Characterization of spike processing and entry mechanisms of seasonal human coronaviruses NL63, 229E and HKU1 (preprint)

Although much has been learned about the entry mechanism of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the details of entry mechanisms of seasonal human coronaviruses (HCoVs) remain less well understood. In the present study, we established that 293T cell lines that stably express angiotensin converting enzyme (ACE2), aminopeptidase N (APN), or transmembrane serine protease 2 (TMPRSS2) support high level transduction of lentiviral pseudoviruses bearing spike proteins of seasonal HCoVs, HCoV-NL63, -229E, or -HKU1, respectively. Our results showed that entry of HCoV-NL63, -229E and -HKU1 pseudoviruses is sensitive to endosomal acidification inhibitors (chloroquine and NH4Cl), indicating virus entry via the endocytosis route. Although HCoV-HKU1 pseudovirus infection requires TMPRSS2 expression on cell surface, endocytosis-mediated HCoV-HKU1 entry requires the serine protease domain but not the serine protease activity of TMPRSS2. We also show that amino acids in the predicted S1/S2 junctions of spike proteins of HCoV-NL63, and -229E are essential for optimal entry but non-essential for spike-mediated entry of HCoV-HKU1. Our findings provide insights into entry mechanism of seasonal HCoVs that may support the development of novel treatment strategies.

Microgliosis, astrogliosis and loss of aquaporin-4 polarity in frontal cortex of COVID-19 patients (preprint)

The severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2), causing human coronavirus disease 2019 (COVID-19), not only affects the respiratory tract, but also impacts other organs including the brain. A considerable number of COVID-19 patients develop neuropsychiatric symptoms that may linger for weeks and months and contribute to \"long-COVID\". While the neurological symptoms of COVID-19 are well described, the cellular mechanisms of neurologic disorders attributed to the infection are still enigmatic. Here, we studied the effect of an infection with SARS-CoV-2 on the structure and expression of marker proteins of astrocytes and microglial cells in the frontal cortex of patients who died from COVID-19 in comparison to non-COVID-19 controls. Most of COVID-19 patients had microglial cells with retracted processes and rounded and enlarged cell bodies in both gray and white matter, as visualized by anti-Iba1 staining and confocal fluorescence microscopy. In addition, gray matter astrocytes in COVID-19 patients were frequently labeled by intense anti-GFAP staining, whereas in non-COVID-19 controls, most gray matter astrocytes expressed little GFAP. The most striking difference between astrocytes in COVID-19 patients and controls was found by anti-aquaporin-4 (AQP4) staining. In COVID-19 patients, a large number of gray matter astrocytes showed an increase in AQP4. In addition, AQP4 polarity was lost and AQP4 covered the entire cell, including the cell body and all cell processes, while in controls, AQP4 immunostaining was mainly detected in endfeet around blood vessels and did not visualize the cell body. In summary, our data suggest neuroinflammation upon SARS-CoV-2 infection including microgliosis and astrogliosis, including loss of AQP4 polarity.

The COVID-19 pandemic immediately increased the suicide rate in Kobe, Japan, especially among populations without psychiatry and primary care physician visits: Interrupted time-series analysis using a population-based database (preprint)

Background Japan has one of the lowest numbers of deaths due to coronavirus disease (COVID-19). However, the annual number of suicides increased for the first time in 2020, after a downward trend since 2007. Objective This study aimed to identify high-risk populations and assess the impact of medical visits on suicide trends in Japan following the COVID-19 pandemic. Method This quasi-experimental study used a population-based database from Hyogo Prefecture between 2012 and 2022. Interrupted time-series analyses were used to identify immediate and slope changes in the monthly number of suicides during the exposure period (2020-2022) compared with the control (2012-2019). The analysis was stratified according to the status of psychiatric department and primary care physician visits. Results: A totalof 2181 cases were included. Two-thirds of the cases were male, with a median age of 54. Primary care physicians and psychiatric history were present in 69% and 40% of the patients, respectively. The level change was 4.46 (95%CI; 1.83, 7.09), 3.04 (95%CI; 1.45, 4.64), and 3.07 (95%CI; 0.60, 5.53), in the overall, no primary care physician visit, and no psychiatric visits groups, respectively, which were significant. The level change was not significant in the groups with primary care and psychiatric department visits, at 1.07 (95%CI; -0.84, 2.98) and 0.64 (95%CI; -1.05, 2.33), respectively. The slope changes were not significant in any group. Conclusion: This study suggests that visits to a medical institution may have helped prevent the rapid increase in suicides during the early stages of the COVID-19 pandemic.

Comparative analysis of serological assays and sero-surveillance for SARS-CoV-2 exposure in US cattle (preprint)

Coronavirus disease-2019 (COVID-19) caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) continues to pose a significant threat to public health globally. Notably, SARS-CoV-2 demonstrates a unique capacity to infect various non-human animal species, documented in captive and free-living animals. However, experimental studies revealed low susceptibility of domestic cattle (Bos taurus) to ancestral B.1 lineage SARS-CoV-2 infection, with limited viral replication and seroconversion. Despite the emergence of viral variants with potentially altered host tropism, recent experimental findings indicate greater permissiveness of cattle to SARS-CoV-2 Delta variant infection compared to other variants, though with limited seroconversion and no clear evidence of transmission. While some studies detected SARS-CoV-2 antibodies in cattle in Italy and Germany, there is no evidence of natural SARS-CoV-2 infection in cattle from the United States or elsewhere. Since serological tests have inherent problems of false positives and negatives, we conducted a comprehensive assessment of multiple serological assays on over 600 cattle serum samples, including pre-pandemic and pandemic cattle sera. We found that SARS-CoV-2 pseudovirus neutralization assays with a luciferase reporter system can produce false positive results, and care must be taken to interpret serological diagnosis using these assays. We found no serological evidence of natural SARS-CoV-2 infection or transmission among cattle in the USA. Hence, it is critical to develop more reliable serological assays tailored to accurately detect SARS-CoV-2 antibodies in cattle populations and rigorously evaluate diagnostic tools. This study underscores the importance of robust evaluation when employing serological assays for SARS-CoV-2 detection in cattle populations.

Exploration of the link between COVID-19 and gastric cancer from the perspective of bioinformatics and systems biology (preprint)

Background: Coronavirus disease 2019 (COVID-19), an infectious disease caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), has caused a global pandemic. Gastric cancer (GC) poses a great threat to people's health, which is a high-risk factor for COVID-19. Previous studies have found some associations between GC and COVID-19, whereas the underlying molecular mechanisms are not well understood. Methods: We used a bioinformatics and systems biology approach to investigate the relationship between GC and COVID-19. The gene expression profiles of COVID-19 (GSE196822) and GC (GSE179252) were downloaded from the Gene Expression Omnibus (GEO) database. After identifying the shared differentially expressed genes (DEGs) for GC and COVID-19, functional annotation, protein-protein interaction (PPI) network, hub genes, transcriptional regulatory networks and candidate drugs were analyzed. Results: A total of 209 shared DEGs were identified to explore the linkages between COVID-19 and GC. Functional analyses showed that Immune-related pathway collectively participated in the development and progression of COVID-19 and GC. In addition, there are selected 10 hub genes including CDK1, KIF20A, TPX2, UBE2C, HJURP, CENPA, PLK1, MKI67, IFI6, and IFIT2. The transcription factor/gene and miRNA/gene interaction networks identified 38 transcription factors (TFs) and 234 miRNAs. More importantly, we identified ten potential therapeutic agents, including ciclopirox, resveratrol, etoposide, methotrexate, trifluridine, enterolactone, troglitazone, calcitriol, dasatinib and deferoxamine, some of which have been reported to improve and treat GC and COVID-19. This study also provides insight into the diseases most associated with mutual DEGs, which may provide new ideas for research on the treatment of COVID-19. Conclusions: This research has the possibility to be contributed to effective therapeutic in COVID-19 and GC.

Mapping immunodominant sites on the MERS-CoV spike glycoprotein targeted by infection-elicited antibodies in humans (preprint)

Middle-East respiratory syndrome coronavirus (MERS-CoV) first emerged in 2012 and causes human infections in endemic regions. Most vaccines and therapeutics in development against MERS-CoV focus on the spike (S) glycoprotein to prevent viral entry into target cells. These efforts, however, are limited by a poor understanding of antibody responses elicited by infection along with their durability, fine specificity and contribution of distinct S antigenic sites to neutralization. To address this knowledge gap, we analyzed S-directed binding and neutralizing antibody titers in plasma collected from individuals infected with MERS-CoV in 2017-2019 (prior to the COVID-19 pandemic). We observed that binding and neutralizing antibodies peak 1 to 6 weeks after symptom onset/hospitalization, persist for at least 6 months, and broadly neutralize human and camel MERS-CoV strains. We show that the MERS-CoV S1 subunit is immunodominant and that antibodies targeting S1, particularly the RBD, account for most plasma neutralizing activity. Antigenic site mapping revealed that polyclonal plasma antibodies frequently target RBD epitopes, particularly a site exposed irrespective of the S trimer conformation, whereas targeting of S2 subunit epitopes is rare, similar to SARS-CoV-2. Our data reveal in unprecedented details the humoral immune responses elicited by MERS-CoV infection, which will guide vaccine and therapeutic design.

Neurological manifestations of acute SARS-CoV-2 infection in pediatric patients: a 3-year study (preprint)

Purpose This study analyzed the neurological manifestation profiles of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection across pandemic waves in pediatric patients.Methods This retrospective study analyzed the data of patients aged 0–18 years who were diagnosed with acute SARS-CoV-2 infection and admitted to a pediatric tertiary hospital between March 1, 2020, and February 28, 2023. The presence of neurological manifestations was established based on the symptoms noted in each patient chart. The relationships between neurological manifestations and pandemic waves or age groups were assessed using the chi-square test.Results This study included 1677 patients. Neurological manifestations were noted in 10% (n = 168) of patients with a 3.2 years median age (interquartile range: 1–11.92). Neurological manifestations were significantly associated with the pandemic waves (p = 0.006) and age groups (p < 0.001). Seizures were noted in 4.2% of cases and reached an increasing frequency over time (p = 0.001), but were not associated with age groups. Febrile seizures accounted for the majority of seizures. Headache was reported in 2.6% of cases and had similar frequencies across the pandemic waves and age groups. Muscular involvement was noted in 2% of cases and reached a decreasing frequency over time (p < 0.001) and showed different frequencies among the age groups.Conclusions Neurological manifestations of acute SARS-CoV-2 infection exhibit distinct patterns, depending on the pandemic wave and patient age group. The Wuhan and Omicron waves involved the nervous system more often than the other waves; however, this was probably through different mechanisms.

Nanoemulsions and nanocapsules as carriers for the development of intranasal mRNA vaccines (preprint)

The global emergency of coronavirus disease 2019 (COVID-19) has spurred extensive worldwide efforts to develop vaccines for protection against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Our contribution to this global endeavor involved the development of a diverse library of nanocarriers, as alternatives to lipid nanoparticles (LNPs), including nanoemulsions (NEs) and nanocapsules (NCs), with the aim of protecting and delivering messenger ribonucleic acid (mRNA) for nasal vaccination purposes. A wide range of prototypes underwent rigorous screening through a series of in vitro and in vivo experiments, encompassing assessments of cellular transfection, cytotoxicity, and intramuscular administration of a model mRNA for protein translation. Consequently, we identified two promising candidates for nasal administration. These candidates include an NE incorporating a combination of an ionizable lipid (C12-200) and cationic lipid (DOTAP) for mRNA entrapment, along with DOPE to facilitate endosomal escape. This NE exhibited a size of 120 nm and a highly positive surface charge (+50 mV). Additionally, an NC formulation comprising the same components with a dextran sulfate shell was identified, with a size of 130 nm and a moderate negative surface charge (-16 mV). Upon intranasal administration of mRNA encoding for ovalbumin (mOVA) associated with optimized versions of NEs and NCs, robust antigen-specific CD8+ T cell responses were observed. These findings underscore the potential of NEs and polymeric NCs in advancing mRNA vaccine development for combating infectious diseases.

SARS-COV-2 RNA removal rates from fresh cabbage during fridge storage, disinfection with neutral electrolyzed water and calcium hypochlorite (preprint)

The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic has raised concerns over viral transmission via contaminated fresh produce. In this study, we evaluated the efficacy of Neutral Electrolyzed Water (NEW) in comparison to calcium hypochlorite (Ca(ClO)2) for the removal of SARS-CoV-2 viral RNA from vegetables stored at refrigeration temperature over a one-week period. Vegetables were inoculated with SARS-CoV-2 viral particles and subsequently treated by immersion in NEW, Ca(ClO)2, or sterile water (control) solutions. Viral RNA was extracted from the produce on days 0, 1, 3, 5, and 7 post-immersion and quantified using RT-qPCR. We demonstrate that NEW solutions reduced over 99% of detectable SARS-CoV-2 RNA on vegetables within the first 24 hours. Comparatively, Ca(ClO)2 treatment resulted in less pronounced viral RNA reduction over the 7-day experiment. At identical concentrations, NEW led to significantly higher SARS-CoV-2 RNA removal from contaminated vegetables versus Ca(ClO)2. Given the on-site production feasibility, lower environmental impacts, and reduced costs associated with NEW solutions, our findings position NEW as a promising alternative to conventional chlorine-based sanitizers for mitigation of SARS-CoV-2 presence on fresh produce.

Abolished frameshifting for predicted structure-stabilizing SARS-CoV-2 mutants: Implications to alternative conformations and their statistical structural analyses (preprint)

The SARS-CoV-2 frameshifting element (FSE) has been intensely studied and explored as a therapeutic target for coronavirus diseases including COVID-19. Besides the intriguing virology, this small RNA is known to adopt many length-dependent conformations, as verified by multiple experimental and computational approaches. However, the role these alternative conformations play in the frameshifting mechanism and how to quantify this structural abundance has been an ongoing challenge. Here, we show by DMS and dual-luciferase functional assays that previously predicted FSE mutants (using the RAG graph theory approach) suppress structural transitions and abolish frameshifting. Furthermore, correlated mutation analysis of DMS data by three programs (DREEM, DRACO, and DANCE-MaP) reveals important differences in their estimation of specific RNA conformations, suggesting caution in the interpretation of such complex conformational landscapes. Overall, the abolished frameshifting in three different mutants confirms that all alternative conformations play a role in the pathways of ribosomal transition.

An outbreak of a pathogenic canine coronavirus type 2 in captive snow leopards (Panthera uncia) in the U.S, with severe gastrointestinal signs (preprint)

The species alphacoronavirus-1 comprises a set of diverse viruses of cats, dogs, and pigs, and is highly recombinogenic. Within this species, canine coronavirus type 2 (CCoV-2) can infect multiple species of canids, causing a range of clinical outcomes. CCoV-2 is genetically related to feline coronavirus type 1 (FCoV-1) and type 2 (FCoV-2), with FCoV-2 being a recombinant genotype of FCoV-1 and CCoV-2. Recently, a novel FCoV (FCoV-23) resulting from recombination with a highly pathogenic (pantropic) CCoV-2 (pCCoV-2) has been identified as the cause of a widespread outbreak among stray/feral cats in Cyprus. To understand the origin of recombinant variants it is crucial to identify hosts that can be infected with viruses in the species alphacoronavirus-1. Experimental evidence indicates that domesticated cats likely play a central role in the emergence of recombinant variants, as they can also be infected with CCoV-2. Wild felids are genetically closely related to domestic cats and may also be susceptible to FCoV and CCoV infection; however, there have been no reports of natural infection with CCoV in domesticated or wild felids. In this study, we retrospectively investigated a localized outbreak of severe enteritis in snow leopards (Panthera uncia) housed in a zoological institute in the U.S. Molecular screening and whole genome sequencing revealed the shedding of CCoV-2 in the feces of the three sick leopards. Phylogenetic analyses of the spike gene revealed it is genetically related to pathogenic variants of CCoV-2 identified in domesticated dogs in the U.S., and to pCCoV-2 CB/05 circulating in Europe. This study provides the first genetic evidence of CCoV-2 infection in a wild felid and highlights the necessity of conducting surveillance of both FCoV and CCoV in domesticated and wild felids.

Analysis of Cell Immunity for Children Infected with COVID-19 and Those Vaccinated against COVID-19 Using T-SPOT®.COVID (preprint)

To elucidate the cellular immune response to coronavirus disease (COVID-19) among children, we assessed cellular immunity in 8 children post-vaccination for COVID-19 and 11 children after severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, using the T-SPOT®.COVID assay for both the spike (S) and nucleocapsid (N) proteins. In the vaccinated group, the T-SPOT®.COVID assay for the S protein yielded positive results in all eight children. In contrast, in the post-infection group, the assay for the N protein was positive in 5 of 11 children, with 3 of these 5 children requiring hospital admission, including 2 who needed mechanical ventilation. Therefore, the T-SPOT®.COVID assay is valuable for assessing cellular immunity against COVID-19, and most children infected with COVID-19 may not develop such immunity unless the disease severity is significant.

High transferability of neutralizing antibodies against SARS-CoV-2 to umbilical cord blood in pregnant women with BNT162b2 XBB.1.5 vaccine - a retrospective cohort study (preprint)

Coronavirus disease 2019 (COVID-19), when contracted by pregnant women, can lead to severe respiratory illness, rapid disease progression, and higher rates of intensive care unit admission. COVID-19 infection during pregnancy is associated with an increased risk of preterm delivery, cesarean section, fetal dysfunction, preeclampsia, and perinatal death. Additionally, vertical transmission of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) from pregnant women to their fetuses has been observed. While severe infections in neonates and infants are rare, newborns can experience serious consequences from COVID-19, despite their suboptimal humoral immune system protection. The amino acids in the structural proteins of SARS-CoV-2 are subjected to constant mutation. Since around January 2023, COVID-19, caused by infection with omicron-type SARS-CoV-2 variants, has been prevalent globally. Omicron-type SARS-CoV-2 variants can evade the immune response triggered by traditional mRNA-based COVID-19 vaccines, such as BNT162b2. Therefore, vaccination with a vaccine (BNT162b2 XBB.1.5) that can provide protection against omicron-type SARS-CoV-2 variants is recommended. Therefore, we examined the titers of anti-spike glycoprotein of SARS-CoV-2 IgG and IgA in the blood and umbilical cord blood obtained from pregnant women vaccinated with BNT162b2 XBB.1.5. The results showed that anti-spike glycoprotein of SARS-CoV-2 IgG and IgA titers were highest in the blood and cord blood obtained from pregnant women vaccinated with BNT162b2 XBB.1.5 at late gestational age (28-34 weeks). No serious side effects or adverse events caused by vaccination of pregnant women with BNT162b2 XBB.1.5 were observed in either pregnant women or newborns. In the future, to validate our findings, large cohort clinical studies involving numerous pregnant women must be conducted.

H2 inhalation therapy in patients with moderate Covid 19 (H2 COVID) : a prospective ascending-dose phase 1 clinical trial (preprint)

Introduction: The Covid-19 pandemic, caused by the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), has triggered a serious global health crisis, resulting in millions of reported deaths since its initial identification in China in November 2019. The global disparities in immunization access emphasize the urgent need for ongoing research into therapeutic interventions. This study focuses on the potential use of molecular dihydrogen (H2) inhalation as an adjunctive treatment for Covid-19. H2 therapy shows promise in inhibiting intracellular signaling pathways associated with inflammation, particularly when administered early in conjunction with nasal oxygen therapy. Methods: This Phase I study, characterized by an open-label, prospective, monocentric, and single ascending dose design, seeks to assess the safety and tolerability of the procedure in individuals with confirmed SARS-CoV-2 infection. Employing a 3+3 design, the study includes three exposure durations (target durations): 1 day (D1), 3 days (D2), and 6 days (D3). Results: We concluded that the Maximum Tolerated Duration is at least three days. Every patient showed clinical improvement and excellent tolerance to H2 therapy. Discussion/conclusion: To the best of our knowledge, this phase 1 clinical trial is the first to establish the safety of inhaling a mixture of H2 (3.6%) and N2 (96.4%) in hospitalized Covid-19 patients. The original device and method employed ensure the absence of explosion risk. The encouraging outcomes observed in the 12 patients included in the study justify further exploration through larger, controlled clinical trials.

Modulation of SARS-CoV-2 spike binding to ACE2 throughconformational selection (preprint)

The first step of SARS-CoV-2 infection involves the interaction between the trimeric viral spike protein () and the host angiotensin-converting enzyme 2 (2). The receptor binding domain () of adopts two conformations: open and closed, respectively, accessible and inaccessible to 2. Therefore, motions are suspected to affect 2 binding; yet a quantitative description of the underlying mechanism has been elusive. Here, using single-molecule approaches, we visualize opening and closing and probe the /2 interaction. Our results show that RBD dynamics affect 2 binding but not unbinding. The resulting modulation is quantitatively predicted by a conformational selection model in which each protomer behaves independently. Our work reveals a general molecular mechanism affecting binding affinity without altering binding strength, helping to understand coronavirus infection and immune evasion.

Cross-reactive antibody responses to coronaviruses elicited by SARS-CoV-2 infection or vaccination (preprint)

Background: The newly emerged SARS-CoV-2 possesses shared antigenic epitopes with other human coronaviruses. We investigated if COVID-19 vaccination or SARS-CoV-2 infection may boost cross-reactive antibodies to other human coronaviruses. Methods Pre- and post-vaccination sera from SARS-CoV-2 naïve healthy subjects who received three doses of the mRNA vaccine (BioNTech, BNT) or the inactivated vaccine (CoronaVac, CV) were used to monitor the level of cross-reactive antibodies raised against other human coronaviruses by enzyme-linked immunosorbent assay. In comparison, convalescent sera from COVID-19 patients with or without prior vaccination history were also tested. Pseudoparticle neutralization assay was performed to detect neutralization antibody against MERS-CoV. Results Among SARS-CoV-2 infection naïve subjects, BNT or CV significantly increased the anti-S2 antibodies against Betacoronaviruses (OC43 and MERS-CoV) but not Alphacoronaviruses (229E). The pre-vaccination antibody response to the common cold human coronaviruses did not negatively impact the post-vaccination antibody response to SARS-CoV-2. Cross-reactive antibodies that binds to the S2 protein of MERS-CoV were similarly detected from the convalescent sera of COVID-19 patients with or without vaccination history. However, these anti-S2 antibodies do not possess neutralizing activity in MERS-CoV pseudoparticle neutralisation tests. Conclusions Our results suggest that SARS-CoV-2 infection or vaccination may potentially modulate population immune landscape against previously exposed or novel human coronaviruses. The findings have implications for future sero-epidemiological studies on MERS-CoV.

Ribavirin’s Daily Dose Growth Determined as the Result of Medicament Testing May Be a Predictor of the Chronic Course of the Disease in Long COVID Syndrome Patients (preprint)

Introduction Long COVID syndrome, a multisystemic condition resulting from severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, affects at least 65 million people worldwide. The disease pathogenesis is unclear, and many different assumptions still exist. This study aimed to explore the use of medicament testing to determine the optimal daily dose of ribavirin through the use of electroacupuncture via the Voll (EAV) diagnostic system for acupuncture points. Materials and methods One hundred one patients (aged 16 to 50) with long COVID syndrome were recruited from the Research Institute of Virology and were eligible according to the inclusion criteria. Patients were randomized to the experimental or the placebo groups. The patients were further examined with EAV diagnostics based on the level of electrodermal activity at the acupuncture points, followed by medicament testing with ribavirin (tablets) to determine the daily doses of the drug. Fifty-two participants were randomized to the experimental group and fourty nine to the placebo group and were considered for data analyses. Results The results of this study demonstrated the feasibility of using EAV to identify meridians with decreased levels of electrodermal activity at acupuncture points, followed by medicament testing with ribavirin to restore the decreased electrodermal conductivity at the studied acupuncture points and to measure the daily dose of the drug. Conclusions The measured daily doses of ribavirin in patients with long COVID syndrome may indirectly serve as a prognostic marker of the course of the disease. However, further clinical and instrumental studies are needed to evaluate the clinical application of medicament testing in assessing long-term COVID syndrome.

Targeted next-generation sequencing of pathogens reveals the profile of secondary infections in COVID-19 patients (preprint)

PURPOSE: To use targeted next-generation sequencing (tNGS) of pathogens for analysing the etiological distribution of secondary infections in patients with severe and critical novel coronavirus pneumonia (COVID-19), to obtain microbial epidemiological data on secondary infections in patients with COVID-19, and to provide a reference for early empirical antibiotic treatment of such patients. METHODS: Patients with infections secondary to severe and critical COVID-19 and hospitalised at the First Affiliated Hospital of Shandong First Medical University between 1 December 2022 and 30 June 2023 were included in the study. The characteristics and etiological distribution of secondary infections in these patients were analysed using tNGS. RESULTS: A total of 95 patients with COVID-19 secondary infections were included in the study, of whom 87.37% had one or more underlying diseases. Forty-eight pathogens were detected, the most common being HSV-4, Candida albicans, Klebsiella pneumoniae, Enterococcus faecium, HSV-1, Staphylococcus aureus, Aspergillus fumigatus, Acinetobacter baumannii, HSV-5, and Stenotrophomonas maltophilia, with Pneumocystis jirovecii being detected in 14.29% of cases. The majority (76.84%) of COVID-19 secondary infections were mixed infections, with mixed viral-bacterial-fungal infections being the most common (28.42%). CONCLUSION: Most secondary infections in severe and critical COVID-19 patients are mixed, with high rates of viral and fungal infections. In clinical settings, monitoring for reactivation or secondary infections by Herpesviridae viruses is crucial; additionally, these patients have a significantly higher rate of P. jirovecii infection. tNGS testing on bronchoalveolar lavage fluid can help determine the aetiology of secondary infections early in COVID-19 patients and assist in choosing appropriate antibiotics.

Impact of the severe acute respiratory syndrome coronavirus 2 pandemic on the incidence of type 1 diabetes mellitus in children in Yamanashi, Japan (preprint)

It is hypothesized that the biopsychosocial stress associated with the severe acute respiratory syndrome coronavirus 2 pandemic (SARS-CoV-2), in combination with the immunologic effects of SARS-CoV-2 and pancreatic β-cell dysfunction, may have contributed to the onset of type 1 diabetes (T1D) in children. We documented the incidence rates of T1D in Yamanashi Prefecture, Japan, from 1986 to 2018 and expanded our analysis to include cases from 2019 to 2022 to assess the influence of coronavirus disease 2019 (COVID-19) on the incidence of T1D. The annual increase in standardized incidences of T1D among 0- to 14-year-olds was 2.089% per year from 1986 to 2019 (p = .0772) and 2.183% per year from 1986 to 2022 (p = .0331). For the 5-9 year age group, the annual increase in crude incidence from 1986 to 2019 was 6.607% per year (p < .01), and from 1986 to 2022, it was 6.270% per year (p < .001). In Yamanashi Prefecture, Japan, the incidence of pediatric T1D increased during the COVID-19 pandemic from 2020 to 2022. However, this trend was an extension of the increase prior to 2019, suggesting that no direct or indirect effect of COVID-19 on this trend was identified.

Differential Patterns of Cross-Protection against Antigenically Distinct Variants in Small Animal Models of SARS-CoV-2 Infection (preprint)

Continuous evolution of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) will likely force more future updates of vaccine composition. Based on a series of studies carried out in human ACE2 transgenic mice (K18-hACE2) and Syrian hamsters, we show that immunity at the respiratory tract, acquired through either previous infection or vaccination with an in-house live attenuate virus, offers protection against antigenically distinct variants in the absence of variant spike-specific neutralizing antibodies. Interestingly, immunity acquired through infection of a modern variant (XBB.1.5) was insufficient in preventing brain infection by the ancestral virus (WA1/2020) in K18-hACE2 mice. Similarly, previous infection with WA1/2020 did not protect against brain infection by XBB.1.5. Our results highlight the importance of immune components other than neutralizing antibodies in maintaining protection against new variants in the respiratory tract, but also paint scenarios where a monovalent vaccine based on a contemporary variant may be less effective against the ancestral strain.