Intranasal administration with recombinant vaccine PRVXJ-delgE/gI/TK-S induces strong intestinal mucosal immune responses against PDCoV.

Porcine deltacoronavirus (PDCoV) is a novel coronavirus that causes enteric diseases in pigs leading to substantial financial losses within the industry. The absence of commercial vaccines and limited research on PDCoV vaccines presents significant challenges. Therefore, we evaluated the safety and immunogenicity of recombinant pseudorabies virus (PRV) rPRVXJ-delgE/gI/TK-S through intranasal mucosal immunization in weaned piglets and SPF mice. Results indicated that rPRVXJ-delgE/gI/TK-S safely induced PDCoV S-specific and PRV gB-specific antibodies in piglets, with levels increasing 7 days after immunization. Virus challenge tests demonstrated that rPRVXJ-delgE/gI/TK-S effectively improved piglet survival rates, reduced virus shedding, and alleviated clinical symptoms and pathological damage. Notably, the recombinant virus reduced anti-inflammatory and pro-inflammatory responses by regulating IFN-γ, TNF-α, and IL-1ß secretion after infection. Additionally, rPRVXJ-delgE/gI/TK-S colonized target intestinal segments infected with PDCoV, stimulated the secretion of cytokines by MLVS in mice, stimulated sIgA secretion in different intestinal segments of mice, and improved mucosal immune function. HE and AB/PAS staining confirmed a more complete intestinal mucosal barrier and a significant increase in goblet cell numbers after immunization. In conclusion, rPRVXJ-delgE/gI/TK-S exhibits good immunogenicity and safety in mice and piglets, making it a promising candidate vaccine for PDCoV.

Macrophage Polarization: An Important Candidate Regulator for Lung Diseases.

Macrophages are crucial components of the immune system and play a critical role in the initial defense against pathogens. They are highly heterogeneous and plastic and can be polarized into classically activated macrophages (M1) or selectively activated macrophages (M2) in response to local microenvironments. Macrophage polarization involves the regulation of multiple signaling pathways and transcription factors. Here, we focused on the origin of macrophages, the phenotype and polarization of macrophages, as well as the signaling pathways associated with macrophage polarization. We also highlighted the role of macrophage polarization in lung diseases. We intend to enhance the understanding of the functions and immunomodulatory features of macrophages. Based on our review, we believe that targeting macrophage phenotypes is a viable and promising strategy for treating lung diseases.

Layered Cu1-zMn1+zO2 Crednerite: Mapping the Phase Stabilization Region via Precise Compositional Control for Optimum Supercapacitor Performance.

CuMnO2 is a prototype ABO2-type crednerite compound featured by transition metal ions of variable valence states essential for creating novel properties and optimum performance. However, the phase stabilization region of CuMnO2 has not yet been well established, restricting one's ability in comprehending this unique structure for functional applications. Here, layered Cu1-zMn1+zO2 crednerite was systematically synthesized and characterized by accurately regulating the reaction parameters of hydrothermal conditions, which led to a first demonstration of the phase diagram for CuMnO2 crednerite. The pure phase layered structure was uncovered to be stabilized under hydrothermal conditions as the temperature varies between 85 and 175 °C and the molar ratio of Cu to (Cu + Mn) varies between 0.45 and 0.55. For Cu1-zMn1+zO2, there appeared non-stoichiometric occupation of transition metal ions. Strikingly, different from many other layered oxides, the samples at a molar ratio of Cu:(Cu + Mn) = 0.55 showed a special structure, in which excess Cu2+ occupied the position of the Mn3+ site to form a Cu2+ (3d9)/Mn4+ (3d3) ionic pair and traces of corresponding cationic ordered phases. Such a configuration (3d9/3d3 ionic pair) gives rise to an optimum super-capacitor performance, as represented by a highest mass specific capacitance of 428.4 F/g at a current density of 1 A/g. The strategy reported in this work for mapping the phase diagram of layered CuMnO2 crednerite is fundamentally important, which may offer guidance to explore the potentials of other ABO2-type compounds for functional applications.

Characterization of B cell receptor H-CDR3 repertoire of spleen in PRV-infected mice.

Pseudorabies virus (PRV), also known as suid Alphaherpesvirus 1 (SuHV-1), which is one of the most devastating infectious pathogen of swine industry worldwide. Vaccination is the safest and most effective PRV prevention and control strategy. B cell receptor (BCR) is membrane-bound immunoglobulin located on the surface of B cells capable of specifically binding foreign antigens, which is one of the most important molecules regulating the proliferation and function of B cells. Here, to assess the molecular diversity of BCR H-CDR3 repertoire after different PRV strains infection, we detected the IGHV, IGHD, IGHJ genes usage and CDR3 sequence changes of mice spleen with PRV vaccine strain (Bartha-K61), variant strain (XJ) and mock infection by high-throughput sequencing. We found that PRV-infected groups shared partial BCR sequences, which are most likely to be PRV-specific BCR candidates. However, there were still differences in the IGHV genes usage as well as the combined usage of IGHV and IGHJ genes between the Bartha-K61 strain and XJ strain infection groups. In addition, the CDR3 sequences exhibited large differences in the types and lengths in PRV infection groups. Our study contributes to a better understanding of the host adaptive immune response to PRV infection and provides a theoretical basis for further research on novel and efficient PRV vaccines in the future.

Antiviral Activity of Porcine IFN-λ3 and IFN-α against Porcine Rotavirus In Vitro.

Interferons (IFNs) play a major role in the host's antiviral innate immunity. In response to viral infection, IFNs bind their receptors and initiate a signaling cascade, leading to the accurate transcriptional regulation of hundreds of IFN-stimulated genes (ISGs). Porcine rotavirus (PoRV) belongs to genus Rotavirus of the Reoviridae family; the infection is a global epidemic disease and a major threat to the pig industry. In this study, we found that IFN-λ3 inhibited the replication of PoRV in both MA104 cells and IPEC-J2 cells, and this inhibition was dose-dependent. Furthermore, the antiviral activity of IFN-λ3 was more potent in IPEC-J2 cells than in MA104 cells. Further research showed that IFN-λ3 and IFN-α might inhibit PoRV infection by activating ISGs, i.e., MxA, OASL and ISG15, in IPEC-J2 cells. However, the co-treatment of IFN-λ3 and IFN-α did not enhance the antiviral activity. Our data demonstrated that IFN-λ3 had antiviral activity against PoRV and may serve as a useful antiviral candidate against PoRV, as well as other viruses in swine.

Cinnamaldehyde Resist Salmonella Typhimurium Adhesion by Inhibiting Type I Fimbriae.

Salmonella Typhimurium (S. Typhimurium), a common foodborne pathogen, severely harms the public and food security. Type I fimbriae (T1F) of S. Typhimurium, plays a crucial role in the pathogenic processes; it mediates the adhesion of bacteria to the mannose receptor on the host cell, assists the bacteria to invade the host cell, and triggers an inflammatory response. Cinnamaldehyde is the main ingredient in cinnamon essential oil. In this study, cinnamaldehyde was demonstrated to inhibit the expression of T1F by hemagglutination inhibition test, transmission electron microscopy, and biofilms. The mechanism of cinnamaldehyde action was studied by proteomics technology, PCR and Western blotting. The results showed that cinnamaldehyde can inhibit T1F in S. typhimurium without the growth of bacteria, by regulating the level of expression and transcription of fimA, fimZ, fimY, fimH and fimW. Proteomics results showed that cinnamaldehyde downregulated the subunits and regulators of T1F. In addition, the invasion assays proved that cinnamaldehyde can indeed reduce the ability of S. typhimurium to adhere to cells. The results of animal experiments showed that the colonization in the intestinal tract and the expression levels of inflammatory cytokine were significantly decreased, and the intestinal mucosal immune factors MUC1 and MUC2 were increased under cinnamaldehyde treatment. Therefore, cinnamaldehyde may be a potential drug to target T1F to treat Salmonella infections.

The Antiviral Activity of Caprylic Monoglyceride against Porcine Reproductive and Respiratory Syndrome Virus In Vitro and In Vivo.

Porcine reproductive and respiratory syndrome (PRRS) is a disease with a major economic impact in the global pig industry, and this study aims to identify potential anti-PRRSV drugs. We examined the cytotoxicity of four medium-chain fatty acids (MCFAs) (caprylic, caprylic monoglyceride, decanoic monoglyceride, and monolaurin) and their inhibition rate against PRRSV. Then the MCFAs with the best anti-PRRSV effect in in vitro assays were selected for subsequent in vivo experiments. Potential anti-PRRSV drugs were evaluated by viral load assay, pathological assay, and cytokine level determination. The results showed that caprylic monoglyceride (CMG) was the least toxic to cells of the four MCFAs, while it had the highest PRRSV inhibition rate. Then the animals were divided into a low-CMG group, a medium-CMG group, and a high-CMG group to conduct the in vivo evaluation. The results indicated that piglets treated with higher concentrations of caprylic monoglyceride were associated with lower mortality and lower viral load after PRRSV infection (p < 0.05). The pulmonary pathology of the piglets also improved after CMG treatment. The levels of pro-inflammatory cytokines (IL-6, IL-8, IL-1ß, IFN-γ, TNF-α) were significantly downregulated, and the levels of anti-inflammatory cytokine (IL-10) were significantly upregulated in the CMG-treated piglets compared to the positive control group (p < 0.05). Taken together, the present study revealed for the first time that caprylic monoglyceride has strong antiviral activity against PRRSV in vitro and in vivo, suggesting that caprylic monoglyceride could potentially be used as a drug to treat PRRS infection.

Infectivity and pathogenesis characterization of getah virus (GETV) strain via different inoculation routes in mice.

In recent years, the epidemiological profile of Getah virus (GETV) has become increasingly serious, posing a huge threat to animal and public health in China. GETV can cause multi-species infection, including horses, pigs, rats, cattle, kangaroos, reptiles and birds. However, there were few reports on the efficiency of the virus entering the host via routes of different systems. In the present study, a GETV strain (SC201807) was obtained from a piglet's blood in 2018 in Sichuan, China. First, we established a quantitative real-time polymerase chain reaction (qRT-PCR) SYBR assay specific to GETV. Then, we evaluated the infection efficiency of different routes using mouse animal model. 108 male mice were randomly divided into four groups as follows: intramuscular, intraoral and intranasal infection routes, and negative control. All mice in the experimental group were inoculated with 4 × 102.85 TCID50 GETV virus. Tissue tropism experiments show that GETV has a wide range of tissue distribution, and intramuscular infection is the first to infect all tissues of the body, and suggest that oral infection may be a new GETV transmission route. Histopathological examination results showed that intramuscular injection of GETV mainly caused different degrees of pathological damage to the tissues, and could rapidly induce a large amount of inflammatory regulatory factors such as IL-6 and TNF-α. Our data may help us to evaluate the risk of transmission of Porcine Getah virus and provide an experimental basis for the prevention and control of Porcine Getah virus.

Isolation and pathogenicity comparison of two novel natural recombinant porcine reproductive and respiratory syndrome viruses with different recombination patterns in Southwest China.

Porcine reproductive and respiratory syndrome virus (PRRSV) causes significant economic losses in the swine industry. Frequent mutations and recombinations account for PRRSV immune evasion and the emergence of novel strains. In this study, we isolated and characterized two novel PRRSV-2 strains from Southwest China exhibiting distinct recombination patterns. They were designated SCABTC-202305 and SCABTC-202309. Phylogenetic results indicated that SCABTC-202305 was classified as lineage 8, and SCABTC-202309 was classified as lineage 1.8. Amino acid mutation analysis identified unique amino acid substitutions and deletions in ORF5 and Nsp2 genes. The results of the recombination analysis revealed that SCABTC-202305 is a recombinant with JXA1 as the major parental strain and NADC30 as the minor parental strain. At the same time, SCABTC-202309 is identified as a recombinant with NADC30 as the major parental strain and JXA1 as the minor parental strain. In this study, we infected piglets with SCABTC-202305, SCABTC-202309, or mock inoculum (control) to study the pathogenicity of these isolates. Although both isolated strains were pathogenic, SCABTC-202305-infected piglets exhibited more severe clinical signs and higher mortality, viral load, and antibody response than SCABTC-202309-infected piglets. SCABTC-202305 also caused more extensive lung lesions based on histopathology. Our findings suggest that the divergent pathogenicity observed between the two novel PRRSV isolates may be attributed to variations in the genetic information encoded by specific genomic regions. Elucidating the genetic determinants governing PRRSV virulence and transmissibility will inform efforts to control this devastating swine pathogen.IMPORTANCEPorcine reproductive and respiratory syndrome virus (PRRSV) is one of the most critical pathogens impacting the global swine industry. Frequent mutations and recombinations have made the control of PRRSV increasingly difficult. Following the NADC30-like PRRSV pandemic, recombination events involving PRRSV strains have further increased. We isolated two novel field PRRSV recombinant strains, SCABTC-202305 and SCABTC-202309, exhibiting different recombination patterns and compared their pathogenicity in animal experiments. The isolates caused higher viral loads, persistent fever, marked weight loss, moderate respiratory clinical signs, and severe histopathologic lung lesions in piglets. Elucidating correlations between recombinant regions and pathogenicity in these isolates can inform epidemiologic tracking of emerging strains and investigations into viral adaptive mechanisms underlying PRRSV immunity evasion. Our findings underscore the importance of continued genomic surveillance to curb this economically damaging pathogen.

Octahedral Nanocrystals of Ru-Doped PtFeNiCuW/CNTs High-Entropy Alloy: High Performance Toward pH-Universal Hydrogen Evolution Reaction.

Integrating high-entropy philosophy and nanocrystal-specific orientation into a single catalyst represents a promising strategy in development of high-performance catalysts. Nonetheless, shape-controlled synthesis of high-entropy alloy (HEA) nanocrystals is challenging owing to the distinct redox potentials and growth dynamics of metal elements. Herein, a one-pot co-reduction method is developed to fabricate ruthenium (Ru)-doped PtFeNiCuW octahedral HEA nanocrystals onto carbon nanotubes (Ru-PtFeNiCuW/CNTs). It is demonstrated that Ru dopants and W(CO)6 promote the concurrent reduction and growth of other metal precursors to obtain higher yield and larger size of HEA nanocrystals, despite low Ru content in Ru-PtFeNiCuW/CNTs. As an electrocatalyst toward hydrogen evolution reaction (HER), Ru-PtFeNiCuW/CNTs exhibits low overpotentials of 9, 16, and 34 mV at a current density of 10 mA cm-2 and Tafel slopes of 19.2, 27.9, and 23.1 mV dec-1 in acidic, alkaline, and neutral electrolytes, respectively. As a cathodic catalyst, Ru-PtFeNiCuW/CNTs operates for up to 1500 and 1200 h in acidic and alkaline electrolyte, respectively, at a current density of 50 mA cm-2 in a two-electrode system for full water splitting. Theoretical calculations reveal accelerated kinetics of H2O dissociation on W sites and *H desorption on hollow Cu-Cu-Cu and Cu-Cu-Pt sites.

Rescue and characterization of PCV4 infectious clones: pathogenesis and immune response in piglets.

Porcine circovirus 4 (PCV4) was first identified in 2019, categorized within the genus Circovirus in the family Circoviridae. To date, the virus has not been isolated from clinical samples. Meanwhile, many aspects of the biology and pathogenic mechanisms of PCV4 infection remain unknown. In this study, PCV4 was successfully rescued from an infectious clone. We utilized a PCV4 virus stock derived from this infectious clone to intranasally inoculate 4-week-old specific-pathogen-free piglets to evaluate PCV4 pathogenesis. The rescued PCV4 was capable of replicating in both PK-15 cells and piglets, with the virus detectable in nearly all collected samples from the challenge groups. Pathological lesions and PCV4-specific antigens were observed in various tissues and organs, including the lungs, kidneys, lymph nodes, spleen, and liver, in the inoculated piglets. Additionally, the levels of pro-inflammatory cytokines in the serum of the PCV4-inoculated group were significantly elevated compared to the control group, indicating that the induced inflammatory response may contribute to tissue damage associated with PCV4 infection. These findings offer new insights into the pathogenesis and inflammatory responses associated with PCV4-related diseases.

The construction and immunogenicity analyses of a recombinant pseudorabies virus with Senecavirus A VP3 protein co-expression.

Senecavirus A (SVA)-associated porcine idiopathic vesicular disease (PIVD) and Pseudorabies (PR) are highly contagious swine disease that pose a significant threat to the global pig industry. In the absence of an effective commercial vaccine, outbreaks caused by SVA have occurred in many parts of the world. In this study, the PRV variant strain PRV-XJ was used as the parental strain to construct a recombinant PRV strain with the TK/gE/gI proteins deletion and the VP3 protein co-expression, named rPRV-XJ-ΔTK/gE/gI-VP3. The results revealed that PRV is a suitable viral live vector for VP3 protein expressing. As a vaccine, rPRV-XJ-ΔTK/gE/gI-VP3 is safe for mice, vaccination with it did not cause any clinical symptoms of PRV. Intranasal immunization with rPRV-XJ-ΔTK/gE/gI-VP3 induced strong cellular immune response and high levels of specific antibody against VP3 and gB and neutralizing antibodies against both PRV and SVA in mice. It provided 100% protection to mice against the challenge of virulent strain PRV-XJ, and alleviated the pathological lesion of heart and liver tissue in SVA infected mice. rPRV-XJ-ΔTK/gE/gI-VP3 appears to be a promising vaccine candidate against PRV and SVA for the control of the PRV variant and SVA.

Construction of recombinant pseudorabies virus expressing PCV2 Cap, PCV3 Cap, and IL-4: investigation of their biological characteristics and immunogenicity.

Background: Porcine circovirus type 2 (PCV2) is a globally prevalent and recurrent pathogen that primarily causes slow growth and immunosuppression in pigs. Porcine circovirus type 3 (PCV3), a recently discovered virus, commonly leads to reproductive disorders in pigs and has been extensively disseminated worldwide. Infection with a single PCV subtype alone does not induce severe porcine circovirus-associated diseases (PCVD), whereas concurrent co-infection with PCV2 and PCV3 exacerbates the clinical manifestations. Pseudorabies (PR), a highly contagious disease in pigs, pose a significant threat to the swine industry in China. Methods: In this study, recombinant strains named rPRV-2Cap/3Cap and rPRV-2Cap/3Cap/IL4 was constructed by using a variant strain XJ of pseudorabies virus (PRV) as the parental strain, with the TK/gE/gI genes deleted and simultaneous expression of PCV2 Cap, PCV3 Cap, and IL-4. The two recombinant strains obtained by CRISPR/Cas gE gene editing technology and homologous recombination technology has genetic stability in baby hamster Syrian kidney-21 (BHK-21) cells and is safe to mice. Results: rPRV-2Cap/3Cap and rPRV-2Cap/3Cap/IL4 exhibited good safety and immunogenicity in mice, inducing high levels of antibodies, demonstrated 100% protection against the PRV challenge in mice, reduced viral loads and mitigated pathological changes in the heart, lungs, spleen, and lymph nodes during PCV2 challenge. Moreover, the recombinant viruses with the addition of IL-4 as a molecular adjuvant outperformed the non-addition group in most indicators. Conclusion: rPRV-2Cap/3Cap and rPRV-2Cap/3Cap/IL4 hold promise as recombinant vaccines for the simultaneous prevention of PCV2, PCV3, and PRV, while IL-4, as a vaccine molecular adjuvant, effectively enhances the immune response of the vaccine.

Porcine reproductive and respiratory syndrome virus infects the reproductive system of male piglets and impairs development of the blood-testis barrier.

Porcine reproductive and respiratory syndrome virus (PRRSV) causes a highly contagious disease that threatens the global swine industry. Recent studies have focused on the damage that PRRSV causes to the reproductive system of male pigs, although pathological research is lacking. Therefore, we examined the pathogenic mechanisms in male piglets infected with PRRSV. Gross and histopathological changes indicated that PRRSV affected the entire reproductive system, as confirmed via immunohistochemical analysis. PRRSV infected Sertoli cells and spermatogonia. To test the new hypothesis that PRRSV infection in piglets impairs blood - testis barrier (BTB) development, we investigated the pathology of PRRSV damage in the BTB. PRRSV infection significantly decreased the quantity and proliferative capacity of Sertoli cells constituting the BTB. Zonula occludens-1 and ß-catenin were downregulated in cell - cell junctions. Transcriptome analysis revealed that several crucial genes and signalling pathways involved in the growth and development of Leydig cells, Sertoli cells, and tight junctions in the testes were downregulated. Apoptosis, necroptosis, inflammatory, and oxidative stress-related pathways were activated, whereas hormone secretion-related pathways were inhibited. Many Sertoli cells and spermatogonia underwent apoptosis during early differentiation. Infected piglets exhibited disrupted androgen secretion, leading to significantly reduced testosterone and anti-Müllerian hormone levels. A cytokine storm occurred, notably upregulating cytokines such as tumour necrosis factor-α and interleukin-6. Markers of oxidative-stress damage (i.e. H2O2, malondialdehyde, and glutathione) were upregulated, whereas antioxidant-enzyme activities (i.e. superoxide dismutase, total antioxidant capacity, and catalase) were downregulated. Our results demonstrated that PRRSV infected multiple organs in the male reproductive system, which impaired growth in the BTB.

Establishment and application of an indirect ELISA for Getah virus E2 antibody detection.

Getah virus (GETV) is a mosquito-transmitted disease that affects animals, causing fever, aseptic meningitis, and abortion. Its prevalence in China poses risks to both animal health and public well-being. Currently, there is a scarcity of seroepidemiological data on GETV due to the absence of commercial antibody detection kits for pigs. The aim of this study is to develop a rapid, accurate, and sensitive ELISA, providing a reliable tool for GETV seroepidemiology and laying the foundation for future commercial assay development. In this study, we removed specific hydrophobic domains and intracellular structures from E2 proteins and constructed the recombinant plasmid pCold-TF-E2. The recombinant protein was expressed using a prokaryotic expression system, and efficient purification of the rE2 protein was achieved using a nickel affinity column. The purified rE2 protein is suitable for the development of an indirect ELISA (rE2 ELISA). Following the optimization of reaction conditions for the rE2-ELISA, the cut-off value was 0.356. Additionally, the rE2-ELISA method showed a positive rate of 37.1% for IgG antibodies against GETV when testing 986 pig clinical serum samples collected from pigs in Sichuan between May 2022 and September 2022. The rE2-ELISA method displayed a 95.1% overall agreement with VNT, boasting a sensitivity of 98.2% and a specificity of 92.6%. These results indicate that IgG ELISA based on rE2 protein is an efficient and economical method for the detection of GETV antibodies in pigs, facilitating the diagnosis and prevention of GETV.

Transcriptome and metabolome analysis reveals PRV XJ delgE/gI/TK protects intracranially infected mice from death by regulating the inflammation.

Pseudorabies virus can cause inflammation in the central nervous system and neurological symptoms. To further investigate the protective mechanism of PRV XJ delgE/gI/TK in the central nervous system, an intracranial PRV-infection mice model was developed. The results demonstrated that immunization with PRV XJ delgE/gI/TK successfully prevented death caused by PRV-intracranial infection. Subsequently, the brains were collected for transcriptome and metabolome analysis. GO and KEGG enrichment analysis indicated that the differentially expressed genes were primarily enriched in pathways such as TNF, NOD-like receptor, JAK-STAT, MAPK, IL-17 and apoptosis signaling. Metabolomics analysis revealed that the differential metabolites were mainly associated with pathways such as fatty acid degradation, arachidonic acid metabolism, linoleic acid metabolism and unsaturated fatty acid biosynthesis. The combined analysis of metabolites and differentially expressed genes revealed a strong correlation between the differential metabolites and TNF, PI3K, and MAPK signaling pathways. Anti-inflammatory metabolites have been shown to inhibit the inflammatory response and prevent mouse death caused by PRV infection. Notably, when glutathione was injected intracranially and dihydroartemisinin was injected intraperitoneally, complete protection against PRV-induced death in mice was observed. Moreover, PRV activates the PI3K/AKT signaling pathway. In conclusion, our study demonstrates that PRV XJ delgE/gI/TK can protects intracranially infected mice from death by regulating various metabolites with anti-inflammatory functions post-immunization.

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.

Rapid, sensitive, and visual detection of swine Japanese encephalitis virus with a one-pot RPA-CRISPR/EsCas13d-based dual readout portable platform.

Japanese encephalitis (JE), a mosquito-borne zoonotic disease caused by the Japanese encephalitis virus (JEV), poses a serious threat to global public health. The low viremia levels typical in JEV infections make RNA detection challenging, necessitating early and rapid diagnostic methods for effective control and prevention. This study introduces a novel one-pot detection method that combines recombinant enzyme polymerase isothermal amplification (RPA) with CRISPR/EsCas13d targeting, providing visual fluorescence and lateral flow assay (LFA) results. Our portable one-pot RPA-EsCas13d platform can detect as few as two copies of JEV nucleic acid within 1 h, without cross-reactivity with other pathogens. Validation against clinical samples showed 100 % concordance with real-time PCR results, underscoring the method's simplicity, sensitivity, and specificity. This efficacy confirms the platform's suitability as a novel point-of-care testing (POCT) solution for detecting and monitoring the JE virus in clinical and vector samples, especially valuable in remote and resource-limited settings.

Histopathological Changes and Inflammatory Response in Specific Pathogen-Free (SPF) with Porcine Circovirus Type 3 Infection.

Since the first report of PCV3 virus infection in 2016, it has been linked to multisystemic inflammation, reproductive failure, cardiac pathology, and clinical indications resembling porcine dermatitis and nephropathy syndrome (PDNS). However, the pathogenesis and clinical significance of PCV3 is still unclear. In this study, a PCV3 infection model was created using SPF pigs, and histopathology and fluorescence quantitative PCR were utilized to examine PCV3's pathogenicity. Reductions in body weight gain and fever were observed during this study. However, other clinical signs such as Dermatitis and Nephropathy Syndrome were not observed through the study. Viremia was detected in the PCV3-inoculated group from 17 days post-inoculation (p.i.) until the end of the study. Nasal shedding was detected from 21 to 35 dpi and fecal shedding was detected during 25-33 days and 39 days, respectively. Gross lesions and histological evaluation were detected in various tissues and organs, including the lung, heart, kidney, lymph nodes, spleen, liver, small intestine, and testis. The heart, lung, liver, kidney, lymph nodes, and spleen showed pathological changes. The pathological features include swelling, inflammation, cell degeneration, necrosis, and hemorrhage. The lesions are consistent with multisystemic inflammation. Tissue viral load results showed only heart, lung, liver, kidney, lymph nodes, and spleen was positive by qRT-PCR. Moreover, the pro-inflammation cytokines in serum increased a lot in the PCV3-inoculated group compared to the control group, demonstrating that the induced inflammation response may be the cause of tissue damage in PCV3-infection. This study demonstrated that PCV3 can produce mild pathological damage to multiple organs, especially multisystemic inflammatory cell infiltration and prolonged viremia, viral shedding in nasal secretions. This is the first in vivo characterization of PCV3 infection in the SPF piglets model using isolated PCV3 strain, and this is also the first time to show the gross and pathological lesion with all tissue and organs in the PCV3-inoculated group. Our findings might serve as a starting point for more investigation into PCV3's pathogenic mechanism.

The first dog-origin porcine circovirus type 4 complete genomic sequence have high homology with that of pig-derived strains.

Introduction: Porcine circovirus 4 (PCV4) was discovered in 2019 and then proved to be pathogenic to piglets. Nevertheless, few studies were currently available about PCV4 infection in species other than pigs and there is no information about the prevalence of PCV4 in dogs. Methods: To fill this gap, 264 dog samples were collected from animal hospitals in the Southwest of China from 2021 to 2022 and screened for PCV4. Moreover, the complete genome of one PCV4 strain (SCABTC-Dog2022) were obtained successfully and shared a high identity (97.9-99.0%) with other PCV4 strains derived from pigs, dairy cows, raccoon dogs and foxes. The SCABTC-Dog2022 were analyzed together with 51 reference sequences. Results and Discussion: The detected results showed a low percentage of PCV-4 DNA (1.14%, 3/264), indicating that PCV4 could be identified in dogs in southwest China. Phylogenetic tree showed that SCABTC-Dog2022 strain derived from dog were clustered in a closed relative and geographically coherent branch with other PCV4 strains collected from four provinces (Sichuan, Fujian, Hunan and Inner Mongolia) of China. To our knowledge, it is the first detection of PCV4 in dogs globally. The association between PCV4 status and clinical syndromes in dogs deserves additional investigations.