A series of COVID-19 autopsies with clinical and pathologic comparisons to both seasonal and pandemic influenza

Abstract

Autopsies of patients who have died from COVID-19 have been crucial in delineating patterns of injury associated with SARS-CoV-2 infection. Despite their utility, comprehensive autopsy studies are somewhat lacking relative to the global burden of disease, and very few comprehensive studies contextualize the findings to other fatal viral infections. We developed a novel autopsy protocol in order to perform postmortem examinations on victims of COVID-19 and herein describe detailed clinical information, gross findings, and histologic features observed in the first 16 complete COVID-19 autopsies. We also critically evaluated the role of ancillary studies used to establish a diagnosis of COVID-19 at autopsy, including immunohistochemistry (IHC), in situ hybridization (ISH), and electron microscopy (EM). IHC and ISH targeting SARS-CoV-2 were comparable in terms of the location and number of infected cells in lung tissue; however, nonspecific staining of bacteria was seen occasionally with IHC. EM was unrevealing in blindly sampled tissues. We then compared the clinical and histologic features present in this series to six archival cases of fatal seasonal influenza and six archival cases of pandemic influenza from the fourth wave of the 'Spanish Flu' in the winter of 1920. In addition to routine histology, the inflammatory infiltrates in the lungs of COVID-19 and seasonal influenza victims were compared using quantitative IHC. Our results demonstrate that the clinical and histologic features of COVID-19 are similar to those seen in fatal cases of influenza, and the two diseases tend to overlap histologically. There was no significant difference in the composition of the inflammatory infiltrate in COVID-19 and influenza at sites of acute lung injury at the time of autopsy. Our study underscores the relatively nonspecific clinical features and pathologic changes shared between severe cases of COVID-19 and influenza, while also providing important caveats to ancillary methods of viral detection.

Keywords: COVID-19; autopsy; diffuse alveolar damage; influenza; viral pneumonia; viral sepsis.

Figure 1

Representative gross and microscopic findings from COVID‐19 autopsies. (A) Spectrum of representative gross changes including diffuse consolidation and variable amounts of hemorrhage. (B,C) DAD (×10 and ×100 magnification, respectively). (D) Interstitial capillary with inflammation (×400) and detachment of alveolar pneumocytes. (E) Hemorrhage (×10 magnification). (F) Thrombosis (×10 magnification). (G) Infarction (×10 magnification). (H) Reactive, detached pneumocytes (×200 magnification). (I) Superimposed bacterial pneumonia (×100 magnification). All magnifications represent total magnification.

Figure 2

Assessment of IHC and ISH for the identification of SARS‐CoV‐2‐infected cells. (A–C) Positive control cell block made of SARS‐CoV‐2‐infected VERO cells, with hematoxylin and eosin (A, ×200 magnification), IHC using the Thermo‐Fisher Scientific monoclonal antibody (B, ×200 magnification), and RNAScope ISH (C, ×200 magnification). (D) Paired region analysis showing the distribution of virus‐infected cells is comparable between IHC (upper panel) and ISH (lower panel) (×100 magnification). (E) Paired region analysis showing that histopathologic changes associated with COVID‐19 (upper panel) may be negative for virus‐infected cells by IHC (lower panels) (×20 magnification). (F) Paired region analysis showing relatively bland areas (upper panel) may have many virus‐infected cells by IHC (lower panel) (F, ×20 magnification; inset ×200 magnification). All magnifications represent total magnification.

Figure 3

The nonspecific cytomorphology of SARS‐CoV‐2‐infected cells in human tissues and demonstration of cross‐reactivity with IHC reagents. (A) Hematoxylin and eosin (H&E)‐stained section (left) with paired IHC (right) of virus‐infected cells showing cytoplasmic vacuolization and reactive nuclear changes in infected cells (×600 magnification). Tissue dissected from this IHC‐positive region of the paraffin block demonstrates candidate virion‐like particles (inset, EM, ×80,000). (B) IHC of a group of cells from the same case (left H&E, right IHC) with identical cytomorphologic features which are negative for virus by IHC (×400 magnification). (C–E) Demonstration of nonspecific staining of bacteria by IHC in a case of necrotizing bacterial pneumonia and influenza from 2015. (C) H&E‐stained image of the region of positive staining (×100 magnification). (D) Positive staining of bacteria and bacteria‐laden macrophages by IHC (×200 magnification). (E) Negative ISH staining of the same region (×200 magnification). All magnifications represent total magnification.

Figure 4

Representative histologic findings in cases of fatal seasonal influenza taken from archival specimens. (A) DAD histologically indistinguishable from COVID‐19 (×50 magnification). (B) Interstitial inflammation with reactive, detached pneumocytes (×400 magnification). (C) Reactive, detached pneumocytes in a background of alveolar hemorrhage (×200 magnification). (D) Thrombosis (×50 magnification). (E) Infarction (×50 magnification). (F) Superimposed bacterial pneumonia (×200 magnification; note the presence of prominent and occasionally multinucleated pneumocytes).

Figure 5

Representative histologic findings in cases of fatal pandemic influenza taken from archival specimens. (A,B) DAD (×100 magnification). (C) DAD, edema, and hemorrhage adjacent to an airway containing inflammatory mucus (×100 magnification). (D) Interstitial capillary inflammation (×200 magnification). (E) Reactive pneumocytes exhibiting nucleoli and cytoplasmic vacuoles, similar to those seen in seasonal influenza and COVID‐19 (×400 magnification). (F) Superimposed bacterial pneumonia (×100 magnification).

Figure 6

Quantitative assessment of inflammation in COVID‐19 and seasonal influenza. Representative paired regions showing hematoxylin and eosin (H&E) staining along with CD20, CD8, and CD3 immunostains (clockwise from H&E) for (A) influenza and (B) COVID‐19. The distribution of T‐cells and relative abundance of CD8‐positive cells relative to CD3 is comparable. (C,D) Representative ROI analysis for CD3 and CD8 immunostains in a case of (C) fatal influenza and (D) COVID‐19. (E,F) Graphical representation of individual CD4:CD8 ratios per ROI (E) and cumulative CD4:CD8 ratios per patient (F). Note that there is no significant difference of CD4:CD8 ratios between COVID‐19 and influenza, although there is a trend toward higher CD4:CD8 for COVID‐19. In Figure 1E, individual patients are identified by a unique shape on the graph for each disease.