Bovine Parainfluenza Virus Type 3 (BPIV3) is a critical pathogen in the Bovine Respiratory Disease Complex (BRDC), leading to significant economic losses in the cattle industry. However, the metabolic reprogramming induced by BPIV3 in cattle remains poorly understood. This study aimed to investigate the impact of BPIV3 infection on the serum metabolome of Simmental cattle using untargeted metabolomics and ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS/MS). The results revealed significant alterations in the lipidome, including the upregulation of phosphatidylcholine (PC) and phosphatidylglycerol (PG), and the downregulation of phosphatidylinositol (PI). Sphingolipid metabolism also showed considerable changes, with increased levels of Trihexosylceramide and D-erythro-Sphingosine C-17. Furthermore, metabolic pathway analysis identified enriched pathways related to lipid metabolism, amino acid metabolism, and energy sensing. These findings suggest that BPIV3 infection induces substantial shifts in lipid metabolism, which may facilitate viral replication and immune evasion. Our results provide a deeper understanding of the metabolic changes in BPIV3-infected cattle and propose potential targets for therapeutic intervention.

This study presents the first molecular epidemiological investigation and genomic characterization of bovine kobuvirus (BKoV) in eastern Inner Mongolia, China. A total of 162 clinical samples were collected in July 2023 and tested for BKoV using nested PCR, yielding an overall detection rate of 18.52%. Differences in BKoV detection rates were observed among sample types, with a significantly higher detection rate in faecal samples than in blood samples. Metagenomic sequencing generated a near-complete genome sequence of strain NM21, which has been submitted to the GenBank database under accession number PV797393. Phylogenetic analyses based on the partial 3D gene sequences and the near-complete genome sequence of strain NM21 showed that all BKoV sequences detected in this study clustered within Clade 1. Amino acid sequence alignment revealed 21 clade-associated amino acid differences that were relatively conserved within each clade; among them, position 66 of the VP0 protein showed a consistent difference between the two clades, with the corresponding amino acid absent in Clade 2 sequences. Bioinformatic analysis of the major capsid protein VP1 predicted three candidate linear B-cell epitopes. Selection pressure analysis showed that the VP1 gene was generally under purifying selection. Recombination analysis suggested that strain NM21 may contain a potential recombination signal in the 2B non-structural protein region. These findings provide baseline data for elucidating the epidemiological characteristics, genetic evolution, and potential antigenic features of BKoV in eastern Inner Mongolia. They may serve as a reference for subsequent molecular surveillance and evolutionary studies.

The early life gut microbial colonization in pre-weaning calves plays a pivotal role in shaping their health, growth, and productivity. This review delves into the dynamic changes of intestinal microbiota during early life, emphasizing key factors such as colostrum management, feeding strategies, roughage supplementation, and microbial interventions including probiotics, prebiotics, and fecal microbiota transplantation (FMT), and non-nutritional stressors that can shape the early life microbial colonization. We highlight the microbiota's critical functions in nutrient metabolism, immune development, gut barrier integrity, and gut-brain axis regulation. Additionally, the consequences of microbial dysbiosis on calf health and its long-term effects on production performance in beef and dairy cattle are discussed. While current research has provided valuable insights, understanding causal mechanisms remains a challenge. This review aims to guide practical strategies for targeted microbial management, offering a pathway to optimize early-life interventions for improved calf health and productivity.

Hybrid capture-based target enrichment prior to sequencing has been shown to significantly improve the sensitivity of detection for genetic regions of interest. In the context of One Health relevant pathogen detection, we present a hybrid capture-based sequencing method that employs an optimized probe set consisting of 149,990 probes, targeting 663 viruses associated with humans and animals. The detection performance was initially assessed using viral reference materials in a background of human nucleic acids. Compared to standard metagenomic next-generation sequencing (mNGS), our method achieved substantial read enrichment, with increases ranging from 143- to 1126-fold, and enhanced detection sensitivity by lowering the limit of detection (LoD) from 103–104 copies to as few as 10 copies based on whole genomes. This method was further validated using infectious samples from both animals and humans, including bovine rectal swabs and throat swabs from SARS-CoV-2 patients across various concentration gradients. In both sample types, our hybrid capture-based sequencing method exhibited heightened sensitivity, increased viral genome coverage, and more comprehensive viral identification and characterization. Our method bridges a critical divide between diagnostic detection and genomic surveillance. These findings illustrate that our hybrid capture-based sequencing method can effectively enhance sensitivity to as few as 10 viral copies and genome coverage to >99% in medium-to-high viral loads. This dual capability is particularly impactful for emerging pathogens like SARS-CoV-2, where early detection and genomic characterization are equally vital, thereby addressing the limitations of metagenomics in the surveillance of emerging infectious diseases in complex samples.

Dear Editor, Tusavirus(TuV),a recently identified member of the genus Proto-parvovirus,was first reported in a Tunisian child with diarrhea in 2014(Phan et al.,2014).Since then,TuV has been sporadically detected in human and animal fecal samples,raising concerns about its zoonotic potential(Reuter et al.,2022;He et al.,2023;Davies et al.,2024).Here,we report the first identification of a TuV strain(GenBank accession number:PP083003)in intestinal tissue from a diarrheic calf in eastern Inner Mongolia,China,and present genomic analysis supporting possible interspecies transmission.

BACKGROUND:Canine distemper virus (CDV) is a pathogen with the capability of cross-species transmission. It has crossed the species barrier to infect many other species, and its host range is expanding. The reverse genetic platform, a useful tool for scientific research, allows the generation of recombinant viruses from genomic cDNA clones in vitro. METHODS:To improve the reverse genetic system of CDV, a plasmid containing three independent expression cassettes was constructed for co-expression of the N, P, and L genes and then transfected with a full-length cDNA clone of CDV into Vero cells. RESULTS:The results indicated that the established rescue system has the advantages of being more convenient, easy to control the transfection ratio, and high rescue efficiency compared with the conventional reverse genetics system. CONCLUSION:This method not only reduces the number of transfection plasmids, but also improves the rescue efficiency of CDV, which could provide a reference for the recovery of other morbilliviruses.

IntroductionPorcine Parvovirus (PPV) is a significant pathogen in the pig industry, with eight genotypes, including PPV7, identified since its emergence in 2016. Co-infections with viruses such as Porcine Circovirus 2 (PCV2) and Porcine Reproductive and Respiratory Syndrome Virus (PRRSV) pose serious risks to swine health. Thus, there is an urgent need for rapid, sensitive, and specific detection methods suitable for use in field settings or laboratories with limited resources.MethodsWe developed a CRISPR/Cas12a-based assay combined with recombinase polymerase amplification (RPA) for the rapid detection of PPV7. Specific RPA primers and five CRISPR RNAs (crRNAs) were designed to target a highly conserved region within the NS1 gene of PPV7. Optimization of crRNA and single-stranded DNA (ssDNA) concentrations was performed to enhance the assay’s performance.ResultsCrRNA optimization identified crRNA-05 as the optimal candidate for Cas12a-based detection of PPV7, as all synthesized crRNAs demonstrated similar performance. The optimal crRNA concentration was determined to be 200 nM, yielding consistent results across tested concentrations. For ssDNA optimization, the strongest fluorescence signal was achieved with 500 nM of the FAM-BHQ ssDNA receptor. The assay showed a minimal detection limit of 100copies/μl for PPV7, confirmed through fluorescence and lateral flow detection methods. Specificity testing indicated that only PPV7 DNA samples returned positive results, confirming the assay’s accuracy. In tests of 50 lung tissue samples from diseased pigs, the RPA-Cas12a assay identified 29 positive samples (58%), surpassing the 22 positive samples (44%) detected by conventional PCR. This highlights the RPA-Cas12a method’s enhanced detection capability and its potential utility in clinical surveillance and management of PPV7 in swine populations.DiscussionThe RPA-Cas12a assay effectively detects PPV7 in clinical samples, enhancing disease surveillance and control in pigs. Its adaptability to resource-limited settings significantly improves PPV7 management and prevention strategies, thereby supporting the overall health and development of the pig industry.

Abstract Background Porcine Parvovirus (PPV) are small, enveloped viruses with single stranded genomic DNA. Till now seven genotypes of PPV have been detected worldwide. They are PPV1 to PPV7 with later was first discovered in 2016 in America and then in Asia and European. It has been reported that PPV7 was a co-infector with Porcine Circovirus 2 (PCV2), PCV3 and Porcine reproductive and respiratory syndrome virus (PRRSV). A rapid, sensitive and specific PPV7 detection method that could be applied in poorly equipped laboratory or event in field could be helpful to reveal its distribution and control the spread of this virus. CRISPR/Cas based systems have exhibited outstanding capacities in the detection of pathogenic microorganisms due to the trans-cleavage activities of the Cas proteins.Results Herein, we established a recombinase polymerase amplification (RPA)-CRISPR/Cas12a based rapid viral detection assay for PPV7. Specific RPA primers and five CRISPR RNAs (crRNAs) were designed and synthesized based on the highly conserved region within the NS1 gene of PPV7. The concentration of crRNA and ssDNA were further optimized. Furthermore, we evaluated the sensitivity, specificity, and clinical effectiveness of the RPA-Cas12a based detection assay. The results indicated that this method could be applied for real-time detection. The detection sensitivity of the novel assay was 100 copies/µl, and there were no cross-reactions with other genotypes of PPV, PCV2, PCV3, PRRSV and pseudorabies virus. The RPA-Cas12a based assay could work well in the detection of clinical samples.Conclusions In summary, we developed a visual, sensitive and specific viral diagnostic method based on CRISPR-Cas12a system for PPV7.

Interferon (IFN), the most effective antiviral cytokine, is involved in innate and adaptive immune responses and is essential to the host defense against virus invasion. Once the host was infected by pathogens, the pathogen-associated molecular patterns (PAMPs) were recognized by the host pattern recognition receptors (PRRs), which activates interferon regulatory transcription factors (IRFs) and nuclear factor-kappa B (NF-κB) signal transduction pathway to induce IFN expression. Pathogens have acquired many strategies to escape the IFN-mediated antiviral immune response. Pestiviruses cause massive economic losses in the livestock industry worldwide every year. The immune escape strategies acquired by pestiviruses during evolution are among the major difficulties in its control. Previous experiments indicated that Erns, as an envelope glycoprotein unique to pestiviruses with RNase activity, could cleave viral ss- and dsRNAs, therefore inhibiting the host IFN production induced by viral ss- and dsRNAs. In contrast, Npro, the other envelope glycoprotein unique to pestiviruses, mainly stimulates the degradation of transcription factor IRF-3 to confront the IFN response. This review mainly summarized the current progress on mechanisms mediated by Npro of pestiviruses to antagonize IFN production.

目的 对猪外周血淋巴细胞(peripheral blood lymphocytes,PBMCs)干扰素诱导跨膜蛋白(IFN-induced transme-mbrane,IFITM)进行拓扑学分析,并检测PBMCs在体外感染猪繁殖与呼吸综合征病毒(porcine reproductive and respi-ratory syndrome virus,PRRSV)后IFITM基因mRNA转录水平的变化.方法 无菌采集PRRSV、猪圆环病毒2(porcine circovirus 2,PCV2)和乙型脑炎病毒(Japanese encephalitis virus,JEV)阴性的仔猪抗凝血,分离PBMCs,PCR扩增猪IFITM CDS序列,测序并进行拓扑学分析.PBMCs体外感染PRRSV,分别于感染后12、24、36和48 h收集细胞样品,RT-PCR法检测PBMCs中PRRSV感染情况,qRT-PCR法检测IFITM1、IFITM2和IFITM3基因mRNA转录水平的变化.结果 成功分离到猪PBMCs,并克隆获得了PBMCs源IFITMCDS全长序列,猪IFITM蛋白可能存在两种拓扑学结构.PBMCs接种PRRSV后24 h可产生明显的细胞病变.PRRSV可在PBMCs中复制.PRRSV感染早期可显著上调PBMCs中IFITM1、IFITM2和IFITM3基因mRNA的转录水平,且均在感染后第12h达高峰,随后逐渐下降;IFITM1基因mRNA的转录水平在病毒感染后第36 h有所回升,然后又迅速回落.结论 PRRSV体外感染PBMCs可显著上调猪IFITM基因mRNA的转录水平,表明IFITM参与PBMCs抗病毒免疫反应.本研究为揭示机体抗PRRSV天然免疫反应提供了参考.

Canine parainfluenza virus (CPIV) is a zoonotic virus that is widely distributed and is the main pathogen causing canine infectious respiratory disease (CIRD), also known as “kennel cough,” in dogs. The CPIV-V protein is the only nonstructural protein of the virus and plays an important role in multiple stages of the virus life cycle by inhibiting apoptosis, altering the host cell cycle and interfering with the interferon response. In addition, studies have shown that the V protein has potential applications in the field of immunotherapy in oncolytic virus therapy or self-amplifying RNA vaccines. In this review, the biosynthesis, structural characteristics and functions of the CPIV-V protein are reviewed with an emphasis on how it facilitates viral immune escape and its potential applications in the field of immunotherapy. Therefore, this review provides a scientific basis for research into the CPIV-V protein and its potential applications.

Since its discovery, Porcine reproductive and respiratory syndrome (PRRS) has had a huge impact on the farming industry. The virus that causes PRRS is Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), and because of its genetic diversity and the complexity of the immune response, the eradication of PRRS has been a challenge. To provide scientific references for PRRSV control and vaccine development, this study describes the processes of PRRSV-induced infection and escape, as well as the host adaptive immune response to PRRSV. It also discusses the relationship between PRRSV and the adaptive immune response.

The interferon-induced transmembrane protein 3 (IFITM3), a small molecule transmembrane protein induced by interferon, is generally conserved in vertebrates, which can inhibit infection by a diverse range of pathogenic viruses such as influenza virus. However, the precise antiviral mechanisms of IFITM3 remain unclear. At least four post-translational modifications (PTMs) were found to modulate the antiviral effect of IFITM3. These include positive regulation provided by S-palmitoylation of cysteine and negative regulation provided by lysine ubiquitination, lysine methylation, and tyrosine phosphorylation. IFITM3 S-palmitoylation is an enzymatic addition of a 16-carbon fatty acid on the three cysteine residues within or adjacent to its two hydrophobic domains at positions 71, 72, and 105, that is essential for its proper targeting, stability, and function. As S-palmitoylation is the only PTM known to enhance the antiviral activity of IFITM3, enzymes that add this modification may play important roles in IFN-induced immune responses. This study mainly reviews the research progresses on the antiviral mechanism of IFITM3, the regulation mechanism of S-palmitoylation modification on its subcellular localization, stability, and function, and the enzymes that mediate the S-palmitoylation modification of IFITM3, which may help elucidate the mechanism by which this IFN effector restrict virus replication and thus aid in the design of therapeutics targeted at pathogenic viruses.

Objective:To explore the molecular mechanism by which oral S2-Ag85DNA vaccines present intestinal antigens. The oral S2-Ag85 vaccine has been shown to protect the human body and effectively improve the titration of the vaccine by acting on intestinal mucosa cells and enhancing their immunogenicity.Method:Mice were immunized with the recombinant S2-Ag85 vaccine, and antibody secretion was then detected in the intestinal tissue. The molecular mechanisms of in vitro detection sensor molecules RIG-1, Pol III, and related conductor transductor molecules DAI, STING, AIM2, IRF3, and IRF7 were determined by separating intestinal IEC, DC, and IELC cells.Results:The S2-Ag85A vaccine was effective in activating dsDNA and RNA transduction pathways in intestinal cells and improving intestinal antigen presentation in mice.

随着信息技术的快速发展,传统的"兽医传染病学"教学方式已经不能满足社会对该领域相关人才的知识储备需求.应对网络时代大趋势,该课程的教学方式也要作出相应的改革."互联网+"混合式教学模式是本研究的主要内容.依托超星学习通平台,从课前预习、课中传授以及课后巩固等3个方面进行线上的内容建设.在课前预习和课后讨论中引入问题导向教学法(Problem-Based Learning,PBL)进行案例讨论,调动学生学习的积极性和参与度.同时,在教学内容中融入耕读文化教育和课程思政教育,培养学生学农爱农情怀,增强学生服务"三农"和农业农村现代化建设的使命感和责任感.

The porcine reproductive and respiratory syndrome virus (PRRSV) is a threat to the health of pigs worldwide, but commercially available vaccines offer limited protection against PRRSV infection. It is necessary to develop a more effective DNA vaccine. The immunological effects of DNA vaccines with three adjuvants were examined in pigs ( Susscrofa domestica ) challenged with PRRSV. These DNA vaccines, which encoded PRRSV GP3 and GP5, were formulated with A1, A2, and A3. Serum specific and neutralizing antibodies, IL-4, IFN-γ, IL-2, IL-10, CD4 + and CD8 + T-lymphocytes, health status, histopathology, and viral loads were determined. The results showed that the use of adjuvant A3 led to higher levels of neutralizing antibodies and a lower viral load in pigs compared to the other adjuvants. The neutralizing antibody titers of the pVAX-GP35+A1 and pVAX-GP35+A3 groups reached a peak of 1:19 at 35 dpi. The maximum concentration of IL-4 was 136.77 pg/mL in the pVAX-GP35+A3 group. At 35 dpi, the IFN-γ concentration in the pVAX-GP35+A1 group was 227.4 pg/mL. pVAX-GP35+A3 group shows the highest IL-2 and IL-10 expression to the peak of 597.6 pg/mL and 189.1 pg/mL, respectively. We found a formulation demonstrated beneficial immune outcomes. This study provides an alternative vaccine to protect pigs from PRRSV.

Brucellosis is a zoonotic disease caused by Gram-negative bacteria. Most of the brucellosis vaccines in the application are whole-bacteria vaccines. Live-attenuated vaccines are widely used for brucellosis prevention in sheep, goats, pigs, and cattle. Thus, there is also a need for an adjuvanted vaccine for human brucellosis, because the attenuated Brucella vaccines now utilized in animals cause human illness. Here, we developed a live-attenuated Brucella suis strain 2 vaccine (S2) adjuvanted with Ag85a (Ag85a-S2). We found that Ag85a-S2 activated cGAS-STING pathways both in intestinal mucosal cells in vivo and in the BMDM and U937 cell line in vitro. We demonstrated that the cGAS knockout significantly downregulated the abundance of interferon and other cytokines induced by Ag85a-S2. Moreover, Ag85a-S2 triggered a stronger cellular immune response compared to S2 alone. In sum, Ag85a-S2-mediated enhancement of immune responses was at least partially dependent on the cGAS-STING pathway. Our results provide a new candidate for preventing Brucella pathogens from livestock, which might reduce the dosage and potential toxicity compared to S2.

Porcine parvoviruses (PPV) and porcine circoviruses type 2 (PCV2) are widespread in the pig population. Recently, it was suggested that PPV7 may stimulate PCV2 and PCV3 replication. The present study aimed to make detection and molecular characterization of PPV7 for the first time in eastern Inner Mongolia Autonomous Region, China. Twenty-seven of ninety-four samples (28.72%) and five in eight pig farms were PPV7 positive. Further detection showed that the co-infection rate of PPV7 and PCV2 was 20.21% (19/94), and 9.59% (9/94) for PPV7 and PCV3. In addition, the positive rate of PPV7 in PCV2 positive samples was higher than that in PCV2 negative samples, supporting that PCV2 could act as a co-factor for PPV7 infection. In total, four PPV7 strains were sequenced and designated as NM-14, NM-19, NM-4, and NM-40. The amplified genome sequence of NM-14 and NM-40 were 3,999nt in length, while NM-19 and NM-4 were 3,996nt with a three nucleotides deletion at 3,097-3,099, resulting in an amino acid deletion in the Cap protein. Phylogenetic analysis based on the capsid amino acid (aa) sequences showed that 52 PPV7 strains were divided into two clades, and the four PPV7 strains in this study were all clustered in clade 1. The genome and capsid amino acid sequence of the four PPV7 strains identified in this study shared 80.0-96.9% and 85.9-100% similarity with that of 48 PPV7 reference strains selected in NCBI. Simplot analysis revealed that NM-19 and NM-4 strains were probably produced by recombination of two PPV7 strains from China. The amino acid sequence alignment analysis of capsid revealed that the four PPV7 strains detected in Inner Mongolia had multiple amino acid mutations in the 6 B cell linear epitopes compared with the reference strains, suggesting that the four PPV7 strains may have different characteristics in receptor binding and immunogenicity. In summary, this paper reported the PPV7 infection and molecular characterization in the eastern of Inner Mongolia Autonomous Region for the first time, which is helpful to understand the molecular epidemic characteristics of PPV7.

This study investigated the effect of colostrum feeding time on the colon digesta microbiome of 2-day-old dairy calves using whole-genome-based metagenome sequencing, aiming to understand the dynamic changes of the colon microbiome when the colostrum feeding is delayed. In total, 24 male Holstein calves were grouped to different pasteurized colostrum feeding time treatments randomly: TRT0h (45 min after birth, n = 7); TRT6h (6 h after birth, n = 8); and TRT12h (12 h after birth, n = 9). Bacteria, archaea, eukaryotes, and viruses were identified in the colon microbiome, with bacteria (99.20%) being the most predominant domain. Streptococcus, Clostridium, Lactobacillus, Ruminococcus, and Enterococcus were the top five abundant bacteria genera. For colon microbiome functions, 114 Kyoto Encyclopedia of Genes and Genomes (KEGG) pathways were identified, with nutrients metabolism-related functions "carbohydrate metabolism," "amino acid metabolism," "metabolism of cofactors and vitamins," "metabolism of terpenoids and polyketides," and "metabolism of other amino acids" being the top five secondary level of KEGG hierarchy functions. When colon microbiomes were compared, they were not affected by delaying first colostrum feeding at both taxonomic and functional levels. However, distinct clusters of colon microbiome profiles were shown based on PERMANOVA analysis despite of different colostrum feeding treatment, suggesting the individualized responses. Moreover, the relative abundance of microbial taxa, microbial functions, and differentially expressed genes was compared between the two distinct clusters, and different relationships were observed among host differentially expressed genes, differential levels of microbial taxa, and microbial functions between the two clusters. Our results suggest that the host may play an important role in shaping the colon microbiome of neonatal dairy calves in response to the early life feeding management. Whether the observed colon microbiome shifts affect gut health and function in the long term requires further research.