Research Review: Mapping SARS-CoV-2 RNA in the Human Body

The Question

Where does SARS-CoV-2 migrate throughout the human body and how is this linked to tissue damage?

As of January 2021, over 92 million people have been infected with COVID-19 globally, resulting in nearly 2 million deaths (https://coronavirus.jhu.edu/). Although most recover, 14% suffer from respiratory distress¹ and 5% become critically ill with diverse complications involving one or multiple organ systems—suggesting COVID-19 is a systemic disease.

Aside from understanding major biological players involved in infection (e.g., host receptor ACE2 and viral Spike protein), the mechanism of viral replication and pathogenesis of COVID-19 are widely unknown. One crucial step needed to elucidate viral pathogenesis in severe cases is understanding the distribution of the virus throughout the entire body.

The Idea

Deinhardt-Emmer et al. hypothesize that, in critically ill patients, SARS-CoV-2 infects multiple organs and that tissue damage increases with organ-specific viral load.

To investigate this idea, Deinhardt-Emmer et al. first had to address two study design challenges:

1. Ensure viral RNA (vRNA) remained intact postmortem prior to analysis
2. Optimize sampling to minimize false negatives

The solutions:

Use very early post-mortem autopsies to reliably analyze COVID-19 RNA, obtain blood serum, and collect tissue samples for ultrastructural analysis.

In previous studies, the time between death and autopsy ranged from 48 h to 5 days. Here, the researchers performed autopsies within 1.5-15 h (5.6 h on average) to minimize vRNA degradation prior to analysis.

Also in previous studies, tissue samples were often limited to only one or a few organs. Here, the researchers took multiple samples per organ for multiple organs, totaling 61 samples per patient. The idea is that the distribution of vRNA is not the same throughout a single organ or organ system, resulting in a different picture depending on where the sample is taken from and making the study vulnerable to false negatives. Robust sampling offers higher resolution of where vRNA is located and minimizes false negatives.

Further, getting a more accurate and thoroughly mapped picture of virus distribution enabled the researchers to see how viral load influenced tissue damage by evaluating organ-specific micro- and macromorphology via comprehensive histopathological investigations.

The Findings

Fascinatingly, you can literally get a front-seat view of the virus particles encased in plasmatic vesicles of alveolar fibrocytes, as the researchers visualized vRNA in lung tissue using transmission electron microscopy.

Patients presented different patterns of vRNA distribution and abundance (n=11; 10 COVID-19-related deaths).

• All patients (with COVID-related death) showed moderate to high levels of vRNA in the lungs (reaching up to 10⁷ RNA copies/ml); low to moderate levels in the lymphatic system; and levels below the detectable limit in the skin, subcutaneous tissue, and skeletal muscle.
• For two patients, vRNA was almost exclusively found in the respiratory system and cervical and hilar lymph nodes (structures topologically related to the respiratory tract).
• Three patients had low to very high vRNA levels in almost every tissue type sampled.
• One individual had high to very high levels of vRNA in almost all organs/systems sampled, but levels were below the detectable limit in the blood.

COVID-19 infection ramped up the proinflammatory and prothrombotic response compared to healthy individuals—This effect was present in all patients (based on post-mortem serum analysis).

Tissue damage was not always present in high-viral load regions.

• The researchers found signs of severe lung damage and a loss of lymph node follicular architecture in all patients, linking tissue damage to viral load.
• In the bone marrow of patient 9, the highest viral loads were found, and significant hemophagocytosis was detectable by microscopy.
• The correlation of high viral load and tissue damage was not found in cardiac tissues or other extra-pulmonary tissues.

The Future

Further work is needed to elucidate SARS-CoV-2 pathological mechanisms, particularly related to replication capabilities in different organs, and the variability between patients to better treat the disease, predict clinical outcomes, and ultimately fight the pandemic and save lives.

Technical Highlight

All 671 tissues were homogenized in RPMI-medium by using the FastPrep-24^TM 5G Instrument. Importantly, homogenization was rapid and thorough enough to efficiently process hundreds of samples while keeping viral particles intact.

Researchers processing tissue samples can utilize MP Bio's FastPrep Instruments and Lysing Matrices with the confidence that their projects are streamlined and their samples are ready to use in downstream analyses.

References:

1. Wu, Z. & McGoogan, J. M. Characteristics of and Important Lessons From the Coronavirus Disease 2019 (COVID-19) Outbreak in China: Summary of a Report of 72□314 Cases From the Chinese Center for Disease Control and Prevention. JAMA 323, 1239–1242, doi: 10.1001/jama.2020.2648 (2020).