SARS-CoV-2 Omicron (B.1.1.529) infection in rhesus macaques, hamsters, and BALB/c mice with severe lung histopathological damage.

Since the first SARS-CoV-2 outbreak in late 2019, the SARS-CoV-2 genome has harbored multiple mutations, especially spike protein mutations. The currently fast-spreading Omicron variant that manifests without symptoms or with upper respiratory diseases has been recognized as a serious global public health problem. However, its pathological mechanism is largely unknown. In this work, rhesus macaques, hamsters, and BALB/C mice were employed as animal models to explore the pathogenesis of Omicron (B.1.1.529). Notably, Omicron (B.1.1.529) infected the nasal turbinates, tracheae, bronchi, and lungs of hamsters and BALB/C mice with higher viral loads than in those of rhesus macaques. Severe histopathological damage and inflammatory responses were observed in the lungs of Omicron (B.1.1.529)-infected animals. In addition, viral replication was found in multiple extrapulmonary organs. Results indicated that hamsters and BALB/c mice are potential animal models for studies on the development of drugs/vaccines and therapies for Omicron (B.1.1.529).

Aryl hydrocarbon receptor is a proviral host factor and a candidate pan-SARS-CoV-2 therapeutic target.

The emergence of a series of SARS-CoV-2 variants has necessitated the search for broad-spectrum antiviral targets. The aryl hydrocarbon receptor (AhR) senses tryptophan metabolites and is an immune regulator. However, the role of AhR in SARS-CoV-2 infection and whether AhR can be used as the target of antiviral therapy against SARS-CoV-2 and its variants are yet unclear. Here, we show that infection with SARS-CoV-2 activates AhR signaling and facilitates viral replication by interfering with IFN-I-driven antiviral immunity and up-regulating ACE2 receptor expression. The pharmacological AhR blockade or AhR knockout reduces SARS-CoV-2 and its variants' replication in vitro. Drug targeting of AhR with AhR antagonists markedly reduced SARS-CoV-2 and its variants' replication in vivo and ameliorated lung inflammation caused by SARS-CoV-2 infection in hamsters. Overall, AhR was a SARS-CoV-2 proviral host factor and a candidate host-directed broad-spectrum target for antiviral therapy against SARS-CoV-2 and its variants, including Delta and Omicron, and potentially other variants in the future.

Aggravated pneumonia and diabetes in SARS-CoV-2 infected diabetic mice.

Multiple clinical and epidemiological studies have shown an interconnection between coronavirus disease 2019 (COVID-19) and diabetes, but experimental evidence is still lacking. Understanding the interplay between them is important because of the global health burden of COVID-19 and diabetes. We found that C57BL/6J mice were susceptible to the alpha strain of SARS-CoV-2. Moreover, diabetic C57BL/6J mice with leptin receptor gene deficiency (db/db mice) showed a higher viral load in the throat and lung and slower virus clearance in the throat after infection than C57BL/6J mice. Histological and multifactor analysis revealed more advanced pulmonary injury and serum inflammation in SARS-CoV-2 infected diabetic mice. Moreover, SARS-CoV-2 infected diabetic mice exhibited more severe insulin resistance and islet cell loss than uninfected diabetic mice. By RNA sequencing analysis, we found that diabetes may reduce the collagen level, suppress the immune response and aggravate inflammation in the lung after infection, which may account for the greater susceptibility of diabetic mice and their more severe lung damage after infection. In summary, we successfully established a SARS-CoV-2 infected diabetic mice model and demonstrated that diabetes and COVID-19 were risk factors for one another.

Histones released by NETosis enhance the infectivity of SARS-CoV-2 by bridging the spike protein subunit 2 and sialic acid on host cells.

Neutrophil extracellular traps (NETs) can capture and kill viruses, such as influenza viruses, human immunodeficiency virus (HIV), and respiratory syncytial virus (RSV), thus contributing to host defense. Contrary to our expectation, we show here that the histones released by NETosis enhance the infectivity of SARS-CoV-2, as found by using live SARS-CoV-2 and two pseudovirus systems as well as a mouse model. The histone H3 or H4 selectively binds to subunit 2 of the spike (S) protein, as shown by a biochemical binding assay, surface plasmon resonance and binding energy calculation as well as the construction of a mutant S protein by replacing four acidic amino acids. Sialic acid on the host cell surface is the key molecule to which histones bridge subunit 2 of the S protein. Moreover, histones enhance cell-cell fusion. Finally, treatment with an inhibitor of NETosis, histone H3 or H4, or sialic acid notably affected the levels of sgRNA copies and the number of apoptotic cells in a mouse model. These findings suggest that SARS-CoV-2 could hijack histones from neutrophil NETosis to promote its host cell attachment and entry process and may be important in exploring pathogenesis and possible strategies to develop new effective therapies for COVID-19.

Three doses of prototypic SARS-CoV-2 inactivated vaccine induce cross-protection against its variants of concern.

Variants are globally emerging very quickly following pandemic prototypic SARS-CoV-2. To evaluate the cross-protection of prototypic SARS-CoV-2 vaccine against its variants, we vaccinated rhesus monkeys with three doses of prototypic SARS-CoV-2 inactivated vaccine, followed by challenging with emerging SARS-CoV-2 variants of concern (VOCs). These vaccinated animals produced neutralizing antibodies against Alpha, Beta, Delta, and Omicron variants, although there were certain declinations of geometric mean titer (GMT) as compared with prototypic SARS-CoV-2. Of note, in vivo this prototypic vaccine not only reduced the viral loads in nasal, throat and anal swabs, pulmonary tissues, but also improved the pathological changes in the lung infected by variants of Alpha, Beta, and Delta. In summary, the prototypic SARS-CoV-2 inactivated vaccine in this study protected against VOCs to certain extension, which is of great significance for prevention and control of COVID-19.

The olfactory route is a potential way for SARS-CoV-2 to invade the central nervous system of rhesus monkeys.

Neurological manifestations are frequently reported in the COVID-19 patients. Neuromechanism of SARS-CoV-2 remains to be elucidated. In this study, we explored the mechanisms of SARS-CoV-2 neurotropism via our established non-human primate model of COVID-19. In rhesus monkey, SARS-CoV-2 invades the CNS primarily via the olfactory bulb. Thereafter, viruses rapidly spread to functional areas of the central nervous system, such as hippocampus, thalamus, and medulla oblongata. The infection of SARS-CoV-2 induces the inflammation possibly by targeting neurons, microglia, and astrocytes in the CNS. Consistently, SARS-CoV-2 infects neuro-derived SK-N-SH, glial-derived U251, and brain microvascular endothelial cells in vitro. To our knowledge, this is the first experimental evidence of SARS-CoV-2 neuroinvasion in the NHP model, which provides important insights into the CNS-related pathogenesis of SARS-CoV-2.

The Gastrointestinal Tract Is an Alternative Route for SARS-CoV-2 Infection in a Nonhuman Primate Model.

BACKGROUND & AIMS: Gastrointestinal (GI) manifestations have been increasingly reported in patients with coronavirus disease 2019 (COVID-19). However, the roles of the GI tract in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection are not fully understood. We investigated how the GI tract is involved in SARS-CoV-2 infection to elucidate the pathogenesis of COVID-19. METHODS: Our previously established nonhuman primate (NHP) model of COVID-19 was modified in this study to test our hypothesis. Rhesus monkeys were infected with an intragastric or intranasal challenge with SARS-CoV-2. Clinical signs were recorded after infection. Viral genomic RNA was quantified by quantitative reverse transcription polymerase chain reaction. Host responses to SARS-CoV-2 infection were evaluated by examining inflammatory cytokines, macrophages, histopathology, and mucin barrier integrity. RESULTS: Intranasal inoculation with SARS-CoV-2 led to infections and pathologic changes not only in respiratory tissues but also in digestive tissues. Expectedly, intragastric inoculation with SARS-CoV-2 resulted in the productive infection of digestive tissues and inflammation in both the lung and digestive tissues. Inflammatory cytokines were induced by both types of inoculation with SARS-CoV-2, consistent with the increased expression of CD68. Immunohistochemistry and Alcian blue/periodic acid-Schiff staining showed decreased Ki67, increased cleaved caspase 3, and decreased numbers of mucin-containing goblet cells, suggesting that the inflammation induced by these 2 types of inoculation with SARS-CoV-2 impaired the GI barrier and caused severe infections. CONCLUSIONS: Both intranasal and intragastric inoculation with SARS-CoV-2 caused pneumonia and GI dysfunction in our rhesus monkey model. Inflammatory cytokines are possible connections for the pathogenesis of SARS-CoV-2 between the respiratory and digestive systems.

Susceptibility of tree shrew to SARS-CoV-2 infection.

Since severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) became a pandemic event in the world, it has not only caused huge economic losses, but also a serious threat to global public health. Many scientific questions about SARS-CoV-2 and Coronavirus disease (COVID-19) were raised and urgently need to be answered, including the susceptibility of animals to SARS-CoV-2 infection. Here we tested whether tree shrew, an emerging experimental animal domesticated from wild animal, is susceptible to SARS-CoV-2 infection. No clinical signs were observed in SARS-CoV-2 inoculated tree shrews during this experiment except the increasing body temperature particularly in female animals. Low levels of virus shedding and replication in tissues occurred in all three age groups. Notably, young tree shrews (6 months to 12 months) showed virus shedding at the earlier stage of infection than adult (2 years to 4 years) and old (5 years to 7 years) animals that had longer duration of virus shedding comparatively. Histopathological examine revealed that pulmonary abnormalities were the main changes but mild although slight lesions were also observed in other tissues. In summary, tree shrew is less susceptible to SARS-CoV-2 infection compared with the reported animal models and may not be a suitable animal for COVID-19 related researches. However, tree shrew may be a potential intermediate host of SARS-CoV-2 as an asymptomatic carrier.

Comparison of nonhuman primates identified the suitable model for COVID-19.

Identification of a suitable nonhuman primate (NHP) model of COVID-19 remains challenging. Here, we characterized severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in three NHP species: Old World monkeys Macaca mulatta (M. mulatta) and Macaca fascicularis (M. fascicularis) and New World monkey Callithrix jacchus (C. jacchus). Infected M. mulatta and M. fascicularis showed abnormal chest radiographs, an increased body temperature and a decreased body weight. Viral genomes were detected in swab and blood samples from all animals. Viral load was detected in the pulmonary tissues of M. mulatta and M. fascicularis but not C. jacchus. Furthermore, among the three animal species, M. mulatta showed the strongest response to SARS-CoV-2, including increased inflammatory cytokine expression and pathological changes in the pulmonary tissues. Collectively, these data revealed the different susceptibilities of Old World and New World monkeys to SARS-CoV-2 and identified M. mulatta as the most suitable for modeling COVID-19.

A vaccine targeting the RBD of the S protein of SARS-CoV-2 induces protective immunity.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a respiratory disease called coronavirus disease 2019 (COVID-19), the spread of which has led to a pandemic. An effective preventive vaccine against this virus is urgently needed. As an essential step during infection, SARS-CoV-2 uses the receptor-binding domain (RBD) of the spike protein to engage with the receptor angiotensin-converting enzyme 2 (ACE2) on host cells1,2. Here we show that a recombinant vaccine that comprises residues 319-545 of the RBD of the spike protein induces a potent functional antibody response in immunized mice, rabbits and non-human primates (Macaca mulatta) as early as 7 or 14 days after the injection of a single vaccine dose. The sera from the immunized animals blocked the binding of the RBD to ACE2, which is expressed on the cell surface, and neutralized infection with a SARS-CoV-2 pseudovirus and live SARS-CoV-2 in vitro. Notably, vaccination also provided protection in non-human primates to an in vivo challenge with SARS-CoV-2. We found increased levels of RBD-specific antibodies in the sera of patients with COVID-19. We show that several immune pathways and CD4 T lymphocytes are involved in the induction of the vaccine antibody response. Our findings highlight the importance of the RBD domain in the design of SARS-CoV-2 vaccines and provide a rationale for the development of a protective vaccine through the induction of antibodies against the RBD domain.