Sigh, yes, the ‘COVID virus’ is real

There has been talk out thar in the wildlands we call Twitter that people can’t find evidence that the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) – also known as ‘the COVID virus’ – is real. Has it ever been isolated in cell culture, visualised by electron microscopy, reacted with antibodies, genetically sequence and otherwise characterised in many samples collected from people with coronavirus disease 19 (COVID-19) all over the world in during the past nine months? Sigh, yes, the ‘COVID virus’ is real. Here are a few of the scientific endeavours that show this virus has been isolated from clinically diagnosed, ill and laboratory-confirmed human COVID-19 cases.

This illustration, created at the Centers for Disease Control and Prevention (CDC), reveals ultrastructural morphology exhibited by coronaviruses. Note the spikes that adorn the outer surface of the virus, which impart the look of a corona surrounding the virion, when viewed electron microscopically. A novel coronavirus, named Severe Acute Respiratory Syndrome coronavirus 2 (SARS-CoV-2), was identified as the cause of an outbreak of respiratory illness first detected in Wuhan, China in 2019. The illness caused by this virus has been named coronavirus disease 2019 (COVID-19).

January 2020 [1]

Peng Zhou and colleagues from the Wuhan Institute of Virology grew a novel coronavirus (later to be called SARS-CoV-2) in cell culture (isolated) from the bronchoalveolar lavage fluid collected from an ill and RT-PCR-positive female patient (ICU-06 in Spike tree=WIV04/2019 virus).

Extended Table 2. The samples from ICU-06 had some of the highest viral loads (estimated from the low threshold cycle values [CT]) among the patients tested.

The virus was called nCoV-2019 BetaCoV/Wuhan/WIV04/2019 and grew in Vero E6 and Huh7 cells. Virus-induced cellular changes (cytopathic effect or CPE) were observed.

Vero E6 cells are shown at 24 h after infection with (a) mock virus) or (b) SARS-CoV-2.
Mock-virus-infected (c) or SARS-CoV-2-infected (d) samples were stained with rabbit serum raised against recombinant SARSr-CoV Rp3N protein (red; able to cross react) and DAPI (blue; a counterstain). The experiment was conducted twice independently but with similar results.

Whole genome sequencing (WGS) was used to identify the unique genetic sequence of the cultured virus and a specific real-time reverse transcription-polymerase chain reaction (RT-rPCR) designed to allow screening of more sensitive and rapid screening of more samples. The RT-rPCR was tested against human endemic CoVs (229E, OC43, HKU1) as well as MERS-CoV, SARS-CoV and others, and was found to be highly specific for SARS-CoV-2.

The authors also saw viral particles. Electron microscopy (EM) was used to visualise virus in cultured Vero E6 cells after they had been inoculated with the patent sample.

Viral particles shown in ultrathin cut sections of prepared virus-positive Vero E6 cell cultures, using electron microscopy at 200 kV. The inset shows the viral particles in an intra-cytosolic vacuole, likley on their way to thw surface and out of the infected cell.[1]

They also showed that the antibodies from five infected people could neutralise (=prevent) infection of Vero E6 after incubated dilutions of patient sera with a cultured virus preparation and added to cultures of uninfected cells. An antibody made in horses against the original SARS-CoV could also neutralise the virus however serum from two healthy people in Wuhan could not.

February 2020 [2]

Jeong-Min Kim and colleagues from South Korea inoculated RT-rPCR-positive patient nasopharyngeal and oropharyngeal samples onto Vero cells. After most inoculated (but not mock-inoculated) cells were showing CPE, they were harvested and tested using the two RT-rPCRs (Corman et al. RdRp and E; these detect viruses in the genus Sarbecovirus). RT-rPCR testing found an increase in viral RNA which the team estimated at 10-70-fold.

The study also examined three-day post-inoculation cells using EM. They found virus-like particles in vesicles within the infected cells.

To confirm that the virus was indeed SARS-CoV-2, the authors conducted WGS, naming the SARS-CoV-2 variant they’d isolated, BetaCoV/Korea/KCDC03/2020

February [4]

This is a method for culturing SARS-CoV-2 from patients samples. It doesn’t delve into virus characterisation though. Nonetheless, the image below shows CPE on a monolayer (=single cell layer) of African green monkey kidney Vero C1008, clone E6 cells (ATCC®-CRL-1586).

An example of a SARS-CoV-2-infected monolayer culture of Vero E6 cells demonstrating focal CPE. Source: Dr. Alyssa Pyke, Public Health Virology Laboratory, Forensic and Scientific Services, Queensland. 08FEB2020

March 2020 [3]

Leon Caly and a crew from Australia isolated the virus from an ill traveller from Wuhan. The virus was detected using an in-house (designed by this team) RT-rPCR. The nasopharyngeal swab was used to inoculate the Vero/SLAM cell line.

Growth was seen both by the CPE induced and the use of that specific RT-rPCR to show an increase in the presence of SARS-CoV-2 RNA (decrease in CT)

Cell-free liquid from infected cultures (called supernatant) and cells from the cultures were each examined using EM, finding particles with characteristic coronavirus morphology (=shape) and the same particles inside vesicles within the cells.

To further characterise the amplified agent, WGS identified the genome and names this virus BetaCoV/Australia/VIC/01/2020.

Evidence of SARS-CoV-2 isolation and visualisation in other ways

Below is a brief list of some major feel-good SARS-CoV-2 discovery announcements that came to us through the mainstream media or University websites.

  1. Melbourne scientists first to grow and share novel coronavirus
    https://about.unimelb.edu.au/newsroom/news/2020/january/melbourne-scientists-first-to-grow-and-share-novel-coronavirus
  2. China coronavirus: Hong Kong researchers have already developed vaccine but need time to test it, expert reveals
    https://www.scmp.com/news/hong-kong/health-environment/article/3047956/china-coronavirus-hong-kong-researchers-have
  3. China CDC developing novel coronavirus vaccine
    http://www.xinhuanet.com/english/2020-01/26/c_138734908.htm
  4. Coronavirus: Scientists isolate virus responsible for deadly Covid-19 outbreak
    https://www.independent.co.uk/news/science/coronavirus-covid-19-virus-isolate-canada-scientists-mutations-strains-a9399226.html
  5. I study viruses: How our team isolated the new coronavirus to fight the global pandemic
    https://theconversation.com/i-study-viruses-how-our-team-isolated-the-new-coronavirus-to-fight-the-global-pandemic-133675

Stock free-to-use images of the SARS-CoV-2 virus

These images are made public by the US Centers for Disease Control ND prevention via their Public Health Image Library (PHIL). A fantastic resource for virus images. The exact details of how the viruses were prepared aren’t present, nor the ways in which SARS-CoV-2 was confirmed. that may be a problem for you. However, I’m sure they were prepared and confirmed using the exact same methods as those listed above.

Transmission electron microscopic image of an isolate from the first U.S. case of COVID-19, formerly known as 2019-nCoV (SARS-CoV-2). The spherical viral particles, colourized blue, contain cross-sections through the viral genome, seen as black dots.
Source: https://phil.cdc.gov/Details.aspx?pid=23354

Electron microscopic image of a negatively stained particle of SARS-CoV-2, causative agent of COVID-19. Note the prominent spikes from which the coronavirus gets its name for “corona”, or “crown-like”.
http://phil.cdc.gov/Details.aspx?pid=23640

Thin section electron microscopic image of SARS-CoV-2, the causative agent of COVID-19. Spherical virus particles contain black dots, which are cross-sections through the viral nucleocapsid. In the cytoplasm of the infected cell, clusters of particles are found within the membrane-bound cisternae of the rough endoplasmic reticulum/Golgi area.
https://phil.cdc.gov/Details.aspx?pid=23640

Electron microscopic image of a negatively stained particle of SARS-CoV-2, causative agent of COVID-19. Note the prominent spikes from which the coronavirus gets its name for “corona”, or “crown-like”.
Source: https://phil.cdc.gov/Details.aspx?pid=23641

Transmission electron microscopic image of an isolate from the first U.S. case of COVID-19, formerly known as 2019-nCoV. The spherical extracellular viral particles contain cross-sections through the viral genome, seen as black dots.
Source: https://phil.cdc.gov/Details.aspx?pid=23336

Some final thoughts

I know there will be comments below (I’ll make sure to publish some of the less offensive ones 😉) to the extent that ‘I don’t care what y’all say, I ain’t seen no evidence, make it empirical, abide by the Koch’…or something.

And that’s all well and good.

There is evidence and some of the most significant parts of it are listed above.

It convinces me. If it doesn’t convince you, there is nothing more I can do except ask what it would take to convince you and in the meantime assure you that you are wrong on this one. Yes, the ‘COVID virus’ is real.

References

  1. Discovery of a novel coronavirus associated with the recent pneumonia outbreak in humans and its potential bat origin
    https://www.biorxiv.org/content/10.1101/2020.01.22.914952v2 and then
    https://www.nature.com/articles/s41586-020-2012-7
  2. Identification of Coronavirus Isolated from a Patient in Korea with COVID-19
    https://ophrp.org/journal/view.php?doi=10.24171/j.phrp.2020.11.1.02
  3. Isolation and rapid sharing of the 2019 novel coronavirus (SARS‐CoV‐2) from the first patient diagnosed with COVID‐19 in Australia
    https://www.mja.com.au/journal/2020/212/10/isolation-and-rapid-sharing-2019-novel-coronavirus-sars-cov-2-first-patient
  4. Culture of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2; f.2019-nCoV)
    https://www.protocols.io/view/culture-of-the-severe-acute-respiratory-syndrome-c-bcduis6w?step=5

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