Little is known regarding anterior uveitis (AU), the most common ocular disease associated with cytomegalovirus (CMV) infection in immunocompetent populations. CMV AU is highly prevalent in Asia, with a higher incidence in men. Clinically, it manifests mainly as anterior chamber inflammation and elevated intraocular pressure (IOP). Acute CMV AU may resemble Posner-Schlossman syndrome with its recurrent hypertensive iritis, while chronic CMV AU may resemble Fuchs uveitis because of its elevated IOP. Without prompt treatment, it may progress to glaucoma; therefore, early diagnosis is critical to prognosis. Knowledge regarding clinical features and aqueous humor analyses can facilitate accurate diagnoses; so, we compared and summarized these aspects. Early antiviral treatment reduces the risk of a glaucoma surgery requirement, and therapeutic effects vary based on drug delivery. Both oral valganciclovir and topical ganciclovir can produce positive clinical outcomes, and higher concentration and frequency are beneficial in chronic CMV retinitis. An extended antiviral course could prevent relapses, but should be limited to 6 months to prevent drug resistance and side effects. In this review, we have systematically summarized the pathogenesis, clinical features, diagnostic and therapeutic aspects, and immunological mechanisms of CMV AU with the goal of providing a theoretical foundation for early clinical diagnosis and treatment.

The influenza neuraminidase (NA) is a promising target for next-generation vaccines. Protection induced by vaccination with the computationally optimized broadly reactive NA antigen (N1-I COBRA NA) was characterized in both influenza serologically naive and pre-immune ferret models following H1N1 (A/California/07/2009, CA/09) or H5N1 (A/Vietnam/1203/2004, Viet/04) influenza challenges. The N1-I COBRA NA vaccine elicited antibodies with neutralizing ELLA activity against both seasonal and pandemic H1N1 influenza, as well as the H5N1 influenza virus. In both models, N1-I COBRA NA-vaccinated ferrets that were challenged with CA/09 virus had similar morbidity (weight loss and clinical symptoms) as ferrets vaccinated with the CA/09 HA control vaccine. There were significantly reduced viral titers compared to the mock-vaccinated control animals. Ferrets vaccinated with N1-I COBRA NA or Viet/04 NA vaccines were protected against the H5N1 virus infection with minimal clinical symptoms and negligible weight loss. In contrast, ferrets vaccinated with the CA/09 NA vaccine lost ~10% of their original body weight with 25% mortality. Vaccination with either HA or NA vaccines did not inhibit contact transmission of CA/09 virus to naïve cage mates. Overall, the N1-I COBRA vaccine elicited protective immune responses against both H1N1 and H5N1 infections and partially mitigated disease in contact-transmission receiving ferrets. These results indicate that the N1-I COBRA NA performed similarly to the CA/09 HA and NA positive controls. Therefore, the N1-I COBRA NA alone induces protection against viruses from both H5N1 and H1N1 subtypes, indicating its value as a vaccine component in broadly protective influenza vaccines.

(1) Background: During the COVID-19 pandemic, rapid and reliable diagnostic tools are needed for detecting SARS-CoV-2 infection in urgent cases at admission to the hospital. We aimed to assess the performances of the rapid molecular VitaPCR™ test (Menarini Diagnostics) in a sample of older adults admitted to the Emergency Department of two Italian hospitals (2) Methods: The comparison between the rapid VitaPCR™ and the RT-PCR was performed in 1695 samples. Two naso-pharyngeal swab samplings from each individual were obtained and processed using the VitaPCR™ and the RT-PCR for the detection of SARS-CoV-2 (3) Results: VitaPCR™ exhibited good precision (<3% CV) and an almost perfect overall agreement (Cohen's K = 0.90) with the RT-PCR. The limit of detection of the VitaPCR™ was 4.1 copies/µL. Compared to the RT-PCR, the sensitivity, the specificity, and the positive and negative predictive values of VitaPCR™ were 83.4%, 99.9%, 99.2% and 98.3%, respectively (4) Conclusions: The VitaPCR™ showed similar sensitivity and specificity to other molecular-based rapid tests. This study suggests that the VitaPCR™ can allow the rapid management of patients within the Emergency Department. Nevertheless, it is advisable to obtain a negative result by a RT-PCR assay before admitting a patient to a regular ward.

The impact of mosquito-borne diseases on human and veterinary health is being exacerbated by rapid environmental changes caused mainly by changing climatic patterns and globalization. To gain insight into mosquito-borne virus circulation from two counties in eastern and southeastern Romania, we have used a combination of sampling methods in natural, urban and peri-urban sites. The presence of 37 mosquito-borne viruses in 16,827 pooled mosquitoes was analyzed using a high-throughput microfluidic real-time PCR assay. West Nile virus (WNV) was detected in 10/365 pools of Culex pipiens (n = 8), Culex modestus (n = 1) and Aedes vexans (n = 1) from both studied counties. We also report the first molecular detection of Sindbis virus (SINV) RNA in the country in one pool of Culex modestus. WNV infection was confirmed by real-time RT-PCR (10/10) and virus isolation on Vero or C6/36 cells (four samples). For the SINV-positive pool, no cytopathic effectwas observed after infection of Vero or C6/36 cells, but no amplification was obtained in conventional SINV RT-PCR. Phylogenetic analysis of WNV partial NS5 sequences revealed that WNV lineage 2 of theCentral-Southeast European clade, has a wider circulation in Romania than previously known.

Although chronic hepatitis C has been effectively treated with direct-acting antivirals (DAAs), the use of conventional therapy with peg-interferon (Peg-IFN) or (predominantly) ribavirin (RBV), remains widespread. R70Q/H and L/C91M amino acid substitutions in the hepatitis C virus (HCV) core protein may modulate responses to IFN and/or RBV, and are associated with cirrhosis, hepatocellular carcinoma (HCC), insulin resistance, and liver steatosis. We evaluated the R70Q/H and L/C91M substitutions, clinical and epidemiological profiles, and risk factors of Brazilian patients chronically infected with HCV subgenotypes 1a and 1b (HCV-GT1a and HCV-GT1b) unresponsive to IFN and/or RBV therapy. Sequencing and pyrosequencing analyses and sociodemographic and clinical predictive variables were used to assess the relationship between R70Q/H and L/C91M substitutions. Leukocyte counts, ALT levels, and ALT/AST ratios were significantly reduced in treated individuals, but more of these patients had advanced fibrosis and cirrhosis. L91M was more prevalent (19.7%), occurring only in HCV-GT1b, followed by R70Q/P (11.5%) and R70P (1.4%). R70Q/P exhibited higher mean AST, ALT, and GGT values, whereas L91M showed higher mean GGT values. Pyrosequencing of the L91M position revealed mutant subpopulations in 43.75% of samples.

More than 40 human cases of severe encephalitis caused by Borna disease virus 1 (BoDV-1) have been reported to German health authorities. In an endemic region in southern Germany, we conducted the seroepidemiological BoSOT study ("BoDV-1 after solid-organ transplantation") to assess whether there are undetected oligo- or asymptomatic courses of infection. A total of 216 healthy blood donors and 280 outpatients after solid organ transplantation were screened by a recombinant BoDV-1 ELISA followed by an indirect immunofluorescence assay (iIFA) as confirmatory test. For comparison, 288 serum and 258 cerebrospinal fluid (CSF) samples with a request for tick-borne encephalitis (TBE) diagnostics were analyzed for BoDV-1 infections. ELISA screening reactivity rates ranged from 3.5% to 18.6% depending on the cohort and the used ELISA antigen, but only one sample of a patient from the cohort with requested TBE diagnostics was confirmed to be positive for anti-BoDV-1-IgG by iIFA. In addition, the corresponding CSF sample of this patient with a three-week history of severe neurological disease tested positive for BoDV-1 RNA. Due to the iIFA results, all other results were interpreted as false-reactive in the ELISA screening. By linear serological epitope mapping, cross-reactions with human and bacterial proteins were identified as possible underlying mechanism for the false-reactive ELISA screening results. In conclusion, no oligo- or asymptomatic infections were detected in the studied cohorts. Serological tests based on a single recombinant BoDV-1 antigen should be interpreted with caution, and an iIFA should always be performed in addition.

The hepatitis C virus (HCV) causes acute and chronic hepatitis, cirrhosis, and hepatocellular carcinoma, as well as extrahepatic manifestations such as malignant lymphoma. Currently, direct-acting antiviral agents (DAAs) against HCV infection can lead to a sustained virological response (SVR) in almost all HCV-infected patients. In this review article, we discuss acute exacerbation and alanine aminotransferase (ALT) flare in patients with chronic HCV infection. Although acute liver failure caused by HCV infection is rare, careful attention should be paid to the cases with ALT elevation during the natural course of chronic HCV infection. HCV genotype 2 infection, the use of rituximab, and a higher dose of corticosteroid are factors associated with HCV acute exacerbation and ALT flare. Treatment regimens for cancer have been interrupted or changed due to ALT flare due to HCV infection in some patients undergoing chemotherapy for cancer. The pathogenesis of HCV acute exacerbation and ALT flare could involve cellular as well as humoral immune responses. In the DAA era, the earlier introduction of DAAs may prevent chronic HCV-infected patients with acute exacerbation and ALT flare from developing into a more severe form, although DAAs may not be effective for all of them.

Grapevine is a widely grown fruit crop that is seriously affected by different viruses, reducing grape yield and quality, as well as threatening profitability. Vineyard disease management requires accurate identification of viral infections. This study aimed to survey the presence of ten grapevine viruses in four geographic sites in the Mendoza province of Argentina. Two hundred twenty-three composite cane samples from 1060 plants of six cultivars were collected from 26 blocks distributed across 11 vineyards. The cane samples were screened by RT-PCR for the following viruses: grapevine leafroll-associated viruses 1-4 (GLRaV 1, 2, 3, and 4), grapevine fanleaf virus (GFLV), grapevine fleck virus (GFkV), grapevine virus A (GVA) and B (GVB), grapevine rupestris stem pitting associated virus (GRSPaV), and arabis mosaic virus (ArMV). The results showed an uneven occurrence of viruses through the sampled regions, with GRSPaV being prevalent (71.1%), followed by GFLV (28.9%), GFkV (20.6%), and GLRaV-2 (14.7%). GVB was not detected. This study revealed a moderate prevalence of viruses associated with economically impactful diseases in the vineyards surveyed.

The clinical course and outcome of COVID-19 are highly variable, ranging from asymptomatic infections to severe disease and death. Understanding the risk factors of severe COVID-19 is relevant both in the clinical setting and at the epidemiological level. Here, we provide an overview of host, viral and environmental factors that have been shown or (in some cases) hypothesized to be associated with severe clinical outcomes. The factors considered in detail include the age and frailty, genetic polymorphisms, biological sex (and pregnancy), co- and superinfections, non-communicable comorbidities, immunological history, microbiota, and lifestyle of the patient; viral genetic variation and infecting dose; socioeconomic factors; and air pollution. For each category, we compile (sometimes conflicting) evidence for the association of the factor with COVID-19 outcomes (including the strength of the effect) and outline possible action mechanisms. We also discuss the complex interactions between the various risk factors.

Human Adenovirus type 6 (HAdV-C6) is a promising candidate for the development of oncolytic vectors as it has low seroprevalence and the intrinsic ability to evade tissue macrophages. However, its further development as a therapeutic agent is hampered by the lack of convenient cloning methods. We have developed a novel technology when a shuttle plasmid carrying the distal genome parts with modified E1A and E3 regions is recombined in vitro with the truncated HAdV-C6 genome. Using this approach, we have constructed a novel Ad6-hT-GM vector controlled by the hTERT promoter and expressing granulocyte-macrophage colony-stimulating factor (GM-CSF) instead of 6.7K and gp19K E3 proteins. We have demonstrated that control by the hTERT promoter may result in delayed viral replication, which nevertheless does not significantly change the cytotoxic ability of recombinant viruses. The insertion of the transgene by displacing the E3-6.7K/gp19K region does not drastically change the expression patterns of E3 genes; however, mild changes in expression from major late promoter were observed. Finally, we have demonstrated that the treatment of human breast cancer xenografts in murine models with Ad6-hT-GM significantly decreased the tumor volume and improved survival time compared to mock-treated mice.

Progressive liver fibrosis after anti-HCV treatment is a risk factor for HCC. Angiopoietin-2 (Ang2) is associated with non-regression of liver fibrosis after direct-acting antiviral (DAA). This study evaluated the predictive value of serum Ang2 levels for HCC occurrence or recurrence after DAA administration. In this retrospective study, 310 HCV-infected patients treated with DAAs in 2014-2020 were screened and evaluated for HCC occurrence or recurrence every three-six months. Multivariate Cox regression analysis revealed that age ≥ 75 years (HR: 2.92, 95% CI: 1.34-6.33; p = 0.007) and baseline Ang2 level ≥ 464 pg/mL (HR: 2.75, 95% CI: 1.18-6.37; p = 0.019) were significantly associated with HCC occurrence after DAA therapy. A high or low risk of HCC after DAA therapy could be distinguished by the combination of age and baseline Ang2 level. The cumulative incidences of de-novo HCC at two and four years were 0.8% and 3.8% in the low-risk group and 22.6% and 27.1% in the high-risk group, respectively. Baseline Ang2 level ≥ 402 pg/mL was significantly associated with HCC recurrence in patients who achieved sustained virological response with DAAs (HR: 3.68). In conclusion, serum Ang2 levels can predict HCC occurrence and recurrence after successful HCV eradication by DAAs.

Oncolytic herpes simplex virus (oHSV) is a type of virus that selectively targets and kills cancer cells, leaving normal cells unharmed. Accurate viral titer is of great importance for the production and application of oHSV products. Droplet digital PCR (ddPCR) is known for having good reproducibility, not requiring a standard curve, not being affected by inhibitors, and being precise even in the detection of low copies. In the present study, we developed a droplet digital PCR assay for the quantification of HSV-1 and applied it in the oHSV production. The established ddPCR showed good specificity, linearity, a low limit of quantification, great reproducibility, and accuracy. The quantification result was well-associated with that of plaque assay and CCID50. Amplification of the purified virus without DNA extraction by ddPCR presented similar results to that from the extracted DNA, confirming the good resistance against PCR inhibitors. With the ddPCR, viral titer could be monitored in real time during the production of oHSV; the optimal harvest time was determined for the best virus yield in each batch. The ddPCR can be used as a useful tool for the quantification of oHSV and greatly facilitate the manufacturing process of oHSV products.

The rise of antimicrobial resistant (AMR) bacteria is a major health concern, especially with regard to members of the ESKAPE group, to which vancomycin-resistant (VRE) Enterococcus faecium belongs. Phage therapy has emerged as a novel alternative for the treatment of AMR infections. This, however, relies on the isolation and characterisation of a large collection of phages. This work describes the exploration of human faeces as a source of new E. faecium-infecting phages. Phage vB_EfaH_163 was isolated and characterised at the microbiological, genomic, and functional levels. vB_EfaH_163 phage, a new member of Herelleviridae, subfamily Brockvirinae, has a dsDNA genome of 150,836 bp that does not harbour any virulence factors or antibiotic resistance genes. It infects a wide range of E. faecium strains of different origins, including VRE strains. Interestingly, it can also infect Enterococcus faecalis strains, even some that are linezolid-resistant. Its capacity to control the growth of a clinical VRE isolate was shown in broth culture and in a Galleria mellonella animal model. The discovery and characterisation of vB_EfaH_163 increases the number of phages that might be used therapeutically against AMR bacteria.

This study describes differences in clinical presentation in hospitalized children with acute COVID-19 and MIS-C between the Delta and Omicron (BA.1.1) waves in a tertiary children's hospital. This retrospective cohort study with case adjudication of hospitalized children with SARS-CoV-2-positive testing or MIS-C diagnosis occurred during the Delta and Omicron waves, from August 2021 until February 2022. There were no differences noted by race, but both waves disproportionally affected black children (24% and 25%). Assigned by a three-person expert panel, incidental diagnoses were higher in the Omicron wave (34% versus 19%). Hospitalization rates of non-incidental cases were higher during Omicron (3.8 versus 5.9 per 1000 PCR-positive community cases). Respiratory-related admissions were prominent during Delta, while Omicron clinical presentations varied, including a high number of cases of croup and seizures. Length of stay and ICU use during Omicron was significantly less than Delta for MIS-C and acute cases. Estimation of vaccination efficacy for preventing hospital admissions was 85.1-91.7% in the early Omicron period. Our estimates suggest that a protective role for vaccination continues into the Omicron wave. The high rate of incidental cases during the Omicron wave should be considered when reviewing more cursory summative data sets. This study emphasizes the need for continued clinical suspicion of COVID-19 even when lower respiratory symptoms are not dominant.

Epstein-Barr virus (EBV) is a gamma-herpesvirus associated with 10% of all gastric cancers (GCs) and 1.5% of all human cancers. EBV-associated GCs (EBVaGCs) are pathologically and clinically distinct entities from EBV-negative GCs (EBVnGCs), with EBVaGCs exhibiting differential molecular pathology, treatment response, and patient prognosis. However, the tumor immune landscape of EBVaGC has not been well explored. In this study, a systemic and comprehensive analysis of gene expression and immune landscape features was performed for both EBVaGC and EBVnGC. EBVaGCs exhibited many aspects of a T cell-inflamed phenotype, with greater T and NK cell infiltration, increased expression of immune checkpoint markers (BTLA, CD96, CTLA4, LAG3, PD1, TIGIT, and TIM3), and multiple T cell effector molecules in comparison with EBVnGCs. EBVaGCs also displayed a higher expression of anti-tumor immunity factors (PDL1, CD155, CEACAM1, galectin-9, and IDO1). Six EBV-encoded miRNAs (miR-BARTs 8-3p, 9-5p, 10-3p, 22, 5-5p, and 14-3p) were strongly negatively correlated with the expression of immune checkpoint receptors and multiple markers of anti-tumor immunity. These profound differences in the tumor immune landscape between EBVaGCs and EBVnGCs may help explain some of the observed differences in pathological and clinical outcomes, with an EBV-positive status possibly being a potential biomarker for the application of immunotherapy in GC.

Virome research is a rapidly growing area in the microbiome field that is increasingly associated with human diseases, such as inflammatory bowel disease (IBD). Although substantial progress has been made, major methodological challenges limit our understanding of the virota. In this review, we describe challenges that must be considered to accurately report the virome composition and the current knowledge on the virome in health and IBD. First, the description of the virome shows strong methodological biases related to wetlab (e.g., VLP enrichment) and bioinformatics approaches (viral identification and classification). Second, IBD patients show consistent viral imbalances characterized by a high relative abundance of phages belonging to the Caudovirales and a low relative abundance of phages belonging to the Microviridae. Simultaneously, a sporadic contraction of CrAss-like phages and a potential expansion of the lysogenic potential of the intestinal virome are observed. Finally, despite numerous studies that have conducted diversity analysis, it is difficult to draw firm conclusions due to methodological biases. Overall, we present the many methodological and environmental factors that influence the virome, its current consensus in health and IBD, and a contributing hypothesis called the "positive inflammatory feedback loop" that may play a role in the pathophysiology of IBD.

This study aimed to evaluate the effectiveness of the phage cocktail to improve the microbiological quality of five different mixed-leaf salads: rucola, mixed-leaf salad with carrot, mixed-leaf salad with beetroot, washed and unwashed spinach, during storage in refrigerated conditions. Enterobacterales rods constituted a significant group of bacteria in the tested products. Selected bacteria were tested for antibiotic resistance profiles and then used to search for specific bacteriophages. Forty-three phages targeting bacteria dominant in mixed-leaf salads were isolated from sewage. Their titer was determined, and lytic activity was assessed using the Bioscreen C Pro automated growth analyzer. Two methods of phage cocktail application including spraying, and an absorption pad were effective for rucola, mixed leaf salad with carrot, and mixed leaf salad with beetroot. The maximum reduction level after 48 h of incubation reached 99.9% compared to the control sample. In washed and unwashed spinach, attempts to reduce the number of microorganisms did not bring the desired effect. The decrease in bacteria count in the lettuce mixes depended on the composition of the autochthonous saprophytic bacteria species. Both phage cocktail application methods effectively improved the microbiological quality of minimally processed products. Whole-spectral phage cocktail application may constitute an alternative food microbiological quality improvement method without affecting food properties.

The rapid emergence of new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants has resulted in the ongoing global coronavirus disease 2019 (COVID-19) pandemic. Thus, the rapid development of a platform to detect a broad range of SARS-CoV-2 variants is essential for successful COVID-19 management. In this study, four SARS-CoV-2 spike protein-specific single-chain variable fragments (scFvs) were isolated from a synthetic antibody library using phage display technology. Following the conversion of these scFvs into monoclonal antibodies (mAbs) (K104.1-K104.4) and production and purification of the mAbs, the antibody pair (K104.1 and K104.2) that exhibited the highest binding affinity (K104.1 and K104.2, 1.3 nM and 1.9 nM) was selected. Biochemical analyses revealed that this antibody pair specifically bound to different sites on the S2 subunit of the spike protein. Furthermore, we developed a highly sensitive sandwich immunoassay using this antibody pair that accurately and quantitatively detected the spike proteins of wild-type SARS-CoV-2 and multiple variants, including Alpha, Beta, Gamma, Delta, Kappa, and Omicron, in the picomolar range. Conclusively, the novel phage display-derived mAbs we have developed may be useful for the rapid and efficient detection of the fast-evolving SARS-CoV-2.

Measles is a highly contagious, but vaccine-preventable disease caused by the measles virus (MeV). Although the administration of two doses of measles vaccines is the most effective strategy to prevent and eliminate measles, MeV continues to spread worldwide, even in 2022. In measles-eliminated countries, preparedness and response to measles outbreaks originating from imported cases are required to maintain elimination status. Under these circumstances, real-time reverse transcription (RT) PCR for MeV could provide a diagnostic method capable of strengthening the subnational capacity for outbreak responses. Real-time RT-PCR can detect MeV RNA from patients with measles at the initial symptomatic stage, which can enable rapid public health responses aimed at detecting their contacts and common sources of infection. Furthermore, low cycle threshold (Ct) values (i.e., high viral load) of throat swabs indicate high infectiousness in patients with measles. The high basic reproduction number of measles suggests that patients with high infectiousness can easily become super-spreaders. This opinion proposes a possible strategy of rapid and intensive responses to counter measles outbreaks caused by super-spreader candidates showing low Ct values in throat swabs. Our strategy would make it possible to effectively prevent further measles transmission, thereby leading to the early termination of measles outbreaks.

Human adenovirus (HAdV) is extremely common and can rapidly spread in confined populations such as daycare centers, hospitals, and retirement homes. Although HAdV usually causes only minor illness in otherwise healthy patients, HAdV can cause significant morbidity and mortality in certain populations, such as the very young, very old, or immunocompromised individuals. During infection, the viral DNA undergoes dramatic changes in nucleoprotein structure that promote the rapid expression of viral genes, replication of the DNA, and generation of thousands of new infectious virions-each process requiring a distinct complement of virus and host-encoded proteins. In this review, we summarize our current understanding of the nucleoprotein structure of HAdV DNA during the various phases of infection, the cellular proteins implicated in mediating these changes, and the role of epigenetics in HAdV gene expression and replication.

Pseudorabies virus (PRV) generally infects pigs and threatens the pig industry. However, recently we have isolated a PRV strain designated hSD-1/2019 from infected humans. In this study, we compared the complete genome sequence of hSD-1/2019 with those of pig-originated PRV strains. Sequence alignments revealed that the genome sequence of hSD-1/2019 was highly homologous to those of the porcine PRV strains. Phylogenetic analyses found that hSD-1/2019 was the closest related to porcine PRV endemic strains in China, particularly the variant strains circulating recently. We also showed that the glycoproteins important for the multiplication and pathogenesis of hSD-1/2019 were highly similar to those of the pig endemic strains. Diversifying selection analyses revealed that hSD-1/2019 and pig variant strains are under diversifying selection. Recombination analysis indicated that hSD-1/2019 was a recombinant of several PRV variant strains and an earlier PRV classic strain. Finally, we found that both human and pig-originated PRV strains could induce cytopathic effects in cells from humans, pigs, and mice, but only the human PRV and pig-variant PRV formed large syncytia in human cell lines. The data presented in this study contribute to our understanding of the molecular basis for the pathogenesis of human PRV from a genomic aspect.

A new HP-PRRSV strain (SD2020) was isolated from pigs with suspected highly pathogenic porcine reproductive and respiratory syndrome disease in a pig farm in Shandong Province, China, and its genome was sequenced. This pig farm has been using the VR-2332 vaccine strain to immunize pigs for a long time. The phylogenic and single nucleotide polymorphisms (SNPs) analysis of the viruses isolated from dead pigs showed that SD2020 was a natural recombinant virus of the VR-2332 vaccine strain and the JXA1 similar strain, and that two splicing fragments highly homologous to JXA1 in the virus genome were probably derived from the JXA1 wild strain and JXA1-R vaccine strain, respectively. Therefore, the possible recombination events of SD2020 and its mutation site might be related to high pathogenicity.

Alphaviruses are arthropod-borne, single-stranded positive sense RNA viruses that rely on the engagement of host RNA-binding proteins to efficiently complete the viral lifecycle. Because of this reliance on host proteins, the identification of host/pathogen interactions and the subsequent characterization of their importance to viral infection has been an intensive area of study for several decades. Many of these host protein interaction studies have evaluated the Protein:Protein interactions of viral proteins during infection and a significant number of host proteins identified by these discovery efforts have been RNA Binding Proteins (RBPs). Considering this recognition, the field has shifted towards discovery efforts involving the direct identification of host factors that engage viral RNAs during infection using innovative discovery approaches. Collectively, these efforts have led to significant advancements in the understanding of alphaviral molecular biology; however, the precise extent and means by which many RBPs influence viral infection is unclear as their specific contributions to infection, as per any RNA:Protein interaction, have often been overlooked. The purpose of this review is to summarize the discovery of host/pathogen interactions during alphaviral infection with a specific emphasis on RBPs, to use new ontological analyses to reveal potential functional commonalities across alphaviral RBP interactants, and to identify host RBPs that have, and have yet to be, evaluated in their native context as RNA:Protein interactors.

Norovirus is the leading viral agent of gastroenteritis in humans. RNA-dependent RNA polymerase (RdRp) is essential in the replication of norovirus RNA. Here, we present a comprehensive evolutionary analysis of the norovirus GI RdRp gene. Our results show that the norovirus GI RdRp gene can be divided into three groups, and that the most recent common ancestor was 1484. The overall evolutionary rate of GI RdRp is 1.821 × 10-3 substitutions/site/year. Most of the amino acids of the GI RdRp gene were under negative selection, and only a few positively selected sites were recognized. Amino acid substitutions in the GI RdRp gene accumulated slowly over time. GI.P1, GI.P3 and GI.P6 owned the higher evolutionary rates. GI.P11 and GI.P13 had the faster accumulation rate of amino acid substitutions. GI.P2, GI.P3, GI.P4, GI.P6 and GI.P13 presented a strong linear evolution. These results reveal that the norovirus GI RdRp gene evolves conservatively, and that the molecular evolutionary characteristics of each P-genotype are diverse. Sequencing in RdRp and VP1 of norovirus should be advocated in the surveillance system to explore the effect of RdRp on norovirus activity.

The COVID-19 pandemic has created significant concern for everyone. Recent data from many worldwide reports suggest that most infections are caused by the Omicron variant and its sub-lineages, dominating all the previously emerged variants. The numerous mutations in Omicron's viral genome and its sub-lineages attribute it a larger amount of viral fitness, owing to the alteration of the transmission and pathophysiology of the virus. With a rapid change to the viral structure, Omicron and its sub-variants, namely BA.1, BA.2, BA.3, BA.4, and BA.5, dominate the community with an ability to escape the neutralization efficiency induced by prior vaccination or infections. Similarly, several recombinant sub-variants of Omicron, namely XBB, XBD, and XBF, etc., have emerged, which a better understanding. This review mainly entails the changes to Omicron and its sub-lineages due to it having a higher number of mutations. The binding affinity, cellular entry, disease severity, infection rates, and most importantly, the immune evading potential of them are discussed in this review. A comparative analysis of the Delta variant and the other dominating variants that evolved before Omicron gives the readers an in-depth understanding of the landscape of Omicron's transmission and infection. Furthermore, this review discusses the range of neutralization abilities possessed by several approved antiviral therapeutic molecules and neutralizing antibodies which are functional against Omicron and its sub-variants. The rapid evolution of the sub-variants is causing infections, but the broader aspect of their transmission and neutralization has not been explored. Thus, the scientific community should adopt an elucidative approach to obtain a clear idea about the recently emerged sub-variants, including the recombinant variants, so that effective neutralization with vaccines and drugs can be achieved. This, in turn, will lead to a drop in the number of cases and, finally, an end to the pandemic.

Cyprinid herpesvirus 3 (CyHV-3) can cause severe disease in koi and common carp (Cyprinus carpio). Currently, no effective treatment is available against CyHV-3 infection in koi. Both LSD1 and JMJD2 are histone demethylases (HD) and are critical for immediate-early (IE) gene activation essential for lytic herpesvirus replication. OG-L002 and ML324 are newly discovered specific inhibitors of LSD1 and JMJD2, respectively. Here, HD inhibitors were compared with acyclovir (ACV) against CyHV-3 infection in vitro and in vivo. ML324, at 20-50 µM, can completely block ~1 × 103 PFU CyHV-3 replication in vitro, while OG-L002 at 20 µM and 50 µM can produce 96% and 98% inhibition, respectively. Only about 94% inhibition of ~1 × 103 PFU CyHV-3 replication was observed in cells treated with ACV at 50 µM. As expected, CyHV-3 IE gene transcription of ORF139 and ORF155 was blocked within 72 h post-infection (hpi) in the presence of 20 µM ML324. No detectable cytotoxicity was observed in KF-1 or CCB cells treated for 24 h with 1 to 50 µM ML324. A significant reduction of CyHV-3 replication was observed in ~6-month-old infected koi treated with 20 µM ML324 in an immersion bath for 3-4 h at 1-, 3-, and 5-days post-infection compared to the control and ACV treatments. Under heat stress, 50-70% of 3-4-month-old koi survived CyHV-3 infection when they were treated daily with 20 µM ML324 in an immersion bath for 3-4 h within the first 5 d post-infection (dpi), compared to 11-19% and 22-27% of koi in the control and ACV treatments, respectively. Our study demonstrates that ML324 has the potential to be used against CyHV-3 infection in koi.

Wastewater surveillance of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has shown to be an important approach to determine early outbreaks of infections. Wastewater-based epidemiology (WBE) is regarded as a complementary tool for monitoring SARS-CoV-2 trends in communities. In this study, the changes in the SARS-CoV-2 RNA levels in wastewater during Easter holidays in 2021 and 2022 in the City of Cape Town were monitored over nine weeks. Our findings showed a statistically significant difference in the SARS-CoV-2 RNA viral load between the study weeks over the Easter period in 2021 and 2022, except for study week 1 and 4. During the Easter week, 52% of the wastewater treatment plants moved from the lower (low viral RNA) category in 2021 to the higher (medium to very high viral RNA) categories in 2022. As a result, the median SARS-CoV-2 viral loads where higher during the Easter week in 2022 than Easter week in 2021 (p = 0.0052). Mixed-effects model showed an association between the SARS-CoV-2 RNA viral loads and Easter week over the Easter period in 2021 only (p < 0.01). The study highlights the potential of WBE to track outbreaks during the holiday period.

Plum pox virus (PPV) is a causal agent of the stone fruit tree sharka disease that often causes enormous economic losses. Due to its worldwide distribution and economic importance, rapid and reliable diagnostic technologies are becoming increasingly important for successful management of sharka disease. In this study, we have produced two super-sensitive and specific anti-PPV monoclonal antibodies (i.e., MAbs 13H4 and 4A11). Using these two MAbs, we have now developed a dot enzyme-linked immunosorbent assay (dot-ELISA) and a colloidal gold immunochromatographic strip (CGICS) assay. These two technologies can be used to quickly and reliably detect PPV. The results of these sensitivity assays confirmed that the dot-ELISA and CGICS assays could detect PPV infection in apricot tree leaf crude extracts diluted up to 1:5120 and 1:6400 (w/v), respectively. Further analyses using field-collected apricot tree leaf samples showed that the detection endpoint of the dot-ELISA was ~26 times above that obtained through RT-PCR, and the CGICS was as sensitive as RT-PCR. This present study is to broaden the knowledge about detection limits of dot-ELISA and CGICS for PPV monitoring. We consider that these newly developed dot-ELISA and CGICS are particularly useful for large scale PPV surveys in fields.

Human endogenous retroviruses (HERVs) are remnants of ancestral germline infections by exogenous retroviruses. Human endogenous retroviruses W family envelope gene (HERV-W env, also called ERVWE1), located on chromosome 7q21-22, encodes an envelope glycoprotein from the HERV-W family. Mounting evidence suggests that aberrant expression of ERVWE1 involves the etiology of schizophrenia. Moreover, the genetic and morphological studies indicate that dendritic spine deficits may contribute to the onset of schizophrenia. Here, we reported that ERVWE1 changed the density and morphology of the dendritic spine through inhibiting Wingless-type (Wnt)/c-Jun N-terminal kinases (JNK) non-canonical pathway via miR-141-3p in schizophrenia. In this paper, we found elevated levels of miR-141-3p and a significant positive correlation with ERVWE1 in schizophrenia. Moreover, serum Wnt5a and actin-related protein 2 (Arp2) levels decreased and demonstrated a significant negative correlation with ERVWE1 in schizophrenia. In vitro experiments disclosed that ERVWE1 up-regulated miR-141-3p expression by interacting with transcription factor (TF) Yin Yang 1 (YY1). YY1 modulated miR-141-3p expression by binding to its promoter. The luciferase assay revealed that YY1 enhanced the promoter activity of miR-141-3p. Using the miRNA target prediction databases and luciferase reporter assays, we demonstrated that miR-141-3p targeted Wnt5a at its 3' untranslated region (3' UTR). Furthermore, ERVWE1 suppressed the expression of Arp2 through non-canonical pathway, Wnt5a/JNK signaling pathway. In addition, ERVWE1 inhibited Wnt5a/JNK/Arp2 signal pathway through miR-141-3p. Finally, functional assays showed that ERVWE1 induced the abnormalities in hippocampal neuron morphology and spine density through inhibiting Wnt/JNK non-canonical pathway via miR-141-3p in schizophrenia. Our findings indicated that miR-141-3p, Wnt5a, and Arp2 might be potential clinical blood-based biomarkers or therapeutic targets for schizophrenia. Our work also provided new insight into the role of ERVWE1 in schizophrenia pathogenesis.

Genome integrity is critical for proper cell functioning, and chromosome instability can lead to age-related diseases, including cancer and neurodegenerative disorders. Chromosome instability is caused by multiple factors, including replication stress, chromosome missegregation, exposure to pollutants, and viral infections. Although many studies have investigated the effects of environmental or lifestyle genotoxins on chromosomal integrity, information on the effects of viral infections on micronucleus formation and other chromosomal aberrations is still limited. Currently, HIV infection is considered a chronic disease treatable by antiretroviral therapy (ART). However, HIV-infected individuals still face important health problems, such as chronic inflammation and age-related diseases. In this context, this article reviews studies that have evaluated genomic instability using micronucleus assays in the context of HIV infection. In brief, HIV can induce chromosome instability directly through the interaction of HIV proteins with host DNA and indirectly through chronic inflammation or as a result of ART use. Connections between HIV infection, immunosenescence and age-related disease are discussed in this article. The monitoring of HIV-infected individuals should consider the increased risk of chromosome instability, and lifestyle interventions, such as reduced exposure to genotoxins and an antioxidant-rich diet, should be considered. Therapies to reduce chronic inflammation in HIV infection are needed.

The order Mononegavirales contains a variety of highly pathogenic viruses that may infect humans, including the families Filoviridae, Bornaviridae, Paramyxoviridae, and Rhabodoviridae. Animal models have historically been important to study virus pathogenicity and to develop medical countermeasures. As these have inherent shortcomings, the rise of microphysiological systems and organoids able to recapitulate hallmarks of the diseases caused by these viruses may have enormous potential to add to or partially replace animal modeling in the future. Indeed, microphysiological systems and organoids are already used in the pharmaceutical R&D pipeline because they are prefigured to overcome the translational gap between model systems and clinical studies. Moreover, they may serve to alleviate ethical concerns related to animal research. In this review, we discuss the value of animal model alternatives in human pathogenic filovirus and bornavirus research. The current animal models and their limitations are presented followed by an overview of existing alternatives, such as organoids and microphysiological systems, which might help answering open research questions.

Aging processes, including immunosenescence, inflammation, inflammasome formation, genomic instability, telomeric attrition, and altered autophagy, are involved in viral infections and they may contribute to increased pathophysiological responses to the SARS-CoV-2 infection in the elderly; this poses additional risks of accelerated aging, which could be found even after recovery. Aging is associated with oxidative damage. Moreover, SARS-CoV-2 infections may increase the production of reactive oxygen species and such infections will disturb the Ca++ balance via an endoplasmic reticulum (ER) stress-mediated unfolded protein response. Although vaccine development and anti-inflammation therapy lower the severity of COVID-19, the prevalence and mortality rates are still alarming in some countries worldwide. In this review, we describe the involvement of viral proteins in activating ER stress transducers and their downstream signals and in inducing inflammation and inflammasome formation. Furthermore, we propose the potential of melatonin as an ER stress modulator, owing to its antioxidant, anti-inflammatory, and immunoregulatory effects in viral infections. Considering its strong safety profile, we suggest that additive melatonin supplementation in the elderly could be beneficial in treating COVID-19.

COVID-19 is still a global public health concern, and the SARS-CoV-2 mutations require more effective antiviral agents. In this study, the antiviral entry activity of thirty-one flavonoids was systematically evaluated by a SARS-CoV-2 pseudovirus model. Twenty-four flavonoids exhibited antiviral entry activity with IC50 values ranging from 10.27 to 172.63 µM and SI values ranging from 2.33 to 48.69. The structure-activity relationship of these flavonoids as SARS-CoV-2 entry inhibitors was comprehensively summarized. A subsequent biolayer interferometry assay indicated that flavonoids bind to viral spike RBD to block viral interaction with ACE2 receptor, and a molecular docking study also revealed that flavonols could bind to Pocket 3, the non-mutant regions of SARS-CoV-2 variants, suggesting that flavonols might be also active against virus variants. These natural flavonoids showed very low cytotoxic effects on human normal cell lines. Our findings suggested that natural flavonoids might be potential antiviral entry agents against SARS-CoV-2 via inactivating the viral spike. It is hoped that our study will provide some encouraging evidence for the use of natural flavonoids as disinfectants to prevent viral infections.

The necrogenic strain N5 of tomato mosaic virus (ToMV-N5) causes systemic necrosis in tomato cultivar Hezuo903. In this work, we mapped the viral determinant responsible for the induction of systemic necrosis. By exchanging viral genes between N5 and a non-necrogenic strain S1, we found that movement protein (MP) was the determinant for the differential symptoms caused by both strains. Compared with S1 MP, N5 MP had an additional ability to increase virus accumulation, which was not due to its functions in viral cell-to-cell movement. Actually, N5 MP, but not S1 MP, was a weak RNA silencing suppressor, which assisted viral accumulation. Sequence alignment showed that both MPs differed by only three amino acid residues. Experiments with viruses having mutated MPs indicated that the residue isoleucine at position 170 in MP was the key site for MP to increase virus accumulation, but also was required for MP to induce systemic necrosis in virus-infected tomato plants. Collectively, the lethal necrosis caused by N5 is dependent on its MP protein that enhances virus accumulation via its RNA silencing suppressor activity, probably leading to systemic necrosis responses in tomato plants.

The Japanese government withdrew its recommendation for human papillomavirus (HPV) vaccination in June 2013 and resumed it in April 2022. This period is known as the vaccine crisis in Japan. This study aimed to elucidate the prevalence and genotype distribution of HPV among Japanese female citizens, and the effect of vaccination against HPV-16/18 in the era of the vaccine crisis. We recruited Japanese female citizens and asked them to provide self-collected samples from the vaginal wall using cotton swabs for HPV genotyping. Furthermore, we collected the participants' characteristics, including lifestyle and experience of vaccination against HPV, to determine the significant association with HPV infection. HPV-16/18 positivity was found in 5.6% (115/2044) of participants. The highest vaccination rate was observed in the age group of 20-24 years (60.6%), whereas the lowest HPV-16/18 positivity was observed in the age group of 45-49 years (2.8%), followed by the age group of 20-24 years (4.0%). Experience with HPV vaccination significantly reduced the risk of HPV-16/18 infection (adjusted odds ratio, 0.047; 95% confidence interval, 0.011-0.196). Vaccinated women were much less likely to be infected by HPV-16/18, regardless of the HPV vaccine type or the vaccination dose.

Viral hemorrhagic septicaemia virus (VHSV) has been demonstrated to cause high mortalities in a wide range of teleosts, farmed as well as wild. In Europe, VHSV of genotypes Ib, Id, II, and III have been detected in wild fish, including Atlantic herring Clupea harengus, but disease outbreaks have not been observed in Atlantic herring and the effects on wild stocks are not well documented. Here, we have tested two VHSV isolates from herring (genotypes Ib and III, from the western coasts of Norway and Denmark, respectively) in a challenge experiment with herring (mean weight 2.59 g, SD 0.71 g) caught on the west coast of Denmark. The Norwegian genotype Ib isolate (NO-F-CH/2009) showed an accumulated mortality of 47% compared to 6% mortality with the Danish genotype III isolate 4p168 and zero in the unchallenged control group. In both groups, we found positive rt-RT-PCR and positive immunohistochemistry of VHSV from days 6 and 8 onward. With both isolates, the organs mainly affected were the heart and kidney. The results demonstrate the susceptibility of Atlantic herring to VHSV, and both genotypes gave pathological findings in several organs. Genotype III showed a low mortality rate, and the importance of this genotype for herring is therefore not determined. Genotype Ib showed both high prevalence and mortality, and this genotype is therefore likely to have a negative effect on wild Atlantic herring stocks. Further examinations to determine how VHSV can affect wild Atlantic herring stocks are needed.

African swine fever (ASF) is a highly contagious hemorrhagic viral disease of domestic and wild pigs of all breeds and ages, caused by African swine fever virus (ASFV). Due to the absence of a safe and efficacious vaccine, accurate laboratory diagnosis is critical for the control of ASF prevention. The p30 protein is immunogenic and stimulates a high level of antibody response to ASFV infection. We developed a panel of 4 monoclonal antibodies (mAbs) against p30 protein, and mAb-2B4 showed the highest percent of inhibition (PI) of 70% in the solid phase blocking ELISA (bELISA). Epitope mapping revealed the mAb-2B4 recognized the epitope of aa 12-18 of p30, which is conserved among various ASFV genotypes. Subsequently, a competitive enzyme-linked immunosorbent assay (cELISA) was established using HRP-labeled mAb-2B4. The cutoff for discrimination between 98 negative sera and 40 positive sera against ASFV was determined by plotting a receiver operating characteristic (ROC) curve. It yielded the area under the curve (AUC) of 0.998, and a diagnostic specificity of 97.96% and a sensitivity of 97.5% were achieved when the cutoff value was determined at 37.1%. Furthermore, the results showed an excellent repeatability of the established cELISA and no cross-reaction to antisera against six other pig pathogens. Additionally, the cELISA detected a titer of 1:256 in the positive standard serum. Overall, mAb-2B4 showed a conserved epitope and high ability to be inhibited by positive sera in ASFV antibody detection. The cELISA based on HRP-labeled mAb-2B4 offers an alternative to other assays for a broader diagnostic coverage of ASFV infection.

Herpes simplex virus (HSV) and varicella zoster virus (VZV) rely on transport of virus particles in neuronal axons to spread from sites of viral latency in sensory ganglia to peripheral tissues then on to other hosts. This process of anterograde axonal transport involves kinesin motors that move virus particles rapidly along microtubules. α-herpesvirus anterograde transport has been extensively studied by characterizing the porcine pseudorabies virus (PRV) and HSV, with most studies focused on two membrane proteins: gE/gI and US9. It was reported that PRV and HSV US9 proteins bind to kinesin motors, promoting tethering of virus particles on the motors, and furthering anterograde transport within axons. Alternatively, other models have argued that HSV and PRV US9 and gE/gI function in the cytoplasm and not in neuronal axons. Specifically, HSV gE/gI and US9 mutants are defective in the assembly of virus particles in the cytoplasm of neurons and the subsequent sorting of virus particles to cell surfaces and into axons. However, PRV US9 and gE/gI mutants have not been characterized for these cytoplasmic defects. We examined neurons infected with PRV mutants, one lacking both gE/gI and US9 and the other lacking just US9, by electron microscopy. Both PRV mutants exhibited similar defects in virus assembly and cytoplasmic sorting of virus particles to cell surfaces. As well, the mutants exhibited reduced quantities of infectious virus in neurons and in cell culture supernatants. We concluded that PRV US9 primarily functions in neurons to promote cytoplasmic steps in anterograde transport.

LAG-3 is a type I transmembrane protein expressed on immune cells, such as activated T cells, and binds to MHC class II with high affinity. LAG-3 is an inhibitory receptor, and its multiple biological activities on T cell activation and effector functions play a regulatory role in the immune response. Immunotherapies directed at immune checkpoints, including LAG-3, have become a promising strategy for controlling malignant tumors and chronic viral diseases. Several studies have suggested an association between the expression of LAG-3 with an inadequate immune response during respiratory viral infections and the susceptibility to reinfections, which might be a consequence of the inhibition of T cell effector functions. However, important information relative to therapeutic potential during acute viral lower respiratory tract infections and the mechanism of action of the LAG-3 checkpoint remains to be characterized. In this article, we discuss the contribution of LAG-3 to the impairment of T cells during viral respiratory infections. Understanding the host immune response to respiratory infections is crucial for developing effective vaccines and therapies.

Lassa fever (LF) is a rodent-borne disease that threatens human health in the sub-region of West Africa where the zoonotic host of Lassa virus (LASV) is predominant. Currently, treatment options for LF are limited and since no preventive vaccine is approved for its infectivity, there is a high mortality rate in endemic areas. This narrative review explores the transmission, pathogenicity of LASV, advances, and challenges of different treatment options. Our findings indicate that genetic diversity among the different strains of LASV and their ability to circumvent the immune system poses a critical challenge to the development of LASV vaccines/therapeutics. Thus, understanding the biochemistry, physiology and genetic polymorphism of LASV, mechanism of evading host immunity are essential for development of effective LASV vaccines/therapeutics to combat this lethal viral disease. The LASV nucleoprotein (NP) is a novel target for therapeutics as it functions significantly in several aspects of the viral life cycle. Consequently, LASV NP inhibitors could be employed as effective therapeutics as they will potentially inhibit LASV replication. Effective preventive control measures, vaccine development, target validation, and repurposing of existing drugs, such as ribavirin, using activity or in silico-based and computational bioinformatics, would aid in the development of novel drugs for LF management.

(1) Background: Largemouth bass virus (LMBV) is a major viral pathogen in largemouth bass (Micropterus salmoides) aquaculture that often causes high mortality and heavy economic losses, thus developing treatments to combat this pathogen is of great commercial importance. Green tea is a well-known medicinal plant that contains active ingredients with antiviral, antibacterial, and other biological activities. The goals of this study were to explore the effect and mechanism of green tea source compounds on LMBV and provide data to serve as the basis for the screening of targeted drugs in the future. In this study, we evaluated the effects of the main component of green tea, epigallocatechin-3-gallate (EGCG), against LMBV infection. (2) Methods: The safe working concentration of EGCG was identified by cell viability detection and light microscopy. The antiviral activity and mechanism of action of EGCG against LMBV infection were evaluated with light microscopy, an aptamer 6-carboxy-fluorescein-based fluorescent molecular probe, and reverse transcription quantitative PCR. (3) Results: The safe working concentration of EGCG was ≤10 μg/mL. EGCG showed significant anti-LMBV infection activity in a concentration-dependent manner, and it also destroyed the structure of virus particles. EGCG impacted the binding of virus particles to cell receptors and virus invasion into the host cells. Inhibitory effects of EGCG on LMBV particles, LMBV binding to the host-cell membrane, and LMBV invasion were 84.89%, 98.99%, and 95.23%, respectively. Meanwhile, the effects of EGCG subsequently were verified in vivo. The fatality rate of the LMBV + EGCG group was significantly lower than that of the LMBV group. (4) Conclusions: Our results suggest that EGCG has effective antiviral properties against LMBV and may be a candidate for the effective treatment and control of LMBV infections in largemouth bass aquaculture.

The COVID-19 pandemic proceeds in waves, with variable characteristics of the clinical picture resulting from the evolution of the SARS-CoV-2 virus. This study aimed to compare the epidemiological characteristics, symptomatology, and outcomes of the disease in patients hospitalized for COVID-19 during periods of different variants dominance. Comparing the periods of dominance of variants preceding the Delta variant, the Delta period was characterized by a higher share of hospitalized females, less frequent comorbidities among patients, and a different age distribution. The lowest need for oxygen therapy and mechanical ventilation was observed under Omicron dominance. The triad of classic COVID-19 symptoms, cough, fever, dyspnoea, and fatigue, were most prevalent during the Delta period, and significantly less common under the Omicron dominance. During the Omicron period, nearly twice as many patients as in the previous periods could be discharged from the hospital within 7 days; the overall 28-day mortality was significantly lower compared to that of the Delta period. It also did not differ between periods that were dominated by the BA.1 and BA.2 subvariants. The study indicates that the Omicron SARS-CoV-2 variant that dominated between January and June 2022 caused a disease which resembled the common cold, and was caused by seasonal alpha and beta-coronaviruses with a low pathogenicity for humans. However, one should note that this effect may not only have been related to biological features of the Omicron lineage, but may additionally have been driven by the increased levels of immunization through natural infections and vaccinations, for which we could not account for due to a lack of sufficient data.

Domestic cat hepadnavirus (DCH) is an emerging virus related to the hepatitis B virus (HBV). The pathogenic potential of DCH in cats remains to be established. The molecular prevalence of DCH varies widely in the regions investigated so far. The aim of this study was to determine the prevalence, load, and risk factors for DCH detection among cats in Hong Kong, and to generate molecular and epidemiological data on the DCH strains circulating in cats in Hong Kong. DCH DNA was detected using DCH-specific qPCR in 57/513 (11.1%) residual diagnostic blood samples from owned cats. The median viral load was 8.85 × 103 copies/mL of whole blood (range for the 5th to the 95th percentile, 3.33 × 103 to 2.2 × 105 copies per mL). Two outliers had higher viral loads of 1.88 × 107 copies/mL and 4.90 × 109 copies/mL. DCH was detected in cats from 3 months to 19 years of age. Sex, age, neuter status, breed, or elevated serum alanine aminotransferase were not statistically associated with DCH DNA detection. On phylogenetic analysis based on 12 complete genome sequences, the Hong Kong DCH viruses clustered in Genotype A with viruses from Australia and Asia (clade A1), distinct from viruses from Europe (clade A2). Sequence analysis found that DCH has similar epsilon and direct repeat regions to human HBV, suggesting a conserved method of replication. Based on our findings, the DCH strains circulating in Hong Kong are a continuum of the Asiatic strains.

The continuously expanding distribution of sand flies, proven vectors of Leishmania and of several phleboviruses, is a growing public health issue in Europe. Especially in Italy, visceral leishmaniasis (VL) is occurring with increasing incidence northward, in previously non-endemic provinces. Around the globe, disease elimination efforts largely focus on sand fly vector insecticidal control, often leading to the development of resistance. In Emilia-Romagna (ER), northern Italy, insecticides are heavily applied for agricultural and mosquito control, but not specifically against sand flies. Here, we investigated the sand fly species composition in certain environmental settings in ER provinces and monitored the presence of pyrethroid resistance mutations and pathogen circulation. Phlebotomus perfiliewi, a dominant vector of Leishmania infantum, was detected almost exclusively in the region. No mutations in the voltage-gated sodium channel gene, e.g., knock-down resistance mutations I1011M, L1014F/S, V1016G, or F1020S, were recorded. Pathogen monitoring revealed that almost 40% of the tested sand fly pools were positive for Leishmania, while the presence of Toscana and Fermo phleboviruses was also observed in much lower frequencies (≤3% positive pools). Regular epidemiological and entomological monitoring, alongside resistance surveillance, is highly recommended to ensure the sustainability and efficiency of vector control interventions.

Zika virus (ZIKV) is an RNA-enveloped virus that belongs to the Flavivirus genus, and ZIKV infections potentially induce severe neurodegenerative diseases and impair male fertility. Palmitoylation is an important post-translational modification of proteins that is mediated by a series of DHHC-palmitoyl transferases, which are implicated in various biological processes and viral infections. However, it remains to be investigated whether palmitoylation regulates ZIKV infections. In this study, we initially observed that the inhibition of palmitoylation by 2-bromopalmitate (2-BP) enhanced ZIKV infections, and determined that the envelope protein of ZIKV is palmitoylated at Cys308. ZDHHC11 was identified as the predominant enzyme that interacts with the ZIKV envelope protein and catalyzes its palmitoylation. Notably, ZDHHC11 suppressed ZIKV infections in an enzymatic activity-dependent manner and ZDHHC11 knockdown promoted ZIKV infection. In conclusion, we proposed that the envelope protein of ZIKV undergoes a novel post-translational modification and identified a distinct mechanism in which ZDHHC11 suppresses ZIKV infections via palmitoylation of the ZIKV envelope protein.

Birds may act as hosts for numerous pathogens, including members of the family Chlamydiaceae, beak and feather disease virus (BFDV), avipoxviruses, Columbid alphaherpesvirus 1 (CoAHV1) and Psittacid alphaherpesvirus 1 (PsAHV1), all of which are a significant biosecurity concern in Australia. While Chlamydiaceae and BFDV have previously been detected in Australian avian taxa, the prevalence and host range of avipoxviruses, CoAHV1 and PsAHV1 in Australian birds remain undetermined. To better understand the occurrence of these pathogens, we screened 486 wild birds (kingfisher, parrot, pigeon and raptor species) presented to two wildlife hospitals between May 2019 and December 2021. Utilising various qPCR assays, we detected PsAHV1 for the first time in wild Australian birds (37/486; 7.61%), in addition to BFDV (163/468; 33.54%), Chlamydiaceae (98/468; 20.16%), avipoxviruses (46/486; 9.47%) and CoAHV1 (43/486; 8.85%). Phylogenetic analysis revealed that BFDV sequences detected from birds in this study cluster within two predominant superclades, infecting both psittacine and non-psittacine species. However, BFDV disease manifestation was only observed in psittacine species. All Avipoxvirus sequences clustered together and were identical to other global reference strains. Similarly, PsAHV1 sequences from this study were detected from a series of novel hosts (apart from psittacine species) and identical to sequences detected from Brazilian psittacine species, raising significant biosecurity concerns, particularly for endangered parrot recovery programs. Overall, these results highlight the high pathogen diversity in wild Australian birds, the ecology of these pathogens in potential natural reservoirs, and the spillover potential of these pathogens into novel host species in which these agents cause disease.

A novel virus with a double-stranded RNA (dsRNA) genome was isolated from Fusarium avenaceum strain GS-WW-224, the causal agent of potato dry rot. The virus has been designated as Fusarium avenaceum alternavirus 1 (FaAV1). Its genome consists of two dsRNA segments, 3538 bp (dsRNA1) and 2477 bp (dsRNA2) in length, encoding RNA-dependent RNA polymerase (RdRp) and a hypothetical protein (HP), respectively. The virions of FaAV1 are isometric spherical and approximately 30 nm in diameter. Multiple sequence alignments and phylogenetic analyses based on the amino acid sequences of RdRp and HP indicated that FaAV1 appears to be a new member of the proposed family Alternaviridae. No significant differences in colony morphology and spore production were observed between strains GS-WW-224 and GS-WW-224-VF, the latter strain being one in which FaAV1 was eliminated from strain GS-WW-224. Notably, however, the dry weight of mycelial biomass of GS-WW-224 was higher than that of mycelial biomass of GS-WW-224-VF. The depth and the width of lesions on potato tubers caused by GS-WW-224 were significantly greater, relative to GS-WW-224-VF, suggesting that FaAV1 confers hypervirulence to its host, F. avenaceum. Moreover, FaAV1 was successfully transmitted horizontally from GS-WW-224 to ten other species of Fusarium, and purified virions of FaAV1 were capable of transfecting wounded hyphae of the ten species of Fusarium. This is the first report of an alternavirus infecting F. avenaceum and conferring hypervirulence.

Scientific progress in understanding, preventing, treating, and managing viral infections and associated diseases exemplifies the extent to which research on small DNA tumor viruses has impacted human health [...].

Vector control methods are considered effective in averting dengue transmission. However, several factors may modify their impact. Of these controls, chemical methods, in the long run, may increase mosquitoes' resistance to chemicides, thereby decreasing control efficacy. The biological methods, which may be self-sustaining and very effective, could be hampered by seasonality or heatwaves (resulting in, e.g., loss of Wolbachia infection). The environmental methods that could be more effective than the chemical methods are under-investigated. In this study, a systematic review is conducted to explore the present understanding of the effectiveness of vector control approaches via dengue transmission models.

Studies on cognitive problems of persons with mild COVID-19 courses are still lacking. This study aimed to determine the frequency and associated factors of subjective and objective cognitive problems after COVID-19 in non-hospitalized persons. Study participants were examined at the University Hospital of Augsburg from 04/11/2020 to 26/05/2021. The Wechsler Adult Intelligence Scale (WAIS) IV digit span, Stroop Color and Word Test (SCWT), Regensburger verbal fluency test (RWT) and, subjective ratings of memory and concentration were applied. Of the 372 participants (mean age 46.8 ± 15.2 years, 54.3% women, median time after infection 9.1 months), 24.9% reported concentration and 21.9% memory problems. Overall, 55.6% of the participants had at least a mild negative alteration in any cognitive test. The strongest impairments were found regarding memory functions (41.1% mild alterations, 6.2% distinct impairments) and verbal fluency (12.4% mild alterations, 5.4% distinct impairments). SCWT showed negative alterations in no more than 3.0% of the participants. Level of school education, age, and depressiveness emerged as significantly related to the cognitive tests. The number of complaints and depressiveness were significantly associated with subjective memory and concentration problems. It is important to identify mild cognitive impairment in non-hospitalized COVID-19 patients early to offer them effective interventions.

Equine viral arteritis is an infectious disease of equids caused by equine arteritis virus (EAV), an RNA virus of the family Arteriviridae. Dendritic cells (DC) are important modulators of the immune response with the ability to present antigen to naïve T cells and can be generated in vitro from monocytes (MoDC). DC are important targets for many viruses and this interaction is crucial for the establishment-or rather not-of an anti-viral immunity. Little is known of the effect EAV has on host immune cells, particularly DC. To study the interaction of eqDC with EAV in vitro, an optimized eqMoDC system was used, which was established in a previous study. MoDC were infected with strains of different genotypes and pathogenicity. Virus replication was determined through titration and qPCR. The effect of the virus on morphology, phenotype and function of cells was assessed using light microscopy, flow cytometry and in vitro assays. This study confirms that EAV replicates in monocytes and MoDC. The replication was most efficient in mature MoDC, but variable between strains. Only the virulent strain caused a significant down-regulation of certain proteins such as CD14 and CD163 on monocytes and of CD83 on mature MoDC. Functional studies conducted after infection showed that EAV inhibited the endocytic and phagocytic capacity of Mo and mature MoDC with minimal effect on immature MoDC. Infected MoDC showed a reduced ability to stimulate T cells. Ultimately, EAV replication resulted in an apoptosis-mediated cell death. Thus, EAV evades the host anti-viral immunity both by inhibition of antigen presentation early after infection and through killing infected DC during replication.

The immune responses of liver transplant (LT) recipients after the third boost of the BNT162b2mRNA vaccine improved. This study evaluates the durability of the immune response of LT recipients after the third boost, its predictors, and the impact of emerging variants. The receptor-binding domain IgG was determined at median times of 22 (first test) and 133 days (second test) after the administration of the third boost. IgG antibody titers > 21.4 BAU/mL were defined as a positive response. The neutralization efficacies of the vaccine against the wild-type, Omicron, and Delta variants were compared in the first test. The 59 LT recipients were of a median age of 61 years (range 25-82); 53.5% were male. Following administration of the third dose, the positive immune response decreased from 81.4% to 76.3% between the first and second tests, respectively, (p < 0.0001). The multivariate analysis identified CNI monotherapy (p = 0.02) and hemoglobin > 12 g/dL (p = 0.02) as independent predictors of a maintained positive immune response 133 days after the third dose. The geometric mean titers of Omicron neutralization were significantly lower than the wild-type and Delta virus (21, 137, 128, respectively; p < 0.0001). The immune response after the third BNT162b2mRNA vaccine dose decreased significantly in LT recipients. Further studies are required to evaluate the efficacy of the fourth vaccine dose and the durability of the immune response.

The emergence of recombinant PRRSV strains has been observed for more than a decade. These recombinant viruses are characterized by a genome that contains genetic material from at least two different parental strains. Due to the advanced sequencing techniques and a growing number of data bank entries, the role of PRRSV recombinants has become increasingly important since they are sometimes associated with clinical outbreaks. Chimeric viruses observed more recently are products of PRRSV wild-type and vaccine strains. Here, we report on three PRRSV-1 isolates from geographically distant farms with differing clinical manifestations. A sequencing and recombination analysis revealed that these strains are crossovers between different wild-type strains and the same modified live virus vaccine strain. Interestingly, the recombination breakpoint of all analyzed isolates appears at the beginning of open reading frame 5 (ORF5). RNA structure predictions indicate a conserved stem loop in close proximity to the recombination hotspot, which is a plausible cause of a polymerase template switch during RNA replication. Further research into the mechanisms of the stem loop is needed to help understand the PRRSV recombination process and the role of MLVs as parental strains.

The real-world benefits of direct-acting antiviral (DAA)-induced sustained virologic response (SVR) on the de novo occurrence and progression of esophageal varices (EV) remain unclear in patients with hepatitis C virus (HCV)-related liver cirrhosis (LC). This is a retrospective cohort study evaluating all patients with Child-Pugh class A HCV-related LC during 2013 to 2020 in the Chang Gung Medical System. A total of 215 patients fit the inclusion criteria and were enrolled. Of them, 132 (61.4%) patients achieved DAA induced-SVR and 83 (38.6%) did not receive anti-viral treatment. During a median follow-up of 18.4 (interquartile range, 10.1-30.9) months, the 2-year incidence of de novo EV occurrence was 8 (7.0%) in the SVR group and 7 (12.7%) in the treatment-naïve group. Compared to the treatment-naïve group, the SVR group was associated with a significantly lower incidence of EV occurrence (adjusted hazard ratio [aHR]: 0.47, p = 0.030) and a significantly lower incidence of EV progression (aHR: 0.55, p = 0.033). The risk of EV progression was strongly correlated with the presence of baseline EV (p < 0.001). To the best of our knowledge, this is the first study to demonstrate that DAA-induced SVR is associated with decreased risk of de novo EV occurrence and progression in the real world.

Bluetongue virus (BTV) is the etiologic agent of bluetongue (BT), a viral WOAH-listed disease affecting wild and domestic ruminants, primarily sheep. The outermost capsid protein VP2, encoded by S2, is the virion's most variable protein, and the ability of reference sera to neutralize an isolate has so far dictated the differentiation of 24 classical BTV serotypes. Since 2008, additional novel BTV serotypes, often referred to as "atypical" BTVs, have been documented and, currently, the full list includes 36 putative serotypes. In March 2015, a novel atypical BTV strain was detected in the blood of asymptomatic goats in Sardinia (Italy) and named BTV-X ITL2015. The strain re-emerged in the same region in 2021 (BTV-X ITL2021). In this study, we investigated the pathogenicity and kinetics of infection of BTV-X ITL2021 following subcutaneous and intravenous infection of small ruminants. We demonstrated that, in our experimental settings, BTV-X ITL2021 induced a long-lasting viraemia only when administered by the intravenous route in goats, though the animals remained healthy and, apparently, did not develop a neutralizing immune response. Sheep were shown to be refractory to the infection by either route. Our findings suggest a restricted host tropism of BTV-X and point out goats as reservoirs for this virus in the field.

Monkeypox virus (MPXV) is a member of the Orthopoxvirus genus and the Poxviridae family, which instigated a rising epidemic called monkeypox disease. Proteinases are majorly engaged in viral propagation by catalyzing the cleavage of precursor polyproteins. Therefore, proteinase is essential for monkeypox and a critical drug target. In this study, high-throughput virtual screening (HTVS) and molecular dynamics simulation were applied to detect the potential natural compounds against the proteinase of the monkeypox virus. Here, 32,552 natural products were screened, and the top five compounds were selected after implementing the HTVS and molecular docking protocols in series. Gallicynoic Acid F showed the minimum binding score of -10.56 kcal/mole in the extra precision scoring method, which reflected the highest binding with the protein. The top five compounds showed binding scores ≤-8.98 kcal/mole. These compound complexes were tested under 100 ns molecular dynamics simulation, and Vaccinol M showed the most stable and consistent RMSD trend in the range of 2 Å to 3 Å. Later, MM/GBSA binding free energy and principal component analysis were performed on the top five compounds to validate the stability of selected compound complexes. Moreover, the ligands Gallicynoic Acid F and H2-Erythro-Neopterin showed the lowest binding free energies of -61.42 kcal/mol and -61.09 kcal/mol, respectively. Compared to the native ligand TTP-6171 (ΔGBind = -53.86 kcal/mol), these two compounds showed preferable binding free energy, suggesting inhibitory application against MPXV proteinase. This study proposed natural molecules as a therapeutic solution to control monkeypox disease.

Infectious diseases, such as COVID-19 [...].

The emergence and spread of antiviral-resistant influenza viruses are of great concern. To minimize the public health risk, it is important to monitor antiviral susceptibilities of influenza viruses. Analyses of the antiviral susceptibilities of influenza A and B viruses have been conducted globally; however, those of influenza C and D viruses are limited. Here, we determined the susceptibilities of influenza C viruses representing all six lineages (C/Taylor, C/Yamagata, C/Sao Paulo, C/Aichi, C/Kanagawa, and C/Mississippi) and influenza D viruses representing four lineages (D/OK, D/660, D/Yama2016, and D/Yama2019) to RNA polymerase inhibitors (baloxavir and favipiravir) by using a focus reduction assay. All viruses tested were susceptible to both drugs. We then performed a genetic analysis to check for amino acid substitutions associated with baloxavir and favipiravir resistance and found that none of the viruses tested possessed these substitutions. Use of the focus reduction assay with the genotypic assay has proven valuable for monitoring the antiviral susceptibilities of influenza C and D viruses as well as influenza A and B viruses. Antiviral susceptibility monitoring of all influenza virus types should continue in order to assess the public health risks posed by these viruses.

The amaranthine scale of the COVID-19 pandemic and unpredictable disease severity is of grave concern. Serological diagnostic aids are an excellent choice for clinicians for rapid and easy prognosis of the disease. To this end, we studied the humoral immune response to SARS-CoV-2 infection to map immunogenic regions in the SARS-CoV-2 proteome at amino acid resolution using a high-density SARS-CoV-2 proteome peptide microarray. The microarray has 4932 overlapping peptides printed in duplicates spanning the entire SARS-CoV-2 proteome. We found 204 and 676 immunogenic peptides against IgA and IgG, corresponding to 137 and 412 IgA and IgG epitopes, respectively. Of these, 6 and 307 epitopes could discriminate between disease severity. The emergence of variants has added to the complexity of the disease. Using the mutation panel available, we could detect 5 and 10 immunogenic peptides against IgA and IgG with mutations belonging to SAR-CoV-2 variants. The study revealed severity-based epitopes that could be presented as potential prognostic serological markers. Further, the mutant epitope immunogenicity could indicate the putative use of these markers for diagnosing variants responsible for the infection.

The emergence of the Coronavirus Disease 2019 (COVID-19) pandemic caused by severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has led to over 6 million deaths. The 3C-like protease (3CLpro) enzyme of the SARS-CoV-2 virus is an attractive druggable target for exploring therapeutic drug candidates to combat COVID-19 due to its key function in viral replication. Marine natural products (MNPs) have attracted considerable attention as alternative sources of antiviral drug candidates. In looking for potential 3CLpro inhibitors, the MNP database (>14,000 molecules) was virtually screened against 3CLpro with the assistance of molecular docking computations. The performance of AutoDock and OEDocking software in anticipating the ligand-3CLpro binding mode was first validated according to the available experimental data. Based on the docking scores, the most potent MNPs were further subjected to molecular dynamics (MD) simulations, and the binding affinities of those molecules were computed using the MM-GBSA approach. According to MM-GBSA//200 ns MD simulations, chetomin (UMHMNP1403367) exhibited a higher binding affinity against 3CLpro than XF7, with ΔGbinding values of -55.5 and -43.7 kcal/mol, respectively. The steadiness and tightness of chetomin with 3CLpro were evaluated, revealing the high stabilization of chetomin (UMHMNP1403367) inside the binding pocket of 3CLpro throughout 200 ns MD simulations. The physicochemical and pharmacokinetic features of chetomin were also predicted, and the oral bioavailability of chetomin was demonstrated. Furthermore, the potentiality of chetomin analogues -namely, chetomin A-D- as 3CLpro inhibitors was investigated. These results warrant further in vivo and in vitro assays of chetomin (UMHMNP1403367) as a promising anti-COVID-19 drug candidate.

PURPOSE: The Omicron subvariant BA.1 of SARS-CoV-2 was first detected in November 2021 and quickly spread worldwide, displacing the Delta variant. In this work, a characterization of the spread of this variant in Mexico is presented. METHODS: The time to fixation of BA.1, the diversity of Delta sublineages, the population density, and the level of virus circulation during the inter-wave interval were determined to analyze differences in BA.1 spread. RESULTS: BA.1 began spreading during the first week of December 2021 and became dominant in the next three weeks, causing the fourth COVID-19 epidemiological surge in Mexico. Unlike previous variants, BA.1 did not exhibit a geographically distinct circulation pattern. However, a regional difference in the speed of the replacement of the Delta variant was observed. CONCLUSIONS: Viral diversity and the relative abundance of the virus in a particular area around the time of the introduction of a new lineage seem to have influenced the spread dynamics, in addition to population density. Nonetheless, if there is a significant difference in the fitness of the variants, or if the time allowed for the competition is sufficiently long, it seems the fitter virus will eventually become dominant, as observed in the eventual dominance of the BA.1.x variant in Mexico.

In human beings, there are five reported variants of concern of severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2). However, in contrast to human beings, descriptions of infections of animals with specific variants are still rare. The aim of this study is to systematically investigate SARS-CoV-2 infections in companion animals in close contact with SARS-CoV-2-positive owners ("COVID-19 households") with a focus on the Delta variant. Samples, obtained from companion animals and their owners were analyzed using a real-time reverse transcriptase-polymerase chain reaction (RT-qPCR) and next-generation sequencing (NGS). Animals were also tested for antibodies and neutralizing activity against SARS-CoV-2. Eleven cats and three dogs in nine COVID-19-positive households were RT-qPCR and/or serologically positive for the SARS-CoV-2 Delta variant. For seven animals, the genetic sequence could be determined. The animals were infected by one of the pangolin lineages B.1.617.2, AY.4, AY.43 and AY.129 and between zero and three single-nucleotide polymorphisms (SNPs) were detected between the viral genomes of animals and their owners, indicating within-household transmission between animal and owner and in multi-pet households also between the animals. NGS data identified SNPs that occur at a higher frequency in the viral sequences of companion animals than in viral sequences of humans, as well as SNPs, which were exclusively found in the animals investigated in the current study and not in their owners. In conclusion, our study is the first to describe the SARS-CoV-2 Delta variant transmission to animals in Switzerland and provides the first-ever description of Delta-variant pangolin lineages AY.129 and AY.4 in animals. Our results reinforce the need of a One Health approach in the monitoring of SARS-CoV-2 in animals.

The transmembrane protein Axl was proposed as an entry receptor for Zika virus (ZIKV) infection in vitro, but conflicting results from in vivo studies have made it difficult to establish Axl as a physiologically relevant ZIKV receptor. Both the functional redundancy of receptors and the experimental model used can lead to variable results. Therefore, it can be informative to explore alternative animal models to analyze ZIKV receptor candidates as an aid in discovering antivirals. This study used chicken embryos to examine the role of chicken Tyro3 (cTyro3), the equivalent of human Axl. Results show that endogenous cTyro3 mRNA expression overlaps with previously described hot spots of ZIKV infectivity in the brain and inner ear. We asked if ectopic expression or knockdown of cTyro3 influenced ZIKV infection in embryos. Tol2 vectors or replication-competent avian retroviruses were used in ovo to introduce full-length or truncated (presumed dominant-negative) cTyro3, respectively, into the neural tube on embryonic day two (E2). ZIKV was delivered to the brain 24 h later. cTyro3 manipulations did not alter ZIKV infection or cell death in the E5/E6 brain. Moreover, delivery of truncated cTyro3 variants to the E3 otocyst had no effect on inner ear formation on E6 or E10.

Migrant populations are more susceptible to viral hepatitis and HIV due to the epidemiology from their country of origin or their social vulnerability when they arrive in Europe. The aims of the study are to explore the incidence of HIV and HCV in low-wage agricultural migrant workers and their knowledge, attitude, and practice with regard to HIV and HCV, as well as their sexual behaviour and risk factors. As part of the mobile clinic services, we performed a screening campaign for HIV-HCV involving migrants living in three Apulian establishments. Results: Between January 2020 and April 2021, 309 migrants (n. 272, 88% male, mean age 28.5 years) were enrolled in the study. Most of the migrants interviewed (n = 297, 96%) reported a stopover in Libya during their trip to Italy. Only 0.9% (n. 3) of migrants reported having been tested for HCV, while 30.7% (n. 95) reported being tested for HIV. Furthermore, screening tests found four migrants (1.3%) to be HIV positive and nine (2.9%) to be HCV positive. The median knowledge score was 1 (IQR 0-3; maximum score: 6 points) for HCV and 3 (IQR 1-4; maximum score: 7 points) for HIV and low use of condoms was 5% (n. 16), while more than 95% show an attitude score of 5 (IQR 5-6; maximum score:6 points) on HIV-HCV education campaigns. In a multivariate analysis, being male (OR = 1.72; 95% CI 1.28-1.92), being single (OR = 1.63; 95% CI 1.20-2.03), being of low educational status (OR = 2.09; 95% CI 1.29-2.21), living in shantytowns for >12 months (OR = 1.95; 95% CI 1.25-2.55), and originating from the African continent (OR = 1.43; 95% CI 1.28-2.01) are significant predictors of poor knowledge on HCV. Our data show low knowledge, especially of HCV, confirming migrants as a population with a higher risk of infection. To develop education programmes, integrated care and screening among migrants could be an effective strategy, considering the high attitude toward these items shown in our study.

ADP-ribosylation (ADPRylation) is a versatile posttranslational modification in eukaryotic cells which is involved in the regulation of a wide range of key biological processes, including DNA repair, cell signalling, programmed cell death, growth and development and responses to biotic and abiotic stresses. Members of the poly(ADP-ribosyl) polymerase (PARP) family play a central role in the process of ADPRylation. Protein targets can be modified by adding either a single ADP-ribose moiety (mono(ADP-ribosyl)ation; MARylation), which is catalysed by mono(ADP-ribosyl) transferases (MARTs or PARP "monoenzymes"), or targets may be decorated with chains of multiple ADP-ribose moieties (PARylation), via the activities of PARP "polyenzymes". Studies have revealed crosstalk between PARylation (and to a lesser extent, MARylation) processes in plants and plant-virus interactions, suggesting that these tight links may represent a novel factor regulating plant antiviral immunity. From this perspective, we go through the literature linking PARylation-associated processes with other plant regulation pathways controlling virus resistance. Once unraveled, these links may serve as the basis of innovative strategies to improve crop resistance to viruses under challenging environmental conditions which could mitigate yield losses.

Heparan sulfate proteoglycans (HSPGs) are a major constituent of the extracellular matrix (ECM) and are found to be implicated in viral infections, where they play a role in both cell entry and release for many viruses. The enzyme heparanase-1 is the only known endo-beta-D-glucuronidase capable of degrading heparan sulphate (HS) chains of HSPGs and is thus important for regulating ECM homeostasis. Heparanase-1 expression is tightly regulated as the uncontrolled cleavage of HS may result in abnormal cell activation and significant tissue damage. The overexpression of heparanase-1 correlates with pathological scenarios and is observed in different human malignancies, such as lymphoma, breast, colon, lung, and hepatocellular carcinomas. Interestingly, heparanase-1 has also been documented to be involved in numerous viral infections, e.g., HSV-1, HPV, DENV. Moreover, very recent reports have demonstrated a role of heparanase-1 in HCV and SARS-CoV-2 infections. Due to the undenied pro-carcinogenic role of heparanase-1, multiple inhibitors have been developed, some reaching phase II and III in clinical studies. However, the use of heparanase inhibitors as antivirals has not yet been proposed. If it can be assumed that heparanase-1 is implicated in numerous viral life cycles, its inhibition by specific heparanase-acting compounds should result in a blockage of viral infection. This review addresses the perspectives of using heparanase inhibitors, not only for cancer treatment, but also as antivirals. Eventually, the development of a novel class antivirals targeting a cellular protein could help to alleviate the resistance problems seen with some current antiretroviral therapies.

Influenza represents a major and ongoing public health hazard. Current collaborative efforts are aimed toward creating a universal flu vaccine with the goals of both improving responses to vaccination and increasing the breadth of protection against multiple strains and clades from a single vaccine. As an intermediate step toward these goals, the current work is focused on evaluating the systemic host response to vaccination in both normal and high-risk populations, such as the obese and geriatric populations, which have been linked to poor responses to vaccination. We therefore employed a metabolomics approach using a time-course (n = 5 time points) of the response to human vaccination against influenza from the time before vaccination (pre) to 90 days following vaccination. We analyzed the urinary profiles of a cohort of subjects (n = 179) designed to evenly sample across age, sex, BMI, and other demographic factors, stratifying their responses to vaccination as "High", "Low", or "None" based on the seroconversion measured by hemagglutination inhibition assay (HAI) from plasma samples at day 28 post-vaccination. Overall, we putatively identified 15,903 distinct, named, small-molecule structures (4473 at 10% FDR) among the 895 samples analyzed, with the aim of identifying metabolite correlates of the vaccine response, as well as prognostic and diagnostic markers from the periods before and after vaccination, respectively. Notably, we found that the metabolic profiles could unbiasedly separate the high-risk High-responders from the high-risk None-responders (obese/geriatric) within 3 days post-vaccination. The purine metabolites Guanine and Hypoxanthine were negatively associated with high seroconversion (p = 0.0032, p < 0.0001, respectively), while Acetyl-Leucine and 5-Aminovaleric acid were positively associated. Further changes in Cystine, Glutamic acid, Kynurenine and other metabolites implicated early oxidative stress (3 days) after vaccination as a hallmark of the High-responders. Ongoing efforts are aimed toward validating these putative markers using a ferret model of influenza infection, as well as an independent cohort of human seasonal vaccination and human challenge studies with live virus.

African swine fever (ASF) is a hemorrhagic viral disease of domestic pigs and wild suids (all Sus scrofa) caused by the ASF virus (ASFV). The disease is spreading worldwide without control, threatening pig production due to the absence of licensed vaccine or commercially available treatments. A thorough understanding of the immunopathogenic mechanisms behind ASFV infection is required to better fight the disease. Cytokines are small, non-structural proteins, which play a crucial role in many aspects of the immune responses to viruses, including ASFV. Infection with virulent ASFV isolates often results in exacerbated immune responses, with increased levels of serum pro-inflammatory interleukins (IL-1α, IL-1β, IL-6), TNF and chemokines (CCL2, CCL5, CXCL10). Increased levels of IL-1, IL-6 and TNF are often detected in several tissues during acute ASFV infections and associated with lymphoid depletion, hemorrhages and oedemas. IL-1Ra is frequently released during ASFV infection to block further IL-1 activity, with its implication in ASFV immunopathology having been suggested. Increased levels of IFN-α and of the anti-inflammatory IL-10 seem to be negatively correlated with animal survival, whereas some correlation between virus-specific IFN-γ-producing cells and protection has been suggested in different studies where different vaccine candidates were tested, although future works should elucidate whether IFN-γ release by specific cell types is related to protection or disease development.

The newly identified porcine Kobuvirus (PKV) has raised concerns owing to its association with diarrheal symptom in pigs worldwide. The process involving the emergence and global spread of PKV remains largely unknown. Here, the origin, genetic diversity, and geographic distribution of PKV were determined based on the available PKV sequence information. PKV might be derived from the rabbit Kobuvirus and sheep were an important intermediate host. The most recent ancestor of PKV could be traced back to 1975. Two major clades are identified, PKVa and PKVb, and recombination events increase PKV genetic diversity. Cross-species transmission of PKV might be linked to interspecies conserved amino acids at 13-17 and 25-40 residue motifs of Kobuvirus VP1 proteins. Phylogeographic analysis showed that Spain was the most likely location of PKV origin, which then spread to pig-rearing countries in Asia, Africa, and Europe. Within China, the Hubei province was identified as a primary hub of PKV, transmitting to the east, southwest, and northeast regions of the country. Taken together, our findings have important implications for understanding the evolutionary origin, genetic recombination, and geographic distribution of PKV thereby facilitating the design of preventive and containment measures to combat PKV infection.

The One Health framework recognizes that human, animal, and environmental health are linked and highly interdependent. Fecal contamination of water, soil, foodstuff, and air may impact many aspects of One Health, and culture, PCR-based, and sequencing methods are utilized in the detection of fecal contamination to determine source, load, and risk to inform targeted mitigation strategies. Viruses, particularly, have been considered as fecal contamination indicators given the narrow host range many exhibit and their association with other biological contaminants. Culture- and molecular-based methods are considered the gold-standards for virus detection and for determining specific sources of fecal contamination via viral indicators. However, viral metagenomics is also being considered as a tool for tracking sources of fecal contamination. In the present review, studies tracking potential sources of fecal contamination in freshwaters, marine waters, foodstuff, soil, and air using viral metagenomics are discussed to highlight the potential of viral metagenomics for optimizing fecal source tracking. Limitations of the use of viral metagenomics to track fecal contamination sources, including sample processing, nucleic acid recovery, sequencing depth, and bioinformatics are also discussed. Finally, the present review discusses the potential of viral metagenomics as part of the toolbox of methods in a One Health approach.

Dengue and obesity are currently highly prevalent conditions worldwide and the association between these two conditions may result in greater risk for DENV infection and disease severity. In this study the association between obesity and recent, inapparent dengue was investigated. Serum DENV IgM and NS1 were evaluated in 49 adult volunteers (15 lean and 34 individuals with obesity, according to body mass index), between September 2017 and June 2018. Adiposity, endocrine, metabolic, and immune data of the participants were also obtained. None of the study participants tested positive for the DENV NS1 antigen. DENV IgM was detected in 33.3% of the lean individuals, and in 44.1% of those with obesity; the presence of DENV IgM was not associated with body mass index (OR = 1.32, 95% CI = 0.59-2.98, p = 0.48). However, body fat index was higher in obese individuals who had recent inapparent dengue (14.7 ± 3.1 versus 12.7 ± 2.1 kg/m2, p = 0.04), as was the expression of CD11b by classical (CD14++CD16-) monocytes (1103.0 ± 311.3 versus 720.3 ± 281.1 mean fluoresce intensity). Our findings suggest an association between adiposity and recent inapparent dengue and the involvement of classical monocytes in this association.

Mosquitoes are vectors of various pathogens that cause diseases in humans and animals. To prevent the outbreak of mosquito-borne diseases, it is essential to control vector populations, as treatment or vaccination for mosquito-borne diseases are often unavailable. Insect-specific viruses (ISVs) have previously been described as being potentially helpful against arboviral disease outbreaks. In this study, we present the first in vivo characterization of the ISV Culex Y virus (CYV). CYV was first isolated from free-living Culex pipiens mosquitoes in 2010; then, it was found in several mosquito cell lines in a further study in 2018. For mammalian cells, we were able to confirm that CYV does not replicate as it was previously described. Additionally, we found that CYV does not replicate in honey bees or locusts. However, we detected replication in the Culex pipiens biotype molestus, Aedes albopictus, and Drosophila melanogaster, thus indicating dipteran specificity. We detected significantly higher mortality in Culex pipiens biotype molestus males and Drosophila melanogaster, but not in Aedes albopictus and female Culex pipiens biotype molestus. CYV could not be transmitted transovarially to offspring, but we detected venereal transmission as well as CYV in mosquitos' saliva, indicating that an oral route of infection would also be possible. CYV's dipteran specificity, transmission routes, and killing effect with respect to Culex males may be used as powerful tools with which to destabilize arbovirus vector populations in the future.

Low-pathogenicity avian influenza viruses (AIV) of the H9N2 subtype can infect and cause disease in chickens. Little is known about the efficacy of immune-based strategies for reducing the transmission of these viruses. The present study investigated the efficacy of Toll-like receptor (TLR) ligands (CpG ODN 2007 and poly(I:C)) to reduce H9N2 AIV transmission from TLR-treated seeder (trial 1) or inoculated chickens (trial 2) to naive chickens. The results from trial 1 revealed that a low dose of CpG ODN 2007 led to the highest reduction in oral shedding, and a high dose of poly(I:C) was effective at reducing oral and cloacal shedding. Regarding transmission, the recipient chickens exposed to CpG ODN 2007 low-dose-treated seeder chickens showed a maximum reduction in shedding with the lowest number of AIV+ chickens. The results from trial 2 revealed a maximum reduction in oral and cloacal shedding in the poly(I:C) high-dose-treated chickens (recipients), followed by the low-dose CpG ODN 2007 group. In these two groups, the expression of type I interferons (IFNs), protein kinase R (PKR), interferon-induced transmembrane protein 3 (IFITM3), viperin, and (interleukin) IL-1β, IL-8, and 1L-18 was upregulated in the spleen, cecal tonsils and lungs. Hence, TLR ligands can reduce AIV transmission in chickens.

Genotype IV Japanese encephalitis (JE) virus (GIV JEV) is the least common and most neglected genotype in JEV. We evaluated the growth and pathogenic potential of the GIV strain 19CxBa-83-Cv, which was isolated from a mosquito pool in Bali, Indonesia, in 2019, and serological analyses were also conducted. The growth ability of 19CxBa-83-Cv in Vero cells was intermediate between that of the genotype I (GI) strain Mie/41/2002 and the genotype V (GV) strain Muar, whereas 19CxBa-83-Cv and Mie/41/2002 grew faster than Muar in mouse neuroblastoma cells. The neuroinvasiveness of 19CxBa-83-Cv in mice was higher than that of Mie/41/2002 but lower than that of Muar; however, there were no significant differences in neurovirulence in mice among the three strains. The neutralizing titers of sera from 19CxBa-83-Cv- and Mie/41/2002-inoculated mice against 19CxBa-83-Cv and Mie/41/2002 were similar, whereas the titers against Muar were lower than those of the other two viruses. The neutralizing titers of JE vaccine-inoculated mouse pool serum against 19CxBa-83-Cv and Muar were significantly lower than those against Mie/41/2002. The neutralizing titers against the three viruses were similar in three out of the five serum samples from GI-infected JE patients, although the titers against Mie/41/2002 were higher than those against 19CxBa-83-Cv and Muar in the remaining two sera samples. In summary, we identified the basic characteristics of 19CxBa-83-Cv, but further studies are needed to better understand GIV JEV.

Different humanized mouse models have been developed to study human diseases such as autoimmune illnesses, cancer and viral infections. These models are based on the use of immunodeficient mouse strains that are transplanted with human tissues or human immune cells. Among the latter, mice transplanted with hematopoietic stem cells have been widely used to study human infectious diseases. However, mouse models built upon the transplantation of donor-specific mature immune cells are still under development, especially in the field of viral infections. These models can retain the unique immune memory of the donor, making them suitable for the study of correlates of protection upon natural infection or vaccination. Here, we will review some of these models and how they have been applied to virology research. Moreover, the future applications and the potential of these models to design therapies against human viral infections are discussed.

Herpes simplex virus (HSV) has persisted within human populations due to its ability to establish both lytic and latent infection. Given this, human hosts have evolved numerous immune responses to protect against HSV infection. Critical in this defense against HSV, the host protein stimulator of interferon genes (STING) functions as a mediator of the antiviral response by inducing interferon (IFN) as well as IFN-stimulated genes. Emerging evidence suggests that during HSV infection, dsDNA derived from either the virus or the host itself ultimately activates STING signaling. While a complex regulatory circuit is in operation, HSV has evolved several mechanisms to neutralize the STING-mediated antiviral response. Within this review, we highlight recent progress involving HSV interactions with the STING pathway, with a focus on how STING influences HSV replication and pathogenesis.

Herpes labialis remains exceedingly prevalent and is one of the most common human viral infections throughout the world. Recurrent herpes labialis evolves from the initial viral infection by herpes simplex virus type 1 (HSV-1) which subsequently presents with or without symptoms. Reactivation of this virus is triggered by psychosocial factors such as stress, febrile environment, ultraviolet light susceptibility, or specific dietary inadequacy. This virus infection is also characterized by uninterrupted transitions between chronic-latent and acute-recurrent phases, allowing the virus to opportunistically avoid immunity and warrant the transmission to other vulnerable hosts simultaneously. This review comprehensively evaluates the current evidence on epidemiology, pathogenesis, transmission modes, clinical manifestations, and current management options of herpes labialis infections.

Infections with the opportunistic Gram-negative bacterium Acinetobacter baumannii pose a serious threat today, which is aggravated by the growing problem of multi-drug resistance among bacteria, caused by the overuse of antibiotics. Treatment of infections caused by antibiotic-resistant A. baumannii strains with the use of phage therapy is not only a promising alternative, but sometimes the only option. Therefore, phages specific for clinical multi-drug resistant A. baumannii were searched for in environmental, municipal, and hospital wastewater samples collected from different locations in Poland. The conducted research allowed us to determine the biological properties and morphology of the tested phages. As a result of our research, 12 phages specific for A. baumannii, 11 of which turned out to be temperate and only one lytic, were isolated. Their lytic spectra ranged from 11 to 75%. The plaques formed by most phages were small and transparent, while one of them formed relatively large plaques with a clearly marked 'halo' effect. Based on Transmission Electron Microscopy (TEM), most of our phages have been classified as siphoviruses (only one phage was classified as a podovirus). All phages have icosahedral capsid symmetry, and 11 of them have a long tail. Optimal multiplicity of infections (MOIs) and the adsorption rate were also determined. MOI values varied depending on the phage-from 0.001 to 10. Based on similarities to known bacteriophages, our A. baumannii-specific phages have been proposed to belong to the Beijerinckvirinae and Junivirinae subfamilies. This study provides an additional tool in the fight against this important pathogen and may boost the interest in phage therapy as an alternative and supplement to the current antibiotics.

Update of medRxiv. 2022 May 26;:

The immunological mechanisms involved in the development of congenital Zika syndrome (CZS) have yet to be fully clarified. This study aims to assess the immuno-inflammatory profile of mothers and their children who have been diagnosed with CZS. Blood samples, which were confirmed clinically using the plaque reduction neutralization test (PRNT), were collected from children with CZS and their mothers (CZS+ group). Samples were also collected from children who did not develop CZS and had a negative PRNT result and from their mothers (CZS- group). The data demonstrated a correlation between the leukocyte profile of CZS+ children and their mothers, more evident in monocytes. Monocytes from mothers of CZS+ children showed low expression of HLA and elevated hydrogen peroxide production. CZS+ children presented standard HLA expression and a higher hydrogen peroxide concentration than CZS- children. Monocyte superoxide dismutase activity remained functional. Moreover, when assessing the monocyte polarization, it was observed that there was no difference in nitrite concentrations; however, there was a decrease in arginase activity in CZS+ children. These data suggest that ZIKV infection induces a maternal immuno-inflammatory background related to the child's inflammatory response after birth, possibly affecting the development and progression of congenital Zika syndrome.

In 2022, Greece was the second most seriously affected European country in terms of the West Nile virus (WNV), after Italy. Specifically, Central Macedonia was the region with the most reported human cases (81.5%). In the present study, 30,816 female Culex pipiens sensu lato mosquitoes were collected from May to September 2022 in the seven regional units of Central Macedonia; they were then grouped into 690 pools and tested for WNV, while next-generation sequencing was applied to the samples, which showed a cycle threshold of Ct < 30 in a real-time RT-PCR test. WNV was detected in 5.9% of pools, with significant differences in the detection rate among regional units and months. It is of interest that in the Thessaloniki regional unit, where most of the human cases were observed, the virus circulation started earlier, peaked earlier, and lasted longer than in the other regional units. All sequences clustered into the Central European subclade of WNV lineage 2, and the virus strain differed from the initial Greek strain of 2010 by 0.52% and 0.27% at the nucleotide and amino acid levels, respectively. Signature substitutions were present, such as S73P and T157A in the prM and E structural proteins, respectively. The screening of mosquitoes provides useful information for virus circulation in a region with a potential for early warning, while the availability of whole-genome sequences is essential for further studies, including virus evolution.

During the coronavirus disease 2019 (COVID-19) pandemic, the influenza virus had a very low prevalence, and in many areas, outbreaks were almost non-existent. In this study, the associations between infection control measures taken for COVID-19 and the global disappearance of the influenza virus were investigated. The detection rate of influenza from baseline was investigated during four seasons (12 weeks from epidemiological week 49 in 2020 and 2021 and 12 weeks from epidemiological week 23 in 2020 and 2021) in each country participating in the surveillance system of the World Health Organization. Three measures of infection control: mask use ratio, social distancing index (an index of human mobility and physical distance obligations), and an index of stringency of measures taken by authorities were studied. In mid-2020, most countries analyzed had high levels of infection control measures, and in most countries, influenza was drastically reduced compared to previous years. Multiple regression analyses compared the study data with data from other seasons. There was an association between high mask use with low influenza detection in all three remaining seasons, an association between a low social distancing index (low mobility and more social contact obligations) with a low influenza detection rate in two seasons, and a marginal significant association of high stringency index with a low influenza detection rate(in 2020-end-seasons). These results support the notion that seasonal influenza is controllable through effective preventive measures, especially those of mask use and human social contact, and these measures should be recommended during future waves of novel influenza virus infection.

COVID-19 severely affected nursing home residents from March 2020 onwards in Belgium. This study aimed to model the impact of vaccination and facility characteristics on cluster occurrence, duration and severity in this setting. Possible clusters were identified between June 2020 and January 2022, based on the Belgian COVID-19 surveillance in nursing homes. Median attack rates (AR) among residents and staff, case hospitalization rates (CHR) and case fatality rates (CFR) were calculated. A negative binomial model was used to identify the association between nursing home characteristics and the number of cases, hospital admissions and deaths and the duration of the cluster. A total of 2239 clusters were detected in more than 80% of nursing homes. Most of these (62%) occurred before the start of COVID-19 vaccination (end of December 2020). After vaccination, the number of clusters, the AR among residents and staff, the CHR and the CFR dropped. Previous cluster(s) and vaccination decreased the number of cases, hospital admissions and deaths among residents. Previous cluster experience and having started vaccination were protective factors. We recommend continued implementation of targeted interventions such as vaccination, large-scale screening and immediate implementation of additional infection prevention and control measures.

The cessation of measles virus (MeV) vaccination in more than 40 countries as a consequence of the COVID-19 pandemic is expected to significantly increase deaths due to measles. MeV can infect the central nervous system (CNS) and lead to lethal encephalitis. Substantial part of virus sequences recovered from patients' brain were mutated in the matrix and/or the fusion protein (F). Mutations of the heptad repeat domain located in the C terminal (HRC) part of the F protein were often observed and were associated to hyperfusogenicity. These mutations promote brain invasion as a hallmark of neuroadaptation. Wild-type F allows entry into the brain, followed by limited spreading compared with the massive invasion observed for hyperfusogenic MeV. Taking advantage of our ex vivo models of hamster organotypic brain cultures, we investigated how the hyperfusogenic mutations in the F HRC domain modulate virus distribution in CNS cells. In this study, we also identified the dependence of neural cells susceptibility on both their activation state and destabilization of the virus F protein. Type I interferon (IFN-I) impaired mainly astrocytes and microglial cells permissiveness contrarily to neurons, opening a new way of consideration on the development of treatments against viral encephalitis.

Genital herpes (GH) has become one of the most common sexually transmitted diseases worldwide, and it is spreading rapidly in developing countries. Approximately 90% of GH cases are caused by HSV-2. Therapeutic HSV-2 vaccines are intended for people already infected with HSV-2 with the goal of reducing clinical recurrences and recurrent virus shedding. In our previous work, we evaluated recombinant adenovirus-based vaccines, including rAd-gD2ΔUL25, rAd-ΔUL25, and rAd-gD2, for their potency as prophylactic vaccines. In this study, we evaluated these three vaccines as therapeutic vaccines against acute and recurrent diseases in intravaginal challenged guinea pigs. Compared with the control groups, the recombinant vaccine rAd-gD2ΔUL25 induced a higher titer of the binding antibody, and rAd-gD2 + rAd-ΔUL25 induced a higher titer of the neutralizing antibody. Both rAd-gD2ΔUL25 and rAd-gD2 + rAd-ΔUL25 vaccines significantly enhanced the survival rate by 50% compared to rAd-gD2 and reduced viral replication in the genital tract and recurrent genital skin disease. Our findings provide a new perspective for HSV-2 therapeutic vaccine research and provide a new technique to curtail the increasing spread of HSV-2.

Human multipotent mesenchymal stromal cells (hMSCs) are of significant therapeutic interest due to their ability to deliver oncolytic adenoviruses to tumors. This approach is also investigated for targeting head and neck squamous cell carcinomas (HNSCCs). HAdV-5-HexPos3, a recently reported capsid-modified vector based on human adenovirus type 5 (HAdV-5), showed strongly improved infection of both hMSCs and the HNSCC cell line UM-SCC-11B. Given that, we generated life cycle-unmodified and -modified replication-competent HAdV-5-HexPos3 vector variants and analyzed their replication within bone marrow- and adipose tissue-derived hMSCs. Efficient replication was detected for both life cycle-unmodified and -modified vectors. Moreover, we analyzed the migration of vector-carrying hMSCs toward different HNSCCs. Although migration of hMSCs to HNSCC cell lines was confirmed in vitro, no homing of hMSCs to HNSCC xenografts was observed in vivo in mice and in ovo in a chorioallantoic membrane model. Taken together, our data suggest that HAdV-5-HexPos3 is a potent candidate for hMSC-based oncolytic therapy of HNSCCs. However, it also emphasizes the importance of generating optimized in vivo models for the evaluation of hMSC as carrier cells.

Hemocytes are essential components of the immune system against invading pathogens in shrimp. Many uncharacterized transcripts exist in hemocytes but the knowledge of them is very limited. In the present study, we identified a novel small protein from the uncharacterized transcripts in hemocytes of Litopenaeus vannamei. This transcript was specifically expressed in hemocytes and encoded a novel secretory protein, which was designated as hemocyte-specific small protein (LvHSSP). The expression level of LvHSSP was significantly up-regulated in the hemocytes of shrimp infected with white spot syndrome virus (WSSV). After knockdown of LvHSSP by RNA interference, the WSSV copy number in shrimp decreased significantly. Conversely, WSSV copy number increased in shrimp when they were infected by WSSV after incubation with recombinant LvHSSP protein. These results suggested that LvHSSP might promote viral infection in shrimp. Immunocytochemical assay showed that the recombinant LvHSSP protein was located on the membrane of hemocytes. Co-IP results showed that LvHSSP could interact with VP26, the main envelope protein of WSSV, suggesting that LvHSSP might mediate WSSV adhesion and entry into host cells by binding to viral envelope protein. Meanwhile, the total hemocyte counts were significantly decreased after LvHSSP knockdown while increased after supplementing with recombinant LvHSSP protein, supporting the idea of hemocytes as the carrier for systemic dissemination of WSSV. This study reported a novel small protein in hemocytes, which modulated the viral infection in shrimp. Our results will enrich the knowledge of invertebrate innate immunity and provide a new field in the study of hemocyte function.

BACKGROUND: Conflicting data regarding the incidence of hepatocellular carcinoma (HCC) after cure of HCV infection with direct-acting antivirals (DAAs) remains. We investigated the incidence and risk factors to HCC after treatment with DAAs followed up for five years. METHODS: A total of 1075 HCV patients ≥ 18 years were treated with DAAs from 2015 to 2019 and followed until 2022. Ultrasonography was performed before DAAs and each 6 months thereafter. RESULTS: Of the total, 51/1075 (4.7%) developed HCC in the median of 40 (IQR 25-58) months: 26/51 (51%) male, median age 60 (IQR 54-66) years, alpha-fetoprotein (AFP) 12.2 (IQR 6.1-18.8) ng/mL, 47/51 (92.1%) cirrhotic 78.7%, 8/51 (15.7%) without sustained virological response (SVR). Seventeen percent had non-characterized nodules before DAAs. Cumulative HCC incidence was 5.9% in 5 years. Overall incidence was 1.46/100 patient-years (PY) (95% CI = 1.09-1.91), being 2.31/100 PY (95% CI = 1.70-3.06), 0.45/100 PY (95% CI = 0.09-1.32) and 0.20/100 PY (95% CI 0.01-1.01) in METAVIR F4, F3 and F2, respectively, and the main risks to HCC were non-characterized nodule, cirrhosis, high AFP values and non-SVR. CONCLUSION: HCV cure reduced risk for HCC, but it still occurred particularly in cirrhotic patients. Some risk factors can be identified to predict early HCC diagnosis.

COVID-19 cases caused by new variants of highly mutable SARS-CoV-2 continue to be identified worldwide. Effective control of the spread of new variants can be achieved through targeting of conserved viral epitopes. In this regard, the SARS-CoV-2 nucleocapsid (N) protein, which is much more conserved than the evolutionarily influenced spike protein (S), is a suitable antigen. The recombinant N protein can be considered not only as a screening antigen but also as a basis for the development of next-generation COVID-19 vaccines, but little is known about induction of antibodies against the N protein via different SARS-CoV-2 variants. In addition, it is important to understand how antibodies produced against the antigen of one variant can react with the N proteins of other variants. Here, we used recombinant N proteins from five SARS-CoV-2 strains to investigate their immunogenicity and antigenicity in a mouse model and to obtain and characterize a panel of hybridoma-derived monoclonal anti-N antibodies. We also analyzed the variable epitopes of the N protein that are potentially involved in differential recognition of antiviral antibodies. These results will further deepen our knowledge of the cross-reactivity of the humoral immune response in COVID-19.

Vaccination has been confirmed to be the safest and, sometimes, the only tool of defense against threats from infectious diseases. The successful history of vaccination is evident in the control of serious viral infections, such as smallpox and polio. Viruses that infect human livers are known as hepatitis viruses and are classified into five major types from A to E, alphabetically. Although infection with hepatitis A virus (HAV) is known to be self-resolving after rest and symptomatic treatment, there were 7134 deaths from HAV worldwide in 2016. In 2019, hepatitis B virus (HBV) and hepatitis C virus (HCV) resulted in an estimated 820,000 and 290,000 deaths, respectively. Hepatitis delta virus (HDV) is a satellite virus that depends on HBV for producing its infectious particles in order to spread. The combination of HDV and HBV infection is considered the most severe form of chronic viral hepatitis. Hepatitis E virus (HEV) is another orally transmitted virus, common in low- and middle-income countries. In 2015, it caused 44,000 deaths worldwide. Safe and effective vaccines are already available to prevent hepatitis A and B. Here, we review the recent advances in protective vaccines against the five major hepatitis viruses.

A genetic analysis of circulating measles virus (MeV) provides strong evidence of an interruption in endemic measles and supports the elimination status of this disease. This study investigated 219 MeVs isolated between 2015 and 2020. Based on the 450 nucleotide sequences of the nucleoprotein gene (N-450), three genotypes of the H1, D8 and B3 with 8, 18 and 6 different N-450 sequences, respectively, were identified. The H1 genotype virus has not circulated in Taiwan since 2017, and the D8 and B3 genotype MeVs became dominant between 2018 and 2019. Different D8 genotype variants were imported from neighboring countries, and the majority of MeV variants were detected only for a short period. However, MVs/Gir Somnath.IND/42.16[D8], a named strain designated by the World Health Organization (WHO), was detected over 2 years. To explore whether the endemic transmission of measles has been underestimated, another sequence window of the hypervariable, noncoding regions between the matrix (M) and fusion (F) genes (MF-NCR) was introduced to clarify the transmission chain. From the chronological sequence analysis of MeVs with N-450 and MF-NCR sequence windows, no endemic MeV variants lasted over 4 weeks, providing strong evidence to support the contention that Taiwan has reached the status for measles elimination.

Giovanni Paolo Martelli passed away on 8 January 2020 [...].

The coronavirus disease (COVID-19) is a pandemic that started in the City of Wuhan, Hubei Province, China, caused by the spread of coronavirus (SARS-CoV-2). Drug discovery teams around the globe are in a race to develop a medicine for its management. It takes time for a novel molecule to enter the market, and the ideal way is to exploit the already approved drugs and repurpose them therapeutically. We have attempted to screen selected molecules with an affinity towards multiple protein targets in COVID-19 using the Schrödinger suit for in silico predictions. The proteins selected were angiotensin-converting enzyme-2 (ACE2), main protease (MPro), and spike protein. The molecular docking, prime MM-GBSA, induced-fit docking (IFD), and molecular dynamics (MD) simulations were used to identify the most suitable molecule that forms a stable interaction with the selected viral proteins. The ligand-binding stability for the proteins PDB-IDs 1ZV8 (spike protein), 5R82 (Mpro), and 6M1D (ACE2), was in the order of nintedanib > quercetin, nintedanib > darunavir, nintedanib > baricitinib, respectively. The MM-GBSA, IFD, and MD simulation studies imply that the drug nintedanib has the highest binding stability among the shortlisted. Nintedanib, primarily used for idiopathic pulmonary fibrosis, can be considered for repurposing for us against COVID-19.

SARS-CoV-2 prevention and control measures did not only impact SARS-CoV-2 circulation, but also the timing and prevalence of other seasonal respiratory viruses. Especially in children, information on exposure and infections to seasonal coronaviruses as well as SARS-CoV-2 in the first year of the pandemic is largely lacking. Therefore, we set up a one-year serological survey in a large tertiary hospital in the Netherlands. We show that seasonal coronavirus seroprevalence significantly decreased in 2021 in children less than one year, most likely due to COVID-19 control measures. The SARS-CoV-2 seroprevalence in children and adolescents increased from 0.4% to 11.3%, the highest in adolescents. This implies higher exposure rates in adolescents as compared to the general population (>18 years old). It is clear that there have been significant changes in the circulation and subsequent immunity against most respiratory pathogens as a result of the mitigation measures. The implications on shorter as well as longer term are still largely unknown, but the impact of the SARS-CoV-2 pandemic and subsequent control measures will continue to affect the dynamics of other pathogens.

Parvovirus B19 (B19V) is a ssDNA human virus, responsible for an ample range of clinical manifestations. Sequencing of B19V DNA from clinical samples is frequently reported in the literature to assign genotype (genotypes 1-3) and for finer molecular epidemiological tracing. The increasing availability of Next Generation Sequencing (NGS) with its depth of coverage potentially yields information on intrinsic sequence heterogeneity; however, integration of this information in analysis of sequence variation is not routinely obtained. The present work investigated genomic sequence heterogeneity within and between B19V isolates by application of NGS techniques, and by the development of a novel dedicated bioinformatic tool and analysis pipeline, yielding information on two newly defined parameters. The first, α-diversity, is a measure of the amount and distribution of position-specific, normalised Shannon Entropy, as a measure of intra-sample sequence heterogeneity. The second, σ-diversity, is a measure of the amount of inter-sample sequence heterogeneity, also incorporating information on α-diversity. Based on these indexes, further cluster analysis can be performed. A set of 24 high-titre viraemic samples was investigated. Of these, 23 samples were genotype 1 and one sample was genotype 2. Genotype 1 isolates showed low α-diversity values, with only a few samples showing distinct position-specific polymorphisms; a few genetically related clusters emerged when analysing inter-sample distances, correlated to the year of isolation; the single genotype 2 isolate showed the highest α-diversity, even if not presenting polymorphisms, and was an evident outlier when analysing inter-sample distance. In conclusion, NGS analysis and the bioinformatic tool and pipeline developed and used in the present work can be considered effective tools for investigating sequence diversity, an observable parameter that can be incorporated into the quasispecies theory framework to yield a better insight into viral evolution dynamics.

In the human gut, temperate bacteriophages interact with bacteria through predation and horizontal gene transfer. Relying on taxonomic data, metagenomic studies have associated shifts in phage abundance with a number of human diseases. The temperate bacteriophage VEsP-1 with siphovirus morphology was isolated from a sample of river water using Enterococcus faecalis as a host. Starting from the whole genome sequence of VEsP-1, we retrieved related phage genomes in blastp searches of the tail protein and large terminase sequences, and blastn searches of the whole genome sequences, with matches compiled from several different databases, and visualized a part of viral dark matter sequence space. The genome network and phylogenomic analyses resulted in the proposal of a novel genus "Vespunovirus", consisting of temperate, mainly metagenomic phages infecting Enterococcus spp.

The β-Coronavirus mouse hepatitis virus (MHV-A59)-RSA59 has a patent stretch of fusion peptide (FP) containing two consecutive central prolines (PP) in the S2 domain of the Spike protein. Our previous studies compared the PP-containing fusogenic-demyelinating strain RSA59(PP) to its one proline-deleted mutant strain RSA59(P) and one proline-containing non-fusogenic non-demyelinating parental strain RSMHV2(P) to its one proline inserted mutant strain RSMHV2(PP). These studies highlighted the crucial role of PP in fusogenicity, hepato-neuropathogenesis, and demyelination. Computational studies combined with biophysical data indicate that PP at the center of the FP provides local rigidity while imparting global fluctuation to the Spike protein that enhances the fusogenic properties of RSA59(PP) and RSMHV2(PP). To elaborate on the understanding of the role of PP in the FP of MHV, the differential neuroglial tropism of the PP and P mutant strains was investigated. Comparative studies demonstrated that PP significantly enhances the viral tropism for neurons, microglia, and oligodendrocytes. PP, however, is not essential for viral tropism for either astroglial or oligodendroglial precursors or the infection of meningeal fibroblasts in the blood-brain and blood-CSF barriers. PP in the fusion domain is critical for promoting gliopathy, making it a potential region for designing antivirals for neuro-COVID therapy.

Initial exposure to influenza virus(es) during early childhood produces protective antibodies that may be recalled following future exposure to subsequent viral infections or vaccinations. Most influenza vaccine research studies use immunologically naïve animal models to assess vaccine effectiveness. However, most people have an extensive influenza immune history, with memory cells produced by viruses or vaccines representing multiple influenza viruses. In this study, we explored the effect influenza seasonal virus-induced immunity has on pre-pandemic influenza virus vaccination. The mice that were pre-immune to historical H1N1 and H3N2 seasonal influenza viruses were vaccinated with adjuvanted pre-pandemic (H2, H5, and H7) HA-based computationally optimized broadly reactive antigen (COBRA) vaccines, and were fully protected from lethal challenge, whereas the mock-vaccinated mice, with or without pre-immunity, were not protected from morbidity or mortality. Detectable antibody titers were present in the pre-immune mice vaccinated with a single dose of vaccine, but not in the immunologically naïve mice. The mice vaccinated twice with the trivalent COBRA HA vaccine had similar antibody titers regardless of their pre-immune status. Overall, seasonal pre-immunity did not interfere with the immune responses elicited by pre-pandemic COBRA HA vaccines or the protection against pre-pandemic viruses.

The adenovirus vector platform remains one of the most efficient toolboxes for generation of transfer vehicles used in gene therapy and virotherapy to treat tumors, as well as vaccines to protect from infectious diseases. The adenovirus genome and capsids can be modified using highly efficient techniques, and vectors can be produced at high titers, which facilitates their rapid adaptation to current needs and disease applications. Over recent years, the adenovirus vector platform has been in the center of attention for vaccine development against the ongoing coronavirus SARS-CoV-2/COVID-19 pandemic. The worldwide deployment of these vaccines has greatly deepened the knowledge on virus-host interactions and highlighted the need to further improve the effectiveness and safety not only of adenovirus-based vaccines but also of gene therapy and oncolytic virotherapy vectors. Based on the current evidence, we discuss here how adenoviral vectors can be further improved by intelligent molecular design. This review covers the full spectrum of state-of-the-art strategies to avoid vector-induced side effects ranging from the vectorization of non-canonical adenovirus types to novel genome engineering techniques.

Porcine parvovirus 1 (PPV1) is recognized as a major cause of reproductive failure in pigs, leading to several clinical outcomes globally known as SMEDI. Despite being known since the late 1960s its circulation is still of relevance to swine producers. Additionally, the emergence of variants such as the virulent 27a strain, for which lower protection induced by vaccines has been demonstrated, is of increasing concern. Even though constant monitoring of PPV1 using molecular epidemiological approaches is of pivotal importance, viral sequence data are scarce especially in low-income countries. To fill this gap, a collection of 71 partial VP2 sequences originating from eight African countries (Burkina Faso, Côte d'Ivoire, Kenya, Mozambique, Namibia, Nigeria, Senegal, and Tanzania) during the period 2011-2021 were analyzed within the context of global PPV1 variability. The observed pattern largely reflected what has been observed in high-income regions, i.e., 27a-like strains were more frequently detected than less virulent NADL-8-like strains. A phylogeographic analysis supported this observation, highlighting that the African scenario has been largely shaped by multiple PPV1 importation events from other continents, especially Europe and Asia. The existence of such an international movement coupled with the circulation of potential vaccine-escape variants requires the careful evaluation of the control strategies to prevent new strain introduction and persistence.

The roles of proteins encoded by members of the genus Ampelovirus, family Closteroviridae are largely inferred by sequence homology or analogy to similarly located ORFs in related viruses. This study employed yeast two-hybrid and bimolecular fluorescence complementation assays to investigate interactions between proteins of grapevine leafroll-associated virus 3 (GLRaV-3). The p5 movement protein, HSP70 homolog, coat protein, and p20B of GLRaV-3 were all found to self-interact, however, the mechanism by which p5 interacts remains unknown due to the absence of a cysteine residue crucial for the dimerisation of the closterovirus homolog of this protein. Although HSP70h forms part of the virion head of closteroviruses, in GLRaV-3, it interacts with the coat protein that makes up the body of the virion. Silencing suppressor p20B has been shown to interact with HSP70h, as well as the major coat protein and the minor coat protein. The results of this study suggest that the virion assembly of a member of the genus Ampelovirus occurs in a similar but not identical manner to those of other genera in the family Closteroviridae. Identification of interactions of p20B with virus structural proteins provides an avenue for future research to explore the mechanisms behind the suppression of host silencing and suggests possible involvement in other aspects of the viral replication cycle.

Multiple lineages of SARS-CoV-2 have been identified featuring distinct sets of genetic changes that confer to the virus higher transmissibility and ability to evade existing immunity. The continuous evolution of SARS-CoV-2 may pose challenges for current treatment options and diagnostic tools. In this study, we have first evaluated the performance of the 14 WHO-recommended real-time reverse transcription (RT)-PCR assays currently in use for the detection of SARS-CoV-2 and found that only one assay has reduced performance against Omicron. We then developed a new duplex real-time RT-PCR assay based on the amplification of two ultra-conserved elements present within the SARS-CoV-2 genome. The new duplex assay successfully detects all of the tested SARS-CoV-2 variants of concern (including Omicron sub-lineages BA.4 and BA.5) from both clinical and wastewater samples with high sensitivity and specificity. The assay also functions as a one-step droplet digital RT-PCR assay. This new assay, in addition to clinical testing, could be adopted in surveillance programs for the routine monitoring of SARS-CoV-2's presence in a population in wastewater samples. Positive results with our assay in conjunction with negative results from an Omicron-specific assay may provide timely indication of the emergence of a novel SARS-CoV-2 variant in a certain community and thereby aid public health interventions.

Oncotoxic proteins such as the non-structural protein 1 (NS1), a constituent of the rodent parvovirus H1 (H1-PV), offer a novel approach for treatment of tumors that are refractory to other treatments. In the present study, mutated NS1 variants were designed and tested with respect to their oncotoxic potential in human hepatocellular carcinoma cell lines. We introduced single point mutations of previously described important residues of the wild-type NS1 protein and a deletion of 114 base pairs localized within the N-terminal domain of NS1. Cell-viability screening with HepG2 and Hep3B hepatocarcinoma cells transfected with the constructed NS1-mutants led to identification of the single-amino acid NS1-mutant NS1-T585E, which led to a 30% decrease in cell viability as compared to NS1 wildtype. Using proteomics analysis, we could identify new interaction partners and signaling pathways of NS1. We could thus identify new oncotoxic NS1 variants and gain insight into the modes of action of NS1, which is exclusively toxic to human cancer cells. Our in-vitro studies provide mechanistic explanations for the observed oncolytic effects. Expression of NS1 variants had no effect on cell viability in NS1 unresponsive control HepG2 cells or primary mouse hepatocytes. The availability of new NS1 variants in combination with a better understanding of their modes of action offers new possibilities for the design of innovative cancer treatment strategies.

BACKGROUND: Since most of the modern human population has no anti-smallpox immunity, it is extremely important to develop and implement effective drugs for the treatment of smallpox and other orthopoxvirus infections. The objective of this study is to determine the main characteristics of the chemical substance NIOCH-14 and its safety and bioavailability in the body of laboratory animals. METHODS: The safety of NIOCH-14 upon single- or multiple-dose intragastric administration was assessed according to its effect on the main hematological and pathomorphological parameters of laboratory mice and rats. In order to evaluate the pharmacokinetic parameters of NIOCH-14 administered orally, a concentration of ST-246, the active metabolite of NIOCH-14, in mouse blood and organs was determined by tandem mass spectrometry and liquid chromatography. RESULTS: The intragastric administration of NIOCH-14 at a dose of 5 g/kg body weight caused neither death nor signs of intoxication in mice. The intragastric administration of NIOCH-14 to mice and rats at doses of 50 and 150 µg/g body weight either as a single dose or once daily during 30 days did not cause animal death or critical changes in hematological parameters and the microstructure of internal organs. The tissue availability of NIOCH-14 administered orally to the mice at a dose of 50 µg/g body weight, which was calculated according to concentrations of its active metabolite ST-246 for the lungs, liver, kidney, brain, and spleen, was 100, 69.6, 63.3, 26.8 and 20.3%, respectively. The absolute bioavailability of the NIOCH-14 administered orally to mice at a dose of 50 µg/g body weight was 22.8%. CONCLUSION: Along with the previously determined efficacy against orthopoxviruses, including the smallpox virus, the substance NIOCH-14 was shown to be safe and bioavailable in laboratory animal experiments.

Microchimerism is the presence of cells in an individual that have originated from a genetically distinct individual. The most common form of microchimerism is fetomaternal microchimerism, i.e., cells from a fetus pass through the placenta and establish cell lineages within the mother. Microchimerism was also described after the transplantation of human organs in human recipients. Consequently, microchimerism may also be expected in xenotransplantation using pig cells or organs. Indeed, microchimerism was described in patients after xenotransplantations as well as in non-human primates after the transplantation of pig organs. Here, for the first time, a comprehensive review of microchimerism in xenotransplantation is given. Since pig cells contain porcine endogenous retroviruses (PERVs) in their genome, the detection of proviral DNA in transplant recipients may be misinterpreted as an infection of the recipient with PERV. To prevent this, methods discriminating between infection and microchimerism are described. This knowledge will be important for the interpretation of screening results in forthcoming human xenotransplantations.

The phage life cycle is a multi-stage process initiated by the recognition and attachment of the virus to its bacterial host. This adsorption step depends on the specific interaction between bacterial structures acting as receptors and viral proteins called Receptor Binding Proteins (RBP). The adsorption process is essential as it is the first determinant of phage host range and a sine qua non condition for the subsequent conduct of the life cycle. In phages belonging to the Caudoviricetes class, the capsid is attached to a tail, which is the central player in the adsorption as it comprises the RBP and accessory proteins facilitating phage binding and cell wall penetration prior to genome injection. The nature of the viral proteins involved in host adhesion not only depends on the phage morphology (i.e., myovirus, siphovirus, or podovirus) but also the targeted host. Here, we give an overview of the adsorption process and compile the available information on the type of receptors that can be recognized and the viral proteins taking part in the process, with the primary focus on phages infecting Gram-positive bacteria.

One of the most pressing and consequential problems in infectious disease research is to better understand the potential of viruses to cause a pandemic, or, in simple terms, determine which virus will cause the next pandemic. We here define pandemics as WHO-declared pandemics, or disease outbreaks commonly referred to as pandemics that predate the WHO pandemic framework. Despite extensive research in the field of infectious diseases in recent decades, all pandemics have found us unprepared, with enormous losses of human lives, tremendous costs for public health, and vast and potentially long-lasting economic losses. Here, we discuss viruses that may cause a pandemic in the future.

Neuraminidase (NA) is an important surface protein on influenza virions, playing an essential role in the viral life cycle and being a key target of the immune system. Despite the importance of NA-based immunity, current vaccines are focused on the hemagglutinin (HA) protein as the target for protective antibodies, and the amount of NA is not standardized in virion-based vaccines. Antibodies targeting NA are predominantly protective, reducing infection severity and viral shedding. Recently, NA-specific monoclonal antibodies have been characterized, and their target epitopes have been identified. This review summarizes the characteristics of NA, NA-specific antibodies, the mechanism of NA inhibition, and the recent efforts towards developing NA-based and NA-incorporating influenza vaccines.

Ilheus virus (ILHV) is a mosquito-borne flavivirus circulating throughout Central and South America and the Caribbean. It has been detected in several mosquito genera including Aedes and Culex, and birds are thought to be its primary amplifying and reservoir host. Here, we describe the genomic and morphologic characterization of ten ILHV strains. Our analyses revealed a high conservation of both the 5'- and 3'-untranslated regions but considerable divergence within the open reading frame. We also showed that ILHV displays a typical flavivirus structural and genomic organization. Our work lays the foundation for subsequent ILHV studies to better understand its transmission cycles, pathogenicity, and emergence potential.

Infection with Aspergillus fumigatus polymycovirus 1 (AfuPmV-1) weakens resistance of Aspergillus fumigatus common reference strain Af293 biofilms in intermicrobial competition with Pseudomonas aeruginosa. We compared the sensitivity of two infected and one virus-free Af293 strains to antifungal drugs. All three were comparably sensitive to drugs affecting fungal membranes (voriconazole, amphotericin) or cell wall glucan synthesis (micafungin, caspofungin). In contrast, forming biofilms of virus-free Af293 were much more resistant than AfuPmV-1-infected Af293 to nikkomycin Z (NikZ), a drug inhibiting chitin synthase. The IC50 for NikZ on biofilms was between 3.8 and 7.5 µg/mL for virus-free Af293 and 0.94-1.88 µg/mL for infected strains. The IC50 for the virus-free A. fumigatus strain 10AF was ~2 µg/mL in most experiments. NikZ also modestly affected the planktonic growth of infected Af293 more than the virus-free strain (MIC 50%, 2 and 4 µg/mL, respectively). Virus-free Af293 biofilm showed increased metabolism, and fungus growing as biofilm or planktonically showed increased growth compared to infected; these differences do not explain the resistance of the virus-free fungus to NikZ. In summary, AfuPmV-1 infection sensitized A. fumigatus to NikZ, but did not affect response to drugs commonly used against A. fumigatus infection. Virus infection had a greater effect on NikZ inhibition of biofilm than planktonic growth.

Adoptive immunotherapy using chimeric antigen receptor (CAR) T cells has been highly successful in treating B cell malignancies and holds great potential as a curative strategy for HIV infection. Recent advances in the use of anti-HIV broadly neutralizing antibodies (bNAbs) have provided vital information for optimal antigen targeting of CAR T cells. However, CD4+ CAR T cells are susceptible to HIV infection, limiting their therapeutic potential. In the current study, we engineered HIV-resistant CAR T cells using CRISPR/Cas9-mediated integration of a CAR cassette into the CCR5 locus. We used a single chain variable fragment (scFv) of the clinically potent bNAb 10-1074 as the antigen-targeting domain in our anti-HIV CAR T cells. Our anti-HIV CAR T cells showed specific lysis of HIV-infected cells in vitro. In a PBMC humanized mouse model of HIV infection, the anti-HIV CAR T cells expanded and transiently limited HIV infection. In conclusion, this study provides proof-of-concept for developing HIV-resistant CAR T cells using CRISPR/Cas9 targeted integration.

BACKGROUND: Chile has achieved the highest coverage for vaccines against the SARS-CoV-2 virus worldwide. OBJECTIVE: To assess the progression of immunity (natural and acquired by vaccine) in a cohort from two Chilean cities. METHODS: Individuals (n = 386) who participated in three phases of population-based serial prevalence studies were included (2020-2021 and 2022). Presence of SARS-CoV-2 antibodies was measured in serum. Data including time of vaccination and type of vaccine received were analysed with descriptive statistics. RESULTS: Seroprevalence was 3.6% in the first round and increased to 96.9% in the second and 98.7% in the third. In the third round, 75% of individuals who had received the basal full scheme were seropositive at 180 days or more since their last dose; 98% of individuals who received one booster dose were seropositive at 180 days or more, and 100% participants who received two boosters were seropositive, regardless of time since their last dose. Participants receiving mRNA vaccines had higher seroprevalence rates over time. CONCLUSIONS: The high vaccination coverage in Chile enabled the population to maintain high levels of antibodies. Vaccination boosters are essential to maintain immunity over time, which also depends on the type of vaccine administered.

Since late 2016, a yellow fever virus (YFV) variant carrying a set of nine amino acid variations has circulated in South America. Three of them were mapped on the methyltransferase (MTase) domain of viral NS5 protein. To assess whether these changes affected viral infectivity, we synthesized YFV carrying the MTase of circulating lineage as well as its isoform with the residues of the previous strains (NS5 K101R, NS5 V138I, and NS5 G173S). We observed a slight difference in viral growth properties and plaque phenotype between the two synthetic YFVs. However, the MTase polymorphisms associated with the Brazilian strain of YFV (2016-2019) confer more susceptibility to the IFN-I. In addition, in vitro MTase assay revealed that the interaction between the YFV MTase and the methyl donor molecule (SAM) is altered in the Brazilian MTase variant. Altogether, the results reported here describe that the MTase carrying the molecular signature of the Brazilian YFV circulating since 2016 might display a slight decrease in its catalytic activity but virtually no effect on viral fitness in the parameters comprised in this study. The most marked influence of these residues stands in the immune escape against the antiviral response mediated by IFN-I.

(1) Background: SARS-CoV-2 infection is notably mild in children, though comorbidities may increase the risk of hospitalization and may represent a risk for increased disease severity. There is an urgent need for targeted therapies with an acceptable efficacy and safety profile. To date, most of the medicines for COVID-19-specific treatment are prescribed off-label for children due to a lack of clinical trials and consequent evidence in this population. (2) Methods: This was a retrospective, observational study investigating the safety of treatments for the prevention of severe COVID-19 in fragile pediatric patients who received monoclonal antibodies and antivirals for mild-to-moderate symptoms between December 2021 and July 2022. (3) Results: Thirty-two patients were included. Monoclonal antibodies were prescribed to 62%, intravenous antivirals to 22%, and oral antivirals to 16% of children. Sotrovimab was the most frequently prescribed drug among monoclonal antibodies and overall (59%). The second most prescribed drug was remdesivir (22%). No severe adverse drug reaction was reported. There was no progression to severe disease and no death cases due to COVID-19 or drug administration. At drug-type stratification, resolution of symptoms and swab positivity time showed no difference between the two groups at 7 and 28 days. Off-label prescriptions were 84% overall, and in similar proportions between the two groups. (4) Conclusions: in this small sample, antivirals seemed safe and showed no differences in efficacy as compared to MAbs for the early treatment of COVID-19 in fragile children, thus representing a valuable choice, even when administered off-label.

The aim of this study was to evaluate whether the presence of anti-hepatitis B (HBV) c antibodies (HBcAb positivity) could influence the control of HIV viremia in patients living with HIV (PLWH) who switch to two-drug antiretroviral therapy (2DR) containing lamivudine (3TC) (2DR-3TC-based). A retrospective multicentre observational study was conducted on 160 PLWH switching to the 2DR-3TC-based regimen: 51 HBcAb-positive and 109 HBcAb-negative patients. The HBcAb-positive PLWH group demonstrated a significantly lower percentage of subjects with HIV viral suppression with target not detected (TND) at all time points after switching (24th month: 64.7% vs. 87.8%, p < 0.0001; 36th month 62.7% vs. 86.8%, p = 0.011; 48th month 57.2% vs. 86.1%, p = 0.021 of the HBcAb-positive and HBcAb-negative groups, respectively). Logistic regression analysis showed that the presence of HBcAb positivity (OR 7.46 [95% CI 2.35-14.77], p = 0.004) could favour the emergence of HIV viral rebound by nearly 54% during the entire study follow-up after switching to 2DR-3TC.

The incidence of oropharyngeal squamous cell carcinoma (OPSCC) has increased in the past decades due to carcinogenic HPV infection. As this patient group suffers from considerable mortality and treatment morbidity it is important to improve prognostic strategies in OPSCC. Inflammation plays a key role in cancer and the neutrophil-to-lymphocyte ratio (NLR) in blood has been suggested as a prognostic factor for OPSCC. This study aimed to investigate the prognostic impact of NLR on overall survival (OS) and recurrence-free survival (RFS) in a retrospective cohort of 1370 patients. Included patients had pretreatment neutrophil and lymphocyte counts available, as well as a known HPV status. Patients were treated with curative intent according to Danish national guidelines. We stratified patients in groups by NLR < 2, NLR 2-4, or NLR > 4 and analyzed the influence of the NLR tertile on OS and RFS. Kaplan-Meier curves illustrated survival probability in OS and RFS in the general cohort and were stratified by HPV status. We found that an increasing NLR was associated with inferior OS (HR = 1.5 for NLR > 4) and RFS (HR = 1.6 for NLR 2-4; HR = 1.8 for NLR > 4) in multivariable analysis. The Kaplan-Meier curves displayed inferior OS and RFS with an increasing NLR for both HPV+ and HPV- patients. In conclusion, we showed that an increasing NLR is prognostic for a worse outcome of OPSCC independently of HPV status. There are possible uses of NLR in prognostication and treatment de-escalation although further studies are warranted to determine the clinical utility.

More than 40 human infections with the zoonotic Borna disease virus 1 (BoDV-1) have been reported to German health authorities from endemic regions in southern and eastern Germany. Diagnosis of a confirmed case is based on the detection of BoDV-1 RNA or BoDV-1 antigen. In parallel, serological assays such as ELISA, immunoblots, and indirect immunofluorescence are in use to detect the seroconversion of Borna virus-reactive IgG in serum or cerebrospinal fluid (CSF). As immunopathogenesis in BoDV-1 encephalitis appears to be driven by T cells, we addressed the question of whether an IFN-γ-based ELISpot may further corroborate the diagnosis. For three of seven BoDV-1-infected patients, peripheral blood mononuclear cells (PBMC) with sufficient quantity and viability were retrieved. For all three patients, counts in the range from 12 to 20 spot forming units (SFU) per 250,000 cells were detected upon the stimulation of PBMC with a peptide pool covering the nucleocapsid protein of BoDV-1. Additionally, individual patients had elevated SFU upon stimulation with a peptide pool covering X or phosphoprotein. Healthy blood donors (n = 30) and transplant recipients (n = 27) were used as a control and validation cohort, respectively. In this pilot study, the BoDV-1 ELISpot detected cellular immune responses in human patients with BoDV-1 infection. Its role as a helpful diagnostic tool needs further investigation in patients with BoDV-1 encephalitis.

Zika virus (ZIKV) causes microcephaly and congenital eye disease. The cellular and molecular basis of congenital ZIKV infection are not well understood. Here, we utilized a biologically relevant cell-based system of human fetal retinal pigment epithelial cells (FRPEs), hiPSC-derived retinal stem cells (iRSCs), and retinal organoids to investigate ZIKV-mediated ocular cell injury processes. Our data show that FRPEs were highly susceptible to ZIKV infection exhibiting increased apoptosis, whereas iRSCs showed reduced susceptibility. Detailed transcriptomics and proteomics analyses of infected FRPEs were performed. Nucleoside analogue drug treatment inhibited ZIKV replication. Retinal organoids were susceptible to ZIKV infection. The Asian genotype ZIKV exhibited higher infectivity, induced profound inflammatory response, and dysregulated transcription factors involved in retinal organoid differentiation. Collectively, our study shows that ZIKV affects ocular cells at different developmental stages resulting in cellular injury and death, further providing molecular insight into the pathogenesis of congenital eye disease.