Identification of porcine PARP11 as a restricted factor for pseudorabies virus.

Introduction: PRV infection in swine can cause devastating disease and pose a potential threat to humans. Advancing the interplay between PRV and host is essential to elucidate the pathogenic mechanism of PRV and identify novel anti-PRV targets. Methods: PARP11-KO PK-15 cells were firstly constructed by CRISPR/Cas9 technology. Next, the effect of PARP11-KO on PRV infection was determined by RT-qPCR, TCID50 assay, RNA-seq, and western blot. Results and discussion: In this study, we identified PARP11 as a host factor that can significantly affect PRV infection. Inhibition of PARP11 and knockout of PARP11 can significantly promoted PRV infection. Subsequently, we further found that PARP11 knockout upregulated the transcription of NXF1 and CRM1, resulting in enhanced transcription of viral genes. Furthermore, we also found that PARP11 knockout could activate the autophagy pathway and suppress the mTOR pathway during PRV infection. These findings could provide insight into the mechanism in which PARP11 participated during PRV infection and offer a potential target to develop anti-PRV therapies.

pUS6 in pseudorabies virus participates in the process of inhibiting antigen presentation by inhibiting the assembly of peptide loading complex.

Pseudorabies virus (PRV) can establish lifelong latent infection in peripheral nervous ganglion, and persistent infections in peripheral blood lymphocytes. Establishing an infection in the lymphocytes does not only enable the PRV to escape host immune surveillance but pass through the placental barrier, leading to fetal death and abortion. Due to the pathogenicity of the PRV, it poses a huge challenge in its prevention and control. The PRV escapes host immunity through downregulation of swine leukocyte antigen class I (SLA I) molecules on infected cells. However, data on the molecular mechanisms of the SLA I suppression remains scant. Here, in order to verify the effect of candidate proteins PRV pUL44 and pUS6 on PRV immune escape related molecules SLA I and peptide loading complex (PLC), we detected the expression of SLA I and PLC components after expressing PRV pUL44 and pUS6. The effects of pUS6 and pUL44 on SLA I and PLC were analyzed by qRT-PCR and Western blot at mRNA and protein level, respectively. Cells expressing pUS6 or pUL44 genes showed a significantly suppressed expression of surface and total SLA I molecules. In addition, unlike UL44, the US6 gene was shown to downregulate the transporter associated with antigen processing 1 (TAP1), TAP2 and Tapasin molecules. The results show that PRV pUS6 may participate in virus immune escape by directly regulating the SLA I, TAP dimer and Tapasin molecules, thus blocking the transportation of TAP-bound peptides to the ER to bind SLA I molecules. We provide a theoretical basis on the mechanism of TAP mediated immune escape by the PRV.

Pseudorabies Virus UL4 protein promotes the ASC-dependent inflammasome activation and pyroptosis to exacerbate inflammation.

Pseudorabies virus (PRV) infection causes systemic inflammatory responses and inflammatory damages in infected animals, which are associated with the activation of inflammasome and pyroptosis in infected tissues. Here, we identified a critical function of PRV non-structural protein UL4 that enhanced ASC-dependent inflammasome activation to promote pyroptosis. Whereas, the deficiency of viral UL4 was able to reduce ASC-dependent inflammasome activation and the occurrences of pyroptosis. Mechanistically, the 132-145 aa of UL4 permitted its translocation from the nucleus to the cytoplasm to interact with cytoplasmic ASC to promote the activation of NLRP3 and AIM2 inflammasome. Further research showed that UL4 promoted the phosphorylation levels of SYK and JNK to enhance the ASC phosphorylation, which led to the increase of ASC oligomerization, thus promoting the activation of NLRP3 and AIM2 inflammasome and enhanced GSDMD-mediated pyroptosis. In vivo experiments further showed that PRV UL4 (132DVAADAAAEAAAAE145) mutated strain (PRV-UL4mut) infection did not lead to a significant decrease in viral titers at 12 h. p. i, but it induced lower levels of IL-1ß, IL-18, and GSDMD-NT, which led to an alleviated inflammatory infiltration and pathological damage in the lungs and brains, and a lower death rate compared with wild-type PRV strain infection. Taken together, our findings unravel that UL4 is an important viral regulator to manipulate the inflammasome signaling and pyroptosis of host cells to promote the pathogenicity of PRV, which might be further exploited as a new target for live attenuated vaccines or therapeutic strategies against pseudorabies in the future.

Salvianolic acid A inhibits pseudorabies virus infection by directly inactivating the virus particle.

BACKGROUND: Pseudorabies virus (PRV), a member of the family Herpesviridae, is responsible for significant economic losses in the pig industry and has recently been associated with human viral encephalitis, leading to severe neurological symptoms post-recovery. Despite the widespread impact of PRV, there are currently no approved effective drugs for treating PRV-related diseases in humans or pigs. Therefore, the exploration and discovery of safe and effective drugs for the prevention and treatment of PRV infection is of paramount importance. PURPOSE: The objective of this study is to screen and identify natural compounds with antiviral activity against PRV. METHODS: First, we used a strain of PRV with green fluorescent protein (PRV-GFP) to screen a natural product chemical library to identify potential antiviral drugs. Next, we assessed the antiviral abilities of salvianolic acid A (SAA) in vitro using virus titer assay, qPCR, and IFA. We investigated the mechanisms of SAA's antiviral activity through viral attachment, internalization, inactivation, and nuclease digestion assay. Finally, we evaluated the efficacy of SAA in inactivating PRV using mice as the experimental subjects. RESULTS: This study screened 206 natural compounds for anti-PRV activity in vitro, resulting in the identification of seven potential antiviral agents. Notably, SAA emerged as a promising candidate with significant anti-PRV activity. The mechanism of action may be that SAA can directly inactivate the virus by disrupting viral envelope. In vivo experiments have shown that pre-incubation of SAA and PRV can effectively inhibit the infectivity and pathogenicity of PRV in mice. CONCLUSION: This study offers valuable insights into the antiviral properties of SAA, potentially informing strategies for controlling PRV epidemics and treating related diseases in both humans and animals.

Immune Responses Induced by a Recombinant Lactiplantibacillus plantarum Surface-Displaying the gD Protein of Pseudorabies Virus.

Pseudorabies virus (PRV) is one of the herpes viruses that can infect a wide range of animals including pigs, cattle, sheep, mice, and wild animals. PRV is a neurotropic alphaherpesvirus capable of infecting a variety of mammals. There is a rising interest in the targeted application of probiotic bacteria to prevent viral diseases, including PRV. In this study, the surface expression of enhanced green fluorescent protein (EGFP) on recombinant Lactiplantibacillus plantarum NC8 (rNC8) through the LP3065 LPxTG motif of Lactobacillus plantarum WCFS1 was generated. The surface expression was observed through confocal microscopy. Dendritic cell targeting peptides (DCpep) were also fused with LPxTG that help to bind with mouse DCs. The PRV-gD was cloned in LP3065 LPxTG, resulting in the generation of rNC8-LP3065-gD. Inactivated rNC8-LP3065-gD was administered intravenously in mice on days 1 and 7 at a dose of 200 µL (109 CFU/mouse) for monitoring immunogenicity. Subsequently, a challenge dose of PRV TJ (104 TCID50) was administered intramuscularly at 14 days post-immunization. The survival rate of the immunized mice reached 80% (4/5) with no significant signs of illness. A significant rise in anti-gD antibodies was detected in the immunized mice by ELISA. Quantitative PCR (qPCR) results showed decreased viral loading in different body tissues. Flow cytometry of lymphocytes derived from mice spleen indicated an increase in CD3+CD4+ T cells, but CD3+CD8+ T cells were not detected. Moreover, it offers a model to delineate immune correlates with rNC8-induced immunity against swine viral diseases.

Effect of pseudorabies virus infection on NMDA receptor expression in mice and its role in immunosuppression.

Pseudorabies virus (PRV), an α-herpesvirus, induces immunosuppression and can lead to severe neurological diseases. N-methyl-D-aspartate receptor (NMDAR), an important excitatory nerve receptor in the central nervous system, is linked to various nervous system pathologies. The link between NMDAR and PRV-induced neurological diseases has not been studied. In vivo studies revealed that PRV infection triggers a reduction in hippocampal NMDAR expression, mediated by inflammatory processes. Extensive hippocampal neuronal degeneration was found in mice on the 6th day by hematoxylin-eosin staining, which was strongly correlated with increased NMDAR protein expression. In vitro studies utilizing the CCK-8 assay demonstrated that treatment with an NMDAR antagonist significantly heightened the cytotoxic effects of PRV on T lymphocytes. Notably, NMDAR inhibition did not affect the replication ability of PRV. However, it facilitated the accumulation of pro-inflammatory cytokines in PRV-infected T cells and enhanced the transcription of the CD25 gene through the secretion of interleukin-2 (IL-2), consequently exacerbating immunosuppression. In this study, we found that NMDAR has functional activity in T lymphocytes and is crucial for the inflammatory and immune responses triggered by PRV infection. These discoveries highlight the significant role of NMDAR in PRV-induced neurological disease pathogenesis.

Analysis of the recombination and evolution of the new type mutant pseudorabies virus XJ5 in China.

Pseudorabies have caused enormous economic losses in China's pig industry and have recurred on many large pig farms since late 2011. The disease is caused by highly pathogenic, antigenic variant pseudorabies virus (vPRV) strains. Our laboratory isolated a pseudorabies virus in 2015 and named it XJ5. The pathogenic ability of this mutant strain was much stronger than that of the original isolate. After we sequenced its whole genome (GenBank accession number: OP512542), we found that its overall structure was not greatly changed compared with that of the previous strain Ea (KX423960.1). The whole genome alignment showed that XJ5 had a strong genetic relationship with the strains isolated in China after 2012 reported in GenBank. Based on the isolation time of XJ5 and the mutation and recombination analysis of programs, we found that the whole genome homology of XJ5 and other strains with Chinese isolates was greater than 95%, while the homology with strains outside Asia was less than 94%, which indicated that there may be some recombination and mutation patterns. We found that virulent PRV isolates emerged successively in China in 2011 and formed two different evolutionary clades from foreign isolates. At the same time, this may be due to improper immunization and the presence of wild strains in the field, and recent reports have confirmed that Bartha vaccine strains recombine with wild strains to obtain new pathogenic strains. We performed genetic evolution analysis of XJ5 isolated and sequenced in our laboratory to trace its possible mutations and recombination. We found that XJ5 may be the result of natural mutation of a virus in a branch of mutant strains widely existing in China.

STRAP upregulates antiviral innate immunity against PRV by targeting TBK1.

Serine/threonine kinase receptor-associated protein (STRAP) serves as a scaffold protein and is engaged in a variety of cellular activities, although its importance in antiviral innate immunity is unknown. We discovered that STRAP works as an interferon (IFN)-inducible positive regulator, facilitating type I IFN signaling during pseudorabies virus infection. Mechanistically, STRAP interacts with TBK1 to activate type I IFN signaling. Both the CT and WD40 7 - 6 domains contribute to the function of STRAP. Furthermore, TBK1 competes with PRV-UL50 for binding to STRAP, and STRAP impedes the degradation of TBK1 mediated by PRV-UL50, thereby increasing the interaction between STRAP and TBK1. Overall, these findings reveal a previously unrecognized role for STRAP in innate antiviral immune responses during PRV infection. STRAP could be a potential therapeutic target for viral infectious diseases.

Pseudorabies virus infection triggers mitophagy to dampen the interferon response and promote viral replication.

Pseudorabies virus (PRV) utilizes multiple strategies to inhibit type I interferon (IFN-I) production and signaling to achieve innate immune evasion. Among several other functions, mitochondria serve as a crucial immune hub in the initiation of innate antiviral responses. It is currently unknown whether PRV inhibits innate immune responses by manipulating mitochondria. In this study, we found that PRV infection damages mitochondrial structure and function, as shown by mitochondrial membrane potential depolarization, reduction in mitochondrial numbers, and an imbalance in mitochondrial dynamics. In addition, PRV infection triggered PINK1-Parkin-mediated mitophagy to eliminate the impaired mitochondria, which resulted in a suppression of IFN-I production, thereby promoting viral replication. Furthermore, we found that mitophagy resulted in the degradation of the mitochondrial antiviral signaling protein, which is located on the mitochondrial outer membrane. In conclusion, the data of the current study indicate that PRV-induced mitophagy represents a previously uncharacterized PRV evasion mechanism of the IFN-I response, thereby promoting virus replication.IMPORTANCEPseudorabies virus (PRV), a pathogen that induces different disease symptoms and is often fatal in domestic animals and wildlife, has caused great economic losses to the swine industry. Since 2011, different PRV variant strains have emerged in Asia, against which current commercial vaccines may not always provide optimal protection in pigs. In addition, there are indications that some of these PRV variant strains may sporadically infect people. In the current study, we found that PRV infection causes mitochondria injury. This is associated with the induction of mitophagy to eliminate the damaged mitochondria, which results in suppressed antiviral interferon production and signaling. Hence, our study reveals a novel mechanism that is used by PRV to antagonize the antiviral host immune response, providing a theoretical basis that may contribute to the research toward and development of new vaccines and antiviral drugs.

The inhibitory effect of swine TAB1 on the replication of pseudorabies virus.

TAK1-binding protein 1 (TAB1) assembles with TAK1 through its C-terminal domain, leading to the self-phosphorylation and activation of TAK1, which plays an important role in the activation of NF-κB and MAPK signaling pathway. Pseudorabies virus (PRV) is the pathogen of Pseudorabies (PR), which belongs to the Alphaherpesvirus subfamily and causes serious economic losses to the global pig industry. However, the impact of swine TAB1 (sTAB1) on PRV infection has not been reported. In this study, evidence from virus DNA copies, virus titer and western blotting confirmed that sTAB1 could inhibit PRV replication and knockout of sTAB1 by CRISPR-Cas9 gene editing system could promote PRV replication. Further mechanistic studies by real-time PCR and luciferase reporter gene assay demonstrated that sTAB1 could enhance the production of inflammatory factors and chemokines, IFN-ß transcription level and IFN-ß promoter activity after PRV infection. In summary, we clarify the underlying mechanism of sTAB1 in inhibiting PRV replication for the first time, which provides a new idea for preventing PRV infection and lays a foundation for PRV vaccine development.

A pseudorabies outbreak in hunting dogs in Campania region (Italy): a case presentation and epidemiological survey.

BACKGROUND: Pseudorabies is an infection of domestic and wild pigs that has occasionally been reported in dogs with fatal encephalitis. Hunting dogs are predisposed to pseudorabies exposure due to incorrect practices (administration of raw infected meat) or close contact with infected wild boars. This study described an outbreak of pseudorabies in two hunting dogs in the Campania region, southern Italy. CASE PRESENTATION: Two hunting dogs were hospitalized after a hunting trip, with fever, itching, and self-inflicted lesions. Laboratory tests showed mild anemia and marked leukocytosis. Despite conservative therapy, both animals died 48 h after the presentation of symptoms. One of the carcasses was sent to the Department of Veterinary Medicine and Animal Production in Naples to confirm the suspicion of pseudorabies. DNA was extracted from different matrices and used as a template for real-time PCR to detect PRV. Several samples (brain, cerebellum, brainstem, lung, and liver) tested positive. Subsequent sequence analyses of glycoprotein E from DNA extracted from the brain stem revealed a sequence similarity to those described in previous cases of pseudorabies in dogs in Italy, France and Belgium. One month after the outbreak, blood samples were collected from 42 dogs belonging to the same hunting team and from 245 dogs (cohort population) living in the Campania region. All samples were tested with two commercial ELISAs to detect seroconversion against glycoproteins B and E. A seroprevalence of 19% was observed in the hunting team affected by the outbreak, while only 0.8% was observed in the regional dog population. CONCLUSIONS: The data reported in this study demonstrate potential exposure to PRV by dead-end hosts, particularly hunting dogs. The sequencing results indicated the homogeneity of PRV strains circulating in the different Italian regions.

Rapid, sensitive, and visual detection of pseudorabies virus with an RPA-CRISPR/EsCas13d-based dual-readout portable platform.

Pseudorabies viruses (PRV) pose a major threat to the global pig industry and public health. Rapid, intuitive, affordable, and accurate diagnostic testing is critical for controlling and eradicating infectious diseases. In this study, a portable detection platform based on RPA-CRISPR/EsCas13d was developed. The platform exhibits high sensitivity (1 copy/µL), good specificity, and no cross-reactivity with common pathogens. The platform uses rapid preamplification technology to provide visualization results (lateral flow assays or visual fluorescence) within 1 h. Fifty pig samples (including tissues, oral fluids, and serum) were tested using this platform and real-time quantitative polymerase chain reaction (qPCR), showing 34.0 % (17 of 50) PRV positivity with the portable CRISPR/EsCas13d dual-readout platform, consistent with the qPCR results. These results highlight the stability, sensitivity, efficiency, and low equipment requirements of the portable platform. Additionally, a novel point-of-care test is being developed for clinical use in remote rural and resource-limited areas, which could be a prospective measure for monitoring the progression of pseudorabies and other infectious diseases worldwide.

Evaluation of the activity and mechanisms of oregano essential oil against PRV in vivo and in vitro.

BACKGROUND: The Pseudorabies Virus (PRV) leading to pseudorabies and causes huge economic losses in pig industry. The development of novel PRV variations has diminished the efficacy of traditional vaccinations, and there is yet no medication that can stop the spread of PRV infection. Therefore, PRV eradication is challenging. Oregano essential oil, the plant-based ingredient for medication feed have been shown to has strong anti-herpesvirus activity, but no anti-PRV function has been reported. RESULTS: The current study assessed the anti-pseudorabies virus (PRV) activity of oregano essential oil and explored its mechanisms and most effective components against PRV. Our in vivo findings demonstrated that oregano essential oil could decrease the PRV load in tissues, mitigate tissue lesions, and enhance the survival rate of mice. The potential antiviral mechanism involves augmenting humoral and cellular immune responses in PRV-infected mice. To further investigate the most effective components of oregano essential oil against PRV, an in vitro study was conducted, revealing that oregano essential oil and its main constituents, carvacrol and thymol, all diminished PRV intracellular proliferation in vitro. Carvacrol exhibited the most potent anti-PRV effect, serving as the primary contributor to oregano essential oil's anti-PRV activity. The mechanisms underlying carvacrol's anti-PRV properties include the upregulation of cytokines TNF-α, IFN-ß, IFN-γ, IL-12, and the inhibition of PRV-induced apoptosis in BHK-21 cells. CONCLUSIONS: Our study provides an effective drug for the prevention and control of PRV infection.

Pseudorabies virus infection increases the permeability of the mammalian respiratory barrier to facilitate Pasteurella multocida infection.

Interaction between viruses and bacteria during the development of infectious diseases is a complex question that requires continuous study. In this study, we explored the interactions between pseudorabies virus (PRV) and Pasteurella multocida (PM), which are recognized as the primary and secondary agents of porcine respiratory disease complex (PRDC), respectively. In vivo tests using mouse models demonstrated that intranasal inoculation with PRV at a sublethal dose induced disruption of murine respiratory barrier and promoted the invasion and damages caused by PM through respiratory infection. Inoculation with PRV also disrupted the barrier function of murine and porcine respiratory epithelial cells, and accelerated the adherence and invasion of PM to the cells. In mechanism, PRV infection resulted in decreased expression of tight junction proteins (ZO-1, occludin) and adherens junction proteins (ß-catenin, E-cadherin) between neighboring respiratory epithelial cells. Additionally, PRV inoculation at an early stage downregulated multiple biological processes contributing to epithelial adhesion and barrier functions while upregulating signals beneficial for respiratory barrier disruption (e.g., the HIF-1α signaling). Furthermore, PRV infection also stimulated the upregulation of cellular receptors (CAM5, ICAM2, ACAN, and DSCAM) that promote bacterial adherence. The data presented in this study provide insights into the understanding of virus-bacteria interactions in PRDC and may also contribute to understanding the mechanisms of secondary infections caused by different respiratory viruses (e.g., influenza virus and SARS-CoV-2) in both medical and veterinary medicine. IMPORTANCE: Co-infections caused by viral and bacterial agents are common in both medical and veterinary medicine, but the related mechanisms are not fully understood. This study investigated the interactions between the zoonotic pathogens PRV and PM during the development of respiratory infections in both cell and mouse models, and reported the possible mechanisms which included: (i) the primary infection of PRV may induce the disruption and/or damage of mammal respiratory barrier, thereby contributing to the invasion of PM; (ii) PRV infection at early stage accelerates the transcription and/or expression of several cellular receptors that are beneficial for bacterial adherence. This study may shed a light on understanding the mechanisms on the secondary infection of PM promoted by different respiratory viruses (e.g., influenza virus and SARS-CoV-2) in both medical and veterinary medicine.

The global landscapes of lysine crotonylation in pseudorabies virus infection.

Lysine crotonylation is a common occurrence in eukaryotic cells, regulating various physiological functions, including chromatin remodeling, cellular growth, and development. However, its involvement in viral infections has rarely been documented. In this study, we reveal that pseudorabies virus (PRV) infection significantly alters the global lysine crotonylation levels in porcine kidney PK-15 cells. Specifically, we identified a few viral proteins, including UL54, gM, gD, UL19, UL37, and UL46, which undergo crotonylation modification. Our observations indicate that at 20 h post-infection (hpi), 551 crotonylation sites were reduced across 345 proteins, while 47 new sites emerged in 37 proteins compared to the control group. By 40 hpi, 263 sites had decreased in 190 proteins, while 389 new sites appeared in 240 proteins. Deeper analysis revealed that the proteins with altered crotonylation levels were primarily involved in binding, catalytic activity, biosynthetic processes, ribosome activity, and metabolic processes. Additionally, our findings underscored the significance of ribosomes and the endoplasmic reticulum (ER), which were enriched with proteins exhibiting altered crotonylation. Overall, our study for the first time offers new insights into the relationship between crotonylation and herpes virus infection, paving the way for future investigations into the role of crotonylation in viral infections.

Pseudorabies virus tegument protein US2 antagonizes antiviral innate immunity by targeting cGAS-STING signaling pathway.

Background: The cGAS-STING axis-mediated type I interferon pathway is a crucial strategy for host defense against DNA virus infection. Numerous evasion strategies developed by the pseudorabies virus (PRV) counteract host antiviral immunity. To what extent PRV-encoded proteins evade the cGAS-STING signaling pathway is unknown. Methods: Using US2 stably expressing cell lines and US2-deficient PRV model, we revealed that the PRV tegument protein US2 reduces STING protein stability and downregulates STING-mediated antiviral signaling. Results: To promote K48-linked ubiquitination and STING degradation, US2 interacts with the LBD structural domain of STING and recruits the E3 ligase TRIM21. TRIM21 deficiency consistently strengthens the host antiviral immune response brought on by PRV infection. Additionally, US2-deficient PRV is less harmful in mice. Conclusions: Our study implies that PRV US2 inhibits IFN signaling by a new mechanism that selectively targets STING while successfully evading the host antiviral response. As a result, the present study reveals a novel strategy by which PRV evades host defense and offers explanations for why the Bartha-K61 classical vaccine strain failed to offer effective defense against PRV variant strains in China, indicating that US2 may be a key target for developing gene-deficient PRV vaccines.

Comparative proteomic analysis of PK-15 cells infected with wild-type strain and its EP0 gene-deleted mutant strain of pseudorabies virus.

IMPORTANCE: As one of the main etiologic agents of infectious diseases in pigs, pseudorabies virus (PRV) infections have caused enormous economic losses worldwide. EP0, one of the PRV early proteins (EP) plays a vital role in PRV infections, but the mechanisms are unclear. OBJECTIVE: This study examined the function of EP0 to provide a direction for its in-depth analysis. METHODS: In this study, the EP0-deleted PRV mutant was obtained, and Tandem Mass Tag-based proteomic analysis was used to screen the differentially expressed proteins (DEPs) quantitatively in EP0-deleted PRV- or wild-type PRV-infected porcine kidney 15 cells. RESULTS: This study identified 7,391 DEPs, including 120 and 21 up-regulated and down-regulated DEPs, respectively. Western blot analysis confirmed the changes in the expression of the selected proteins, such as speckled protein 100. Comprehensive analysis revealed 141 DEPs involved in various biological processes and molecular functions, such as transcription regulator activity, biological regulation, and localization. CONCLUSIONS AND RELEVANCE: These results holistically outlined the functions of EP0 during a PRV infection and might provide a direction for more detailed function studies of EP0 and the stimulation of lytic PRV infections.

Pseudorabies virus VHS protein abrogates interferon responses by blocking NF-κB and IRF3 nuclear translocation.

Herpesviruses antagonize host antiviral responses through a myriad of molecular strategies culminating in the death of the host cells. Pseudorabies virus (PRV) is a significant veterinary pathogen in pigs, causing neurological sequalae that ultimately lead to the animal's demise. PRV is known to trigger apoptotic cell death during the late stages of infection. The virion host shutdown protein (VHS) encoded by UL41 plays a crucial role in the PRV infection process. In this study, we demonstrate that UL41 inhibits PRV-induced activation of inflammatory cytokine and negatively regulates the cGAS-STING-mediated antiviral activity by targeting IRF3, thereby inhibiting the translocation and phosphorylation of IRF3. Notably, mutating the conserved amino acid sites (E192, D194, and D195) in the RNase domain of UL41 or knocking down UL41 inhibits the immune evasion of PRV, suggesting that UL41 may play a crucial role in PRV's evasion of the host immune response during infection. These results enhance our understanding of how PRV structural proteins assist the virus in evading the host immune response.

Rehmmannia glutinosa polysaccharide exerts antiviral activity against pseudorabies virus and antioxidant activity.

Pseudorabies virus (PRV) is an important pathogen harming the global pig industry. Vaccines available for swine cannot protect against PRV completely. Furthermore, no antiviral drugs are available to treat PRV infections. Rehmmannia glutinosa polysaccharide (RGP) possesses several medicinal properties. However, its antiviral activity is not reported. In the present study, we found that RGP can inhibit PRV/XJ5 infection by western blotting, immunofluorescent assay (IFA), and TCID50 assay quantitative polymerase chain reaction (qPCR). We revealed RGP can inhibit virus adsorption and invasion into PK-15 cells in a dose-dependent manner via western blotting, IFA, TCID50 assay, and quantitative polymerase chain reaction (qPCR), and suppressed PRV/XJ5 replication through western blotting, and qPCR. Additionally, it also reduced PRV/XJ5-induced ROS, lipid oxidation, and improved SOD levels in PK-15 cells, which was observed by using corresponding test kits. To conclude, our findings suggest that RGP might be a novel therapeutic agent for preventing and controlling PRV infection and antioxidant agent.

Novel Multiparametric Bioelectronic Measurement System for Monitoring Virus-Induced Alterations in Functional Neuronal Networks.

Development and optimisation of bioelectronic monitoring techniques like microelectrode array-based field potential measurement and impedance spectroscopy for the functional, label-free and non-invasive monitoring of in vitro neuronal networks is widely investigated in the field of biosensors. Thus, these techniques were individually used to demonstrate the capabilities of, e.g., detecting compound-induced toxicity in neuronal culture models. In contrast, extended application for investigating the effects of central nervous system infecting viruses are rarely described. In this context, we wanted to analyse the effect of herpesviruses on functional neuronal networks. Therefore, we developed a unique hybrid bioelectronic monitoring platform that allows for performing field potential monitoring and impedance spectroscopy on the same microelectrode. In the first step, a neuronal culture model based on primary hippocampal cells from neonatal rats was established with reproducible and stable synchronised electrophysiological network activity after 21 days of cultivation on microelectrode arrays. For a proof of concept, the pseudorabies model virus PrV Kaplan-ΔgG-GFP was applied and the effect on the neuronal networks was monitored by impedance spectroscopy and field potential measurement for 72 h in a multiparametric mode. Analysis of several bioelectronic parameters revealed a virus concentration-dependent degeneration of the neuronal network within 24-48 h, with a significant early change in electrophysiological activity, subsequently leading to a loss of activity and network synchronicity. In conclusion, we successfully developed a microelectrode array-based hybrid bioelectronic measurement platform for quantitative monitoring of pathologic effects of a herpesvirus on electrophysiological active neuronal networks.