culprit, and that is enough to treat medically.
Viruses, rule of thumb; but bacteria, toxins, etc., NO. Commonly you can see multiple bacteria as pathogens, and multiple toxins causing injury. We will ask Dr. Fauci.
Here is your viral cell culture lysis experiment. The little circles are areas of viral cell lysis, these will be rich in the virus, Touch your sterile loop to the spot, and transfer it to your ID mechanism, PCR, ELISA, etc. The upper arrow is the cell culture medium, the lower arrow is the lytic lesion caused by the growth of a single virus. (Obviously there are many other methods available, but this is a basic one, because the question was asked above)
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