Search Results (292)

Background and Clinical Significance: Congenital malformations of the right atrium are rare. Their clinical presentation varies widely, from the absence of symptoms to sudden death, often being diagnosed incidentally by cardiac imaging. First described in 1955, the right atrial diverticulum is usually characterized as a pouch-like structure originating from the free atrial wall, or right atrial appendage. The prevalence of congenital malformations of the right atrium is unknown because few clinical cases have been reported. Associated complications include arrhythmias, pulmonary thromboembolism, progressive dilatation marked by a high risk of compression and rupture. In these cases, the optimal therapeutic approach is surgical resection. Case Presentation: We present the case of a 58-year-old, hypertensive female with a history of COVID-19 (Coronavirus Disease 2019), who was admitted for persistent dyspnea and chest pain. An electrocardiogram on arrival showed no arrhythmias or ischemic changes, and echocardiography revealed severe systolic dysfunction—a left ventricular ejection fraction (LVEF) of 20%, moderate mitral and tricuspid regurgitations, and a pericardial collection, adjacent to the right atrium, considered to be a localized pericardial effusion. Coronary angiography excluded ischemic etiology and a viral myocarditis was further suspected. Cardiac magnetic resonance imaging (IRM) showed a non-ischemic scar pattern in the interventricular septum, but also detected a well-defined large mass, which communicated with the right atrium through a 20 mm opening, suggestive of a right atrial diverticulum. Contrast echocardiography confirmed the communication between the cavity and the right atrium. A surgical resection of the large diverticulum was performed. Conclusions: The particularity of this case consists in the incidental identification of a rare cardiac malformation in an adult patient. Full article

In this study, astaxanthin, which has previously been shown to have antiviral effects, was examined for its dose-dependent potency to inhibit cellular SARS-CoV-2 infections. Naturally occurring astaxanthin is obtained and orally administered as ASX-oleoresin, a composition of different astaxanthin fatty acid esters. We therefore hypothesized that the compound’s beneficial effects are not only related to astaxanthin. Thus, a “green” algae extract (i.e., poor astaxanthin content < 0.2%; ASX^p) of the microalgae Haematococcus pluvialis, as well as an astaxanthin-rich algae extract (astaxanthin content = 20%; ASX^r), were tested in in vitro cellular viral infection assays. Thereby, it was found that both extracts reduced viral infections significantly. As a potential mode of inhibitory action, the binding of ASX-oleoresin to the viral spike protein was investigated by isothermal fluorescence titration, revealing binding affinities of Kd = 1.05 µM for ASX^r and Kd = 1.42 µM for ASX^p. Based on our data, we conclude that several ASX-oleoresin fractions from H. pluvialis exhibit antiviral activity, which extends beyond the known antioxidant activity of astaxanthin. From a molecular dynamic simulation of ASX-oleoresin, fatty acid domains could be considered as activity-chaperoning factors of ASX. Therefore, microalgae biomass should be considered in the future for further antiviral activities. Full article

Vaccine adjuvants are non-immunogenic agents that enhance or modulate immune responses to co-administered antigens and are essential to modern vaccines. Despite their importance, few are approved for human use. The rise of new pathogens and limited efficacy of some existing vaccines underscore the need for more advanced and effective formulations, particularly for vulnerable populations. Aluminum-based adjuvants are commonly used in vaccines and effectively promote humoral immunity. However, they mainly induce a Th2-biased response, making them suboptimal for diseases requiring cell-mediated immunity. In contrast, saponin-based adjuvants from the Quillajaceae family elicit a more balanced Th1/Th2 response and generate antigen-specific cytotoxic T cells (CTL). Due to ecological damage and limited availability caused by overharvesting Quillaja saponaria Molina barks, efforts have intensified to identify alternative plant-derived saponins with enhanced efficacy and lower toxicity. Quillaja brasiliensis (A.St.-Hil. and Tul.) Mart. (syn. Quillaja lancifolia D.Don), a related species native to South America, is considered a promising renewable source of Quillajaceae saponins. In this review, we highlight recent advances in vaccine adjuvant research, with a particular focus on saponins extracted from Q. brasiliensis leaves as a sustainable alternative to Q. saponaria saponins. These saponin fractions are structurally and functionally comparable, exhibiting similar adjuvant activity when they were formulated with different viral antigens. An alternative application involves formulating saponins into nanoparticles known as ISCOMs (immune-stimulating complexes) or ISCOM-matrices. These formulations significantly reduce hemolytic activity while preserving strong immunoadjuvant properties. Therefore, research advances using saponin-based adjuvants (SBA) derived from Q. brasiliensis and their incorporation into new vaccine platforms may represent a viable and sustainable solution for the development of more less reactogenic, safer, and effective vaccines, especially for diseases that require a robust cellular immunity. Full article

Background/Objectives: Co-transfection of multiple DNAs is important to many research and therapeutic applications. While the optimization of single plasmid transfection is common, multiple plasmid co-transfection analyses are limited. Here we provide empirical data regarding multiple plasmid co-transfection while altering the number of species of plasmids transfected (up to four different plasmids) and the amount of plasmids/cell using the two most common non-viral techniques, electroporation and lipofection. Methods: A549 human lung epithelial cells were transfected using lipofectamine 2000 or electroporation with combinations of plasmids, each expressing one of four different fluorescent proteins from the CAGG promoter. Twenty-four hours later, cells were analyzed by spectral flow cytometry to determine the number of cells expressing each fluorescent protein and the amount of fluorescent signal of each protein in a cell. Results and Conclusions: For electroporation, while the fraction of cells expressing plasmids increased with increasing amounts of DNA, increasing the number of plasmid species did not alter the fraction of expressing cells and had no effect on levels of expression in individual cells. By contrast, for lipofection, the fraction of cells expressing plasmids was not affected by the amount of DNA added but both the fraction of cells expressing and the level of protein produced in these cells decreased for each plasmid species as the number of delivered species increased. For both lipofection and electroporation after single plasmid transfection, the expressing cells had greater numbers of plasmid copies/cell than non-expressing cells. Multiple plasmid lipofection resulted in more plasmid copies/cell in co-expressing than non-expressing cells. Multiple plasmid electroporation was the inverse of this with fewer plasmid copies/cell in co-expressing than non-expressing cells. Full article

The most common marker used to monitor autophagy is the microtubule-associated protein light chain 3 (LC3). Upon induction of autophagy, LC3 is conjugated to phosphatidylethanolamine and targeted to autophagic membranes, which can be easily detected by immunofluorescence. However, this technique has some limitations. During the early stages of HSV-1 infection, strong LC3B nuclear staining is observed within the viral replication compartments. This staining is only detected when using polyclonal antibodies. It is noteworthy that monoclonal antibodies or the GFP-LC3 plasmid do not reveal any nuclear LC3 staining. Interestingly, LC3B is not detected in the nuclear fraction of infected cells by Western blotting, even when polyclonal antibodies are used. In infected LC3B knockout cells, nuclear staining is still observed when using polyclonal LC3B antibodies. This suggests that polyclonal LC3B antibodies generate non-specific nuclear staining in infected cells, which could result in misinterpretation and erroneous conclusions. These findings raise questions about the reliability of LC3-immunofluorescence assays in herpesvirus infections. It is imperative that the methodology employed for monitoring autophagy by immunofluorescence in viral infections be reviewed and updated, and that the specificity of anti-LC3B antibodies be tested before use. To ensure the accuracy of the results, it is essential to validate this technique with additional assays, such as by immunoblot analysis or via the use of autophagy-deficient cell lines. Full article

GW4869, a cell-permeable, selective inhibitor of neutral sphingomyelinase is a pharmacological agent that blocks the production and release of extracellular vesicles (EVs). Our previous studies have shown that GW4869 inhibits flaviviral loads in tick, mosquito and mammalian cells, including murine cortical neurons. Yet the mechanism(s) of GW4869 inhibitor upon viral infections were not addressed. In the current study, we focused on how GW4869 interferes with Langat Virus (LGTV, a tick-borne flavivirus) replication in ISE6 tick cells. First, we found that GW4869 is neither cytotoxic at tested doses of 50, 100, and 150 µM in tick cells, nor does it directly bind to the free LGTV present in cell culture supernatants. When tick cells were treated with GW4869, followed by infection with viral stock at dilutions of 10⁻², 10⁻³, 10⁻⁴ (the infectious dose determination by viral dilution assay), it affected LGTV replication in tick cells. A reduction in viral burden was noted in GW4869-treated tick cells, which constituted more than half the amount of decrease when compared to the mock control. Next, GW4869 treatment not only resulted in decreased LGTV transcript levels in tick cells and EVs derived from these infected cells, but also revealed diminished EVs concentrations. Enhanced IsSMase transcripts in the LGTV-infected group was noted upon GW4869 treatment, thus suggesting a host response to perhaps inhibit virus replication. In addition, GW4869 treatment reduced LGTV loads in density gradient EVs fractions, which correlated with decreased EVs concentration in those fractions. These data not only indicate that GW4869 affects LGTV replication, but that it also interferes with EV secretion and release from tick cells. Lastly, we found that GW4869 inhibits LGTV replication in tick cells but does not directly affect the infectivity of LGTV viral particles. Overall, our study suggests that GW4869 is a potential therapeutic inhibitor in controlling tick-borne diseases. Full article

Asthma represents a heterogeneous disorder characterized by a dynamic balance between pro-inflammatory and anti-inflammatory forces, with allergic sensitization contributing substantially to airway hyperresponsiveness and remodeling. Central to its pathogenesis are cytokines such as IL-4, IL-5, IL-13, IL-17, and IL-33, which drive recruitment of eosinophils, neutrophils, and other effector cells, thereby precipitating episodic exacerbations in response to viral and environmental triggers. Conventional biomarkers, including blood and sputum eosinophil counts, IgE levels, and fractional exhaled nitric oxide, facilitate phenotypic classification and guide the emerging biologic era. Monoclonal antibodies targeting IgE (omalizumab) and IL-5 (mepolizumab, benralizumab, reslizumab, depemokimab) have demonstrated the ability to reduce exacerbation frequency and improve lung function, with newer agents such as depemokimab offering extended dosing intervals. Itepekimab, an anti-IL-33 antibody, effectively engages its target and mitigates tissue eosinophilia, while CM310-stapokibart, tralokinumab, and lebrikizumab inhibit IL-4/IL-13 signaling with variable efficacy depending on patient biomarkers. Comparative analyses of these biologics, encompassing affinity, dosing regimens, and trial outcomes, underscore the imperative of personalized therapy to optimize disease control in severe asthma. Full article

Background: Olfactory dysfunction (OD)—including anosmia and hyposmia—is a common and often persistent outcome of viral infections. This systematic review consolidates findings from structural and functional MRI studies to explore how COVID-19 SARS-CoV-2-induced smell loss alters the brain. Considerable heterogeneity was observed across studies, influenced by differences in methodology, population characteristics, imaging timelines, and OD classification. Methods: Following PRISMA guidelines, we conducted a systematic search of PubMed/MEDLINE, Scopus, and Web of Science to identify MRI-based studies examining COVID-19’s SARS-CoV-2 OD. Twenty-four studies were included and categorized based on imaging focus: (1) olfactory bulb (OB), (2) olfactory sulcus (OS), (3) grey and white matter changes, (4) task-based brain activation, and (5) resting-state functional connectivity. Demographic and imaging data were extracted and analyzed accordingly. Results: Structural imaging revealed consistent reductions in olfactory bulb volume (OBV) and olfactory sulcus depth (OSD), especially among individuals with OD persisting beyond three months, suggestive of inflammation and neurodegeneration in olfactory-associated regions like the orbitofrontal cortex and thalamus. Functional MRI studies showed increased connectivity in early-stage OD within regions such as the piriform and orbitofrontal cortices, possibly reflecting compensatory activity. In contrast, prolonged OD was associated with reduced activation and diminished connectivity, indicating a decline in olfactory processing capacity. Disruptions in the default mode network (DMN) and limbic areas further point to secondary cognitive and emotional effects. Diffusion tensor imaging (DTI) findings—such as decreased fractional anisotropy (FA) and increased mean diffusivity (MD)—highlight white matter microstructural compromise in individuals with long-term OD. Conclusions: COVID-19’s SARS-CoV-2 olfactory dysfunction is associated with a range of cerebral alterations that evolve with the duration and severity of smell loss. Persistent dysfunction correlates with greater neural damage, underscoring the need for longitudinal neuroimaging studies to better understand recovery dynamics and guide therapeutic strategies. Full article

Influenza A virus (IAV) poses significant public health challenges due to its rapid mutation and drug resistance, necessitating novel antiviral strategies. Rhododendron brachycarpum, traditionally employed in folk medicine to treat inflammatory conditions, contains bioactive flavonoids with potential antiviral effects. In this study, we investigated the anti-influenza activity of R. brachycarpum leaf extract and identified hyperoside (quercetin-3-O-galactoside) as the active constituent. The crude extract and its n-butanol fraction markedly reduced IAV replication in Madin–Darby canine kidney (MDCK) cells, with IC50/CC50 values of 74.51/201.09 μg/mL and 24.5/113.1 μg/mL, respectively. Hyperoside, purified via bioactivity-guided fractionation and HPLC analysis, demonstrated potent antiviral activity, with an IC50 of 66.59 μM (30.92 μg/mL) and a CC50 of 318.9 μM (148.1 μg/mL), indicating a favorable selectivity index. It significantly suppressed viral mRNA and protein expression in infected cells. Time-of-addition and hemagglutination inhibition assays suggested that hyperoside exerts antiviral effects during early infection stages, likely interfering with viral entry. In silico molecular docking analysis further supported this mechanism, revealing that hyperoside binds strongly to the receptor-binding domain of hemagglutinin (−11.5 kcal/mol), potentially blocking viral attachment. These findings reveal that hyperoside is a major antiviral component of R. brachycarpum and underscore its therapeutic potential as a natural antiviral candidate against IAV infections. Full article

The Baculovirus Expression Vector System (BEVS) is an important protein and complex biologics production platform. The baculovirus GP64 protein is the major envelope glycoprotein that aids in virus entry and is required for cell-to-cell transmission in cell culture. Several studies have developed strategies around gp64 gene disruption in an attempt to minimize baculovirus co-production. Here, we investigate the result of transiently targeting the baculovirus gp64 gene with CRISPR-Cas9 during infection. Because not all genomes are effectively disrupted, we describe a variant calling methodology that allows the detection of the targeted mutations in gp64 even though these mutations are not the dominant sequences. Using a transfection-infection assay (T-I assay), the AcMNPV gp64 gene was targeted at six different locations to evaluate the effects of single and multiple targeting sites, and we demonstrated a reduction in the levels of baculovirus vectors while maintaining or enhancing foreign protein production when protein was driven by a p6.9 promoter. Viral genomes were subsequently isolated from the supernatant and cell pellet fractions, and our sequencing pipeline successfully detected indel mutations within gp64 for most of the single-guide RNA (sgRNA) targets. We also observed that 68.8% of variants found in the virus stock were conserved upon virus propagation in cell culture, thus indicating that they are not detrimental to viral fitness. This work provides a comprehensive assessment of CRISPR-Cas9 genome editing of baculovirus vectors, with potential applications in enhancing the efficiency of the BEVS. Full article

Respiratory syncytial virus (RSV) is the leading cause of respiratory infections in children. Extracellular vesicles (EVs), released by airway epithelial cells, contain proteins and different families of non-coding RNAs (EV cargo) that can modulate the responses of target cells to viral infection. Nasal mucosa is a primary site of viral entry and the source of EVs present in the upper airway secretions. In this study we characterized proteins, including inflammatory mediators and cytokines, and the piwi-interacting RNA (piRNAs) cargo of EVs isolated from pediatric human nose organoids (HNO) and nasopharyngeal secretions (NPS) positive for RSV. Using Proximity Extension Assay (PEA) and Luminex multi-target arrays, we found significant enrichment in several chemokines and other mediators/biomarkers, including CCL2, CCL20, CXCL5, CX3CL1, CXCL6, MMP-1, MMP-10, uPA, Flt3L, ARNT and CD40 in EVs secreted by RSV-infected HNO compared to control mock HNO. Analysis of NPS samples from RSV infected children revealed that CCL3, CCL20, CXCL8, uPA, VEGFA, were concentrated in the NPS-EV fraction. LC-MS/MS and Gene Ontology indicated that RSV positive NPS-EVs originate from different cellular sources, with the most abundant proteins from neutrophils and epithelial cells. A total of 490 piRNAs were detected by NGS sequencing of small RNA libraries obtained from NPS-EVs, which has not been reported prior to this study. Identification of inflammatory mediators and small non-coding RNAs which are compartmentalized in EVs contributes to understanding mechanisms of virus-mediated pathogenesis in RSV infections. Full article

Pigs are affected by a variety of pathogenic agents that need to be identified correctly and diagnosed even when co-infections may complicate the application of specific and targeted assays. Next-generation sequencing can provide new perspective to monitor viruses infecting or co-infecting diseased pigs. In this study, we tested, for the first time for diagnostic purposes in a livestock species, a new method based on whole-genome sequencing of all the DNAs extracted from the blood of nine pigs sampled from a farm where there was a suspected outbreak of Post-weaning Multisystemic Wasting Syndrome. We then used unmapped reads on the porcine reference genome to mine for viral sequences using a specifically designed bioinformatic pipeline. Within this fraction of reads, viral sequences ranged from 0.002% to 4.4% of the total unmapped reads and were derived from twelve different viruses known to infect pigs, where three were herpesviruses, eight were parvoviruses, and one was a circovirus. All pig sequencing datasets were positive for one or more viruses, with various potential viral loads. Suid betaherpesvirus 2, also known as Porcine cytomegalovirus (PCMV), was the most frequently identified virus as five out of the nine pig sequencing datasets contained viral sequences from this virus. The results may suggest a heterogeneous viral profile of the diseased pigs that may be derived from potential secondary infections or co-infections. This pilot application demonstrated that a whole-genome sequencing approach can complement other routine diagnostic assays in veterinary virology. Other studies and improvements are needed to validate the results and apply this approach in routine monitoring applications. Full article

Human T cell leukemia virus type 1 (HTLV 1) is an oncogenic retrovirus responsible for the development of adult T cell leukemia (ATL). The minus strand of HTLV-1 provirus encodes an oncoprotein named HTLV-1 bZIP factor (HBZ), which plays a pivotal role in viral replication and T cell proliferation. Of particular interest is the spliced HBZ isoform (sHBZ), which is predominantly expressed in ATL cells and localizes within the nucleolus, conferring immortalizing properties to T cells. Our previous study has shown that sHBZ colocalizes and associates with Nucleophosmin/B23, a nucleolar phosphoprotein with multiple functions. In this study, through an optimized nucleolar isolation method, we first confirmed sHBZ’s nucleolar localization via Western blotting in transfected HEK293T cells, chronically HTLV-1-infected T cell lines, and freshly infected HeLa cells. We further demonstrated that the sHBZ/B23 association predominantly occurs in the nucleolus by co-immunoprecipitation of cell fractions. Our study highlights the nucleolar localization of sHBZ and its possibly essential interaction with this nucleolar-residing protein, leading to cell immortalization. Full article

Flaviviruses are usually transmitted to humans via mosquito or tick bites, whose infections may lead to severe diseases and fatality. During intracellular infection, they remodel the endoplasmic reticulum (ER) membrane to generate compartments scaffolding the replication complex (RC) where replication of the viral genome takes place. In this study, we purified the ER membrane fraction of virus infected cells to identify the proteins that were enriched during flavivirus infection. We found that tripartite motif-containing proteins (TRIMs) including TRIM38, TRIM21, and TRIM14 were significantly enriched during infection with mosquito-borne (West Nile virus strain Kunjin and Zika virus (ZIKV)) and tick-borne (Langat virus (LGTV)) flaviviruses. Further characterizations showed that TRIM21 and TRIM14 act as restriction factors against ZIKV and LGTV, while TRIM38 hinders ZIKV infection. These TRIMs worked as interferon-stimulated genes to mediate IFN-I response against LGTV and ZIKV infections. Restriction of ZIKV by TRIM14 and TRIM38 coincides with their colocalization with ZIKV NS3. TRIM14-mediated LGTV restriction coincides with its colocalization with LGTV NS3 and NS5 proteins. However, TRIM21 did not colocalize with ZIKV and LGTV NS3 or NS5 protein suggesting its antiviral activity is not dependent on direct targeting the viral enzyme. Finally, we demonstrated that overexpression of TRIM21 and TRIM14 restricted LGTV replication. Full article

Micropeptides (miPEPs), encoded by short open reading frames (sORFs) within various genomic regions, have recently emerged as critical regulators of multiple biological processes. In particular, these small molecules are now increasingly being recognized for their role in modulating viral replication, pathogenesis, and host immune responses. Both host miPEPs and virus-derived miPEPs have been noted for their ability to regulate virus–host interactions through diversified mechanisms such as altering protein stability and modulating protein–protein interactions. Although thousands of sORFs have been annotated as having the potential to encode miPEPs, only a small number have been experimentally validated so far, with some directly linked to virus–host interactions and a small subset associated with immune modulation, indicating that the investigation of miPEPs is still in its infancy. The systematic identification, translational status assessment, in-depth characterization, and functional analysis of a substantial fraction of sORFs encoding miPEPs remain largely underexplored. Further studies are anticipated to uncover the intricate mechanisms underlying virus–host interactions, host immune modulation, and the broader biological functions of miPEPs. This article will review the emerging roles of miPEPs in virus–host interactions and host immunity, and discuss the challenges and future perspectives of miPEP studies. Full article