In Situ Hybridization Confirms SARS-CoV-2 RNA in Retinal Tissues After COVID-19

A postmortem examination of eyes from patients with COVID-19 identified specific ocular abnormalities consistent with viral infection and, using in situ hybridization, was able to locate SARS-CoV-2 RNA in multiple neuronal cell layers of the retina. These discoveries confirm previous PCR-based findings and contribute valuable information to the growing database about the possible ocular impacts of SARS-CoV-2 infection.

Study Design

This is a retrospective study observing postmortem ocular tissues from 25 patients (25 eyes) with confirmed COVID-19 at autopsy between April 2020 and December 2021. The objectives were to evaluate macroscopic and microscopic ocular changes and to investigate the location of SARS-CoV-2 in these tissues. Prior to this study, SARS-CoV-2 nucleocapsid gene RNA had been quantified by droplet digital PCR (ddPCR) from one eye of each patient. In this study, the contralateral eyes of 21 patients were fixed in formalin for histopathologic examination. In situ hybridization of sections of the ddPCR-positive eyes from the 4 other patients was used to determine the cellular location of SARS-CoV-2 spike gene RNA.

Outcomes

Ninety-six percent of patients had at least 1 comorbidity; 60% had three or more. Many patients had cardiovascular comorbidities. Abnormalities suggestive of indirect viral effect—including cytoid bodies, vascular changes, and retinal edema—were observed in all 21 eyes that underwent histopathologic evaluation, irrespective of illness duration before death. All 21 eyes had minimal or no inflammation in ocular tissues, although 67% of the contralateral eyes were positive for SARS-CoV-2 by ddPCR. Platelet thrombi, sometimes associated with fibrin and mononuclear cells, were observed in the retinal vascular wall in 5 eyes, all of which had cytoid bodies in the retinal nerve fiber layer and positive ddPCR in the contralateral eye. One eye also showed retinal vascular occlusion and neovascularization. In situ hybridization localized SARS-CoV-2 RNA to neuronal cells of the retinal inner and outer layers, ganglion cells, corneal epithelium, scleral fibroblasts, and oligodendrocytes of the optic nerve. No cytomegalovirus positivity was detected in any patient.

Limitations

Since ocular history and clinical examinations from the donors were not available, it is not known whether any of these observed changes preceded SARS-CoV-2 infection. The diverse comorbidities of the donor cohort could contribute to the observed ocular pathology. Consequently, it is difficult to draw clear conclusions about whether COVID-19, the host immune response, and/or preexisting comorbidities were the cause of the observed pathology.

Clinical Significance

The study uncovers important new insights into SARS-CoV-2 ocular pathogenesis. This study reflects the first reported localization of SARS-CoV-2 to the retinal inner and outer nuclear cells, retinal ganglion cells, and ocular surface by in situ hybridization, validating previous studies that have exclusively used PCR-based methods. These observations also highlight the need to better understand the mechanisms of SARS-CoV-2 infection and persistence in the eye, associated pathogenesis, and long-term ocular sequelae among COVID-19 survivors.

Financial Disclosures: Dr. Maya Eiger-Moscovich discloses no financial relationships.