Severe COVID-19 patients develop acute respiratory distress syndrome that may progress to cytokine storm syndrome, organ dysfunction, and death. Considering that neutrophil extracellular traps (NETs) have been described as important mediators of tissue damage in inflammatory diseases, we investigated whether NETs would be involved in COVID-19 pathophysiology. A cohort of 32 hospitalized patients with a confirmed diagnosis of COVID-19 and healthy controls were enrolled. The concentration of NETs was augmented in plasma, tracheal aspirate, and lung autopsies tissues from COVID-19 patients, and their neutrophils released higher levels of NETs. Notably, we found that viable SARS-CoV-2 can directly induce the release of NETs by healthy neutrophils. Mechanistically, NETs triggered by SARS-CoV-2 depend on angiotensin-converting enzyme 2, serine protease, virus replication, and PAD-4. Finally, NETs released by SARS-CoV-2–activated neutrophils promote lung epithelial cell death in vitro. These results unravel a possible detrimental role of NETs in the pathophysiology of COVID-19. Therefore, the inhibition of NETs represents a potential therapeutic target for COVID-19.

The ability of the new coronavirus SARS‐CoV‐2 to spread and contaminate is one of the determinants of the COVID‐19 pandemic status. SARS‐CoV‐2 has been detected in saliva consistently, with similar sensitivity to that observed in nasopharyngeal swabs. We conducted ultrasound‐guided postmortem biopsies in COVID‐19 fatal cases. Samples of salivary glands (SGs; parotid, submandibular, and minor) were obtained. We analyzed samples using RT‐qPCR, immunohistochemistry, electron microscopy, and histopathological analysis to identify SARS‐CoV‐2 and elucidate qualitative and quantitative viral profiles in salivary glands. The study included 13 female and 11 male patients, with a mean age of 53.12 years (range 8–83 years). RT‐qPCR for SARS‐CoV‐2 was positive in 30 SG samples from 18 patients (60% of total SG samples and 75% of all cases). Ultrastructural analyses showed spherical 70–100 nm viral particles, consistent in size and shape with the Coronaviridae family, in the ductal lining cell cytoplasm, acinar cells, and ductal lumen of SGs. There was also degeneration of organelles in infected cells and the presence of a cluster of nucleocapsids, which suggests viral replication in SG cells. Qualitative histopathological analysis showed morphologic alterations in the duct lining epithelium characterized by cytoplasmic and nuclear vacuolization, as well as nuclear pleomorphism. Acinar cells showed degenerative changes of the zymogen granules and enlarged nuclei. Ductal epithelium and serous acinar cells showed intense expression of ACE2 and TMPRSS receptors. An anti‐SARS‐CoV‐2 antibody was positive in 8 (53%) of the 15 tested cases in duct lining epithelial cells and acinar cells of major SGs. Only two minor salivary glands were positive for SARS‐CoV‐2 by immunohistochemistry. Salivary glands are a reservoir for SARS‐CoV‐2 and provide a pathophysiological background for studies that indicate the use of saliva as a diagnostic method for COVID‐19 and highlight this biological fluid's role in spreading the disease. © 2021 The Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Background: Multi‐organ damage is a common feature of severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) infection, going beyond the initially observed severe pneumonia. Evidence that the testis is also compromised is growing.Objective: To describe the pathological findings in testes from fatal cases of COVID‐19, including the detection of viral particles and antigens, and inflammatory cell subsets.Materials and methods: Postmortem testicular samples were obtained by percutaneous puncture from 11 deceased men and examined by reverse‐transcription polymerase chain reaction (RT‐PCR) for RNA detection and by light and electron microscopy (EM) for SARS‐CoV‐2. Immunohistochemistry (IHC) for the SARS‐CoV‐2 N‐protein and lymphocytic and histiocytic markers was also performed.Results: Eight patients had mild interstitial orchitis, composed mainly of CD68+ and TCD8+ cells. Fibrin thrombi were detected in five cases. All cases presented congestion, interstitial edema, thickening of the tubular basal membrane, decreased Leydig and Sertoli cells with reduced spermatogenesis, and strong expression of vascular cell adhesion molecule (VCAM) in vessels. IHC detected SARS‐Cov‐2 antigen in Leydig cells, Sertoli cells, spermatogonia, and fibroblasts in all cases. EM detected viral particles in the cytoplasm of fibroblasts, endothelium, Sertoli and Leydig cells, spermatids, and epithelial cells of the rete testis in four cases, while RT‐PCR detected SARS‐CoV‐2 RNA in three cases.Discussion and conclusion: The COVID‐19‐associated testicular lesion revealed a combination of orchitis, vascular changes, basal membrane thickening, Leydig and Sertoli cell scarcity, and reduced spermatogenesis associated with SARS‐CoV‐2 local infection that may impair hormonal function and fertility in men.

The largest outbreak of yellow fever of the 21^st century in the Americas began in 2016, with intense circulation in the southeastern states of Brazil, particularly in sylvatic environments near densely populated areas including the metropolitan region of São Paulo city (MRSP) during 2017–2018. Herein, we describe the origin and molecular epidemiology of yellow fever virus (YFV) during this outbreak inferred from 36 full genome sequences taken from individuals who died following infection with zoonotic YFV. Our analysis revealed that these deaths were due to three genetic variants of sylvatic YFV that belong the South American I genotype and that were related to viruses previously isolated in 2017 from other locations in Brazil (Minas Gerais, Espírito Santo, Bahia and Rio de Janeiro states). Each variant represented an independent virus introduction into the MRSP. Phylogeographic and geopositioning analyses suggested that the virus moved around the peri-urban area without detectable human-to-human transmission, and towards the Atlantic rain forest causing human spill-over in nearby cities, yet in the absence of sustained viral transmission in the urban environment.

Although increasing evidence confirms neuropsychiatric manifestations associated mainly with severe COVID-19 infection, the long-term neuropsychiatric dysfunction has been frequently observed after mild infection. Here we show the spectrum of the cerebral impact of SARS-CoV-2 infection ranging from long-term alterations in mildly infected individuals (orbitofrontal cortical atrophy, neurocognitive impairment, excessive fatigue and anxiety symptoms) to severe acute damage confirmed in brain tissue samples extracted from the orbitofrontal region (via endonasal trans-ethmoidal approach) from individuals who died of COVID-19. We used surface-based analyses of 3T MRI and identified orbitofrontal cortical atrophy in a group of 81 mildly infected patients (77% referred anosmia or dysgeusia during acute stage) compared to 145 healthy volunteers; this atrophy correlated with symptoms of anxiety and cognitive dysfunction. In an independent cohort of 26 individuals who died of COVID-19, we used histopathological signs of brain damage as a guide for possible SARS-CoV-2 brain infection, and found that among the 5 individuals who exhibited those signs, all of them had genetic material of the virus in the brain. Brain tissue samples from these 5 patients also exhibited foci of SARS-CoV-2 infection and replication, particularly in astrocytes. Supporting the hypothesis of astrocyte infection, neural stem cell-derived human astrocytes in vitro are susceptible to SARS-CoV-2 infection through a non-canonical mechanism that involves spike-NRP1 interaction. SARS-CoV-2-infected astrocytes manifested changes in energy metabolism and in key proteins and metabolites used to fuel neurons, as well as in the biogenesis of neurotransmitters. Moreover, human astrocyte infection elicits a secretory phenotype that reduces neuronal viability. Our data support the model in which SARS-CoV-2 reaches the brain, infects astrocytes and consequently leads to neuronal death or dysfunction. These deregulated processes are also likely to contribute to the structural and functional alterations seen in the brains of COVID-19 patients.