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Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has continued to circulate in humans since its emergence in 2019. While infection in humans continues, numerous spillover events to at least 32 animal species, including companion and zoo animals, have been reported. Since dogs and cats are highly susceptible to SARS-CoV-2 and have direct contact with their owners and other household members, it is important to know the prevalence of SARS-CoV-2 in dogs and cats. Here, we established an ELISA to detect serum antibodies against the receptor-binding domain and the ectodomain of the SARS-CoV-2 spike and nucleocapsid proteins. Using this ELISA, we assessed seroprevalence in 488 dog serum samples and 355 cat serum samples that were collected during the early pandemic period (between May and June of 2020) and 312 dog serum samples and 251 cat serum samples that were collected during the mid-pandemic period (between October 2021 and January 2022). We found that two dog serum samples (0.41%) collected in 2020, one cat serum sample (0.28%) collected in 2020, and four cat serum samples (1.6%) collected in 2021 were positive for antibodies against SARS-CoV-2. No dog serum samples collected in 2021 were positive for these antibodies. We conclude that the seroprevalence of SARS-CoV-2 antibodies in dogs and cats in Japan is low, suggesting that these animals are not a major SARS-CoV-2 reservoir.

Broadly protective antibodies targeting influenza viruses are of interest as potential therapeutics or to inform vaccine development. Here the authors characterize a human mAb (DA03E17) with heterosubtypic binding to neuraminidases from IAVs and IBVs that provides broad protection in vitro and in vivo.

Japan has reported a relatively small number of COVID-19 cases. Because not all infected persons receive diagnostic tests for COVID-19, the reported number must be lower than the actual number of infections. We assessed SARS-CoV-2 seroprevalence by analyzing >60,000 samples collected in Japan (Tokyo Metropolitan Area and Hokkaido Prefecture) during February 2020–March 2022. The results showed that ≈3.8% of the population had become seropositive by January 2021. The seroprevalence increased with the administration of vaccinations; however, among the elderly, seroprevalence was not as high as the vaccination rate. Among children, who were not eligible for vaccination, infection was spread during the epidemic waves caused by the SARS-CoV-2 Delta and Omicron variants. Nevertheless, seroprevalence for unvaccinated children

The rapid evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variants has emphasized the need to identify antibodies with broad neutralizing capabilities to inform future monoclonal therapies and vaccination strategies. Herein, we identified S728-1157, a broadly neutralizing antibody (bnAb) targeting the receptor-binding site (RBS) that was derived from an individual previously infected with WT SARS-CoV-2 prior to the spread of variants of concern (VOCs). S728-1157 demonstrated broad cross-neutralization of all dominant variants, including D614G, Beta, Delta, Kappa, Mu, and Omicron (BA.1/BA.2/BA.2.75/BA.4/BA.5/BL.1/XBB). Furthermore, S728-1157 protected hamsters against in vivo challenges with WT, Delta, and BA.1 viruses. Structural analysis showed that this antibody targets a class 1/RBS-A epitope in the receptor binding domain via multiple hydrophobic and polar interactions with its heavy chain complementarity determining region 3 (CDR-H3), in addition to common motifs in CDR-H1/CDR-H2 of class 1/RBS-A antibodies. Importantly, this epitope was more readily accessible in the open and prefusion state, or in the hexaproline (6P)-stabilized spike constructs, as compared with diproline (2P) constructs. Overall, S728-1157 demonstrates broad therapeutic potential and may inform target-driven vaccine designs against future SARS-CoV-2 variants.

Endogenous ACE2 is a receptor for SARS-CoV-2 and a recombinant soluble ACE2 protein can inhibit SARS-CoV-2 infection acting as a decoy. Here the authors show that B38-CAP, an ACE2-like enzyme but not a decoy for the virus, is protective against SARS-CoV-2-induced lung injury in animal models.

ABSTRACT The role of the intestinal microbiota in coronavirus disease 2019 (COVID-19) is being elucidated. Here, we analyzed the temporal changes in microbiota composition and the correlation between inflammation biomarkers/cytokines and microbiota in hospitalized COVID-19 patients. We obtained stool specimens, blood samples, and patient records from 22 hospitalized COVID-19 patients and performed 16S rRNA metagenomic analysis of stool samples over the course of disease onset compared to 40 healthy individual stool samples. We analyzed the correlation between the changes in the gut microbiota and plasma proinflammatory cytokine levels. Immediately after admission, differences in the gut microbiota were observed between COVID-19 patients and healthy subjects, mainly including enrichment of the classes Bacilli and Coriobacteriia and decrease in abundance of the class Clostridia. The bacterial profile continued to change throughout the hospitalization, with a decrease in short-chain fatty acid-producing bacteria including Faecalibacterium and an increase in the facultatively anaerobic bacteria Escherichia-Shigella. A consistent increase in Eggerthella belonging to the class Coriobacteriia was observed. The abundance of the class Clostridia was inversely correlated with interferon-γ level and that of the phylum Actinobacteria, which was enriched in COVID-19, and was positively correlated with gp130/sIL-6Rb levels. Dysbiosis was continued even after 21 days from onset. The intestines tended to be an aerobic environment in hospitalized COVID-19 patients. Because the composition of the gut microbiota correlates with the levels of proinflammatory cytokines, this finding emphasizes the need to understand how pathology is related to the temporal changes in the specific gut microbiota observed in COVID-19 patients. IMPORTANCE There is growing evidence that the commensal microbiota of the gastrointestinal and respiratory tracts regulates local and systemic inflammation (gut-lung axis). COVID-19 is primarily a respiratory disease, but the involvement of microbiota changes in the pathogenesis of this disease remains unclear. The composition of the gut microbiota of patients with COVID-19 changed over time during hospitalization, and the intestines tended to be an aerobic environment in hospitalized COVID-19 patients. These changes in gut microbiota may induce increased intestinal permeability, called leaky gut, allowing bacteria and toxins to enter the circulatory system and further aggravate the systemic inflammatory response. Since gut microbiota composition correlates with levels of proinflammatory cytokines, this finding highlights the need to understand how pathology relates to the gut environment, including the temporal changes in specific gut microbiota observed in COVID-19 patients.

Background: To develop an effective vaccine against a novel viral pathogen, it is important to understand the longitudinal antibody responses against its first infection. Here we performed a longitudinal study of antibody responses against SARS-CoV-2 in symptomatic patients. Methods: Sequential blood samples were collected from 39 individuals at various timepoints between 0 and 154 days after onset. IgG or IgM titers to the receptor binding domain (RBD) of the S protein, the ectodomain of the S protein, and the N protein were determined by using an ELISA. Neutralizing antibody titers were measured by using a plaque reduction assay. Findings: The IgG titers to the RBD of the S protein, the ectodomain of the S protein, and the N protein peaked at about 20 days after onset, gradually decreased thereafter, and were maintained for several months after onset. Extrapolation modeling analysis suggested that the IgG antibodies were maintained for this amount of time because the rate of reduction slowed after 30 days post-onset. IgM titers to the RBD decreased rapidly and disappeared in some individuals after 90 days post-onset. All patients, except one, possessed neutralizing antibodies against authentic SARS-CoV-2, which they retained at 90 days after onset. The highest antibody titers in patients with severe infections were higher than those in patients with mild or moderate infections, but the decrease in antibody titer in the severe infection cohort was more remarkable than that in the mild or moderate infection cohort. Interpretation: Although the number of patients is limited, our results show that the antibody response against the first SARS-CoV-2 infection in symptomatic patients is typical of that observed in an acute viral infection. Funding: The Japan Agency for Medical Research and Development and the National Institutes of Allergy and Infectious Diseases.

To clarify the threat posed by emergence of highly pathogenic influenza A(H7N9) virus infection among humans, we characterized the viral polymerase complex. Polymerase basic 2–482R, polymerase basic 2–588V, and polymerase acidic–497R individually or additively enhanced virus polymerase activity, indicating that multiple replication-enhancing mutations in 1 isolate may contribute to virulence.

Many broadly reactive human monoclonal antibodies against the hemagglutinin (HA) stem of influenza A virus have been developed for therapeutic applications. These antibodies typically inhibit viral entry steps, especially the HA conformational change that is required for membrane fusion. To better understand the mechanisms by which such antibodies inhibit viral replication, we established broadly reactive human anti-HA stem antibodies and determined the properties of these antibodies by examining their reactivity with 18 subtypes of HA, evaluating their in vivo protective efficacy, identifying their epitopes, and characterizing their inhibitory mechanisms. Among the eight human monoclonal antibodies we generated, which recognized at least 3 subtypes of the soluble HA antigens tested, clone S9-1-10/5-1 reacted with 18 subtypes of HA and protected mice from lethal infection with H1N1pdm09, H3N2, H5N1, and H7N9 viruses. This antibody recognized the HA2 helix A in the HA stem, and inhibited virus particle release from infected cells but did not block viral entry completely. These results show that broadly reactive human anti-HA stem antibodies can exhibit protective efficacy by inhibiting virus particle release. These findings expand our knowledge of the mechanisms by which broadly reactive stem-targeting antibodies inhibit viral replication and provide valuable information for universal vaccine development.

Currently, influenza vaccines are produced using embryonated chicken eggs. Recently, recombinant influenza vaccines have been developed as a potential alternative to egg-grown vaccines. In this study, we evaluated the efficacy of soluble recombinant hemagglutinin (HA) protein produced in human cell culture (Expi293F cells) as an influenza vaccine against homosubtypic and heterosubtypic influenza virus challenges in mice. Mice were immunized intramuscularly with purified soluble HA protein of H1N1pdm09 virus and then challenged with a lethal dose of H1N1pdm09, seasonal H3N2, or highly pathogenic avian influenza (HPAI) H5N1 virus. Vaccinated mice showed better morbidity than mock-vaccinated mice following H1N1pdm09 challenge. By contrast, all mice died following H3N2 challenge. Interestingly, all vaccinated mice survived challenge with H5N1 virus, whereas all mock-vaccinated mice died. These results suggest that intramuscular immunization with recombinant HA proteins produced in Expi 293F cells could be of value in influenza vaccine strategies.

We describe two adults with A/H3N2 influenza with (patient 1), and without (patient 2) polymerase acidic (PA) subunit I38 T substitution during the same season. Patient 1 had a reduced clinical response to baloxavir, a cap-dependent endonuclease inhibitor (CEI), but was cured by peramivir, a neuraminidase inhibitor. Baloxavir was clinically effective for patient 2, for whom peramivir had been ineffective. Susceptibility to baloxavir can be decreased by a PA unit mutation, but response to treatment can be increased by switching and/or combination with a neuraminidase inhibitor, even though CEI are clinically effective against influenza in adults. Keywords: Baloxavir, Polymerase, Peramivir, Viral infection, Polymerase acidic subunit

The influenza A virus rapidly mutates to escape from antibodies. Here, we isolated and characterized three human monoclonal antibodies (mAbs) that neutralize A(H1N1)pdm09 viruses. Generation of escape mutant viruses suggested that these antibodies recognized conserved residues of the receptor-binding site (RBS) of hemagglutinin (HA) and that mutant viruses that escaped from these mAbs rarely appeared. Moreover, the escape mutant viruses grew significantly slower than wild-type virus, indicating their reduced fitness. These results indicate that these three human mAbs against the RBS of HA have the potential to be anti-influenza agents with a low propensity for the development of resistant viruses.

Reverse transcription-quantitative PCR (RT-qPCR)-based tests are widely used to diagnose coronavirus disease 2019 (COVID-19). As a result that these tests cannot be done in local clinics where RT-qPCR testing capability is lacking, rapid antigen tests (RATs) for COVID-19 based on lateral flow immunoassays are used for rapid diagnosis. However, their sensitivity compared with each other and with RT-qPCR and infectious virus isolation has not been examined. Here, we compared the sensitivity among four RATs by using severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) isolates and several types of COVID-19 patient specimens and compared their sensitivity with that of RT-qPCR and infectious virus isolation. Although the RATs read the samples containing large amounts of virus as positive, even the most sensitive RAT read the samples containing small amounts of virus as negative. Moreover, all RATs tested failed to detect viral antigens in several specimens from which the virus was isolated. The current RATs will likely miss some COVID-19 patients who are shedding infectious SARS-CoV-2.

The A(H1N1)pdm09 virus emerged in 2009 and continues to circulate in human populations. Recent A(H1N1)pdm09 viruses, that is, A(H1N1)pdm09 viruses circulating in the post-pandemic era, can cause more or less severe infections than those caused by the initial pandemic viruses. To evaluate the changes in pathogenicity of the A(H1N1)pdm09 viruses during their continued circulation in humans, we compared the nucleotide and amino acid sequences of ten A(H1N1)pdm09 viruses isolated in Japan between 2009 and 2015, and experimentally infected mice with each virus. The severity of infection caused by these Japanese isolates ranged from milder to more severe than that caused by the prototypic pandemic strain A/California/04/2009 (CA04/09); however, specific mutations responsible for their pathogenicity have not yet been identified.

The rapid evolution of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variants has emphasized the need to identify antibodies with broad neutralizing capabilities to inform future monoclonal therapies and vaccination strategies. Herein, we identified S728-1157, a broadly neutralizing antibody (bnAb) targeting the receptor-binding site (RBS) that was derived from an individual previously infected with WT SARS-CoV-2 prior to the spread of variants of concern (VOCs). S728-1157 demonstrated broad cross-neutralization of all dominant variants, including D614G, Beta, Delta, Kappa, Mu, and Omicron (BA.1/BA.2/BA.2.75/BA.4/BA.5/BL.1/XBB). Furthermore, S728-1157 protected hamsters against in vivo challenges with WT, Delta, and BA.1 viruses. Structural analysis showed that this antibody targets a class 1/RBS-A epitope in the receptor binding domain via multiple hydrophobic and polar interactions with its heavy chain complementarity determining region 3 (CDR-H3), in addition to common motifs in CDR-H1/CDR-H2 of class 1/RBS-A antibodies. Importantly, this epitope was more readily accessible in the open and prefusion state, or in the hexaproline (6P)-stabilized spike constructs, as compared with diproline (2P) constructs. Overall, S728-1157 demonstrates broad therapeutic potential and may inform target-driven vaccine designs against future SARS-CoV-2 variants. [ABSTRACT FROM AUTHOR]