Differential beta-coronavirus infection dynamics in human bronchial epithelial organoids.

The lower respiratory system serves as the target and barrier for beta-coronavirus (beta-CoV) infections. In this study, we explored beta-CoV infection dynamics in human bronchial epithelial (HBE) organoids, focusing on HCoV-OC43, SARS-CoV, MERS-CoV, and SARS-CoV-2. Utilizing advanced organoid culture techniques, we observed robust replication for all beta-CoVs, particularly noting that SARS-CoV-2 reached peak viral RNA levels at 72 h postinfection. Through comprehensive transcriptomic analysis, we identified significant shifts in cell population dynamics, marked by an increase in goblet cells and a concurrent decrease in ciliated cells. Furthermore, our cell tropism analysis unveiled distinct preferences in viral targeting: HCoV-OC43 predominantly infected club cells, while SARS-CoV had a dual tropism for goblet and ciliated cells. In contrast, SARS-CoV-2 primarily infected ciliated cells, and MERS-CoV showed a marked affinity for goblet cells. Host factor analysis revealed the upregulation of genes encoding viral receptors and proteases. Notably, HCoV-OC43 induced the unfolded protein response pathway, which may facilitate viral replication. Our study also reveals a complex interplay between inflammatory pathways and the suppression of interferon responses during beta-CoV infections. These findings provide insights into host-virus interactions and antiviral defense mechanisms, contributing to our understanding of beta-CoV infections in the respiratory tract.

Coinfection with SARS-CoV-2 and Influenza A Virus Increases Disease Severity and Impairs Neutralizing Antibody and CD4+ T Cell Responses.

Given the current coronavirus disease 2019 (COVID-19) pandemic, coinfection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A virus (IAV) is a major concern for public health. However, the immunopathogenic events occurring with coinfections of SARS-CoV-2 and IAV remain unclear. Here, we report the pathogenic and immunological consequences of SARS-CoV-2 and IAV H1N1 coinfection in the K18-hACE2 transgenic mouse model. Compared with a single infection with SARS-CoV-2 or IAV, coinfections not only prolonged the primary virus infection period but also increased immune cell infiltration and inflammatory cytokine levels in bronchoalveolar lavage fluid leading to severe pneumonia and lung damage. Moreover, coinfections caused severe lymphopenia in peripheral blood, resulting in reduced total IgG, neutralizing antibody titers, and CD4+ T cell responses against each virus. This study sheds light on the immunopathogenesis of SARS-CoV-2 and IAV coinfection, which may guide the development of effective therapeutic strategies for the treatment of patients coinfected with these viruses. IMPORTANCE The cocirculation of influenza virus merging with the COVID-19 pandemic raises a potentially severe threat to public health. Recently, increasing numbers of SARS-CoV-2 and influenza virus coinfection have been reported from many countries. It is a worrisome issue that SARS-CoV-2 coinfection with other pathogens may worsen the clinical outcome and severity of COVID-19 and increase fatality. Here, we evaluated SARS-CoV-2 and IAV coinfection using the K18-hACE2 mouse model. Coinfected mice exhibited increased mortality with prolonged IAV shedding. Furthermore, coinfected mice showed a higher level of cytokines and chemokines than a single infection condition. Interestingly, our data show that coinfected mice showed significantly fewer virus-specific and neutralizing antibodies than the mice with a single infection. Overall, this study suggests that coinfection aggravates viral pathology by impaired neutralizing antibody response.

Seasonal dynamics of Haemaphysalis tick species as SFTSV vectors in South Korea.

Ticks pose a significant public health threat due to their ability to transmit various pathogens, including emerging tick-borne diseases. This study conducted a comprehensive surveillance of Haemaphysalis tick species and their severe fever with thrombocytopenia syndrome virus (SFTSV) infection rates in South Korea throughout the year 2023, from January to December. To ensure accurate and rapid identification of the prevalent Haemaphysalis tick species in South Korea, we designed PCR primer sets targeting the ITS1 gene, specifically distinguishing Haemaphysalis longicornis from Haemaphysalis flava. Among the 10,343 ticks collected from wild animals, H. longicornis constituted the majority, accounting for 65.5% (6,784/10,343 ticks), followed by H. flava at 33.8% (3,491/10,343 ticks), and Ixodes nipponensis at 0.7% (68/10,343 ticks). These identified ticks were then categorized into 811 pools, with 63 pools testing positive for SFTSV. Remarkably, the prevalence of SFTSV-positive H. longicornis ticks peaked during the summer months, aligning with heightened human outdoor activities and, consequently, an increased risk of human exposure. Conversely, it is noteworthy that H. flava exhibited a higher prevalence during the winter season, reaching its peak in January, with an SFTSV minimum infection rate similar to that of H. longicornis. These findings underscore the year-round presence of Haemaphysalis ticks as potential vectors for SFTSV, extending the temporal window for potential human exposure. Consequently, these results emphasize the necessity for active and continuous field surveillance to comprehensively understand and mitigate the public health risks associated with these tick-borne pathogens. IMPORTANCE: To date, the majority of tick surveillance studies have primarily focused on warmer seasons, which are considered optimal periods for ticks to actively seek hosts and transmit pathogens through blood-feeding activities. Consequently, tick species active during winter have often been overlooked, leading to an underestimation of their significance in transmitting severe fever with thrombocytopenia syndrome virus (SFTSV). In this study, we aimed to examine year-round tick prevalence with SFTSV and illuminate the role of the winter-dominant species, Haemaphysalis flava, in South Korea. Through rigorous identification facilitated by a primer set designed specifically for this purpose, we emphasize that H. flava, a competent vector species, harbors SFTSV in the winter season, thereby acting as an overwintering reservoir for the virus. This phenomenon may contribute to a higher infection rate among ticks in the following year.

Enhancing Omicron Sublineage Neutralization: Insights From Bivalent and Monovalent COVID-19 Booster Vaccines and Recent SARS-CoV-2 Omicron Variant Infections.

BACKGROUND: Omicron variants have rapidly diversified into sublineages with mutations that enhance immune evasion, posing challenges for vaccination and antibody responses. This study aimed to compare serum cross-neutralizing antibody responses against various SARS-CoV-2 Omicron sublineages (BA.1, BA.5, XBB.1.17.1, FK.1.1, and JN.1) in recipients of monovalent COVID-19 boosters, bivalent booster recipients, and individuals who had recovered from Omicron BA.5 infections. METHODS: We conducted a micro-neutralization assay on serum samples from monovalent BNT162b2 booster recipients (N = 54), bivalent BNT162b2 booster recipients (N = 24), and SARS-CoV-2 Omicron BA.5-recovered individuals (N = 13). The history of SARS-CoV-2 Omicron infection was assessed using ELISA against the SARS-CoV-2 NP protein. RESULTS: Bivalent booster recipients exhibited significantly enhanced neutralization efficacy against Omicron sublineages compared to those who had received monovalent booster vaccinations. Omicron BA.5-recovered individuals displayed similar neutralizing antibodies (NAbs) to the bivalent booster recipients. Despite the improved neutralization in bivalent recipients and BA.5-recovered individuals, there were limitations in neutralization against the recently emerged Omicron subvariants: XBB.1.17.1 FK.1.1, and JN.1. In both monovalent and bivalent booster recipients, a history of Omicron breakthrough infection was associated with relatively higher geometric mean titers of NAbs against Omicron BA.1, BA.5, and XBB.1.17.1 variants. CONCLUSION: This study underscores the intricate interplay between vaccination strategies, immune imprinting, and the dynamic landscape of SARS-CoV-2 variants. Although bivalent boosters enhance neutralization, addressing the challenge of emerging sublineages like XBB.1.17.1, FK.1.1, and JN.1 may necessitate the development of tailored vaccines, underscoring the need for ongoing adaptation to effectively combat this highly mutable virus.

Evolutional dynamics of highly pathogenic avian influenza H5N8 genotypes in wintering bird habitats: Insights from South Korea's 2020-2021 season.

The winter of 2020-2021 in South Korea witnessed severe outbreaks of Highly Pathogenic Avian Influenza (HPAI) viruses, specifically multiple genotypes of the H5N8 subtype. These outbreaks prompted an extensive investigation into the genetic characteristics and evolutionary dynamics of these viruses. Under the auspices of the National Institute of Wildlife Disease Control and Prevention (NIWDC), we conducted a nationwide surveillance program, collecting 7588 specimens from diverse wild bird habitats. Influenza A viruses were isolated at a rate of 5.0%, with HPAI H5N8 viruses accounting for 38.5% of isolates, predominantly found in wild bird carcasses (97.3%). Genetic analysis revealed the emergence of novel HPAI genotypes due to genetic reassortment events. G1 and G2 viruses were separately introduced into Korea, with G1 viruses displaying dynamic behavior, resulting in diverse sub-genotypes (G1-1 to G1-5) and mainly isolated from clinical specimens. Conversely, the G2 virus, introduced later, became the dominant strain consistently isolated mainly from bird carcasses (88.9%). These findings underscore the emergence of numerous novel HPAI genotypes shaped by multiple reassortment events in high-density wintering grounds of migratory birds. These sites act as hotspots for genetic exchanges, significantly influencing avian ecology, including resident bird species, and contributing to HPAI H5N8 evolution. The genetic diversity and ongoing evolution of these viruses highlight the need for vigilant surveillance and adaptive control measures. Recognizing the potential spillover to human populations, a One Health approach is essential to mitigate the evolving threats posed by avian influenza.

Deciphering the evolutionary landscape of severe fever with thrombocytopenia syndrome virus across East Asia.

Severe fever with thrombocytopenia syndrome virus (SFTSV) poses a significant public health challenge in East Asia, necessitating a deeper understanding of its evolutionary dynamics to effectively manage its spread and pathogenicity. This study provides a comprehensive analysis of the genetic diversity, recombination patterns, and selection pressures across the SFTSV genome, utilizing an extensive dataset of 2041 sequences from various hosts and regions up to November 2023. Employing maximum likelihood and Bayesian evolutionary analysis by sampling trees (BEAST), we elucidated the phylogenetic relationships among nine distinct SFTSV genotypes (A, B1, B2, B3, B4, C, D, E, and F), revealing intricate patterns of viral evolution and genotype distribution across China, South Korea, and Japan. Furthermore, our analysis identified 34 potential reassortments, underscoring a dynamic genetic interplay among SFTSV strains. Genetic recombination was observed most frequently in the large segment and least in the small segment, with notable recombination hotspots characterized by stem-loop hairpin structures, indicative of a structural propensity for genetic recombination. Additionally, selection pressure analysis on critical viral genes indicated a predominant trend of negative selection, with specific sites within the RNA-dependent RNA polymerase and glycoprotein genes showing positive selection. These sites suggest evolutionary adaptations to host immune responses and environmental pressures. This study sheds light on the intricate evolutionary mechanisms shaping SFTSV, offering insights into its adaptive strategies and potential implications for vaccine development and therapeutic interventions.

HA N193D substitution in the HPAI H5N1 virus alters receptor binding affinity and enhances virulence in mammalian hosts.

During the 2021/2022 winter season, we isolated highly pathogenic avian influenza (HPAI) H5N1 viruses harbouring an amino acid substitution from Asparagine(N) to Aspartic acid (D) at residue 193 of the hemagglutinin (HA) receptor binding domain (RBD) from migratory birds in South Korea. Herein, we investigated the characteristics of the N193D HA-RBD substitution in the A/CommonTeal/Korea/W811/2021[CT/W811] virus by using recombinant viruses engineered via reverse genetics (RG). A receptor affinity assay revealed that the N193D HA-RBD substitution in CT/W811 increases α2,6 sialic acid receptor binding affinity. The rCT/W811-HA193N virus caused rapid lethality with high virus titres in chickens compared with the rCT/W811-HA193D virus, while the rCT/W811-HA193D virus exhibited enhanced virulence in mammalian hosts with multiple tissue tropism. Surprisingly, a ferret-to-ferret transmission assay revealed that rCT/W811-HA193D virus replicates well in the respiratory tract, at a rate about 10 times higher than that of rCT/W811-HA193N, and all rCT/W811-HA193D direct contact ferrets were seroconverted at 10 days post-contact. Further, competition transmission assay of the two viruses revealed that rCT/W811-HA193D has enhanced growth kinetics compared with the rCT/W811-HA193N, eventually becoming the dominant strain in nasal turbinates. Further, rCT/W811-HA193D exhibits high infectivity in primary human bronchial epithelial (HBE) cells, suggesting the potential for human infection. Taken together, the HA-193D containing HPAI H5N1 virus from migratory birds showed enhanced virulence in mammalian hosts, but not in avian hosts, with multi-organ replication and ferret-to-ferret transmission. Thus, this suggests that HA-193D change increases the probability of HPAI H5N1 infection and transmission in humans.

SARS-CoV-2 variants with NSP12 P323L/G671S mutations display enhanced virus replication in ferret upper airways and higher transmissibility.

With the emergence of multiple predominant SARS-CoV-2 variants, it becomes important to have a comprehensive assessment of their viral fitness and transmissibility. Here, we demonstrate that natural temperature differences between the upper (33°C) and lower (37°C) respiratory tract have profound effects on SARS-CoV-2 replication and transmissibility. Specifically, SARS-CoV-2 variants containing the NSP12 mutations P323L or P323L/G671S exhibit enhanced RNA-dependent RNA polymerase (RdRp) activity at 33°C compared with 37°C and high transmissibility. Molecular dynamics simulations and microscale thermophoresis demonstrate that the NSP12 P323L and P323L/G671S mutations stabilize the NSP12-NSP7-NSP8 complex through hydrophobic effects, leading to increased viral RdRp activity. Furthermore, competitive transmissibility assay reveals that reverse genetic (RG)-P323L or RG-P323L/G671S NSP12 outcompetes RG-WT (wild-type) NSP12 for replication in the upper respiratory tract, allowing markedly rapid transmissibility. This suggests that NSP12 P323L or P323L/G671S mutation of SARS-CoV-2 is associated with increased RdRp complex stability and enzymatic activity, promoting efficient transmissibility.

Transmissibility and pathogenicity of SARS-CoV-2 variants in animal models.

As of February 2022, SARS-CoV-2 is still one of the most serious public health threats due to its high mortality rate and rapid spread of novel variants. Since the first outbreak in 2019, general understanding of SARS-CoV-2 has been improved through basic and clinical studies; however, knowledge gaps still exist in our understanding of the emerging novel SARSCoV-2 variants, which impacts the corresponding development of vaccines and therapeutics. Especially, accumulation of mutations in SARS-CoV-2 and rapid spread in populations with previous immunity has resulted in selection of variants that evade the host immune response. This phenomenon threatens to render current SARS-CoV-2 vaccines ineffective for controlling the pandemic. Proper animal models are essential for detailed investigations into the viral etiology, transmission and pathogenesis mechanisms, as well as evaluation of the efficacy of vaccine candidates against recent SARS-CoV-2 variants. Further, the choice of animal model for each research topic is important for researchers to gain better knowledge of recent SARS-CoV-2 variants. Here, we review the advantages and limitations of each animal model, including mice, hamsters, ferrets, and non-human primates, to elucidate variant SARS-CoV-2 etiology and transmission and to evaluate therapeutic and vaccine efficacy.

Inhibition of a broad range of SARS-CoV-2 variants by antiviral phytochemicals in hACE2 mice.

Although several vaccines and antiviral drugs against SARS-CoV-2 are currently available, control and prevention of COVID-19 through these interventions is limited due to inaccessibility and economic issues in some regions and countries. Moreover, incomplete viral clearance by ineffective therapeutics may lead to rapid genetic evolution, resulting in the emergence of new SARS-CoV-2 variants that may escape the host immune system as well as currently available COVID-19 vaccines. Here, we report that phytochemicals extracted from Chlorella spp. and Psidium guajava possess broad-spectrum antiviral activity against a range of SARS-CoV-2 variants. Through chromatography-based screening, we identified four bioactive compounds and subsequently demonstrated their potential antiviral activities in vivo. Interestingly, in hACE2 mice, treatment with these compounds significantly attenuates SARS-CoV-2-induced proinflammatory responses, demonstrating their potential anti-inflammatory activity. Collectively, our study suggests that phytochemicals from edible plants may be readily available therapeutics and prophylactics against multiple SARS-CoV-2 strains and variants.

Infection Route Impacts the Pathogenesis of Severe Fever with Thrombocytopenia Syndrome Virus in Ferrets.

The threat of severe fever with thrombocytopenia syndrome (SFTS) to public health has been increasing due to the rapid spread of the ticks that carry the causative viral agent. The SFTS virus (SFTSV) was first identified in China and subsequently detected in neighboring countries, including South Korea, Japan, and Vietnam. In addition to the tick-mediated infection, human-to-human transmission has been recently reported with a high mortality rate; however, differential study of the pathogen has been limited by the route of infection. In this study, we investigated the pathogenic potential of SFTSV based on the infection route in aged ferrets, which show clinical signs similar to that of human infections. Ferrets inoculated with SFTSV via the intramuscular and subcutaneous routes show clinical signs comparable to those of severe human infections, with a mortality rate of 100%. Contrastingly, intravascularly infected ferrets exhibit a comparatively lower mortality rate of 25%, although their early clinical signs are similar to those observed following infection via the other routes. These results indicate that the infection route could influence the onset of SFTS symptoms and the pathogenicity of SFTSV. Thus, infection route should be considered in future studies on the pathogenesis of SFTSV infection.

Age-dependent pathogenic characteristics of SARS-CoV-2 infection in ferrets.

While the seroprevalence of SARS-CoV-2 in healthy people does not differ significantly among age groups, those aged 65 years or older exhibit strikingly higher COVID-19 mortality compared to younger individuals. To further understand differing COVID-19 manifestations in patients of different ages, three age groups of ferrets are infected with SARS-CoV-2. Although SARS-CoV-2 is isolated from all ferrets regardless of age, aged ferrets (≥3 years old) show higher viral loads, longer nasal virus shedding, and more severe lung inflammatory cell infiltration, and clinical symptoms compared to juvenile (≤6 months) and young adult (1-2 years) groups. Furthermore, direct contact ferrets co-housed with the virus-infected aged group shed more virus than direct-contact ferrets co-housed with virus-infected juvenile or young adult ferrets. Transcriptome analysis of aged ferret lungs reveals strong enrichment of gene sets related to type I interferon, activated T cells, and M1 macrophage responses, mimicking the gene expression profile of severe COVID-19 patients. Thus, SARS-CoV-2-infected aged ferrets highly recapitulate COVID-19 patients with severe symptoms and are useful for understanding age-associated infection, transmission, and pathogenesis of SARS-CoV-2.

SARS-CoV-2 variants show temperature-dependent enhanced polymerase activity in the upper respiratory tract and high transmissibility.

With the convergent global emergence of SARS-CoV-2 variants of concern (VOC), a precise comparison study of viral fitness and transmission characteristics is necessary for the prediction of dominant VOCs and the development of suitable countermeasures. While airway temperature plays important roles in the fitness and transmissibility of respiratory tract viruses, it has not been well studied with SARS-CoV-2. Here we demonstrate that natural temperature differences between the upper (33°C) and lower (37°C) respiratory tract have profound effects on SARS-CoV-2 replication and transmission. Specifically, SARS-COV-2 variants containing the P323L or P323L/G671S mutation in the NSP12 RNA-dependent RNA polymerase (RdRp) exhibited enhanced RdRp enzymatic activity at 33°C compared to 37°C and high transmissibility in ferrets. MicroScale Thermophoresis demonstrated that the NSP12 P323L or P323L/G671S mutation stabilized the NSP12-NSP7-NSP8 complex interaction. Furthermore, reverse genetics-derived SARS-CoV-2 variants containing the NSP12 P323L or P323L/G671S mutation displayed enhanced replication at 33°C, and high transmission in ferrets. This suggests that the evolutionarily forced NSP12 P323L and P323L/G671S mutations of recent SARS-CoV-2 VOC strains are associated with increases of the RdRp complex stability and enzymatic activity, promoting the high transmissibility.

Enhanced antibody responses in fully vaccinated individuals against pan-SARS-CoV-2 variants following Omicron breakthrough infection.

Omicron has become the globally dominant severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant, creating additional challenges due to its ability to evade neutralization. Here, we report that neutralizing antibodies against Omicron variants are undetected following COVID-19 infection with ancestral or past SARS-CoV-2 variant viruses or after two-dose mRNA vaccination. Compared with two-dose vaccination, a three-dose vaccination course induces broad neutralizing antibody responses with improved durability against different SARS-CoV-2 variants, although neutralizing antibody titers against Omicron remain low. Intriguingly, among individuals with three-dose vaccination, Omicron breakthrough infection substantially augments serum neutralizing activity against a broad spectrum of SARS-CoV-2 variants, including Omicron variants BA.1, BA.2, and BA.5. Additionally, after Omicron breakthrough infection, memory T cells respond to the spike proteins of both ancestral and Omicron SARS-CoV-2 by producing cytokines with polyfunctionality. These results suggest that Omicron breakthrough infection following three-dose mRNA vaccination induces pan-SARS-CoV-2 immunity that may protect against emerging SARS-CoV-2 variants of concern.

Experimental Animal Models of Coronavirus Infections: Strengths and Limitations.

Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Since the emergence of SARS-CoV-2 in the human population in late 2019, it has spread on an unprecedented scale worldwide leading to the first coronavirus pandemic. SARS-CoV-2 infection results in a wide range of clinical manifestations from asymptomatic to fatal cases. Although intensive research has been undertaken to increase understanding of the complex biology of SARS-CoV-2 infection, the detailed mechanisms underpinning the severe pathogenesis and interactions between the virus and the host immune response are not well understood. Thus, the development of appropriate animal models that recapitulate human clinical manifestations and immune responses against SARS-CoV-2 is crucial. Although many animal models are currently available for the study of SARS-CoV-2 infection, each has distinct advantages and disadvantages, and some models show variable results between and within species. Thus, we aim to discuss the different animal models, including mice, hamsters, ferrets, and non-human primates, employed for SARS-CoV-2 infection studies and outline their individual strengths and limitations for use in studies aimed at increasing understanding of coronavirus pathogenesis. Moreover, a significant advantage of these animal models is that they can be tailored, providing unique options specific to the scientific goals of each researcher.

Severe fever with thrombocytopenia syndrome virus: emerging novel phlebovirus and their control strategy.

An emerging infectious disease first identified in central China in 2009, severe fever with thrombocytopenia syndrome (SFTS) was found to be caused by a novel phlebovirus. Since SFTSV was first identified, epidemics have occurred in several East Asian countries. With the escalating incidence of SFTS and the rapid, worldwide spread of SFTSV vector, it is clear this virus has pandemic potential and presents an impending global public health threat. In this review, we concisely summarize the latest findings regarding SFTSV, including vector and virus transmission, genotype diversity and epidemiology, probable pathogenic mechanism, and clinical presentation of human SFTS. Ticks most likely transmit SFTSV to animals including humans; however, human-to-human transmission has been reported. The majority of arbovirus transmission cycle includes vertebrate hosts, and potential reservoirs include a variety of both domestic and wild animals. Reports of the seroprevalence of SFTSV in both wild and domestic animals raises the probability that domestic animals act as amplifying hosts for the virus. Major clinical manifestation of human SFTS infection is high fever, thrombocytopenia, leukocytopenia, gastrointestinal symptoms, and a high case-fatality rate. Several animal models were developed to further understand the pathogenesis of the virus and aid in the discovery of therapeutics and preventive measures.

Development of Spike Receptor-Binding Domain Nanoparticles as a Vaccine Candidate against SARS-CoV-2 Infection in Ferrets.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a causative agent of the CoV disease 2019 (COVID-19) pandemic, enters host cells via the interaction of its receptor-binding domain (RBD) of the spike protein with host angiotensin-converting enzyme 2 (ACE2). Therefore, the RBD is a promising vaccine target to induce protective immunity against SARS-CoV-2 infection. In this study, we report the development of an RBD protein-based vaccine candidate against SARS-CoV-2 using self-assembling Helicobacter pylori-bullfrog ferritin nanoparticles as an antigen delivery system. RBD-ferritin protein purified from mammalian cells efficiently assembled into 24-mer nanoparticles. Sixteen- to 20-month-old ferrets were vaccinated with RBD-ferritin nanoparticles (RBD nanoparticles) by intramuscular or intranasal inoculation. All vaccinated ferrets with RBD nanoparticles produced potent neutralizing antibodies against SARS-CoV-2. Strikingly, vaccinated ferrets demonstrated efficient protection from SARS-CoV-2 challenge, showing no fever, body weight loss, or clinical symptoms. Furthermore, vaccinated ferrets showed rapid clearance of infectious virus in nasal washes and lungs as well as of viral RNA in respiratory organs. This study demonstrates that spike RBD-nanoparticles are an effective protein vaccine candidate against SARS-CoV-2.

Pathogenic assessment of avian influenza viruses in migratory birds.

ABSTRACTSeveral subtypes of avian influenza (AI) viruses have caused human infections in recent years; however, there is a severe knowledge gap regarding the capacity of wild bird viruses to infect mammals. To assess the risk of mammalian infection by AI viruses from their natural reservoirs, a panel of isolates from 34 wild birds was examined in animal models. All selected AI virus subtypes were found to predominantly possess Eurasian lineage, although reassortment with North American lineage AI viruses was also noted in some isolates. When used to infect chickens, 20 AI isolates could be recovered from oropharyngeal swabs at 5 days post-infection (dpi) without causing significant morbidity. Similarly, mild to no observable disease was observed in mice infected with these viruses although the majority replicated efficiently in murine lungs. As expected, wild bird AI isolates were found to recognize avian-like receptors, while a few strains also exhibited detectable human-like receptor binding. Selected strains were further tested in ferrets, and 15 out of 20 were found to shed the virus in the upper respiratory tract until 5 dpi. Overall, we demonstrate that a diversity of low-pathogenic AI viruses carried by wild migratory birds have the capacity to infect land-based poultry and mammalian hosts while causing minimal signs of clinical disease. This study reiterates that there is a significant capacity for interspecies transmission of AI viruses harboured by wild aquatic birds. Thus, these viruses pose a significant threat to human health underscoring the need for continued surveillance.

Development of spike receptor-binding domain nanoparticle as a vaccine candidate against SARS-CoV-2 infection in ferrets.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a causative agent of COVID-19 pandemic, enters host cells via the interaction of its Receptor-Binding Domain (RBD) of Spike protein with host Angiotensin-Converting Enzyme 2 (ACE2). Therefore, RBD is a promising vaccine target to induce protective immunity against SARS-CoV-2 infection. In this study, we report the development of RBD protein-based vaccine candidate against SARS-CoV-2 using self-assembling H. pylori -bullfrog ferritin nanoparticles as an antigen delivery. RBD-ferritin protein purified from mammalian cells efficiently assembled into 24-mer nanoparticles. 16-20 months-old ferrets were vaccinated with RBD-ferritin nanoparticles (RBD-nanoparticles) by intramuscular or intranasal inoculation. All vaccinated ferrets with RBD-nanoparticles produced potent neutralizing antibodies against SARS-CoV-2. Strikingly, vaccinated ferrets demonstrated efficient protection from SARS-CoV-2 challenge, showing no fever, body weight loss and clinical symptoms. Furthermore, vaccinated ferrets showed rapid clearance of infectious viruses in nasal washes and lungs as well as viral RNA in respiratory organs. This study demonstrates the Spike RBD-nanoparticle as an effective protein vaccine candidate against SARS-CoV-2.

Differences in seroprevalence between epicenter and non-epicenter areas of the COVID-19 outbreak in South Korea.

To compare the standardized severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) seroprevalence of high epicenter region with non-epicenter region, serological studies were performed with a total of 3,268 sera from Daegu City and 3,981 sera from Chungbuk Province. Indirect immunofluorescence assay (IFA) for SARS-CoV-2 IgG results showed a high seroprevalence rate in the Daegu City (epicenter) compared with a non-epicenter area (Chungbuk Province) (1.27% vs. 0.91%, P = 0.0358). It is noteworthy that the highest seroprevalence in Daegu City was found in elderly patients (70's) whereas young adult patients (20's) in Chungbuk Province showed the highest seroprevalence. Neutralizing antibody (NAb) titers were found in three samples from Daegu City (3/3, 268, 0.09%) while none of the samples from Chungbuk Province were NAb positive. These results demonstrated that even following the large outbreak, the seropositive rate of SARS-CoV-2 in the general population remained low in South Korea.