SARS-CoV-2 infection in hamsters and humans results in lasting and unique systemic perturbations post recovery (2022, Frere et al)

SNT Gatchaman

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SARS-CoV-2 infection in hamsters and humans results in lasting and unique systemic perturbations post recovery
Justin J. Frere, Randal A. Serafini, Kerri D. Pryce, Marianna Zazhytska, Kohei Oishi, Ilona Golynker, Maryline Panis, Jeffrey Zimering, Shu Horiuchi, Daisy A. Hoagland, Rasmus Møller, Anne Ruiz, Albana Kodra, Jonathan B. Overdevest, Peter D. Canoll, Alain C. Borczuk, Vasuretha Chandar, Yaron Bram, Robert Schwartz, Stavros Lomvardas, Venetia Zachariou, Benjamin R. tenOever

The host response to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection can result in prolonged pathologies collectively referred to as post-acute sequalae of COVID-19 (PASC) or long COVID.

To better understand the mechanism underlying long COVID biology, we compared the short- and long-term systemic responses in the golden hamster following either SARS-CoV-2 or influenza A virus (IAV) infection.

Results demonstrated that SARS-CoV-2 exceeded IAV in its capacity to cause permanent injury to the lung and kidney and uniquely impacted the olfactory bulb (OB) and epithelium (OE). Despite a lack of detectable infectious virus, the OB and OE demonstrated myeloid and T cell activation, proinflammatory cytokine production, and an interferon response that correlated with behavioral changes extending a month post viral clearance.

These sustained transcriptional changes could also be corroborated from tissue isolated from individuals who recovered from COVID-19. These data highlight a molecular mechanism for persistent COVID-19 symptomology and provide a small animal model to explore future therapeutics.

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Pyrrhus

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Benjamin tenOever and colleagues have been working very hard over the last two years to generate this animal model of Long Covid. They are to be applauded for their hard work and diligence.

However, there is a cloud that has lingered over their research.

When they began their research, Benjamin tenOever publicly declared that the coronavirus was incapable of persisting in a human body for more than two weeks. So, even before they had established their animal model of Long Covid, they had ruled out viral persistence as a potential factor in Long Covid.

Their preliminary results seemed to confirm tenOever's public statement: they did not find infectious virus in their hamsters after 2 weeks. But they did find sub-genomic viral RNA, which tenOever dismissed as "viral remnants", and not indicative of viral persistence.

I asked tenOever if they had used an RNA stabilizer to preserve the viral RNA, since there was no mention in their preliminary results that they had made an attempt to preserve the full genome of viral RNA. Perhaps they did the tissue homogenization, RNA extraction, and RT-PCR within minutes of killing the hamsters, in which case there would be no need for an RNA stabilizer.

As expected I did not receive a reply, but tenOever publicly stated that they would run a parallel experiment to confirm or refute the possibility of viral persistence. In this parallel experiment, they would administer molnupiravir, an anti-coronaviral drug, to a second set of hamsters a few days after infection. If the sub-genomic viral RNA indeed represented viral persistence, then the molnupiravir group should show no sub-genomic viral RNA after 2 weeks, unlike the untreated group.

When the NIH found out about this parallel experiment, they pulled tenOever's funding because the NIH program officer said that it was absurd to look for viral persistence after two weeks. When tenOever went public about the NIH's action, people were confused and asked tenOever to clarify: did the NIH refuse to fund the experiment in the first place, or did the NIH fund the experiment and then retract the funding once they found out that the experiment was designed to look for viral persistence?

Benjamin tenOever was clear: the NIH agreed to fund the experiment and then later retracted the funding once they found out that the experiment was designed to look for viral persistence. The current fate of this parallel experiment remains unclear.

In the current paper, this is the problematic part:
Frere et al. 2022 said:
Collected tissues were homogenized with PBS or Trizol (Thermo Fisher Scientific) in Lysing Matrix A homogenization tubes (MP Biomedicals) for 40 s at 6 m/s for 2 cycles in a FastPrep 24 5G bead grinder and lysis system (MP Biomedicals) for plaque assay or RNA isolation, respectively. Additional tissues were fixed in 4% PFA for at least 72 hours prior to embedding in paraffin wax blocks for histology. [...] RNA was isolated from homogenized samples by TRIzol/phenol-chloroform extraction.

Unfortunately, there is no mention of how much time elapsed between killing the hamsters and tissue homogenization, nor is there any mention of an RNA stabilizing agent. (Unless you consider Trizol to be a sufficiently stabilizing agent.)

Other than the above, it's a very impressive body of research.
 

SNT Gatchaman

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When they began their research, Benjamin tenOever publicly declared that the coronavirus was incapable of persisting in a human body for more than two weeks. So, even before they had established their animal model of Long Covid, they had ruled out viral persistence as a potential factor in Long Covid.
That seems like an unfortunate preconception, particularly when studies in the last two years are showing evidence of viral persistence in gut and brain (your posts :)).

Also, why even start with that idea? Isn't it better to assume nothing? Surely the approach should be to 1) make an animal model to see what is happening and 2) progressively refine experiments to see why it's happening.
 

SNT Gatchaman

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Benjamin tenOever was clear: the NIH agreed to fund the experiment and then later retracted the funding once they found out that the experiment was designed to look for viral persistence.
As quoted in the Atlantic's article from May 22 —

Benjamin tenOever, a virologist at NYU, told me that he recently had NIH funding pulled from a project that would have investigated whether antivirals could combat long-COVID symptoms in a hamster model. His contact at the agency said the study had “no merit,” tenOever told me. “They were like, This doesn’t make sense, because why would Paxlovid ever help long COVID? The virus is long gone.