Emerging respiratory viruses - pathogenesis and countermeasures

National Institute Of Allergy And Infectious Diseases

Investigators

Linked publications & trials

Abstract

In any given year, lower respiratory tract infections are the leading cause of infectious disease deaths worldwide, and the fifth most important cause of death overall. The emergence of SARS-CoV-2 and the resulting COVID-19 pandemic highlighted how devastating the effect of emerging respiratory viruses on global public health and economies can be. The COVID-19 pandemic also showed the difficulty of effectively treating severe viral lower respiratory tract infections, once again highlighting that our current understanding of the pathogenesis of viral lower respiratory tract infections is insufficient to drive the development of effective treatments. Many emerging respiratory viruses cause neurological disease besides respiratory tract disease. The main goal of the Molecular Pathogenesis Section is to contribute to our understanding of the pathogenesis of emerging viruses that cause severe lower respiratory tract disease such as Nipah virus, influenza A virus and coronaviruses on the level of the host and individual cell. We are investigating the pathogenesis of these viruses in the respiratory tract and the brain. Ultimately, our goal is to identify common pathways involved in lower respiratory tract and neurological disease progression and druggable targets within those pathways. Model development to study the pathogenesis of emerging respiratory viruses We invest a lot of time and effort into identifying, developing, and establishing in vivo and in vitro model systems that are optimized to address the particular aspect of disease pathogenesis we are interested in. Human organoid systems offer an exciting opportunity to study emerging respiratory viruses in the relevant tissue of the human host (Flagg and de Wit, Curr Opin Virol, 2024). We have established human lung organoids (hLO) derived from adult stem cells and induced pluripotent stem cells (iPSC). These organoids exist represent the distal lung, or alveoli, where gas exchange takes place. Our organoid cultures consist of alveolar type 2 cells (AT2). These AT2 cells can be differentiated into alveolar type 1 cells (AT1) in air-liquid interface cultures, using a protocol we optimized in the lab. Together, these two cell types enable us to study the pathogenesis of emerging respiratory viruses in an area of the lung that is extremely important in severe respiratory disease, i.e. the alveoli, and in a human host. We continued the development of model systems to study pathogenesis of emerging respiratory viruses in the central nervous system (CNS). For one of our viruses of interest where the majority of patients develops neurological complications, Nipah virus, we developed a new Syrian model to mimic neurological disease in humans. Hamsters were inoculated intracranially in the cerebellomedullary cistern with different doses of Nipah virus. Intracranial inoculation resulted in a rapid progression towards neurological disease. High Nipah viral loads were detected in the brains, and the virus spread from the CNS to the lungs. Histopathologic examination of the brain showed ischemic necrosis, often accompanied by marked edema and hemorrhage. Nipah viral antigen was detected primarily in meninges and cerebellum, but rarely observed in brain parenchyma. These histological lesions were different from the typical lesions observed in Nipah virus-infected humans and hamsters (and other animal models) inoculated via different routes. Thus, despite the consistent development of neurological disease, intracranial inoculation does not result in a model representative of Nipah virus neurological disease. Thus, we e continued our attempts to develop a new Syrian hamster model of neurological disease using a combination of Nipah virus inoculation and suboptimal remdesivir treatment to skew the disease in these animals to neurological signs rather than respiratory disease. We have also started to optimize methods to culture human cerebral organoids in our lab. We found that the recommended methods did not result in consistent production of neuronal tissue. In the meantime, we have relied on our collaborator Dr. Haigh to occasionally supply us with human cerebral organoids to continue our research of the neuropathogenesis of Nipah virus and highly pathogenic avian influenza (HPAI) H5N1 virus. Pathogenesis We completed the project on the evolution of SARS-CoV-2 Omicron introduced in our FY24 report. Briefly, we used our human lung organoids and upper respiratory tract air-liquid interface cultures to show that recent Omicron variants replicate efficiently in the upper respiratory tract, failed to replicate in human lung organoids. These data were combined with in vivo studies in the Syrian hamster models, where the recent Omicron variants displayed limited pathology in the lungs. Together, our data in vivo and in vitro data demonstrate that Omicron lineage evolution has favored increased fitness in the upper respiratory tract, i.e. SARS-CoV-2 is becoming more and more like a ‘common cold’ virus and highlight the strength and synergism of our approach combining organoid models with in vivo studies. Emergence of HPAI H5N1 virus in dairy cows Since the emergence of HPAI H5N1 virus in dairy cows in the US in March 2024, we have initiated several studies on this virus. We used our human lung organoids to assess the risk of the clade 2.3.4.4b HPAI H5N1 viruses currently circulating in the US to human health. We compared virus replication and host responses in human alveolar epithelium infected with contemporary HPAI H5N1 viruses to those of a historical HPAI H5N1 isolate from a fatal human case in 2004. The historical isolate, A/Vietnam/1203/2004, replicated most efficiently, followed by A/Texas/37/2024, recently isolated from a human case, then A/bovine/Ohio/B24OSU-342/2024, isolated from a dairy cow in the US in 2024. Cell survival data matched replication kinetics, with A/Vietnam/1203/2004 causing cell death earlier than the other two viruses; however, similar levels of cell death were measured by the end of the time course for all three isolates. Induction of interferon-stimulated genes was lower with A/Texas/37/2024 and A/bovine/Ohio/B24OSU-342/2024 than with A/Vietnam/1203/04. An overly exuberant immune response, including cytokine storm, is known to play a role in the high mortality from HPAI H5N1 virus infections in humans. Limited innate immune activation elicited by the A/Texas/37/2024 and A/bovine/Ohio/B24OSU-342/2024 isolates, together with their reduced virus replication capacity in the human alveolar epithelium, suggests a significantly reduced pathogenicity of the contemporary HPAI H5N1 clade 2.3.4.4b viruses. We are currently expanding these studies to include the D1.1 genotype. Human infections with this genotype were more severe; however, only a few cases have been detected so far with this genotype and confounding factors such as age and comorbidities of the patients may have played a role in disease development. Our organoid systems will help to determine whether the increased pathogenicity observed in human cases of D1.1 HPAI H5N1 are virus-intrinsic or not. In a collaborative study in C57BL/6J mice, we compared infection with A/Vietnam/1203/2004 and A/Bovine/Ohio/B24osu-342/2024. Infection with both HPAI H5N1 isolates caused severe disease requiring euthanasia. However, the tissue tropism of these two isolates differed significantly: while the virus from 2004 was largely restricted to the respiratory tract, the virus from 2024 replicated in various regions of the brain as well as in the respiratory tract. Notably, in addition to abundant evidence of CNS infection was also detected in the choroid plexus, retina and inner ear. The significance of the difference in neurotropism between these two HPAI H5N1 isolates is currently unclear, as both isolates are neurotropic in other animal models. In cows infected with HPAI H5N1, high virus titers are detected in milk, raising concerns of exposure to humans through consumption of milk. We showed that pasteurization inactivates HPAI H5N1 in milk, but infection via raw milk, including via drinking, could still occur. To assess the risk of exposure to HPAI H5N1, we compared the effect of different routes of infection with bovine HPAIV H5N1 clade 2.3.4.4b in cynomolgus macaques. While intranasal or intratracheal inoculation of macaques caused systemic infection, resulting in mild and severe respiratory disease, respectively, orogastric inoculation resulted in limited infection and seroconversion of macaques that did not show overt signs of disease. Thus, exposure to contaminated liquid products and raw milk consumption are a risk for HPAIV H5N1 infection of primates.