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The mutational drift of SARS-CoV-2 and the appearance of multiple variants, including the latest Omicron variant and its sub-lineages, has significantly reduced (and in some cases abolished) the protective efficacy of Wuhan spike-antigen-based vaccines and therapeutic antibodies. One of the most functionally constrained and thus largely invariable regions of the spike protein is the one involved in the interaction with the ACE2 receptor mediating the cellular entry of SARS-CoV-2. Engineered ACE2, both as a full-length protein or as an engineered polypeptide fragment, has been shown to be capable of preventing the host-cell binding of all viral variants and to be endowed with potent SARS-CoV-2 neutralization activity both in vitro and in vivo. Here, we report on the biochemical and antiviral properties of rationally designed ACE2 N-terminal, three-helix fragments that retain a native-like conformation. One of these fragments, designated as PRP8_3H and produced in recombinant form, bears structure-stabilizing and binding-affinity enhancing mutations in α-helix-I and in both α-helix I and II, respectively. While the native-like, unmodified three α-helices ACE2 fragment proved to be thermally unstable and without any detectable pseudovirion neutralization capacity, PRP8_3H was found to be highly thermostable and capable of binding to the SARS-CoV-2 spike receptor-binding domain with nanomolar affinity and to neutralize both Wuhan and Omicron spike-expressing pseudovirions at (sub)micromolar concentrations. PRP8_3H thus lends itself as a highly promising ACE2 decoy prototype suitable for a variety of formulations and prophylactic applications. [ABSTRACT FROM AUTHOR]

The spread of SARS-CoV-2 has led to an interest in using ivermectin (a potent antiparasitic agent) as an antiviral agent despite the lack of convincing in vivo clinical data for its use against COVID-19. The off-target prophylactic use of ivermectin adds a substantial risk of drug–drug interactions with pharmaceutical medications used to treat chronic conditions like diabetes and hypertension (metformin and hydrochlorothiazide, respectively). Therefore, this study aims to evaluate the potential drug–drug interactions between ivermectin with either metformin or hydrochlorothiazide. In silico experiments and high-throughput screening assays for CYP3A4 were conducted to understand how metformin and hydrochlorothiazide might affect CYP3A4's role in metabolizing ivermectin. The study findings indicated that hydrochlorothiazide is more stable than both ivermectin and metformin. This conclusion was further supported by root mean square fluctuation analysis, which showed that hydrochlorothiazide is more flexible. The variation in the principal component analysis scatter plot across the first three normal modes suggests hydrochlorothiazide has a more mobile conformation than ivermectin and metformin. Additionally, a strong inhibition of CYP3A4 by hydrochlorothiazide was observed, suggesting that hydrochlorothiazide's regulatory effects could significantly impede CYP3A4 activity, potentially leading to a reduced metabolism and clearance of ivermectin in the body. Concurrent administration of these drugs may result in drug–drug interactions and hinder the hepatic metabolism of ivermectin. [ABSTRACT FROM AUTHOR]

Alpha-synuclein (α-syn) is a 140-amino-acid, intrinsically disordered, soluble protein that is abundantly present in the brain. It plays a crucial role in maintaining cellular structures and organelle functions, particularly in supporting synaptic plasticity and regulating neurotransmitter turnover. However, for reasons not yet fully understood, α-syn can lose its physiological role and begin to aggregate. This altered α-syn disrupts dopaminergic transmission and causes both presynaptic and postsynaptic dysfunction, ultimately leading to cell death. A group of neurodegenerative diseases known as α-synucleinopathies is characterized by the intracellular accumulation of α-syn deposits in specific neuronal and glial cells within certain brain regions. In addition to Parkinson's disease (PD), these conditions include dementia with Lewy bodies (DLBs), multiple system atrophy (MSA), pure autonomic failure (PAF), and REM sleep behavior disorder (RBD). Given that these disorders are associated with α-syn-related neuroinflammation—and considering that SARS-CoV-2 infection has been shown to affect the nervous system, with COVID-19 patients experiencing neurological symptoms—it has been proposed that COVID-19 may contribute to neurodegeneration in PD and other α-synucleinopathies by promoting α-syn misfolding and aggregation. In this review, we focus on whether SARS-CoV-2 could act as an environmental trigger that facilitates the onset or progression of α-synucleinopathies. Specifically, we present new evidence on the potential role of SARS-CoV-2 in modulating α-syn function and discuss the causal relationship between SARS-CoV-2 infection and the development of parkinsonism-like symptoms. [ABSTRACT FROM AUTHOR]

Respiratory infections caused by RNA viruses are a major contributor to respiratory disease due to their ability to cause annual epidemics with profound public health implications. Influenza A virus (IAV) infection can affect a variety of host signaling pathways that initiate tissue regeneration with hyperplastic and/or dysplastic changes in the lungs. Although these changes are involved in lung recovery after IAV infection, in some cases, they can lead to serious respiratory failure. Despite being ubiquitously observed, there are limited data on the regulation of long-term recovery from IAV infection leading to normal or dysplastic repair represented by inflammation-to-metaplasia transition in mice or humans. To address this knowledge gap, we used integrative bioinformatics analysis with further verification in vivo to elucidate the dynamic molecular changes in IAV-infected murine lung tissue and identified the core genes (Birc5, Cdca3, Plk1, Tpx2, Prc1. Rrm2, Nusap1, Spag5, Top2a, Mcm5) and transcription factors (E2F1, E2F4, NF-YA, NF-YB, NF-YC) involved in persistent lung injury and regeneration processes, which may serve as gene signatures reflecting the long-term effects of IAV proliferation on the lung. Further analysis of the identified core genes revealed their involvement not only in IAV infection but also in COVID-19 and lung neoplasm development, suggesting their potential role as biomarkers of severe lung disease and its complications represented by abnormal epithelial proliferation and oncotransformation. [ABSTRACT FROM AUTHOR]

Studies suggest that the dynamic changes in cellular response might correlate with disease severity and outcomes in SARS-CoV-2 patients. The study aimed to investigate the dynamic changes of lymphocyte subsets in patients with COVID-19. In this regard, 53 patients with COVID-19 were prospectively included, classified as mild, moderate, and severe. The peripheral lymphocyte profiles (LyT, LyB, and NK cells), as well as CD4+/CD8+, CD3+/CD19+, CD3+/NK and CD19+/NK ratios, and their dynamic changes during hospitalization and correlation with disease severity and outcome were assessed. We found significant differences in CD3+ lymphocytes between severity groups (p < 0.0001), with significantly decreased CD3+CD4+ and CD3+CD8+ in patients with severe disease (p < 0.0001 and p = 0.048, respectively). Lower CD3+/CD19+ and CD3+/NK ratios among patients with severe disease (p = 0.019 and p = 0.010, respectively) were found. The dynamic changes of lymphocyte subsets showed a significant reduction in NK cells (%) and a significant increase in CD3+CD4+ and CD3+CD8+ cells in patients with moderate and severe disease. The ROC analysis on the relationship between CD3+ cells and fatal outcome yielded an AUC of 0.723 (95% CI 0.583–0.837; p = 0.007), while after addition of age and SpO2, ferritin and NLR, the AUC significantly improved to 0.927 (95%CI 0.811–0.983), p < 0.001 with a sensitivity of 90.9% (95% CI 58.7–99.8%) and specificity of 85.7% (95% CI 69.7–95.2%). The absolute number of CD3+ lymphocytes might independently predict fatal outcomes in COVID-19 patients and T-lymphocyte subset evaluation in high-risk patients might be useful in estimating disease progression. [ABSTRACT FROM AUTHOR]

Background/Objectives: Extracorporeal membrane oxygenation (ECMO) has been widely used as a life support technique in COVID-19 acute respiratory distress syndrome (ARDS). The use of anticoagulation during ECMO support remains a topic of debate. The primary aim of this study is to demonstrate the safety and efficacy of using argatroban as an anticoagulant instead of heparin in patients with heparin-associated thrombocytopenia. Methods: 40 patients were enrolled and initially treated with unfractionated heparin for anticoagulation during ECMO, composing the UFH group. Twenty-one of these patients experienced a drop in platelet count to below 100,000 cells/mm3 and, after testing negative for IgG anti-PF4/heparin, the anticoagulation was switched to argatroban, composing the ARG group. Hemorrhagic events were recorded along with blood chemistry parameters. Results: Bleedings were significantly more frequent in the UFH group than in ARG group (58/579 days vs. 21/357 days, p = 0.041). No significant differences were observed in hemorrhagic episodes for each bleeding site, except for tracheal stoma (14 vs. 1, p = 0.011). No differences in activated partial thromboplastin time (aPTT) values were found between the two groups (aPTT 42.65 s vs. 44.70 s, p = 0.443). Linear regression analysis revealed that the platelet count on day 5 was correlated with the initial platelet count but not with the type of anticoagulant used (p = 0.001, CI 0.55, 0.69 and p = 0.078). Linear regression analysis in both groups showed a correlation between the duration of ECMO support and intensive care unit stay for the median aPTT and median platelet count. Furthermore, no major systemic thrombotic events or circuit clotting were observed in this patient cohort. Conclusions: Argatroban seems to be safe in patients with persistent heparin-associated thrombocytopenia undergoing ECMO. [ABSTRACT FROM AUTHOR]

Background/Objectives: COVID-19 led to a pandemic that has brought misery to millions of people but more so to those with pre-existing conditions. For this infection, several antiviral drugs were employed, including remdesivir (R) and Paxlovid (nirmatrelvir/ritonavir (NR)). Methods: The current study compared the effectiveness of remdesivir and Paxlovid treatment for COVID-19 patients with comorbid conditions. Data from a cohort of 151 adult patients with COVID-19 who also had associated comorbidities were used in this study. These patients were treated with antivirals according to local guidelines. The subjects included 78 case-patients assigned to group R and 73 to group NR. Results: In group NR, a considerable improvement in oxygen saturation was seen in the first 24 h of treatment (p = 0.010), but the levels were significantly higher from the second day of treatment (p < 0.001) in group R of patients. At the end of the 5 days of treatment, the oxygen saturation improved statistically significantly compared to the admission day, but only in the R group (95.11 ± 1.80; 91.76 ± 1.80; p < 0.001). Conclusions: Both drugs can be considered a breakthrough in the current treatment approach to the COVID-19 disease since they provide readily available options that can alleviate the severity of the disease and, hence, the prognosis of patients. That is why their effectiveness relies on the correct administration time and choosing the patient with suitable characteristics regarding the presence of comorbidities and the likelihood of the critical further development of the process. [ABSTRACT FROM AUTHOR]

Background: During the COVID-19 pandemic, paracetamol was widely recommended in different clinical settings, and sometimes advised over non-steroidal anti-inflammatory drugs (NSAIDs). These recommendations sparked a strong debate, with reports suggesting either potential benefits or harms for the individuals infected with SARS-CoV-2. As no systematic review is available, we performed a meta-analysis to estimate the impact of paracetamol on COVID-19 clinical outcomes compared to a placebo, no use, or NSAIDs. Methods: We searched PubMed, Scopus, Web of Science, and ClinicalTrials.gov for randomized trials or observational studies evaluating any COVID-19 clinical outcome. Data were combined using a generic inverse-variance approach. The Grading of Recommendations, Assessment, Development, and Evaluations (GRADE) approach was used to determine the certainty of evidence for each outcome. Results: One randomized trial and five observational studies, enrolling over 34,000 patients, were included. Overall, as compared to the patients using NSAIDs or receiving no treatment, the individuals who received paracetamol showed no significant differences in the risk of death (summary relative risks 0.93 and 0.91, respectively: both p > 0.05), need to transfer to the intensive care unit, need for respiratory support, or cardiovascular or renal complications. All studies showed a high risk of bias, with a low overall quality of evidence. Conclusions: This meta-analysis found no evidence of harmful or beneficial effects of paracetamol on main COVID-19-related outcomes. Also, the current literature does not provide sufficient data to support a preferential choice between paracetamol and NSAIDs for COVID-19 symptoms management. Further research is needed to confirm the present findings and provide critical insights on the policies to adopt in the case of future pandemics. [ABSTRACT FROM AUTHOR]

Introduction: Healthcare workers (HCWs) are at higher risk of SARS-CoV-2 infection. Viral surveillance for early detection of COVID-19 is a critical strategy to understand this population's infection dynamics and prevent transmission. The study examines SARS-CoV-2 infection and reinfection among HCWs vaccinated against COVID-19 working at a primary healthcare unit serving a disenfranchised community in Brazil. Methods: The study was conducted in Cidade Estrutural, Federal District, Brazil, between February and October 2021. Participants were interviewed and provided samples. A prospective open cohort study was used to analyze the frequency of SARS-CoV-2 infection and reinfection, and the vaccine-induced seroconversion. Nasopharyngeal swab specimen was collected from workers presenting with flu-like symptoms and subjected to RT-qPCR. Peripheral blood samples were also collected every 30 ± 2 days for eight months, starting from the day participants received their first dose of COVID-19 vaccine, and submitted to serological testing (IgM and IgG chemiluminescence). The frequencies of infection and reinfection (RT-qPCR positive results 90 days after the infection) were calculated along with their respective confidence intervals (95% CI). Results: Of the 128 workers, 61 (47.65%; CI: 39.19–56.25) reported probable SARS-CoV-2 infection before vaccination and 50 (39.06%; CI: 31.04–47.71) had SARS-CoV-2 infection after vaccination, confirmed by molecular test. Reinfection was identified in seven workers (7/50, 14%; CI: 6.95–26.18) based on the 90-day interval between results. The serological data from the 128 workers during the cohort indicated that 68 (53.12%; CI: 44.5–61.5) had IgG antibodies and 46 had IgM antibodies (35.93%; CI: 28.14–44.54) against SARS-CoV-2. SARS-CoV-2 infection was common in 56% of the community health workers (CHWs), 50% of registered nurses, and licensed vocational nurses (33%). Following the COVID-19 vaccination, the percentage of infections among HCWs decreased from 47.83% to 4.35%. Conclusion: These results demonstrate that (i) approximately 40% of the workers were infected with SARS-CoV-2 in 2021 and (ii) reinfections confirmed by RT-qPCR occurred in 14% of the HCWs after vaccination. The results provide valuable insights into the circulation of SARS-CoV-2 among HCWs in a primary care unit serving a minoritized community. [ABSTRACT FROM AUTHOR]

Background and Objectives: The COVID-19 pandemic posed significant challenges to healthcare systems worldwide, and mortality rates were driven by a complex interaction of patient-specific factors, one of the most important being those related to the scheduling of invasive mechanical ventilation. This study examined the sociodemographic, clinical, and ventilatory factors associated with mortality in COVID-19 patients admitted to the ICU of a hospital in Colombia. Methods: A retrospective cohort study was conducted, involving 116 patients over the age of 18 who were admitted to the ICU with a confirmed diagnosis of COVID-19 between March 2020 and May 2021. Data were collected from the patients' medical records. Statistical analysis was performed using SPSS version 24®. Odds ratios (OR) and 95% confidence intervals were calculated to identify factors associated with COVID-19 mortality, followed by adjustment through binary logistic regression. Results: It was found that 65.5% of the patients were male, with a mean age of 64 ± 14 years, and the overall mortality rate was 49%. Factors significantly associated with higher mortality included male sex (OR: 6.9, 95% CI: 1.5–31.7), low oxygen saturation on admission (OR: 7.6, 95% CI: 1.1–55), and PEEP settings at 96 h (OR: 8, 95% CI: 1.4–45). Mortality was not influenced by socioeconomic status or health system affiliation. Conclusions: This study identified male sex, age over 65 years, PEEP greater than 10 cmH2O at 96 h of mechanical ventilation, and low oxygen saturation as significant factors associated with higher mortality in COVID-19 patients, while no significant associations were found with socioeconomic status or health system affiliation. These findings highlight the importance of focusing on clinical management and ventilatory strategies in reducing mortality, particularly for high-risk groups, rather than relying on socioeconomic factors as predictors of outcomes. [ABSTRACT FROM AUTHOR]

Graphene-enhanced surface plasmon resonance (SPR) biosensors offer promising advancements in viral detection, particularly for SARS-CoV-2. This study presents the design and optimization of a multilayer SPR biosensor incorporating silver, silicon nitride, single-layer graphene, and thiol-tethered ssDNA to achieve high sensitivity and specificity. Key metrics, including SPR angle shift (Δθ), sensitivity (S), detection accuracy (DA), and figure of merit (FoM), were assessed across SARS-CoV-2 concentrations from 150 to 525 mM. The optimized biosensor achieved a sensitivity of 315.91°/RIU at 275 mM and a maximum Δθ of 4.2° at 400 mM, demonstrating strong responsiveness to virus binding. The sensor maintained optimal accuracy and figure of merit at lower concentrations, with a linear sensitivity response up to 400 mM, after which surface saturation limited further responsiveness. These results highlight the suitability of the optimized biosensor for real-time, point-of-care SARS-CoV-2 detection, particularly at low viral loads, supporting its potential in early diagnostics and epidemiological monitoring. [ABSTRACT FROM AUTHOR]

A list of diagnosis codes mapped to CDC-defined high-risk conditions for severe COVID-19 outcomes is currently not available in the literature. We reviewed the CDC list of underlying conditions associated with severe COVID-19 and a coding expert and two clinicians mapped the relevant high-risk conditions to the appropriate ICD-10-CM codes. We additionally assessed the prevalence of these conditions in the Optum de-identified-Clinformatics® Data Mart Database and the Optum de-identified Electronic Health Record dataset. A comprehensive list of approximately 8200 codes were mapped to the CDC-defined high-risk underlying conditions; these ICD-10-CM codes were stratified into three groups corresponding with the CDC strength of evidence category (conclusive, suggestive, or mixed evidence). Applying these codes to administrative claims and EHR datasets demonstrated a consistent prevalence of high-risk conditions over four years (2018–2021). These findings present a comprehensive list of codes that can be used by clinicians and researchers to identify and characterize patients at high risk for severe COVID-19 outcomes. [ABSTRACT FROM AUTHOR]

Over 80% of the world's deadliest pandemics are caused by viral infections, and vaccination remains the most effective way to prevent these infections from spreading. Since the discovery of the first vaccine over two centuries ago, several vaccine design technologies have been developed. Next-generation vaccines, based on mRNA and viral vector technologies, have recently emerged as alternatives to traditional vaccines. Adenoviral vector-based vaccines against coronavirus disease 2019 have demonstrated a more sustained antibody response as compared to mRNA vaccines. However, this has not been without complications, with a few cases of severe adverse events identified in vaccinated individuals, and the underlying mechanism is the subject of intense investigation. Adeno-associated viral vectors induce a weaker cellular immune response compared to adenoviral vectors, and it is mainly for this reason that there has been a diminished interest in exploring them as a vaccine platform until recently. This review will discuss recent developments and the potential of adeno-associated viral vectors as anti-viral vaccines. [ABSTRACT FROM AUTHOR]

Objectives: SARS-CoV-2 caused the COVID-19 pandemic, which overwhelmed global healthcare systems. Over 776 million COVID-19 cases and more than 7 million deaths were reported by WHO in September 2024. COVID-19 vaccination is crucial for preventing infection and controlling the pandemic. Here, we describe the design and development of a next-generation multi-epitope vaccine for SARS-CoV-2, consisting of T cell epitopes. Methods: Immunoinformatic methods were used to derive models for the selection of MHC binders specific for the mouse strain used in this study among a set of human SARS-CoV-2 T cell epitopes identified in convalescent patients with COVID-19. The immunogenicity of the vaccine prototype was tested on humanized-ACE2 transgenic B6.Cg-Tg(K18-ACE2)2Prlmn/J mice by in vitro, in vivo, and ex vivo immunoassays. Results: Eleven binders (two from the Envelope (E) protein; two from the Membrane (M) protein; three from the Spike (S) protein; and four from the Nucleocapsid (N) protein) were synthesized and included in a multi-epitope vaccine prototype. The animals were immunized with a mix of predicted MHC-I, MHC-II, or MHC-I/MHC-II peptide epitopes in Complete Freund's Adjuvant, and boosted with peptides in Incomplete Freund's Adjuvant. Immunization with SARS-CoV-2 epitopes remodeled the lymphocyte profile. A weak humoral response and the significant production of IL-4 and IFN-γ from T cells were found after the vaccination of the animals. Conclusions: The multi-epitope vaccine prototype presented in this study demonstrates immunogenicity in mice and shows potential for human vaccine construction. [ABSTRACT FROM AUTHOR]

Long COVID or Post-Acute Sequelae of SARS-CoV-2 infection (PASC) is a condition characterized by numerous lingering symptoms that persist for weeks to months following the viral illness. While treatment for PASC is still evolving, several therapeutic approaches beyond traditional antiviral therapies are being investigated, such as immune-modulating agents, anti-inflammatory drugs, and various supportive interventions focusing at alleviating symptoms and enhancing recovery. We aimed to summarize the breadth of available evidence, identify knowledge gaps, and highlight promising non-antiviral therapies for Long COVID/PASC. We followed the framework of a scoping methodology by mapping existing evidence from a range of studies, including randomized clinical trials, observational research, and case series. Treatments evaluated include metformin, low-dose naltrexone (LDN), dexamethasone, statins, omega-3 fatty acids, L-arginine, and emerging therapies like intravenous immunoglobulin (IVIg) and therapeutic apheresis. Early findings suggest that metformin has the strongest clinical evidence, particularly from large phase 3 trials, while LDN and dexamethasone show potential based on observational studies. However, many treatments lack robust, large-scale trials. This review emphasizes the need for further research to confirm the efficacy of these treatments and guide clinical practice for Long COVID management. [ABSTRACT FROM AUTHOR]

Increasing evidence suggests bats are the ancestral hosts of the majority of coronaviruses. In general, coronaviruses primarily target the gastrointestinal system, while some strains, especially Betacoronaviruses with the most relevant representatives SARS-CoV, MERS-CoV, and SARS-CoV-2, also cause severe respiratory disease in humans and other mammals. We previously reported the susceptibility of Rousettus aegyptiacus (Egyptian fruit bats) to intranasal SARS-CoV-2 infection. Here, we compared their permissiveness to an oral infection versus respiratory challenge (intranasal or orotracheal) by assessing virus shedding, host immune responses, tissue-specific pathology, and physiological parameters. While respiratory challenge with a moderate infection dose of 1 × 104 TCID50 caused a systemic infection with oral and nasal shedding of replication-competent virus, the oral challenge only induced nasal shedding of low levels of viral RNA. Even after a challenge with a higher infection dose of 1 × 106 TCID50, no replication-competent virus was detectable in any of the samples of the orally challenged bats. We postulate that SARS-CoV-2 is inactivated by HCl and digested by pepsin in the stomach of R. aegyptiacus, thereby decreasing the efficiency of an oral infection. Therefore, fecal shedding of RNA seems to depend on systemic dissemination upon respiratory infection. These findings may influence our general understanding of the pathophysiology of coronavirus infections in bats. [ABSTRACT FROM AUTHOR]

In the frame of the I-MOVE-COVID-19 project, a cohort of 2050 patients admitted in two Spanish reference hospitals between March 2020 and December 2021 was selected and a range of clinical factor data were collected at admission to assess their impact on the risk COVID-19 severity outcomes through a multivariate adjusted analysis and nomograms. The need for ventilation and intensive care unit (ICU) admission were found to be directly associated with a higher death risk (OR 6.9 and 3.2, respectively). The clinical predictors of death were the need for ventilation and ICU, advanced age, neuromuscular disorders, thrombocytopenia, hypoalbuminemia, dementia, cancer, elevated creatin phosphokinase (CPK), and neutrophilia (OR between 1.8 and 3.5), whilst the presence of vomiting, sore throat, and cough diminished the risk of death (OR 0.5, 0.2, and 0.1, respectively). Admission to ICU was predicted by the need for ventilation, abdominal pain, and elevated lactate dehydrogenase (LDH) (OR 371.0, 3.6, and 2.2, respectively) as risk factors; otherwise, it was prevented by advanced age (OR 0.5). In turn, the need for ventilation was predicted by low oxygen saturation, elevated LDH and CPK, diabetes, neutrophilia, obesity, and elevated GGT (OR between 1.7 and 5.2), whilst it was prevented by hypertension (OR 0.5). These findings could enhance patient management and strategic interventions to combat COVID-19. [ABSTRACT FROM AUTHOR]

SARS-CoV-2 utilizes the angiotensin-converting enzyme 2 (ACE2) receptors located on membranes to enter host cells. Nevertheless, the ACE2 gene primarily encodes for a zinc metalloproteinase, which is a part of the renin–angiotensin system (RAS). ACE2 downregulation results in the deregulation of RAS in favor of pro-fibrosis, pro-apoptosis, oxidative stress, pro-inflammation, aldosterone production and release, and blood vessel contraction axis. ACE2 is highly expressed in the placenta. There are both axes of the RAS system in the placenta. This study aims to assess the perinatal outcomes with ACE2 receptor polymorphisms in pregnant women infected with SARS-CoV-2 during pregnancy. The case-control study was conducted to determine the association of ACE2 single-nucleotide polymorphisms in 171 COVID-19-positive pregnant subjects and 112 control subjects. The recessive mutations of rs2158082 and rs4830974 were associated with an increased risk of low birthweight and preterm birth, whereas the dominant mutation of rs2285666 (CT + TT) was associated with decreased odds of low birthweight. COVID-19 was not a significant factor contributing to the adverse perinatal outcomes in our sampling. These findings may help to clarify the controversy regarding the increased risk of adverse perinatal outcomes observed during COVID-19 as well as provide new perspectives for research on the genetic factors associated with a higher risk of adverse perinatal outcomes. [ABSTRACT FROM AUTHOR]

The coronavirus disease 2019 (COVID-19) pandemic, driven by the emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has been characterized by the virus's ongoing evolution, leading to the appearance of more transmissible variants that have often triggered infection surges. In this study, we analyzed the SARS-CoV-2 epidemic in Cyprus, utilizing 1627 viral sequences from infected individuals between November 2022 and February 2024. Over this period, 251 distinct lineages and sublineages were identified, predominantly categorized into three groups: Omicron 5, XBB, and JN.1 (parental lineage BA.2.86), all of which harbor S protein mutations linked to enhanced transmissibility and immune escape. Despite the relatively low numbers of new infections during this period, and the lack of any major waves, unlike earlier phases of the pandemic, these lineages demonstrated varying periods of dominance, with Omicron 5 prevailing from November 2022 to February 2023, XBB variants leading from March to November 2023, and JN.1 generating a wavelet from December 2023 to February 2024. These findings suggest that the SARS-CoV-2 epidemic in Cyprus has reached endemicity, with new variants gradually replacing previously circulating variants irrespective of seasonal patterns. This study highlights the critical importance of ongoing surveillance of SARS-CoV-2 evolution in Cyprus and emphasizes the role of preventive measures in limiting virus transmission, providing valuable insights for safeguarding public health. [ABSTRACT FROM AUTHOR]

During the COVID-19 pandemic, veterinary diagnostic laboratories tested both human and animal samples and needed to ensure that they could accurately perform large numbers of diagnostic tests in a timely manner. Sample pooling, a methodology used effectively for over 80 years as a surveillance tool for screening large numbers of potentially infected individuals, was employed. Given its sensitivity, real-time polymerase chain reaction (PCR) is more suitable for employing this strategy, as compared to other less sensitive testing methods. In this study, we evaluated the capability of detecting SARS-CoV-2 in both 5-sample and 10-sample pools of feces using real-time reverse transcriptase polymerase chain reaction (rRT-PCR) as well as determined the level of sensitivity. A blinded method test (BMT) by an independent laboratory was conducted to assess the five-sample fecal pool. To complement detection capability, the stability of the genome within a PBS fecal suspension was measured under various time and temperature conditions across a 28-day period. Our results showed that the limit of detection for 5-sample and 10-sample fecal pools is 12.8 and 6.4 genome copies in a 25 µL PCR, respectively. The 5-sample and 10-sample pooling resulted in a cycle threshold (Ct) value loss of 2.35 and 3.45, as compared to Ct values of known positive individual samples, but consistent detection was still achieved in pools containing positive samples with an original Ct below 36 and 34, respectively. The simulation of clinical five-sample pooling showed that all positive samples could be detected regardless of the number (1–3) of positive samples in each pool. The BMT results demonstrated excellent sensitivity (100 copies/reaction) in five-sample pools for the detection of SARS-CoV-2 RNA even though a fecal matrix effect was observed. Finally, our results show that the SARS-CoV-2 genome remains stable over a wide range of time and temperature variations. Overall, our findings provide solid data to scale up SARS-CoV-2 testing capacity in veterinary diagnostic laboratories. [ABSTRACT FROM AUTHOR]

This retrospective study analysed the routine data obtained by multiplex real-time RT-qPCR methods for respiratory virus detection. A total of 4814 respiratory specimens collected during 1 September 2022–31 August 2023 were included in the study. A total of 38% of the specimens were positive for at least one target, with the incidence maximum (82%) for the small children (age group 0–4 years). The five dominant virus groups were rhinovirus (RV, 12%), influenza virus A (IAV, 7%), adenovirus (AdV, 6%), respiratory syncytial virus (RSV, 5%), and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2, 5%). The specimens with multi-detections represented 19% of the positives, unevenly distributed (n = 225, 56, 43, 24) among the age groups 0–4, 5–14, 15–64, and 65< years, respectively. The dominant virus groups in multi-positive specimens were RV (53%), AdV (43%), and bocavirus (BoV, 35%)—in mutual pairs as well as all three together—followed by RSV (21%), and IAV (15%). Our study focused on the specimens with codetections and provides an insight into the variety of the respiratory virus interactions in Latvia during the first year since pandemic-related social restriction measures were eased. The observations also emphasise the need to consider the differentiation between rhinoviruses and enteroviruses, especially for the youngest patients in the age group 0–4. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was responsible for causing the COVID-19 pandemic. Intensive research has illuminated the complex biology of SARS-CoV-2 and its continuous evolution during and after the COVID-19 pandemic. While much attention has been paid to the structure and functions of the viral spike protein and the entry step of viral infection, partly because these are targets for neutralizing antibodies and COVID-19 vaccines, the later stages of SARS-CoV-2 replication, including the assembly and egress of viral progenies, remain poorly characterized. This includes insight into how the activities of the viral structural proteins are orchestrated spatially and temporally, which cellular proteins are assimilated by the virus to assist viral assembly, and how SARS-CoV-2 counters and evades the cellular mechanisms antagonizing virus assembly. In addition to becoming infectious, SARS-CoV-2 progenies also need to survive the hostile innate and adaptive immune mechanisms, such as recognition by neutralizing antibodies. This review offers an updated summary of the roles of SARS-CoV-2 structural proteins in viral assembly, the regulation of assembly by viral and cellular factors, and the cellular mechanisms that restrict this process. Knowledge of these key events often reveals the vulnerabilities of SARS-CoV-2 and aids in the development of effective antiviral therapeutics. [ABSTRACT FROM AUTHOR]

Background The development of vaccines against SARS-CoV-2 has proved to be a highly successful strategy. In this work, the aim is to study the effects of the SARS-CoV-2 vaccine on the production of inflammatory markers and how this affect morbidity and mortality. Electronic medical record (EMR) data from 210 patients diagnosed with COVID-19 from November 2020 to June 2021 were collected. The admitted patients were divided into three groups, the one-dose vaccinated, two-dose vaccinated, and the non-vaccinated. All patients were moderate or severe in disease level as defined by the WHO classification. The results show that CRP was 101 ± 5.3, 97 ± 10.8, and 145 ± 17.3 (p < 0.05), fibrinogen 529 ± 16.3, 397 ± 33.8, and 610 ± 15 (p < 0.05), D-dimer 1244 ± 89, 1279 ± 297, and 1615 ± 224 (p < 0.05), ferritin was 1170 ± 122, 999 ± 202, and 1663 ± 409 (p < 0.05), IL-6 was 196 ± 12, 96 ± 5, and 580 ± 402 (NS), for the non-vaccinated, one-dose vaccinated, and two-dose vaccinated groups, respectively. The high level of CRP up to 150–200 mg/dL was more common among the surviving vaccinated patients. Oxygen supplementation, mechanical ventilation, and mortality were higher in the non-vaccinated group. Blood urea nitrogen (BUN) level was higher in the vaccinated patients, 25 ± 0.14 vs. 33 ± 6.15, respectively (p < 0.05). Inflammation markers were significantly higher in the vaccinated groups compared to non-vaccinated groups. On the other hand, extremely high levels of CRP (>200 mg/dL) were correlated with high mortality incidence. [ABSTRACT FROM AUTHOR]

The SARS-CoV-2 outbreak has resulted in a considerable number of deaths worldwide. The virus damages the pulmonary artery endothelium, leading to a condition known as microvascular pulmonary inflammatory thrombotic syndrome (MPITS), which can be fatal and cause multiple organ failure. The presence of preexisting comorbidities has been shown to significantly impact the severity and prognosis of patients with SARS-CoV-2 infection. The objective of this study was to compare the age groups of patients with coronavirus disease 2019 (COVID-19) and to identify the prevalence of comorbidities associated with death and survival in an area of southern Italy. The data set consisted of 1985 patients with confirmed cases of SARS-CoV-2 infection who were admitted to the A.O.U. San Giovanni di Dio e Ruggi d'Aragona Hospital in Salerno between January 2021 and December 2022. The results were presented for the overall population and stratified by outcome and age group. All analyses were performed using the XLSTAT (Lumivero, 2024, Paris, France) and STATA software (release 16.1, StataCorp LLG, College Station, TX, USA, 2019) packages. In the study, population, 636 cases (32%) resulted in death, with a higher prevalence in the 60–79 age group, followed by the ≥80 and 30–59 age groups. The most prevalent diseases among deceased and surviving patients with confirmed cases of SARS-CoV-2 infection were those affecting the circulatory system (61.5% vs. 55.5%), the respiratory system (55.8% vs. 26.2%), and the metabolic system (25.9% vs. 25.4%). In patients aged 30–79, respiratory diseases were the primary cause of mortality, whereas in those aged ≥80, circulatory system diseases were more prevalent. Among survivors, cardiovascular diseases were the most common comorbidities across all age groups, followed by respiratory diseases and endocrine, metabolic, and immune disorders. Moreover, these comorbidities were associated with an elevated risk of mortality. The study emphasizes the substantial influence of age and comorbidities on the mortality associated with SARS-CoV-2 infection. These findings highlight the necessity for targeted interventions to manage comorbid conditions in patients with SARS-CoV-2 infection, particularly in older adults. [ABSTRACT FROM AUTHOR]

Background/Objectives: COVID-19 is characterised by a wide variety of clinical manifestations, and clinical tests and genetic analysis might help to predict patient outcomes. Methods: In the current study, the expression of genes related to immune response (CCL5, IFI6, OAS1, IRF9, IL1B, and TGFB1) was analysed in the upper airway and paired-blood samples from 25 subjects infected with SARS-CoV-2. Relative gene expression was determined by RT-qPCR. Results: CCL5 expression was higher in the blood than in the upper airway (p < 0.001). In addition, a negative correlation was found between IFI6 and viral load (p = 0.033) in the upper airway, suggesting that the IFI6 expression inhibits the viral infection. Concerning sex, women expressed IL1B and IRF9 in a higher proportion than men at a systemic level (p = 0.008 and p = 0.049, respectively). However, an increased expression of IRF9 was found in men compared to women in the upper airway (p = 0.046), which could be due to the protective effect of IRF9, especially in men. Conclusions: The higher expression of CCL5 in blood might be due to the key role of this gene in the migration and recruitment of immune cells from the systemic circulation to the lungs. Our findings confirm the existence of sex differences in the immune response to early stages of the infection. Further studies in a larger cohort are necessary to corroborate the current findings. [ABSTRACT FROM AUTHOR]

Simple Summary: This review explores recent findings on the role of neurotrophins, particularly NGF and BDNF, in the neurological and immune-related consequences of SARS-CoV-2 infection and its long-term outcomes. Neurotrophins (NTs) constitute a family of small protein messengers that play a fundamental role in both the central and peripheral nervous systems. In particular, the nerve growth factor (NGF) and the brain-derived neurotrophic factor (BDNF) play a subtle role in the survival, differentiation, and functioning of neuronal populations, as well as in the fine regulation of immune functions. The SARS-CoV-2 infection was characterized by a sequela of symptoms (serious respiratory pathology, inflammatory storm, neurological discomfort, up to the less serious flu-like symptoms), which caused, at the end of 2023, more than 7 million deaths worldwide. Despite the official end of the pandemic, the physical and psychological consequences are currently the object of scientific research, both acute and chronic/long-lasting (Long-COVID-19). Given the multifactorial nature of the outcomes of SARS-CoV-2 infection in adults and children, several studies have investigated the potential involvement of the NGF and BDNF systems in the pathology. This narrative review aims to summarize the most recent evidence on this crucial topic. [ABSTRACT FROM AUTHOR]

The gut microbiota metabolizes flavonoids, amino acids, dietary fiber, and other components of foods to produce a variety of gut microbiota-derived metabolites. Flavonoids are the largest group of polyphenols, and approximately 7000 flavonoids have been identified. A variety of phenolic acids are produced from flavonoids and amino acids through metabolic processes by the gut microbiota. Furthermore, these phenolic acids are easily absorbed. Phenolic acids generally represent phenolic compounds with one carboxylic acid group. Gut microbiota-derived phenolic acids have antiviral effects against several viruses, such as SARS-CoV-2 and influenza. Furthermore, phenolic acids influence the immune system by inhibiting the secretion of proinflammatory cytokines, such as interleukin-1β and tumor necrosis factor-α. In the nervous systems, phenolic acids may have protective effects against neurodegenerative diseases, such as Alzheimer's and Parkinson's diseases. Moreover, phenolic acids can improve levels of blood glucose, cholesterols, and triglycerides. Phenolic acids also improve cardiovascular functions, such as blood pressure and atherosclerotic lesions. This review focuses on the current knowledge of the effects of phenolic acids produced from food-derived flavonoids and amino acids by the gut microbiota on health and disease. [ABSTRACT FROM AUTHOR]

The design and synthesis of antiviral compounds were guided by computationally predicted data against highly conserved non-structural proteins (Nsps) of the SARS-CoV-2 coronavirus. Chromenephenylmethanone-1 (CPM-1), a novel biphenylpyran (BPP), was selected from a unique natural product library based on in silico docking scores to coronavirus Nsps with high specificity to the methyltransferase protein (2′-O-MTase, Nsp10–16), which is responsible for viral mRNA maturation and host innate immune response evasion. To target the 2′-O-MTase, CPM-1, along with intermediate BPP regioisomers, tetrahydrophenylmethanones (TPMs), were synthesized and structurally validated via nuclear magnetic resonance (NMR) data and DP4+ structure probability analyses. To investigate the activity of these BPPs, the following in vitro assays were conducted: SARS-CoV-2 inhibition, biochemical target validation, mutagenicity, and cytotoxicity. CPM-1 possessed notable activity against SARS-CoV-2 with 98.9% inhibition at 10 µM and an EC50 of 7.65 µM, as well as inhibition of SARS-CoV-2's 2′-O-MTase (expressed and purified) with an IC50 of 1.5 ± 0.2 µM. In addition, CPM-1 revealed no cytotoxicity (CC50 of >100 µM) or mutagenicity (no frameshift or base-pair mutations). This study demonstrates the potential of computational modeling for the discovery of natural product prototypes followed by the design and synthesis of drug leads to inhibit the SARS-CoV-2 2′-O-MTase. [ABSTRACT FROM AUTHOR]

Although a fourth dose of SARS-CoV-2 vaccine was shown to be effective, the immunogenicity of a fifth dose in immunocompetent individuals had not been well described. This was a prospective observational cohort study of previously vaccinated healthcare workers at a single tertiary hospital in Israel. Individuals were administered up to three booster doses of the BNT162b2 mRNA vaccine (i.e., up to five overall doses), during the period between July 2021 and January 2023. Immunogenicity was assessed using the SARS-CoV-2 IgG (sCOVG) semi-quantitative assay, performed at several time points. The cohort consisted of 162 individuals (median age 69 years, 62% female). Of these, 104 (64%) received four doses and 58 (36%) received five doses. Anti-SARS-CoV-2 antibody levels increased in all cases, regardless of the baseline levels. The fold-change increase in the mean sCOVG index was 29.2 (SD 2.6) after the third vaccine, 3.8 (SD 2.4) after the fourth vaccine, and 3.6 (SD 3.0) after the fifth vaccine. A waning effect over time was seen in 78% and 43% of participants for the third and fourth doses, respectively. Adverse events following the fifth dose were limited and mild. Similar to previous booster vaccines, a fifth dose of BNT162b2 is immunogenic and safe in healthy individuals, although the clinical implications remain unclear. [ABSTRACT FROM AUTHOR]

Background/Objectives: Severe clinical course and mortality from COVID-19 are mostly associated with increased concentrations of IL-6 and IL-10. Findings from clinical trials suggest that both statins and renin–angiotensin–aldosterone system inhibitors (RAASI) might have the potential to reduce unfavorable outcomes in patients with COVID-19. The aim of this study was to evaluate the effect of statins and RAASI on the cytokine concentrations in COVID-19 patients. Methods: SARS-CoV-2 infected patients were enrolled in this study, and demographic, clinical, and routine laboratory data were evaluated. Plasma cytokine levels were measured by multiplex assay. Results: COVID-19 patients with chronic cardiovascular diseases (CVD) had significantly lower median plasma IL-6 levels than COVID-19 patients with no co-morbidities (26 vs. 53 pg/mL, p = 0.021). COVID-19 patients with CVD who were taking statins had significantly lower median concentrations of IL-6 (21 vs. 44 pg/mL, p = 0.027), TNFα (21 vs. 39.5 pg/mL, p = 0.036), and IL-10 (19 vs. 25.5 pg/mL, p = 0.025) compared to COVID-19 patients with no CVD. In a binary logistic regression model, IL-6 was a significant variable, with an odds ratio value of 0.961 (95% CI 0.929–0.995). Regarding RAASI, only plasma IL-6 (22 vs. 44 pg/mL, p = 0.012) levels were found to be significantly lower in COVID-19 patients with CVD consuming these medications compared to patients who did not have any CVD. Conclusions: COVID-19 patients who had chronic cardiovascular co-morbidities and who were administered statins or RAASI had significantly lower concentrations of IL-6 than COVID-19 patients who did not have any co-morbidities. These findings suggest that the use of statins or RAASI may be of value in COVID-19 patients. [ABSTRACT FROM AUTHOR]

Isothermal amplification (IA) techniques like rolling circle amplification (RCA) and loop-mediated isothermal amplification (LAMP) have gained significant attention in recent years due to their ability to rapidly amplify DNA or RNA targets at a constant temperature without the need for complex thermal cycling equipment. Such technologies, combined with colorimetric systems rendering visual confirmation of the amplification event, are ideal for the development of point-of-need detection methods suitable for field settings where access to specialized laboratory equipment is limited. The utility of these technologies, thus far limited to DNA and RNA targets, could be broadened to a wide range of targets by using aptamers. Composed of DNA or RNA themselves, aptamers can bind to substances, including proteins, metabolites, and inorganic substances. Their nucleic acid nature can potentially allow them to serve as a bridge, extending the reach of DNA/RNA-centric technologies to the broader molecular world. Indeed, the change in aptamer conformation occurring during ligand interaction can be used to elaborate ligand-responding RCA or LAMP templates. By using an existing aptamer targeting SARS-CoV-2 Spike protein as a model, we explored the possibility of establishing ligand-responsive IA systems. Our study used aptamers with simple sequence modifications as templates in LAMP assays and hyperbranched RCA (HRCA) by exploiting the dynamic nature of the model aptamer to trigger these IA systems. Importantly, our work uniquely demonstrates that this aptamer's dynamic response to ligand binding can regulate both RCA and LAMP processes. This novel approach of using aptamer conformational changes to trigger LAMP paves the way for new aptamer-based detection assays. Our system detects 50 nM of Spike protein, with LAMP occurring within 30 min in the presence of Spike. The colorimetric readout showed clear results, allowing for the detection of Spike protein presence. [ABSTRACT FROM AUTHOR]

Chronic myeloid leukemia (CML) is a myeloproliferative neoplasm characterized by excessive proliferation of myeloid cells. The COVID-19 pandemic has raised concerns regarding the impact of SARS-CoV-2 on patients with malignancies, particularly those with CML. This study aimed to investigate the effects of SARS-CoV-2 proteins on CML cell viability and the protective role of vitamin K2 (VK2) in conjunction with imatinib. Experiments conducted on K562 CML cells demonstrated that the SARS-CoV-2 envelope protein induces cytotoxicity and activates caspase 3/7, which are key markers of apoptosis. VK2 mitigated these cytotoxic effects and decreased cytokine production while inhibiting colony formation. Furthermore, the combination of VK2 with imatinib significantly reduced cellular proliferation, diminished mitochondrial membrane potential, and markedly suppressed colony formation. These findings suggest that VK2 protects CML cells from SARS-CoV-2-induced cytotoxicity and enhances the therapeutic efficacy of imatinib, presenting a potential strategy to improve CML treatment during the COVID-19 pandemic. [ABSTRACT FROM AUTHOR]

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), a novel betacoronavirus, is the causative agent of COVID-19, which has caused economic and social disruption worldwide. To date, many drugs and vaccines have been developed for the treatment and prevention of COVID-19 and have effectively controlled the global epidemic of SARS-CoV-2. However, SARS-CoV-2 is highly mutable, leading to the emergence of new variants that may counteract current therapeutic measures. Electron microscopy (EM) is a valuable technique for obtaining ultrastructural information about the intracellular process of virus replication. In particular, EM allows us to visualize the morphological and subcellular changes during virion formation, which would provide a promising avenue for the development of antiviral agents effective against new SARS-CoV-2 variants. In this review, we present our recent findings using transmission electron microscopy (TEM) combined with electron tomography (ET) to reveal the morphologically distinct types of SARS-CoV-2 particles, demonstrating that TEM and ET are valuable tools for visually understanding the maturation status of SARS-CoV-2 in infected cells. This review also discusses the application of EM analysis to the evaluation of genetically engineered RNA viruses. [ABSTRACT FROM AUTHOR]

Extensive research has been conducted on the SARS-CoV-2 virus in association with various infectious diseases to understand the pathophysiology of the infection and potential co-infections. In tropical countries, exposure to local viruses may alter the course of SARS-CoV-2 infection and coinfection. Notably, only a portion of the antibodies produced against SARS-CoV-2 proteins demonstrate neutralizing properties, and the immune response following natural infection tends to be temporary. In contrast, long-lasting IgG antibodies are common after dengue virus infections. In cases where preexisting antibodies from an initial dengue virus infection bind to a different dengue serotype during a subsequent infection, there is a potential for antibody-dependent enhancement (ADE) and the formation of immune complexes associated with disease severity. Both SARS-CoV-2 and dengue infections can result in immunodeficiency. Viral proteins of both viruses interfere with the host's IFN-I signaling. Additionally, a cytokine storm can occur after viral infection, impairing a proper response, and autoantibodies against a wide array of proteins can appear during convalescence. Most of the reported autoantibodies are typically short-lived. Vaccines against both viruses alter the immune response, affecting the course of viral infection and enhancing clearance. A comprehensive analysis of both viral infections and pathogenicity is revisited to prevent infection, severity, and mortality. [ABSTRACT FROM AUTHOR]

Codon bias analysis of SARS-CoV-2 reveals suboptimal adaptation for translation in human cells it infects. The detailed examination of the codons preferentially used by SARS-CoV-2 shows a strong preference for LysAAA, GlnCAA, GluGAA, and ArgAGA, which are infrequently used in human genes. In the absence of an adapted tRNA pool, efficient decoding of these codons requires a 5-methoxycarbonylmethyl-2-thiouridine (mcm5s2) modification at the U34 wobble position of the corresponding tRNAs (tLysUUU; tGlnUUG; tGluUUC; tArgUCU). The optimal translation of SARS-CoV-2 open reading frames (ORFs) may therefore require several adjustments to the host's translation machinery, enabling the highly biased viral genome to achieve a more favorable "Ready-to-Translate" state in human cells. Experimental approaches based on LC-MS/MS quantification of tRNA modifications and on alteration of enzymatic tRNA modification pathways provide strong evidence to support the hypothesis that SARS-CoV-2 induces U34 tRNA modifications and relies on these modifications for its lifecycle. The conclusions emphasize the need for future studies on the evolution of SARS-CoV-2 codon bias and its ability to alter the host tRNA pool through the manipulation of RNA modifications. [ABSTRACT FROM AUTHOR]

The high variability observed in the clinical symptoms of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infections has been attributed to the presence, in a proportion of infection-naive subjects, of pre-existing cross-reactive immune responses. Here, we demonstrate that the bovine coronavirus spike protein (BoS) may represent a source of protective immunity to SARS-CoV-2. Indeed, vaccination of BALB/c mice with a Bovine herpesvirus 4 (BoHV-4)-based vector expressing BoS induced both cell-mediated and humoral immune responses that cross-react with SARS-CoV-2 spike protein. Although the spike-specific antibodies induced by BoS did not neutralize SARS-CoV-2, the T lymphocytes activated by BoS were able to induce cytotoxicity of cells expressing spike proteins derived from several SARS-CoV-2 variants. These results demonstrate that immunization with BoS may represent a source of cross-reactive immunity to SARS-CoV-2, and that these cross-reactive immune responses may exert protective functions. These results contribute to deciphering the mechanisms responsible for lack or mildness of symptoms observed in many individuals upon SARS-CoV-2 infection and may open new ways for the development of new vaccines for coronaviruses. [ABSTRACT FROM AUTHOR]

The SARS-CoV-2 pandemic has highlighted the challenges posed by the emergence and rapid global spread of previously unknown viruses. Early investigations on the pathogenesis of newly identified viruses are often hampered by a lack of appropriate sample material and conventional detection methods. In this study, viral replication within the lungs of SARS-CoV-2-infected Syrian golden hamsters was assessed by immunolabeling dsRNA intermediates with three different monoclonal antibodies in formalin-fixed, paraffin-embedded tissue samples. The presence of dsRNA was compared to viral antigen levels, viral titers, and genomic RNA replicates using three different variants of concern and an ancestral virus strain at a single time point and during the course of infection with an ancestral variant, and then validated using fluorescent 2-plex in situ hybridization. The results indicate that the detection of viral infection using anti-dsRNA antibodies is restricted to an early phase of infection with high viral replication activity. Additionally, the combined detection of dsRNA intermediates and viral antigens may help to bridge the interpretation gaps between viral antigen levels and viral titers at a single time point. Further testing in other viral infections or species is needed to assess the potential of dsRNA as an early marker for viral infections. [ABSTRACT FROM AUTHOR]

Objectives: Following the COVID-19 pandemic, global epidemiological trends demonstrate a return to pre-pandemic levels of respiratory syncytial virus (RSV) and influenza (Flu) A/B viruses. For the appropriate clinical management of viral infections, reliable and timely diagnosis is crucial. The clinical presentation of these respiratory viral infections shows significant overlaps; thus, the syndromic diagnosis of these infections becomes challenging. The goal of this study was to compare the performance of three multiplex real-time PCR-based platforms for the detection of SARS-CoV-2, Flu A, Flu B, and RSV. Materials and Methods: A retrospective study was performed on 200 de-identified nasopharyngeal and oropharyngeal specimens. All samples were tested simultaneously on three PCR-based platforms for the detection of SARS-CoV-2, Flu A, Flu B, and RSV: HealthTrackRx's real-time PCR Open Array® respiratory panel, TrueMark™ SARS-CoV-2, Flu A, Flu B, RSV Select Panel, and BioFire® RP2.1 Panel. The positive and negative predictive value of each test was evaluated at a 95% confidence interval. Results: Among the 200 tested samples, the TrueMark™ and OpenArray® laboratory-developed tests (LDTs) showed a 100% concordance for the detection of SARS-CoV-2, Flu A, Flu B, and RSV. Overall agreement of 100% was observed for nasopharyngeal samples between the laboratory-developed tests and FDA-approved BioFire® RP2.1 Panel. Diagnostic results for these four respiratory viruses, in clinical samples, between the LDTs and the FDA-approved comparator demonstrated full concordance. Conclusions: Respiratory viral infections represent one of the major global healthcare burdens. Consequently, the accurate detection and surveillance of these viruses are critical, particularly when these viruses are known to co-circulate. The excellent performance and full concordance of the LDTs, with the BioFire® Respiratory RP2.1 panel, in detecting SARS-CoV-2, Flu A, Flu B, and RSV shows that these tests can be confidently implemented for the clinical testing of respiratory viral infections. [ABSTRACT FROM AUTHOR]

Since the onset of the COVID-19 pandemic, a variety of diagnostic approaches, including RT-qPCR, RAPID, and LFA, have been adopted, with RT-qPCR emerging as the gold standard. However, a significant challenge in COVID-19 diagnostics is the wide range of symptoms presented by patients, necessitating early and accurate diagnosis for effective management. Although RT-qPCR is a precise molecular technique, it is not immune to false-negative results. In contrast, CRISPR-based detection methods for SARS-CoV-2 offer several advantages: they are cost-effective, time-efficient, highly sensitive, and specific, and they do not require sophisticated instruments. These methods also show promise for scalability, enabling diagnostic tests. CRISPR technology can be customized to target any genomic region of interest, making it a versatile tool with applications beyond diagnostics, including therapeutic development. The CRISPR/Cas systems provide precise gene targeting with immense potential for creating next-generation diagnostics and therapeutics. One of the key advantages of CRISPR/Cas-based therapeutics is the ability to perform multiplexing, where different sgRNAs or crRNAs can target multiple sites within the same gene, reducing the likelihood of viral escape mutants. Among the various CRISPR systems, CRISPR/Cas13 and CARVER (Cas13-assisted restriction of viral expression and readout) are particularly promising. These systems can target a broad range of single-stranded RNA viruses, making them suitable for the diagnosis and treatment of various viral diseases, including SARS-CoV-2. However, the efficacy and safety of CRISPR-based therapeutics must be thoroughly evaluated in pre-clinical and clinical settings. While CRISPR biotechnologies have not yet been fully harnessed to control the current COVID-19 pandemic, there is an optimism that the limitations of the CRISPR/Cas system can be overcome soon. This review discusses how CRISPR-based strategies can revolutionize disease diagnosis and therapeutic development, better preparing us for future viral threats. [ABSTRACT FROM AUTHOR]

The development of novel tools to tackle viral processes has become a central focus in global health, during the COVID-19 pandemic. The spike protein is currently one of the main SARS-CoV-2 targets, owing to its key roles in infectivity and virion formation. In this context, exploring innovative strategies to block the activity of essential factors of SARS-CoV-2, such as spike proteins, will strengthen the capacity to respond to current and future threats. In the present work, we developed and tested novel bispecific molecules that encompass: (i) oligonucleotide aptamers S901 and S702, which bind to the spike protein through its S1 domain, and (ii) hydrophobic tags, such as adamantane and tert-butyl-carbamate-based ligands. Hydrophobic tags have the capacity to trigger the degradation of targets recruited in the context of a proteolytic chimera by activating quality control pathways. We observed that S901-adamantyl conjugates promote the degradation of the S1 spike domain, stably expressed in human cells by genomic insertion. These results highlight the suitability of aptamers as target-recognition molecules and the robustness of protein quality control pathways triggered by hydrophobic signals, and place aptamer-Hytacs as promising tools for counteracting coronavirus progression in human cells. [ABSTRACT FROM AUTHOR]

The onset of SARS-CoV-2 infection in 2019 sparked a global COVID-19 pandemic. This infection is marked by a significant rise in both viral and host kinase activity. Our primary objective was to identify a pivotal host kinase essential for COVID-19 infection and the associated phenomenon of the cytokine storm, which may lead to long-term COVID-19 complications irrespective of viral genetic variations. To achieve this, our study tracked kinase phosphorylation dynamics in RAW264.7 macrophages following SPIKE transfection over time. Among the kinases surveyed, p70S6K (RPS6KB1) exhibited a 3.5-fold increase in phosphorylation at S418. This significant change prompted the selection of p70S6K for in silico investigation, utilizing its structure bound to M2698 (PDB: 7N93). M2698, an oral dual Akt/p70S6K inhibitor with an IC50 of 1.1 nM, exhibited psychosis side effects in phase I clinical trials, potentially linked to its interaction with Akt2. Our secondary objective was to discover a small-molecule analogue of M2698 that exhibits a distinct binding preference for p70S6K over Akt2 through computational modeling and analysis. The in silico part of our project began with validating the prediction accuracy of the docking algorithm, followed by an OCA analysis pinpointing specific atoms on M2698 that could be modified to enhance selectivity. Subsequently, our investigation led to the identification of an analog of M2698, designated as S34, that showed a superior docking score towards p70S6K compared to Akt2. To further assess the stability of S34 in its protein–ligand (PL) complexes with p70S6K and Akt2, MD simulations were conducted. These simulations suggest that S34, on average, forms two hydrogen bond interactions with p70S6K, whereas it only forms one hydrogen bond interaction with Akt2. This difference in hydrogen bond interactions likely contributed to the observed larger root mean square deviation (RMSD) of 0.3 nm in the S34-Akt2 complex, compared to 0.1 nm in the S34-p70S6K complex. Additionally, we calculated free binding energy to predict the strength of the binding interactions of S34 to p70S6K and Akt2, which showed ~2-fold favorable binding affinity of S34 in the p70S6K binding pocket compared to that in the Akt2 binding pocket. These observations may suggest that the S34-p70S6K complex is more stable than the S34-Akt2 complex. Our work focused on identifying a host kinase target and predicting the binding affinity of a novel small molecule to accelerate the development of effective treatments. The wet bench results specifically highlight p70S6K as a compelling anti-COVID-19 target. Meanwhile, our in silico investigations address the known off-target effects associated with M2698 by identifying a close analog called S34. In conclusion, this study presents novel and intriguing findings that could potentially lead to clinical applications with further investigations. [ABSTRACT FROM AUTHOR]

The COVID-19 pandemic, caused by SARS-CoV-2, prompted global efforts to develop vaccines to control the disease. Various vaccines, including mRNA (BNT162b2, mRNA-1273), adenoviral vector (ChAdOx1, Ad26.COV2.S), and inactivated virus platforms (BBIBP-CorV, CoronaVac), elicit high-titer, protective antibodies against the virus, but long-term antibody durability and effectiveness vary. The objective of this study is to elucidate the factors that influence vaccine effectiveness (VE) and the longevity of humoral immune responses to COVID-19 vaccines through a review of the relevant literature, including clinical and real-world studies. Here, we discuss the humoral immune response to different COVID-19 vaccines and identify factors influencing VE and antibody longevity. Despite initial robust immune responses, vaccine-induced immunity wanes over time, particularly with the emergence of variants, such as Delta and Omicron, that exhibit immune escape mechanisms. Additionally, the durability of the humoral immune responses elicited by different vaccine platforms, along with the identification of essential determinants of long-term protection—like pre-existing immunity, booster doses, hybrid immunity, and demographic factors—are critical for protecting against severe COVID-19. Booster vaccinations substantially restore neutralizing antibody levels, especially against immune-evasive variants, while individuals with hybrid immunity have a more durable and potent immune response. Importantly, comorbidities such as diabetes, cardiovascular disease, chronic kidney disease, and cancer significantly reduce the magnitude and longevity of vaccine-induced protection. Immunocompromised individuals, particularly those undergoing chemotherapy and those with hematologic malignancies, have diminished humoral responses and benefit disproportionately from booster vaccinations. Age and sex also influence immune responses, with older adults experiencing accelerated antibody decline and females generally exhibiting stronger humoral responses compared to males. Understanding the variables affecting immune protection is crucial to improving vaccine strategies and predicting VE and protection against COVID-19. [ABSTRACT FROM AUTHOR]

Background: European countries have included healthcare workers (HCWs) among priority groups for COVID-19 vaccination. We established a multi-country hospital network to measure the SARS-CoV-2 incidence and effectiveness of COVID-19 vaccines among HCWs against laboratory-confirmed SARS-CoV-2 infection. Methods: HCWs from 19 hospitals in 10 countries participated in a dynamic prospective cohort study, providing samples for SARS-CoV-2 testing at enrolment and during weekly/fortnightly follow-up. We measured the incidence during pre-Delta (2 May–6 September 2021), Delta (7 September–14 December 2021), and Omicron (15 December 2021–2 May 2023) waves. Using Cox regression, we measured the relative vaccine effectiveness (rVE) of the first COVID-19 booster dose versus primary course alone during Delta and Omicron waves. Results: We included a total of 3015 HCWs. Participants were mostly female (2306; 79%), with a clinical role (2047; 68%), and had a median age of 44 years. The overall incidence of SARS-CoV-2 infection was 3.01/10,000 person-days during pre-Delta, 4.21/10,000 during Delta, and 23.20/10,000 during Omicron waves. rVE was 59% (95% CI: −25; 86) during Delta and 22% (1; 39) during Omicron waves. rVE was 51% (30; 65) 7–90 days after the first booster dose during the Omicron wave. Conclusions: The incidence of SARS-CoV-2 infection among HCWs was higher during the Omicron circulation period. The first COVID-19 vaccine booster provided additional protection against SARS-CoV-2 infection compared to primary course vaccination when recently vaccinated <90 days. This multi-country HCW cohort study addressing infection as the main outcome is crucial for informing public health interventions for HCWs. [ABSTRACT FROM AUTHOR]

The emergence of SARS-CoV-2 has necessitated the development of versatile vaccines capable of addressing evolving variants. Prime-2-CoV_Beta, a novel Orf virus-based COVID-19 vaccine, was developed to express the SARS-CoV-2 spike and nucleocapsid antigens. This first-in-human, phase I, dose-finding clinical trial was conducted to assess the safety, reactogenicity, and immunogenicity of Prime-2-CoV_Beta as a booster in healthy adults. From June 2022 to June 2023, 60 participants in Germany received varying doses of Prime-2-CoV_Beta. The study demonstrated a favorable safety profile, with no serious adverse events (AEs) reported. All AEs were mild (107) or moderate (10), with the most common symptoms being pain at the injection site, fatigue, and headache. Immunogenicity assessments revealed robust vaccine-induced antigen-specific immune responses. High doses notably elicited significant increases in antibodies against the spike and nucleocapsid proteins as well as neutralizing antibodies against SARS-CoV-2 and its variants. Additionally, the vaccine did not induce ORFV-neutralizing antibodies, indicating the potential for repeated administration. In conclusion, Prime-2-CoV_Beta was safe, well tolerated, and immunogenic, demonstrating potential as a broadly protective vaccine against SARS-CoV-2 and its variants. These promising results support further evaluation of higher doses and additional studies to confirm efficacy and long-term protection. This trial was registered at ClinicalTrials, NCT05389319. [ABSTRACT FROM AUTHOR]

Background: Multiple myeloma (MM) patients are at high risk of severe infections including COVID-19 due to an immune dysregulation affecting both innate and adaptive immune responses. However, our understanding of the immune responses to infection and vaccination in MM patients is limited. To gain more detailed insights into infection- and vaccine-elicited T cell immunity in MM, we studied the CD8+ T cell response on the single-epitope level in SARS-CoV-2 convalescent and mRNA-vaccinated MM patients. Methods: We compared peptide/MHC class I tetramer-enriched SARS-CoV-2-specific CD8+ T cells and antibody responses in MM patients (convalescent: n = 16, fully vaccinated: n = 5, vaccinated convalescent: n = 5) and healthy controls (HCs) (convalescent: n = 58, fully vaccinated: n = 7) either after infection with SARS-CoV-2 alone, complete mRNA vaccination or SARS-CoV-2 infection and single-shot mRNA vaccination (hybrid immunity). Results: MM patients have lower frequencies and a lower proportion of fully functional virus-specific CD8+ T cells compared to HCs, after both SARS-CoV-2 infection and vaccination. CD8+ T cell memory subset distribution in MM patients is skewed towards reduced frequencies of central memory (TCM) T cells and higher frequencies of effector memory 1 (TEM1) T cells. In contrast, the humoral immune response was comparable in both cohorts after viral clearance. Notably, CD8+ T cell frequencies as well as the humoral immune response were improved by a single dose of mRNA vaccine in convalescent MM patients. Conclusions: MM patients have relative immunological deficiencies in SARS-CoV-2 immunity but benefit from hybrid immunity. These findings underline the relevance of vaccinations in this vulnerable patient group. [ABSTRACT FROM AUTHOR]

Background: Patients with chronic liver disease (CLD) have impaired vaccine immunogenicity and an excess risk of severe COVID-19. While variant-adapted COVID-19 mRNA vaccines are recommended for vulnerable individuals, their efficacy in patients with CLD has not been studied. Methods: We present the first evaluation of XBB.1.5 COVID-19 vaccine immunogenicity against the SARS-CoV-2 JN.1 variant in patients with CLD. Serum anti-receptor binding domain (RBD) IgG, neutralization, and saliva anti-RBD IgG and IgA against wild-type SARS-CoV-2 (WT) and the XBB.1.5, EG.5.1, BA.2.86, and JN.1 variants were quantified before and 2–4 weeks following a fourth dose of XBB.1.5 mRNA vaccines. Results: Vaccination boosted anti-RBD IgG and neutralization against all tested variants including JN.1 (each p < 0.001). Following immunization, neutralization was lower against JN.1 compared to WT, XBB.1.5, and EG.5.1 (p < 0.001, p < 0.001, and p < 0.01, respectively). Vaccination reduced neutralization failure rates against BA.2.86 and JN.1 (each p < 0.05). The evasion of vaccine-induced antibodies by the tested variants was low, indicated by the positive correlation between anti-RBD IgG and neutralization. At mucosal sites, vaccination boosted anti-RBD IgG (each p < 0.01) but failed to induce infection-blocking IgA (each p > 0.05). Conclusion: XBB.1.5 vaccines protect CLD patients against recent SARS-CoV-2 variants, but developing vaccines with optimized mucosal immunogenicity is required to prevent SARS-CoV-2 transmission and recurrent seasonal COVID-19 outbreaks. [ABSTRACT FROM AUTHOR]

Background/Objectives: The receptor binding motif (RBM) of the SARS-CoV-2 spike protein is critical for viral entry into host cells. Development of a vaccine targeting this region is a promising strategy for COVID-19 prevention. To enhance the immunogenicity of SARS-CoV-2 vaccines, we developed an adenoviral vector expressing the RBM from the SARS-CoV-2 spike protein that fused to the human Fc (hFc) domain. Methods: The recombinant RBM_hFc fusion protein was successfully cloned into the pacAd5CMV-N-pA (pAd5) vector and expressed in HEK293 cells as a ~40 kDa protein. A recombinant adenovirus encoding RBM_hFc was subsequently generated and confirmed by cytopathic effect assay. Results: Western blot analysis verified the expression of RBM_hFc in the adenovirus (AdV). ELISA assays, validated for IgG detection, demonstrated a twofold increase in IgG antibody levels (M–1.090 at 450 nm; SD—±0.326; and 95% CI—0.250 [0.839 to 1.340]) in sera from BALB/c mice immunized with Ad/RBM_hFc, compared to the negative control group. Result suggests a robust humoral immune response induced by the Ad/RBM_hFc vaccine. Moreover, ELISpot assays demonstrated a tenfold increase in IFN-γ -producing cells (M—440 spot-forming cells; SD—±124.976; and 95% CI—75.522 [364.478 to 515.522]) in mice immunized with AdV/RBM_hFc compared to the negative control group. Result proved that AdV/RBM_hFc-stimulated a robust cellular immune response in animal model. Conclusions: Our findings indicate that the RBM_hFc fusion protein enhances both humoral and cellular immune responses. These results suggest the potential of adenoviral vectors carrying RBM_hFc as vaccine candidates. However, comprehensive evaluation of the protective efficacy of these adenoviral vectors will necessitate rigorous experimental studies. [ABSTRACT FROM AUTHOR]

Objective: COVID-19 vaccine-neutralizing antibodies provide early data on potential vaccine effectiveness, but their usefulness depends on study reliability and reporting quality. Methods: We systematically evaluated 50 published post-vaccination neutralizing antibody studies for key parameters that determine study and data quality regarding sample size, SARS-CoV-2 infection, vaccination regimen, sample collection period, demographic characterization, clinical characterization, experimental protocol, live virus and pseudo-virus details, assay standardization, and data reporting. Each category was scored from very high to low or unclear quality, with the lowest score determining the overall study quality score. Results: None of the studies attained an overall high or very high score, 8% (n = 4) attained moderate, 42% (n = 21) low, and 50% (n = 25) unclear. The categories with the fewest studies assessed as ≥ high quality were SARS-CoV-2 infection (42%), sample size (30%), and assay standardization (14%). Overall quality was similar over time. No association between journal impact factor and quality score was found. Conclusions: We found that reporting in neutralization studies is widely incomplete, limiting their usefulness for downstream analyses. [ABSTRACT FROM AUTHOR]

Background. The combined or multivalent vaccines are actively used in pediatric practice and offer a series of advantages, including a reduced number of injections and visits to the doctor, simplicity of the vaccination schedule and minimization of side effects, easier vaccine monitoring and storage, and lower vaccination costs. The practice of widespread use of the combined vaccines has shown the potential to increase vaccination coverage against single infections. The mRNA platform has been shown to be effective against the COVID-19 pandemic and enables the development of combined vaccines. There are currently no mRNA-based combined vaccines approved for use in humans. Some studies have shown that different mRNA components in a vaccine can interact to increase or decrease the immunogenicity and efficacy of the combined vaccine. Objectives. In the present study, we investigated the possibility of combining the mRNA vaccines, encoding seasonal influenza and SARS-CoV-2 antigens. In our previous works, both vaccine candidates have shown excellent immunogenicity and efficacy profiles in mice. Methods. The mRNA-LNPs were prepared by microfluidic mixing, immunogenicity in mice was assessed by hemagglutination inhibition assay, enzyme-linked immunoassay and virus neutralization assay. Immunological efficacy was assessed in a mouse viral challenge model. Results. In this work, we demonstrated that the individual mRNA components of the combined vaccine did not affect the immunogenicity level of each other. The combined vaccine demonstrated excellent protective efficacy, providing a 100% survival rate when mice were infected with the H1N1 influenza virus and reducing the viral load in the lungs. Four days after the challenge with SARS-CoV-2 EG.5.1.1., no viable virus and low levels of detectable viral RNA were observed in the lungs of vaccinated mice. Conclusions. The combination does not lead to mutual interference between the individual vaccines. We believe that such a combined mRNA-based vaccine could be a good alternative to separated human vaccinations for the prevention of COVID-19 and influenza. [ABSTRACT FROM AUTHOR]

Background: Since 2019, the SARS-CoV-2 virus has been responsible for the global spread of respiratory illness. As of 1 September 2024, the cumulative number of infections worldwide exceeded 776 million. There are many structural proteins of the virus, among which the SARS-CoV-2 nucleocapsid (N) protein plays a pivotal role in the viral life cycle, participating in a multitude of essential activities following viral invasion. An important antiviral immune response is the major histocompatibility complex (MHC)-restricted differentiation cluster 8 (CD8+) T cell cytotoxicity. Therefore, understanding the immunogenicity of SARS-CoV-2 NP-specific MHC-I-restricted epitopes is highly important. Methods: MHC-I molecules from 11 human leukocyte antigen I (HLA-I) superfamilies with 98% population coverage and 6 mouse H2 alleles were selected. The affinity were screened by IEDB, NetMHCpan, SYFPEITHI, SMMPMBEC and Rankpep. Further immunogenicity and conservative analyses were performed using VaxiJen and BLASTp, respectively. EpiDock was used to simulate molecular docking. Cluster analysis was performed. Selective epitopes were validated by enzyme-linked immunospot (ELISpot) assay and flow cytometry in the mice with pVAX-NPSARS-CoV-2 immunization. Enzyme-Linked Immunosorbent Assay (ELISA) was used to detect whether the preferred epitope induced humoral immunity. Results: There were 64 dominant epitopes for the H-2 haplotype and 238 dominant epitopes for the HLA-I haplotype. Further analysis of immunogenicity and conservation yielded 8 preferred epitopes, and docking simulations were conducted with corresponding MHC-I alleles. The relationships between the NP peptides and MHC-I haplotypes were then determined via two-way hierarchical clustering. ELISA, ELISpot assay, and flow cytometry revealed that the preferred epitope stimulated both humoral and cellular immunity and enhanced cytokine secretion in mice. Conclusions: our study revealed the general patterns among multiple haplotypes within the humans and mice superfamily, providing a comprehensive assessment of the pan-MHC-I immunoreactivity of SARS-CoV-2 NP. Our findings would render prospects for the development and application of epitope-based immunotherapy in lasting viral epidemics. [ABSTRACT FROM AUTHOR]