Search Results (457)

Respiratory viruses such as SARS-CoV-2, influenzas A and B, respiratory syncytial virus (RSV), human metapneumovirus (hMPV), human parainfluenza virus type 3 (HPIV-3), and rhinoviruses remain major causes of global morbidity. Their rapid evolution, high transmissibility, and limited therapeutic options, together with the absence of approved vaccines for several pathogens, highlight the need for broad-acting and pathogen-independent antiviral strategies. Nitric oxide exhibits antiviral activity through redox-dependent mechanisms, including S-nitrosylation of cysteine-containing viral proteins and disruption of redox-sensitive structural domains. Clinical studies conducted during the SARS-CoV-2 pandemic demonstrated that a nitric oxide nasal spray (NONS) rapidly reduced nasal viral load and transmission. In this study, we evaluated the in vitro virucidal activity of the NONS against a panel of clinically relevant respiratory viruses representing four major virus families. Virus suspensions of approximately 10⁴ CCID₅₀ were exposed to a full-strength NONS for contact times ranging from 5 s to 2 min at room temperature, followed by neutralization and quantification of residual infectivity using endpoint dilution assays. The NONS rapidly reduced viral infectivity across all viruses tested, achieving >3 log₁₀ reductions within 2 min. SARS-CoV-2 variants including Alpha, Beta, Gamma, Delta, Omicron BA.1, and XBB 2.0 were reduced to levels at or below the assay detection limit within 30 s to 2 min. Influenza A and B viruses showed the fastest loss of infectivity, reaching detection limits within 10–15 s. RSV, hMPV, HPIV-3, and human rhinovirus 14 were similarly inactivated within 1–2 min. These findings demonstrate that the NONS exhibits rapid and broad-spectrum virucidal activity against diverse respiratory viruses and supports its potential role in pandemic preparedness but also seasonal use. Full article

Five years after the onset of the SARS-CoV-2 pandemic, post-COVID-19 condition continues to affect millions of subjects with persistent symptoms that significantly impair quality of life. Post-COVID-19 pain, particularly in women, has emerged as a frequent yet underestimated symptom. The validation and identification of animal models that reproduce persistent symptoms after an acute SARS-CoV-2 infection is crucial for a better understanding of the underlying mechanisms. The aim of the current study was to evaluate thermal nociception, biomarkers of inflammation, and nerve tissue damage in a female animal model of post-COVID-19 condition. A SARS-CoV-2 infection model was established by intranasal administration of the Omicron variant (BA.1.17 lineage) in transgenic female C57BL/6 mice expressing the human ACE2 receptor (hACE2). Nociception was assessed using the hot-plate test for 28 days post-infection. Afterwards, animals were sacrificed to analyze plasma inflammatory biomarkers by multiplex analysis. In addition, IL-6, IL-18, and IL-1β expression were evaluated by immunohistochemistry to analyze neural inflammation in the saphenous nerve. The results revealed that heat nociceptive thresholds in infected mice did not significantly differ from those of non-infected, but a trend toward lower thresholds was observed in the infected group (days 14 and 28 post-infection). In addition, a slight modification in pro- and anti-inflammatory cytokines/chemokines in plasma was detected, but no changes in the expression of IL-6, IL-1β, or IL-18 were observed in the saphenous nerve. Based on all the analyses conducted, infection with the Omicron variant of SARS-CoV-2 did not induce thermal sensitization in animals nor alterations in the expression of inflammatory biomarkers in the saphenous nerve. Finally, a slight state of systemic inflammation was present in the infected animals. The absence of detectable changes in this animal model underscores the need for further research to clarify the discrepancies observed in human patients and to explore alternative pathways potentially involved in post-COVID-19 pain syndromes. Full article

We describe RNA-Seq analyses conducted on nasopharyngeal swabs collected from 37 patients admitted to an Australian intensive care unit from October 2022 to August 2023. During this time, the dominant omicron sublineage infections broadly progressed from BA.5 to BA.2-like, to XBB-like, then XBC, consistent with global trends. Viral load and patient metadata correlations indicated this cohort was broadly representative of severe COVID-19 patients. Human gene expression analyses were complicated by the large range (>5 log) and variability in viral reads. Nevertheless, the comparison of XBC and BA.5 samples that had comparable viral read counts, revealed differentially expressed genes and a cellular deconvolution signature that indicated increased targeting of ciliated epithelial cells by XBC. To obtain more evidence for increased targeting of ciliated epithelial cells by the later omicron sublineage viruses, a series of mouse strains were infected with a BA.5 or a XBB isolate. Increased infection of the nasal turbinates and ciliated epithelial cells by XBB was demonstrated by viral titrations and immunohistochemistry, respectively. Compared with previous lineages, the omicron lineage showed increased targeting of ciliated epithelia in the upper respiratory tract, with the data presented herein suggesting this trend continued for the omicron sublineages. Full article

Post-COVID cardiac complications have emerged as a significant and persistent clinical concern, yet their underlying mechanisms remain poorly understood. Animal models can act as proxies to investigate the pathophysiology of the human, post-acute sequelae of SARS-CoV-2 infection (PASC). The aim of this experimental study was to evaluate the expression of inflammatory biomarkers in cardiac tissue 28 days after SARS-CoV-2 infection in a female hACE2 mouse model, with a focus on chemokine-mediated immune activation. Twelve female C57BL/6 hACE2 mice were infected with the Omicron variant (BA.1.17 lineage) of SARS-CoV-2, and eleven non-infected mice served as controls. Cardiac tissue was analyzed via Western blot for markers of innate immune activation (TLR4, MyD88, NF-κB) and pro-inflammatory cytokines (IL-6, IL-18, IL-1β, TNF-α, CD11d). Cardiac tissue injury markers (iNOS, PAI-1 and Connexin43) were also analyzed. Compared to non-infected mice, cardiac tissue from infected mice showed significantly higher expression of IL-6 (p = 0.028), indicating an inflammatory state, and CD11d (p = 0.016), suggesting an inflammatory stage accompanied by sustained activation of chemokine-mediated inflammatory signaling. No significant differences in TLR4 (p = 0.340), MyD88 (p = 0.410), NF-κB p65 (p = 0.780), IL-18 (p = 0.548), IL-1β (p = 0.455), and TNF-α (p = 0.125) expressions were observed Similarly, no changes in cardiac damage markers (iNOS: p = 0.4684; PAI-1: p = 0.5345; Connexin 43: p = 0.2879) were found. The results of this experimental study would support the hypothesis of persistent low-grade inflammation as a contributor to post-COVID cardiac sequelae in females that is not accompanied by severe tissue damage, as also observed in clinical studies. This study also reinforces the need for studies evaluating the functional and structural evolution of the myocardium after an acute SARS-CoV-2 infection. Full article

Background/Objectives: SARS-CoV-2 continues to generate antigenically divergent variants that reduce the breadth of existing vaccine-induced antibody responses. Fc-fusion subunit vaccines offer advantages in stability, antigen display, and Fc-mediated immune engagement. This study aimed to design and evaluate a tetravalent RBD–Fc fusion construct incorporating RBDs from Wuhan-Hu-1 and Omicron BA.4/BA.5 and to determine whether this configuration can induce broad antibody recognition across SARS-CoV-2 variants. The objective was to assess its feasibility, biochemical properties, and initial immunogenicity. Methods: Immune responses to the construct were first assessed using the C-ImmSim simulation platform. The full-length fusion was synthesized, subcloned into pcDNA3.1(+), expressed in HEK293 cells, and purified by Protein G affinity chromatography. Protein integrity was evaluated by reducing SDS–PAGE. BALB/c mice (female, 8 weeks) were immunized with a prime–boost–boost schedule, and sera were analyzed by ELISA, considering binding to Wuhan-Hu-1, Omicron BA.4/BA.5, and a panel of RBD variants. Results: In silico analysis predicted coordinated antigen clearance, class switching, memory B- and CD4⁺ T-cell formation, and transient cytokine induction. The recombinant protein was expressed efficiently, yielding a major ~56 kDa band and a ~23 kDa RBD fragment. Vaccinated mice generated strong IgG responses to Wuhan-Hu-1 and BA.4/BA.5 RBDs and showed broad binding to major variant RBDs. Conclusions: The tetravalent RBD–Fc fusion vaccine was successfully produced and elicited broad antibody binding across SARS-CoV-2 variants, supporting its potential as a versatile protein-based vaccine platform. Full article

Background/Objectives: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can naturally infect a broad spectrum of animal species, with cats, minks, and ferrets being highly susceptible. There is a potential risk that infected animals could transmit viruses to humans. Moreover, SARS-CoV-2 continues to evolve via mutation and genetic recombination, resulting in the continuous emergence of new variants that have triggered a wave of reinfection. Therefore, safe and effective corona virus disease 2019 (COVID-19) vaccines for animals are still being sought. Methods: We generated three recombinant Modified vaccinia virus Ankara (MVAs) expressing the prefusion-stabilized S proteins, S6P, DS6P, and BA2S6P, targeting the full-length S protein genes of the ancestral, Delta, and Omicron BA.2 strains of SARS-CoV-2. Subsequently, the safety, immunogenicity, and protective efficacy of these MVA-based oral COVID-19 vaccine candidates were assessed in mice, minks, and cats. Results: These recombinant MVAs are safe in mice, minks, and cats. Oral or intramuscular vaccination with rMVA-S6P induced a robust SARS-CoV-2 neutralizing antibody (NA) response and conferred complete protection against the SARS-CoV-2 challenge in mice. Meanwhile, oral or intramuscular administration of these recombinant MVAs in combination induced a potent and durable NA response against homotypic SARS-CoV-2 pseudovirus in mice, minks, and cats, respectively. Conclusions: These findings suggest that the MVA-vectored vaccines are promising oral COVID-19 vaccine candidates for animals, and that the combined vaccination approach is an effective administration strategy for such vaccines. Full article

Background: With the emergence of SARS-CoV-2 Omicron sub-variants exhibiting increased transmissibility and immune escape, booster immunization is recommended. Ideally, vaccination across all age groups, including children and adolescents, is critical to control viral spread and reduce variant emergence. The heterologous scheme consisting of two doses of SOBERANA^® 02 followed by a third dose of SOBERANA^® Plus, which are recombinant protein subunit vaccines constructed from the ancestral RBD, has proven safety, immunogenicity, and effectiveness in pediatric populations as primary series. This study evaluated the safety and immunogenicity of a SOBERANA^® Plus booster dose administered six months after primary vaccination in individuals aged 3–18 years. Methods: In this follow-up analysis of a phase I/II trial, 244 participants received the booster. Safety was monitored via active surveillance at 1 h, 24 h, and over 28 days post-vaccination. Humoral responses were assessed 28 days post-booster. Antibody responses to the SARS-CoV-2 nucleocapsid (N) protein were assessed in all collected serum samples. Results: Adverse events occurred in 18% of participants, predominantly local (85.2%) versus systemic (14.8%); no serious or severe adverse events were reported. All humoral response parameters increased significantly post-booster, including neutralizing antibodies against D614G (24.7-fold increase) and Omicron BA.1 (55.9-fold increase), with similar responses in N-negative and N-positive individuals. Importantly, cross-neutralizing activity against recent Omicron sub-variants (XBB.1.5 and EG.5.1) was also detected. Conclusions: A SOBERANA^® Plus booster is safe and significantly enhances cross-neutralizing immunity against evolving Omicron sub-variants in children and adolescents. These results highlight the potential of first-generation RBD-based vaccines to maintain broad immunity despite viral evolution. Full article

Background/Objectives: Since its emergence in late 2019, SARS-CoV-2 has demonstrated remarkable genetic diversity driven by mutations and recombination events that shaped the course of the COVID-19 pandemic. Continuous genomic monitoring is essential to track viral evolution, assess the spread of variants of concern (VOCs), and inform public health strategies. The present study aimed to characterize the molecular epidemiology of SARS-CoV-2 in northern Greece from the first national case in February 2020 through early 2025. Methods: A total of 66 respiratory samples collected from hospitalized patients across Northern Greece were subjected to whole-genome sequencing using Oxford Nanopore Technologies’ MinION Mk1C platform and the ARTIC protocol. Sequences were analyzed with PANGO, Nextclade, and GISAID nomenclature systems for lineage and clade assignment, and the WHO nomenclature for VOCs. Results: Across 66 genomes, 34 PANGO lineages were identified. Early introductions included B.1 (2/66), B.1.177 (3/66), and B.1.258 (1/66). Alpha (5/66) and Beta (5/66) circulated in February–June 2021. Delta (AY.43) was detected in early 2022 (2/66; Jan–Feb) but was rapidly displaced by Omicron and reached 100% of the sequences by May 2022. Omicron diversified into BA.1/BA.1.1 (3/66), BA.2 (6/66), BA.4/BA.5 (14/66), BF.5 (1/66), EG.5 (1/66; designated a WHO Variant of Interest in 2023), JN.1 (4/66; globally dominant lineage prompting vaccine updates in 2024–2025), KS.1 (2/66; together with KS.1.1 are recognized PANGO lineages that were tracked internationally but remained less prevalent), KP.3 (5/66; together with KP.3.1.1, prominent “FLiRT” descendants circulating in 2024), and recombinants XDK, XDD, and XEC (5/66), reported by their PANGO names in accordance with the WHO’s current framework, which reserves Greek letters only for newly designated VOCs. Conclusions: This five-year genomic analysis provides an insight into the continuous evolution of SARS-CoV-2 in northern Greece. The findings underscore the importance of sustained genomic surveillance, integrated with epidemiological data, to detect emerging variants, monitor recombination, and strengthen preparedness for future coronavirus threats. Full article

Background: The Delta variant of SARS-CoV-2 virus, one of the alarming variants of concern (VOC) with a distinct mutation characteristic, was immensely detrimental and a significant cause of the prolonged pandemic waves. This study aimed to analyze the genetic characteristics of the predominant Delta variant in acute febrile illness (AFI) patients in Ethiopia. Method: Nasopharyngeal swab samples were collected from AFI patients in four hospitals from February 2021 to June 2022 and tested for SARS-CoV-2 by using RT-qPCR. Of 101 positive samples, 48 stored specimens were re-tested, and 26 with sufficient RNA quality (Ct < 30) were sequenced using whole-genome sequencing to identify variants of concern, specific virus lineages and mutation features. Result: Delta variants (21J clade) were found predominant among all the sequenced SARS-CoV-2 isolate (80.8%, 21/26). AY.120 (46.2%) and B.1.617.2 (26.9%) were the predominant sub-lineages of the Delta variant. Omicron (21k, Pango BA.1.1/BA.1.17/BA.1) and Alpha (20I, Pango B.1.1.7) variants accounted for 11.5% and 7.7% of the total sequenced samples. Phylogenetic analysis showed evidence of local transmission and possible multiple introductions of SARS-CoV-2 VOCs in Ethiopia. The number of mutations increases dramatically from Alpha (~35 avg) to Delta (~42 avg) to Omicron (~56 avg). The Delta variant revealed a spike mutation on L452R and T478K and P681R, and was characterized by the double deletion E156-F157- in Spike protein. Conclusions: The findings are indicative of a gradual change in the genetic coding of the virus underscoring the importance of ongoing genomic surveillance to track the evolution and spread of SARS-CoV-2 and other emerging virus. Full article

Neutralizing antibodies (nAbs) are key indicators of protection against SARS-CoV-2, and their measurement remains essential for monitoring vaccine responses and population immunity. While the plaque reduction neutralization test (PRNT) is the gold standard, it relies on replicative viruses and is not suited for high-throughput applications. Here, both an in-house and a commercial pseudovirus-based neutralization (PBN) assay were standardized and compared with PRNT to assess performance and concordance. The in-house PBN employed a VSV-ΔG pseudovirus encoding NanoLuc and displaying the SARS-CoV-2 Spike from the Wuhan or Omicron BA.1 variants in HEK293T-hACE2 cells, whereas the commercial assay (Integral Molecular, Philadelphia, PA, USA) used a lentiviral backbone with Renilla or GFP reporters and Wuhan or Omicron XBB.1.5/XBB.1.9 Spikes in Vero E6-ACE2-TMPRSS2 cells. Both assays showed strong correlations with PRNT, the commercial assay; moreover, they offered superior reproducibility and scalability, while the in-house version provided a cost-effective alternative suitable for BSL-2 settings. A total of 600 serum samples from vaccinated individuals were analyzed by commercial PBN at collection time points, from pre-vaccination to twelve months post–second dose, enabling large-scale screening, revealing marked differences in neutralization between Wuhan and Omicron XBB.1.5/1.9, and allowing unbiased classification of low, medium, and high responders using k-means clustering. The geometric mean titers (log₁₀ GMT) highlighted a ~1.5 log₁₀ (eightfold) reduction in neutralizing activity against Omicron, reflecting antibody waning and antigenic drift. Altogether, this study integrates assay standardization, PRNT comparison, and large-scale immune profiling, establishing a robust framework for harmonized pseudovirus-based neutralization testing. Full article

Immunosuppressive therapies used in clinics to reduce the risk of rejection in transplanted patients unfortunately also decrease the response of the immune system to the pathogens. Previous data has shown that the most diffuse SARS-CoV-2 variants of concern between 2020 and 2021 showed a different modulation of the host immune response in healthy subjects, with the Delta B.1.617.2 variant leading to a failure in the activation of the adaptive immune response. In this study, the transcriptomic profiles of monocyte-derived macrophages (MDM), isolated from four immunosuppressed kidney transplant patients and exposed to SARS-CoV-2 VOCs, were analyzed and compared with previously published data gathered from immune-competent subjects. Human monocytes were isolated from peripheral blood mononuclear cells (PBMCs) of four kidney transplant patients admitted to the IRCCS Policlinico San Matteo of Pavia (Italy), differentiated into macrophages, and exposed to the active and the UV-inactivated particles of the different SARS-CoV-2 VOCs (D614G, Alpha B.1.1.7, Gamma P.1, Delta B.1.617.2 and Omicron BA.1). Bulk RNA-Seq was performed and significant transcripts were assessed based on Student’s t-test (p-value < 0.05) and Fold change > 2. RNA-Seq data analyses of immunosuppressed MDMs showed that SARS-CoV-2 VOCs, although transcriptionally active, did not induce strong alterations in the transcriptomic profiles of these cells, while a strong down-regulation of key genes involved in the innate immunity pathways was observed when comparing these data to the ones obtained from immunocompetent participants. Overall, this study suggests that patients under immunosuppressive therapies do have an altered macrophage response to SARS-CoV-2 viral infection. Full article

The SARS-CoV-2 Omicron variant caused a global surge in COVID-19 cases following its emergence in November 2021, rapidly diversifying in the subsequent months. Although many studies have documented Omicron’s diversification, few have explored its impact on pediatric populations or the seasonality of other respiratory viruses in children. This study aims to investigate the diversity and circulation patterns of SARS-CoV-2 Omicron sublineages in pediatric patients in São Paulo, Brazil, and assess their co-circulation with other respiratory pathogens. Respiratory samples collected from patients under 18 years old across five hospitals between January 2022 and April 2023 were tested for different respiratory viruses using real-time RT-PCR. Whole-genome sequencing was performed on SARS-CoV-2-positive samples. Among the 7868 pediatric respiratory samples tested, 3902 were positive for viral pathogens. Respiratory Syncytial Virus accounted for the highest number of positive cases (n = 1248), exhibiting an atypical off-season peak in November 2022. SARS-CoV-2 was detected in 297 samples, of which 103 were sequenced. BA.1 and BA.5 sublineages had predominant genomic diversity and circulation time. These findings highlight the Omicron variant’s significant impact on the epidemiology and seasonal distribution of respiratory viruses in children, emphasizing the ongoing need for vaccination and robust surveillance efforts in pediatric populations. Full article

Throughout the 2019 coronavirus disease pandemic, various vaccine regimens were implemented. Real-world data comparing their effectiveness during the BA.1/BA.2 Omicron wave remain limited. We prospectively enrolled healthcare workers who had completed two doses of mRNA or ChAdOx1 (A) vaccine and received an mRNA vaccine booster (BNT162b2 (P) or mRNA-1273 (M)). Neutralizing antibody levels were measured 6 months after the primary vaccinations and 1 month post-booster vaccination using a surrogate virus neutralization assay. Breakthrough infections were identified through institutional surveillance and the national reporting system. Among 318 participants (P-P-P: 71; A-A-P: 205; A-P-P: 19; M-M-M: 23), pre-booster neutralizing activity was lowest in the ChAdOx1-primed groups. One month post-booster vaccination, the neutralizing activity exceeded 97% across all regimens. The cumulative incidence of breakthrough infection varied significantly from 43.7% (P-P-P) to 84.2% (A-P-P). In adjusted Cox models, A-P-P showed the highest infection risk (HR 2.99, 95% CI 1.65–5.42). In summary, mRNA boosters restored neutralizing activity, but during the early BA.1/BA.2 Omicron wave they were less effective in preventing infections regardless of disease severity. Therefore, antibody titers alone are insufficient for evaluating protection, underscoring the need for continuous monitoring to support timely policy decisions during epidemic surges. Full article

Background: The continuous evolution of SARS-CoV-2 necessitates the development of targeted strategies based on the immunological profiles of distinct age groups. Despite this imperative, comprehensive insights into the dynamics and broad-spectrum efficacy of neutralizing antibodies (NAbs) against emerging variants across different age groups, particularly in children, remain inadequate. Methods: Following the termination of China’s dynamic ‘zero-COVID-19’ policy in January 2023, which coincided with a widespread Omicron outbreak and numerous breakthrough infections, a longitudinal cohort study was established encompassing all age groups in Hubei, China. Follow-up assessments were conducted in March (Visit 1), June (Visit 2), and October (Visit 3) 2023. A total of 320 individuals were randomly selected and stratified into three age categories: children (<18 years, n = 80), adults (18–59 years, n = 167), and the elderly (≥60 years, n = 73). The NAbs against emerging SARS-CoV-2 variants BA.5, XBB.1.5, EG.5, and JN.1 were evaluated for each group. Trajectory modeling was employed to classify antibody trends into five categories: low-level stability, median-level stability, high-level stability, early increase, and late increase. Results: In March 2023, children exhibited significantly higher NAb levels against BA.5, XBB.1.5, EG.5, and JN.1 compared to adults and the elderly. However, these levels rapidly declined. From June to October 2023, no significant difference in NAb levels was observed between children and the other age groups. Regarding the broad-spectrum effectiveness of NAbs, the effectiveness in children was comparable to that of adults and the elderly in March 2023. However, from June to October 2023, children’s effectiveness became significantly lower than that of the other age groups. Trajectory analysis revealed that the highest proportions of high-level stability (31.3%) and median-level stability (42.5%) were observed among children. In contrast, adults and the elderly were most commonly categorized into the early increase (adult 46.7%, elderly 49.3%) and median-level stability (adult 22.1%, elderly 20.5%) categories. Conclusions: Although children initially demonstrate higher levels of NAbs, these levels decrease more rapidly than in adults and the elderly, eventually equalizing in later stages of recovery. Furthermore, the broad-spectrum effectiveness of NAbs in children is narrower than in other age groups. These findings suggest that children are at an elevated risk of infection with newly emerging variants, underscoring the urgent need to intensify focus on reinfections among children and develop tailored strategies to protect this vulnerable population. Full article

COVID-19 has triggered the rapid adoption of human organoid-based tissue culture models to overcome the limitations of the commonly used Vero cell line that did not fully recapitulate SARS-CoV-2 infection of human tissues. As the primary site of SARS-CoV-2 infection, the human nasal epithelium (HNE) cultivated in vitro and differentiated at air–liquid interface (ALI) is an ideal model to study infection processes and for testing anti-viral antibodies and drugs. However, the need for primary basal cells to establish the ALI-HNE limits the scalability of this model system. To try and bypass this bottleneck, we devised an ALI-differentiated form of the human adenocarcinoma cell line Calu-3, reported to model most aspects of authentic SARS-CoV-2 infection, including viral entry. The ALI-Calu-3 were tested for infection by a panel of SARS-CoV-2 variants, including ancestral (VIC01) and early pandemic lineages (VIC2089, Beta, Delta), and Omicron subvariants (BA2.75, BA4, BA5, XBB1.5). All tested lineages infected the ALI-HNE. In stark contrast, infection of the ALI-Calu-3 by Omicron subvariants BA4 and XBB1.5 was reduced. These data support the use of ALI-Calu-3 as a complementary, intermediary model for most but not all SARS-CoV-2 lineages, and places the ALI-HNE as the benchmark culture model for SARS-CoV-2 infection. Full article