Virus Isolation Part I

Has the SARS-CoV-2 Virus Been Isolated? Yes and No

According to virology and their modified definition of isolation

(actually that is being kind, they have completely inverted the word to mean mixture which is adding things, not separating one thing from others),

yes the virus has been isolated many times the world over. They take a sample from a patient, add fetal bovine serum and antibiotics to a cell line - typically African green monkey kidney cells (Vero-CCL-81).

Us regular folk would simply call that a cell culture. Calling it a virus isolation is presuming the virus is present in the sample. How can a researcher know that a virus is present until they examine the culture and separate those things in the culture so they can be characterized?

They commonly use gentamicin as the antibiotic which induces nephrotoxicity (Nephrotoxicity is defined as rapid deterioration in the kidney due to toxic effect of medications and chemicals.) by inhibiting protein synthesis in renal (kidney) cells.

Then when they observe CPE (cytopathic effects) or in other words cell breakdown, they claim proof of a virus. Then they take the DNA/RNA fragments and construct a genome with computer software. They also make use of databases of genetic sequences. One such database is GenBank.

GenBank ® is the NIH genetic sequence database, an annotated collection of all publicly available DNA sequences (Nucleic Acids Research, 2013 Jan;41(D1):D36-42).

The question that must be asked is why don't they sequence the full entire genome of the virus rather than piecing it together from fragments? How can you determine the genome of a thing if you don't isolate (or separate that thing) from all other things in the patient sample? Even if you separate out the bacteria, yeasts, fungi, etc. with high speed centrifugation how do you know that any fragments that remain are only virus fragments? They could be fragments from human cells, monkey cells, bovine cells, yeast cells, bacterial cells, etc.

So according to the usual definition of isolation, no they have not isolated it. If someone knows of a paper in the literature that describes isolation, purification and sequencing the entire genome without using metagenomic software to assemble or align from fragments, I would like to see it.

Questions

• Why do they always have to culture the virus? Wouldn’t there be enough virus particles on a mask that an infected person wears for a few minutes, an hour? If they can map a human’s DNA from a mouth swab, why would this be any different? Culturing could possibly be useful to try and determine viral behavior, but that is riddled with problems and assumptions

• A cell culture in a lab is missing blood circulation, the microbiome, the electrical currents in the body, the pH of the body, the temperature, the ratio of water, access to the mineral stores, the hormonal system, and probably other factors we don’t know about. It is assumed that the cell will respond in vitro (in a test tube or outside the body) the same way as it would in vivo (in the body)

• The fetal bovine serum is intended to promote rapid cell growth and the gentamicin is intended to prevent growth of bacteria. Gentamicin is toxic to kidney cells. This would stress the cells. They are also both substances that would not be found in the body, so in addition to the previously mentioned factors, these push the experiment even further from the conditions found in the body

• Why do they use kidney cells of an African green monkey? Why not use cells from human lung tissue if this is a respiratory virus that is highly infectious?