Search

Classical swine fever (CSF), caused by CSF virus (CSFV), is a highly contagious disease affecting pigs and causing massive pig production losses with severe global economic recession. The immunization of live-attenuated vaccines is still one of the key measures to CSFV management in endemic countries. However, there are also strong controversies about the usage of live-attenuated vaccines, particularly in pregnant sows and young pigs, such as in Europe, where domestic pigs are routinely not vaccinated until severe outbreaks occur. Here, we report a CSF outbreak in a pig farm in China, which affected more than 90% of the delivery sows and led to ∼45% birth loss. Surprisingly, phylogenetic analysis showed that the CSFV isolate (named CSFV/HeNLY2022, GenBank No. OR195698) was clustered into subgenotype 1.1a, closely together with the live-attenuated vaccine strains. Further genomic analysis also revealed that the isolate CSFV/HeNLY2022 shared the highest nucleotide identity of 99.7% with the C/HVRI vaccine strain (C-strain, GenBank No. AY805221). Moreover, compared to the C/HVRI strain, a total of eight amino acid mutations, distributed in Erns (H436thY and S476thR), E1 (T502thI and P581thT), E2 (M979thK and A1061thS), NS5A (A2980thT), and NS5B (I3818thM), were characterized in the CSFV/HeNLY2022 isolate. Our results suggested that the CSF outbreak was most likely caused by the live-attenuated CSFV vaccine or its derivative. It raises concern that the unscientific application of CSFV vaccines could potentially lead to CSFV spread in pigs. It is needed to perform a more rigorous evaluation of the safety of the C-strain-derived vaccines in combination with other different live-attenuated vaccines. [ABSTRACT FROM AUTHOR]

Pseudorabies virus (PRV) is an economically important viral agent affecting the swine industry in China. Accurate, rapid and simple detection is critical to PRV control and eradication. In the present study, a visible and low equipment-dependent recombinase-aid amplification assay integrated with lateral flow assay (RAA-LFA) was successfully developed to detect the PRV against the gE gene. The RAA-LFA did not react with the other swine pathogens, indicating the method has a good specificity. The limit of detection (LOD) for this RAA-LFA method was 21 copies per reaction against standard plasmids containing gE gene. Notably, the RAA-LFA can detect as low as 6.0 × 100 50 % tissue culture infective dose (TCID 50) viral titer per reaction under nucleic-acid-extraction free condition. Clinical detection showed that the results detected by RAA-LFA were completely consistent with that of the qPCR assay. Taken together, the developed PRV RAA-LFA method provides approachability, comparable accuracy and sensitivity tool for PRV point-of-care testing (POCT), which is valuable to PRV control in areas where equipment and personnel resources are scarce. • A RAA-LFA detection method was successfully developed against PRV gE gene. • The RAA-LFA method was tolerance to crude samples without extraction of viral DNA. • The RAA-LFA method is suitable for the application of POCT. [ABSTRACT FROM AUTHOR]

With the continuous development of China's economy and society, people and the government have higher and higher requirements for food safety. Testing for food dopants and toxins can prevent the occurrence of various adverse health phenomena in the world's population. By deploying new and powerful sensors that enable rapid sensing processes, the food industry can help detect trace adulteration and toxic substances. At present, as a common food safety detection method, lateral flow immunochromatography (LFI) is widely used in food safety testing, environmental testing and clinical medical treatment because of its advantages of simplicity, speed, specificity and low cost, and plays a pivotal role in ensuring food safety. This paper mainly focuses on the application of lateral flow immunochromatography and new technologies combined with test strips in food safety detection, such as aptamers, surface-enhanced Raman spectroscopy, quantum dots, electrochemical test strip detection technology, biosensor test strip detection, etc. In addition, sensing principles such as fluorescence resonance energy transfer can also more effective. Different methods have different characteristics. The following is a review of the application of these technologies in food safety detection. [ABSTRACT FROM AUTHOR]

BACKGROUND Ochratoxin A ( OTA) and zearalenone ( ZEN) are natural products of filamentous fungi that are harmful to humans and animals exposed to them even in extremely low concentration. The immunochromatographic test strip has become a popular diagnostic tool for detecting analytes. Its major advantages are that results can be obtained within 5-10 min, all needed reagents are included in the strip and it can be used to detect OTA and ZEN contamination in spots. In this study a colloidal gold-based immunochromatographic test strip of competitive format was developed for the rapid simultaneous qualitative and quantitative detection of OTA and ZEN in corn and other cereals. RESULTS The test strip results with the naked eye showed that the sensitivities were 6 µg kg−1 OTA and 20 µg kg−1 ZEN in cereal, while those with a TSR3000 membrane strip reader showed that the IC50 values of OTA and ZEN were 1.7905 and 4.3514 ng mL−1 and the lower detection limit ( LDL) values were 0.7697 and 1.2000 µg kg−1 respectively. These results were confirmed by high-performance liquid chromatography. CONCLUSION The immunochromatographic test strip developed in this study could be used for the rapid simultaneous, qualitative and quantitative screening of OTA and ZEN in corn samples. © 2015 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

An abrupt decrease in egg production and quality in laying hens can be brought on by the avian adenovirus known as the egg drop syndrome virus (EDSV). So far, there are no efficient commercial diagnostic approaches for EDSV. In the present study, we first purified two monoclonal antibodies (mAbs 5G4 and 6G6) specifically against the EDSV fibre protein. The mAb 5G4 was then employed as capture antibody. Additionally, the detection antibody 6G6 (HRP-6G6) was conjugated with horseradish peroxidase. Consequently, based on these two mAbs, we developed a sandwich enzyme-linked immunosorbent assay (ELISA) for EDSV detection. Specificity evaluation demonstrated that only EDSV gave a positive result while other avian viruses showed negative results. Sensitivity investigation showed that the limit of detection of EDSV was 102.9 TCID50/ml and the limit of detection of the purified His-Fiber was 5 ng/ml. Furthermore, the screening of samples from infected chickens showed that this method was very effective and could be applied to the detection of EDSV in infected samples. In summary, the sandwich enzyme-linked immunosorbent assay developed here provides an efficient, sensitive, and large-scale method for the specific diagnosis of EDSV. A sandwich ELISA was developed to detect EDSV using the mAbs 5G4 and HRP-6G6. The sandwich ELISA maintained high specificity and sensitivity. The sandwich ELISA had equivalent consistency with real-time PCR assay. [ABSTRACT FROM AUTHOR]

Wheat gliadin is thought to be the main cause of wheat allergy leading to celiac disease. In this study, we reported an electrochemical immunosensor based on AuNPs/Zn/Ni-ZIF-8-800@graphene for simple, efficient, and sensitive detection of gliadin. A monoclonal antibody against gliadin were prepared by hybridoma technique. Zn/Ni-ZIF-8-800 with high stability was prepared by using zinc–nickel bimetallic organic framework MOFs, and graphene and gold nanoparticles with large surface area and excellent conductivity were introduced. Chitosan was used as a binder to fixed the composite on the surface of glassy carbon electrode. The fabricated materials and the corresponding sensors were comprehensively characterized by X-ray diffractometer, scanning electron microscope, transmission electron microscopy, Fourier transform infrared spectroscopy, Brunauer–Emmett–Teller, and cyclic voltammetry analyses. Under the optimal conditions, the linear range of the electrochemical immunosensor was 0.1–100 μg mL−1, and the detection limit was 0.950 μg mL−1. In addition, the fabricated sensor exhibited high selectivity and long-term stability. The recovery test showed that the prepared sensor was suitable for the detection of gliadin in real food and feed, exhibiting great promising of the developed sensor in practical application. [ABSTRACT FROM AUTHOR]

Background: Porcine deltacoronavirus (PDCoV) is one of the emerging swine enteric coronaviruses (SECoVs), which has been widely prevalent in the North America and Asia. In addition to causing severe diarrhea in piglets, PDCoV also shows the potential to infect diverse host species, including calves, chickens, turkey poults, and humans. However, the clinical pathogenicity and genetic evolution of PDCoV is still not fully understood. Results: Here, we recorded an outbreak of a novel recombinant PDCoV strain (CHN-HeN06-2022) in a large nursery fattening pig farm. Genomic analysis showed that the CHN-HeN06-2022 strain shared 98.3-98.7% sequence identities with the Chinese and American reference strains. To clarify the evolutionary relationships, phylogenetic analysis was performed using the PDCoV genome sequences available in the GenBank database. Based on genetic distance and geographical distribution, the phylogenetic tree clearly showed that all the PDCoV sequences could be divided into lineage 1 and lineage 2, which were further classified into sublineage 1.1 (Chinese strains), 1.2 (the North American strains), 2.1 (the Southeast Asian strains), and 2.2 (Chinese strains). Corresponding to the evolutionary tree, we found that, compared to lineage 1, lineage 2 strains usually contain a continuous 6-nt deletion in Nsp2 and a 9-nt deletion in Nsp3, respectively. Furthermore, recombination analysis suggested that the CHN-HeN06-2022 occurred segments exchange crossed Nsp2 and Nsp3 region between sublineage 1.1 and sublineage 2.1. Combined with previously reported recombinant strains, the highest recombination frequency occurred in Nsp2, Nsp3, and S gene. Additionally, we identified a total of 14 amino acid sites under positive selection in spike protein, most of which are located in the regions related with the viral attachment, receptor binding, and membrane fusion. Conclusions: Taken together, our studies provide novel insights into the genetic diversity and adaptive evolution of PDCoV. It would be helpful to the development of vaccine and potential antiviral agent. [ABSTRACT FROM AUTHOR]

The affinity of peptides is a crucial factor in studying peptide–protein interactions. Despite the development of various techniques to evaluate peptide–receptor affinity, the results may not always reflect the actual affinity of the peptides accurately. The current study provides a free tool to assess the actual peptide affinity based on virtual docking data. This study employed a dataset that combined actual peptide affinity information (active and inactive) and virtual peptide–receptor docking data, and different machine learning algorithms were utilized. Compared with the other algorithms, the random forest (RF) algorithm showed the best performance and was used in building three RF models using different numbers of significant features (four, three, and two). Further analysis revealed that the four-feature RF model achieved the highest Accuracy of 0.714 in classifying an independent unknown peptide dataset designed with the PEDV spike protein, and it also revealed overfitting problems in the other models. This four-feature RF model was used to evaluate peptide affinity by constructing the relationship between the actual affinity and the virtual docking scores of peptides to their receptors. [ABSTRACT FROM AUTHOR]

Outbreaks of hydropericardium hepatitis syndrome caused by fowl adenovirus serotype 4 (FAdV-4) with a novel genotype have been reported in China since 2015, with significant economic losses to the poultry industry. Fiber2 is one of the important structural proteins on FAdV-4 virions. In this study, the C-terminal knob domain of the FAdV-4 Fiber2 protein was expressed and purified, and its trimer structure (PDB ID: 7W83) was determined for the first time. A series of affinity peptides targeting the knob domain of the Fiber2 protein were designed and synthesized on the basis of the crystal structure using computer virtual screening technology. A total of eight peptides were screened using an immunoperoxidase monolayer assay and RT-qPCR, and they exhibited strong binding affinities to the knob domain of the FAdV-4 Fiber2 protein in a surface plasmon resonance assay. Treatment with peptide number 15 (P15; WWHEKE) at different concentrations (10, 25, and 50 μM) significantly reduced the expression level of the Fiber2 protein and the viral titer during FAdV-4 infection. P15 was found to be an optimal peptide with antiviral activity against FAdV-4 in vitro with no cytotoxic effect on LMH cells up to 200 μM. This study led to the identification of a class of affinity peptides designed using computer virtual screening technology that targeted the knob domain of the FAdV-4 Fiber2 protein and may be developed as a novel potential and effective antiviral strategy in the prevention and control of FAdV-4. [ABSTRACT FROM AUTHOR]

Due to the rapid mutation of porcine epidemic diarrhea virus (PEDV), existing vaccines cannot provide sufficient immune protection for pigs. Therefore, it is urgent to design the affinity peptides for the prevention and control of this disease. In this study, we made use of a molecular docking technology for virtual screening of affinity peptides that specifically recognized the PEDV S1 C-terminal domain (CTD) protein for the first time. Experimentally, the affinity, cross-reactivity and sensitivity of the peptides were identified by an enzyme-linked immunosorbent assay (ELISA) and a surface plasmon resonance (SPR) test, separately. Subsequently, Cell Counting Kit-8 (CCK-8), quantitative real-time PCR (qRT-PCR), Western blot and indirect immunofluorescence were used to further study the antiviral effect of different concentrations of peptide 110766 in PEDV. Our results showed that the P/N value of peptide 110766 at 450 nm reached 167, with a KD value of 216 nM. The cytotoxic test indicated that peptide 110766 was not toxic to vero cells. Results of the absolute quantitative PCR revealed that different concentrations (3.125 μM, 6.25 μM, 12.5 μM, 25 μM, 50 μM, 100 μM, 200 μM) of peptide 110766 could significantly reduce the viral load of PEDV compared with the virus group (p < 0.0001). Similarly, results of Western blot and indirect immunofluorescence also suggested that the antiviral effect of peptide 110766 at 3.125 is still significant. Based on the above research, high-affinity peptide 110766 binding to the PEDV S1-CTD protein was attained by a molecular docking technology. Therefore, designing, screening, and identifying affinity peptides can provide a new method for the development of antiviral drugs for PEDV. [ABSTRACT FROM AUTHOR]

Gram-positive enhancer matrix (GEM) nanoparticles are often used in mucosal immunity, preparation of subunit vaccines or as an immune adjuvant due to its good immunological activities in recent years. Here, we designed and screened out a high affinity peptide ligand PL23, which could specifically target the non-epitope region of Classic Swine Fever Virus (CSFV) E2 protein, by virtual screening technology, enzyme linked immunosorbent assay (ELISA) and surface plasmon resonance (SPR) test. The OD value of PL23 at 450 nm was reached 1.982, and the K D value of it was 90.12 nM. Its binding capacity to protein was verified by SDS-PAGE as well. PL23 was subsequently conjugated to GEM nanoparticles by dehydration synthesis generating GEM-PL23 particles, and the GEM-PL-E2 particles were assembled after incubated with CSFV E2 protein. The cytotoxic test indicated that PL23, CSFV E2 protein, GEM nanoparticles, GEM-PL23 particles and GEM-PL-E2 particles were not toxic to cells and GEM nanoparticles could significantly promote the growth of APCs at high concentration for 1 h, p<0.001. In addition, GEM nanoparticles could promote the uptake of antigen by APCs. The cytokines tests suggested that GEM-PL-E2 particles could promote innate immune responses, regulate adaptive immune responses generated by T cells and APCs, and promote the differentiation and maturation of dendritic cells without producing inflammasomes. The results of immunological activity identification showed GEM-PL-E2 particles induced higher levels of both neutralizing antibodies and anti-CSFV antibodies than CSFV E2 protein in mice. This strategy provided a new, simpler, faster and cheaper method for assembling GEM nanoparticles, using an affinity peptide ligand replaced the protein anchor (PA), and provided a better application prospect for the application of GEM particles. [ABSTRACT FROM AUTHOR]

Abstract: Previous studies have shown that porcine reproductive and respiratory syndrome virus (PRRSV) nonstructural protein 1α (nsp1α) was the interferon (IFN) antagonist. However, the mechanism was unclear. In the present study, deletion of the carboxyl-terminal extension (CTE) (167–180 amino acid (aa)) made nsp1α lose its inhibitory ability to the induction of IFN-β. And a series of C-terminal truncated mutants for nsp1α showed that 1–176 aa of nsp1α was able to inhibit the induction of IFN-β and deleting or mutating the amino acid F176 made nsp1α not inhibit the induction of IFN-β. In conclusion, the CTE and the amino acid F176 were critical for nsp1α as the IFN antagonist and the region representing 167–176 was the minimal subunit of the CTE for nsp1α to retain its suppressive activity to the induction of IFN-β. [Copyright &y& Elsevier]

Background: Porcine reproductive and respiratory syndrome virus (PRRSV) infection can cause severe reproductive failure in sows and respiratory distress in pigs of all ages, leading to major economic losses. To date, there are still no effective strategies to prevent and control PRRSV. Antibody-dependent enhancement (ADE), a phenomenon in which preexisting non-neutralizing antibodies or sub-neutralizing antibodies facilitate virus entry and replication, may be a significant obstacle in the development of effective vaccines for many viruses, including PRRSV. However, the contribution of ADE to PRRSV infection remains controversial, especially in vivo. Whether attenuated PRRSV vaccines prevent or worsen subsequent disease in pigs infected by novel PRRSV strains requires more research. In the present study, in vivo experiments were conducted to evaluate ADE under different immune statuses, which were produced by waiting different lengths of time after vaccination with a commercially available attenuated highly pathogenic PRRSV (HP-PRRSV) vaccine (JXA1-R) before challenging the pigs with a novel heterologous NADC30-like strain. Results: Piglets that were vaccinated before being challenged with PRRSV exhibited lower mortality rates, lower body temperatures, higher bodyweight gain, and lower viremia. These results demonstrate that vaccination with JXA1-R alleviated the clinical signs of PRRSV infection in all vaccinated groups. Conclusions: The obtained data indicate that the attenuated vaccine test here provided partial protection against the NADC30-like strain HNhx. No signs of enhanced PRRSV infection were observed under the applied experimental conditions. Our results provide some insight into the molecular mechanisms underlying vaccine-induced protection or enhancement in PRRSV. [ABSTRACT FROM AUTHOR]

Purification of the enveloped virus poses a challenge as one must retain viral infectivity to preserve immunogenicity. The traditional process of virus purification is time‐consuming, laborious and hard to scale up. Here, a rapid, simple and extensible laboratory program for the purification of Japanese encephalitis virus (JEV) was developed by using differential centrifugation, ultrafiltration, Sepharose 4 fast flow gel chromatography, and CaptoTM Core 700 chromatography. The entire process recovered 61.64% of the original virus, and the purified virus particles maintained good activity and immunogenicity. The purification process described has potential application in large‐scale production of high‐purity JEV. [ABSTRACT FROM AUTHOR]

Classical swine fever (CSF) caused by the classical swine fever virus (CSFV) is a highly contagious swine disease resulting in large economical losses worldwide. The viral envelope glycoprotein E2 and Erns are major targets for eliciting antibodies against CSFV in infected animals. In this report, the glycoprotein E2 and Erns were expressed using the baculovirus system and their protective immunity in rabbits were tested. Twenty CSFV seronegative rabbits were randomly divided into five groups. Each rabbit was intramuscularly immunized with CSFV-E2, CSFV-Erns, or their combination (CSFV-E2 + Erns). Besides, a commercial CSFV vaccine (C-strain) and PBS were used as positive or negative controls, respectively. Four weeks after the second immunization, all the rabbits were challenged with 100 RID50 of CSFV C-strain. High levels of CSFV E2-specific antibody, neutralizing antibody and cellular immune responses to CSFV were elicited in the rabbits inoculated with C-strain, CSFV-E2, and CSFV-E2 + Erns. And the rabbits inoculated with the three vaccines received complete protection against CSFV C-strain. However, no neutralizing antibody was detected in the Erns vaccinated rabbits and the rabbits exhibited fever typical of CSFV, suggesting the Erns alone is not able to induce a protective immune response. Taken together, while the Erns could not confer protection against CSFV, E2 and E2 + Erns could not only elicit humoral and cell-mediated immune responses but also confer complete protection against CSFV C-strain in rabbits. [ABSTRACT FROM AUTHOR]

Introduction: Epidemic Japanese encephalitis is one of the most important zoonotic diseases that cause central nervous system damage. The vaccination has become the most effective and economical measure for its control. Hence, real-time monitoring of Japanese encephalitis virus (JEV) proliferation is crucial to optimize virus inoculation, culturing conditions, and virus harvest time. Methods: The proliferation dynamics of JEV in BHK-21 cells was studied by combining the established quantitative PCR method with the conventional TCID50 assay in this study. Results: The proliferation curve determined by the 2 methods has a definite parallel relationship, but the quantitative real-time PCR method (4 h) is faster and more sensitive than the TCID50 method (3–4 days). The determination results of TCID50 showed that the highest viral titer was 105.44 TCID50/0.1 mL and 104.86 TCID50/0.1 mL in cell suspension and culture supernate, respectively, while the virus RNA copies reached the peak at 1.0 × 107.5 copies/µL and 1.0 × 105.6 copies/µL in cell suspension and culture supernate, respectively. Conclusion: The comprehensive analysis showed that the best time for JEV proliferation in BHK-21 cell was 60 h post infection. [ABSTRACT FROM AUTHOR]

The nucleocapsid (N) protein is a suitable candidate for early diagnosis of porcine epidemic diarrhea virus (PEDV). Here, we identified the linear B-cell epitopes of the PEDV N-protein by integrating a computational-experimental framework and constructed three-dimensional (3D) structure model of the N protein using the ColabFold program in Google Colaboratory. Furthermore, we prepared the monoclonal antibodies against the predicted epitopes and recombinant N protein, respectively, and selected pairing mAbs (named 9C4 and 3C5) to develop a double-antibody sandwich immunochromatographic test strip using CdSe/ZnS quantum dots (QDs)-labelled 9C4 and 3C5 as capture and detection antibodies, respectively. This strip can specifically detect PEDV within 10 min with a detection limit of less than 6.25 × 103 TCID 50 /mL. In comparison with RT-PCR for testing 90 clinical samples, the relative sensitivity and specificity of the strip were found to be 98.0% and 100%, respectively, with a concordance rate of 98.9% and a kappa value of 0.978, indicating that QDs-ICTS is a reliable method for the application of PEDV detection in clinical samples. • Discovery of novel B-cell liner epitopes in the N protein of PEDV in silico. • Preparation of mAbs against predicted epitopes and recombination N protein of PEDV. • A quantum dot-based immunochromatographic test strip was developed to detect PEDV. [ABSTRACT FROM AUTHOR]

Carprofen (CAR) is a non-steroidal anti-inflammatory drug commonly used to treat various musculoskeletal diseases and infections in food animals. While CAR residues in food animal tissues caused by veterinary medicine abuse are a potential danger to human health. In this study, a high-affinity monoclonal antibody (mAb) against CAR was prepared, and then an immunochromatographic strip for rapidly screening CAR in bovine muscle was developed to help enforce regulatory compliance. The visual detection limit (cut-off value) by the naked eye of the fabricated test strip was 12.5 ng g−1. Scanning by the strip reader, the IC50 of the test strip was calculated as 1.743 ng g−1, and the limit of detection (LOD, IC10) was 0.283 ng g−1. The proposed method provided an efficient tool for monitoring CAR in bovine muscle. Meantime, our work suggested great potential for the analysis of CAR residues in other food matrixes. [ABSTRACT FROM AUTHOR]

Vesicular stomatitis (VS), characterized by vesicular lesions, produces significant economic losses in livestock industry. Infection by its causative agent, VS virus (VSV), has been previously shown to be mediated by the glycoprotein (G) during attachment, endocytosis and membrane fusion. In the current study, we revealed a novel role of VSV G protein in negative regulation of host cell pro-inflammatory responses. We determined that VSV G protein inhibited lipopolysaccharide (LPS)-induced pro-inflammatory responses as naïve VSV virions in murine peritoneal macrophage-like cell line RAW 264.7. Furthermore, we identified that VSV G protein suppressed nuclear factor kappa-B (NF-κB) and mitogen-activated protein kinase (MAPK)-mediated pro-inflammatory pathways in a dose-dependent manner. Moreover, we demonstrated that α2-3-linked sialic acids on VSV G protein were involved in antagonizing NF-κB- and MAPK-mediated pro-inflammatory responses. All these results expand the knowledge of VSV pathogenesis and strengthen the importance of VSV G protein in host innate immunity, which support implications for the development of VSV-based vaccination and oncolysis. [ABSTRACT FROM AUTHOR]

Valnemulin (VAL) and tiamulin (TIA) are pleuromutilin antibiotics used primarily for treating bacterial infections in swine or other food animals. Furthermore, VAL and TIA are also employed as feed additives to promote animal growth. However, the illegal use of VAL and TIA could cause a series of hazards to consumers. Here, VAL was designed to be conjugated with bovine serum protein to prepare immunogen. A highly sensitive monoclonal antibody that recognized both VAL and TIA has been successfully produced. Moreover, an immunochromatographic strip assay for rapidly screening VAL and TIA in porcine liver was established with visual detection limits (cutoff values) of 50 and 25 ng/g, respectively. The IC50 values calculated from the equation of the standard curve were 6.06 and 3.45 ng/g and the limits of detection were 0.96 and 0.29 ng/g for VAL and TIA. According to the recovery experiment results, the test strip exhibited acceptable accuracy and precision. Generally, the proposed strip provided a practical tool for the detection of VAL and TIA. Practical Application: We produced a highly sensitive monoclonal antibody and developed an immunoassay strip for simultaneously monitoring TIA and VAL. Additionally it was preliminarily confirmed that the rapid detection tool was suitable for screening TIA and VAL in porcine liver. [ABSTRACT FROM AUTHOR]

BACKGROUND: Nitroxynil (NIT) is a veterinary drug against hepatic fluke disease for food‐producing cattle and sheep. NIT has a long half‐life time in animals since it is highly bound to plasma protein. Therefore NIT possibly remains in animal edible tissues or milk due to drug abuse. In this study, a specific murine monoclonal antibody (mAb) against NIT was prepared and an immunochromatographic strip assay based on the mAb was developed for screening NIT in milk. RESULTS: The affinity constant of the anti‐NIT mAb was 2.93 × 1010 and the anti‐NIT mAb had almost no cross‐reactivity with other analogs, so that it showed good specificity. The cutoff value of this test strip was considered to be 50 ng mL−1 by the naked eye. When detected by the strip reader, the half maximum inhibition concentration (IC50) of the immunoassay strip was calculated to be 5.716 ng mL−1 and the limit of detection (LOD) was 1.146 ng mL−1. Intra‐assay recoveries from 88.80 to 97.13% were obtained, with the highest coefficient of variation (CV) at 9.01%; inter‐assay recoveries ranged from 84.60 to 106.87%, with the highest CV at 9.93%. CONCLUSION: The operative procedure of the proposed method can be completed within 10 min. The strip developed in this study was a practical tool for rapid semiquantitative and quantitative detection of NIT in milk. This study suggested great potential for analytically monitoring NIT in other food samples. © 2019 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Background: Porcine circovirus type 2 (PCV2) is the pathogen of porcine circovirus associated diseases (PCVAD) and one of the main pathogens in the global pig industry, which has brought huge economic losses to the pig industry. In recent years, there has been limited research on the prevalence of PCV2 in Henan Province. This study investigated the genotype and evolution of PCV2 in this area. Results: We collected 117 clinical samples from different regions of Henan Province from 2015 to 2018. Here, we found that the PCV2 infection rate of PCV2 was 62.4%. Thirty-seven positive clinical samples were selected to amplify the complete genome of PCV2 and were sequenced. Based on the phylogenetic analysis of PCV2 ORF2 and complete genome, it was found that the 37 newly detected strains belonged to PCV2a (3 of 37), PCV2b (21 of 37) and PCV2d (13 of 37), indicating the predominant prevalence of PCV2b and PCV2d strains. In addition, we compared the amino acid sequences and found several amino acid mutation sites among different genotypes. Furthermore, the results of selective pressure analysis showed that there were 5 positive selection sites. Conclusions: This study indicated the genetic diversity, molecular epidemiology and evolution of PCV2 genotypes in Henan Province during 2015–2018. [ABSTRACT FROM AUTHOR]

DPL (http://www.peptide-ligand.cn/) is a comprehensive database of peptide ligand (DPL). DPL1.0 holds 1044 peptide ligand entries and provides references for the study of the polypeptide platform. The data were collected from PubMed-NCBI, PDB, APD3, CAMPR3, etc. The lengths of the base sequences are varied from 3 to78. DPL database has 923 linear peptides and 88 cyclic peptides. The functions of peptides collected by DPL are very wide. It includes 540 entries of antiviral peptides (including SARS-CoV-2), 55 entries of signal peptides, 48 entries of protease inhibitors, 45 entries of anti-hypertension, 37 entries of anticancer peptides, etc. There are 270 different kinds of peptide targets. All peptides in DPL have clear binding targets. Most of the peptides and receptors have 3D structures experimentally verified or predicted by CYCLOPS, I-TASSER and SWISS-MODEL. With the rapid development of the COVID-2019 epidemic, this database also collects the research progress of peptides against coronavirus. In conclusion, DPL is a unique resource, which allows users easily to explore the targets, different structures as well as properties of peptides. [ABSTRACT FROM AUTHOR]

Egg drop syndrome virus (EDSV) is an avian adenovirus that causes markedly decrease in egg production, and in the quality of the eggs when it infects chickens. Until now, EDSV virus-cell interactions are poorly understood, and the cellular receptor is still unknown. In the present study, we determined the atomic structure of the fiber head of EDSV (residues 377–644) at 2.74 Å resolution. Structure comparison with the (chick embryo lethal orphan) CELO long fiber head and human adenovirus fiber heads reveals that the avian adenovirus may interact with the same attachment factor in a unique fashion. Based on the previous studies of CELO virus, we assumed that the chicken coxsackievirus and adenovirus receptor (CAR) may be the attachment factor. We then demonstrate that the chicken CAR serves as a cellular attachment factor for EDSV based on three lines of evidences. Taken together, the results presented here are helpful for further exploring the pathogenesis related to the interaction between EDSV and host cells, and may be used for vaccine development and intervention strategies against EDSV infection. [ABSTRACT FROM AUTHOR]

• Generation a double deletion mutant Marek's disease virus absence of both meq and lorf9 genes. • Double deletion mutant Marek's disease virus has similar growth characteristics with parental virus in vitro. • Double deletion mutant Marek's disease virus significantly reduced replication in vivo. • Double deletion of meq and lorf9 genes overcome lymphoid organ atrophy induced by meq-deletion Marek's disease virus. • Double deletion mutant Marek's disease virus is fully attenuated. Marek's disease virus (MDV) is a highly contagious alphaherpesvirus that causes rapid onset of T cell lymphomas in chickens. MDV continues to break through vaccinal immunity due to the emergence of highly virulent field strains. Earlier studies revealed that deletion of the meq gene from MDV results in attenuated vaccines that protect against disease when chickens are infected with highly virulent strains. However, meq -deleted viruses still retain the ability to induce lymphoid organ atrophy, which raises safety concerns. In an earlier study, we found that deletion of lorf9 counteracts this lymphoid organ atrophy. Here, we describe the generation of a double deletion mutant virus lacking virus-encoded meq and lorf9. In vitro studies revealed that during replication, the mutant virus had kinetic characteristics similar to the parental virus; however, in vivo the replication capability was significantly reduced. Results of animal studies revealed no obvious MDV-specific symptoms and lesions. Importantly, the double deletion mutant virus lost the capacity to induce lymphoid organ atrophy, which has been the main obstacle during development of a good vaccine candidate. [ABSTRACT FROM AUTHOR]

Graphical abstract Highlights • A novel electrochemical immunosensor was developed using nanohybrid materials and anti-monensin monoclonal antibody (Ab). • Zn/Ni-ZIF-8 800@graphene/AuNPs nanocomposites were used to modify glassy carbon electrode (GCE) for the first time. • The synergistic effect of Zn/Ni-ZIF-8 800, graphene and AuNPs had enhanced the conductivity and sensitivity of the biosensor. • The nanocomposites exhibited high analytical performance for detecting monensin in milk sample. Abstract A facile, efficient and sensitive electrochemical immunosensor based on Au nanoparticles/Zn/Ni-ZIF-8-800@graphene composites was developed to detect the monensin in milk. Zn/Ni-ZIF-8-800 with high stability was obtained via pyrolysis of the Zn/Ni-bimetallic metal-organic frameworks (MOFs) precursor Zn/Ni-ZIF-8. Benefitting from the synergistic effects including the high porosity of the Zn/Ni-ZIF-8-800 and the excellent electrical conductivity of graphene, the electrochemical signal got greatly enhanced. The AuNPs were electrodeposited on the Zn/Ni-ZIF-8-800@graphene surface for antibody-antigen capture. Then the constructed hybrid materials not only possess high specific surface area, but also exhibit excellent biocompatibility and electron transfer ability. Morphology and elemental analyse of these composites were conducted via scanning electron microscopy (SEM) and energy dispersive spectroscopy (EDS). In addition, the crystal structure was analyzed by X-ray diffraction (XRD). Afterwards, the preparation process of the biosensor was investigated by cyclic voltammetry (CV), and the detection procedure was evaluated by differential pulse voltammetry (DPV). Under the optimal conditions, the achieved linear range was 0.25–100 ng mL−1, and the calculated detection limit was 0.11 ng mL−1. The as-prepared biosensor in this work offers an effective approach to measure monensin and may be a promising candidate for detecting other veterinary drugs in food products. [ABSTRACT FROM AUTHOR]

Abstract: BACKGROUND: The importance of peptides in regulatory interactions has caused peptide‐protein docking to attract the attention of many researchers. A variety of methods for molecular modeling of peptide‐protein docking, such as local search and global search, are currently used. RESULTS: The interactions of 11 peptides and CSFV E2 protein were evaluated by the GalaxyPepDock and FlexX/ SYBYL programs, respectively. The assessment scores of all the peptides were correlated with their KD values. The final results showed that a moderate correlation coefficient was represented between KD values and CScores of predicted models by FlexX/ SYBYL. CONCLUSION: Our results demonstrate that considering the flexibility of the peptide is better than searching for more potential binding sites on the target protein surface while performing peptide‐protein molecular docking. These data provide reasonable evidence for the molecular design of peptides and guidance for the functional assignment of target proteins. © 2018 Society of Chemical Industry [ABSTRACT FROM AUTHOR]

Two different genotypes of porcine epidemic diarrhea virus (PEDV), the classical and variant strains, are classified by multiple insertions and deletions in their S genes. It is critical to detect and differentiate two genotypes in the pork industry to prevent PEDV outbreaks. In the present study, a novel duplex TaqMan RT-PCR was developed for detecting and differentiating PEDV strains in China. There was no cross-amplification between the two probes when using standard recombinant plasmids, and the specificity was further confirmed by using other seven non-PEDV swine pathogens. The minimum copies required for the detection of both classical and variant PEDV were 4.8 × 10 2 DNA copies/reaction. The repeatability of TaqMan RT-PCR was evaluated using standard recombinant plasmids and gave coefficients of variation 0.19–4.93. In recent 5 years, 79 clinical samples were collected from piglets with severe diarrhea in the Central China. Among these clinical samples, 51 were confirmed as PEDV positive by conventional RT-PCR, whereas 63 variant PEDV, 3 co-infections and 1 classical PEDV were confirmed by this duplex TaqMan RT-PCR, with viral loads of 10 2 –10 8 , 10 2 –10 3 , and 10 4 copies/reaction, respectively. Therefore, the duplex TaqMan RT-PCR could be a useful method for detecting and differentiating variant and classical PEDV strains. The results showed that variant PEDV was prevalent in clinical samples in central China. Moreover, in this study, co-infection by PEDV strains was detected for the first time and might help explain the emergence of the novel recombinant PEDV in recent years. [ABSTRACT FROM AUTHOR]

Abstract: Porcine reproductive and respiratory syndrome virus (PRRSV) causes an economically important disease in swine-producing area, and interferon beta (IFN-β) is the first responder against the animal virus infection. However, whether PRRSV could induce the production of IFN-β is controversial. In this paper, we first time found that PRRSV could phosphorylate IFN-regulatory factor 3 (IRF-3) and weakly activate the IFN-β promoter in MARC-145 cells in early infection, but the activations of IRF-3 and IFN-β promoter were rapidly inhibited in the following infection. Furthermore, which components or products of the invading PRRSV cause PRRSV to inhibit IFN-β promoter activity attracted our attentions. The obtained results showed that PRRSV nsp1 could inhibit Poly(I:C)-induced IFN-β promoter activity in MARC-145 cells by down-regulating the protein level of IRF-3 and inhibiting the phosphorylation of IRF-3. In conclusion, our results suggested that PRRSV could be sensed by professional IFN-β-producing system and had mechanisms to inhibit this action in MARC-145 cells. [Copyright &y& Elsevier]

Abstract: An immunochromatographic strip was developed for the serological detection of type O foot-and-mouth disease (FMD) in swine. In the strip, the expressed protein of VP1, the main protective antigen of FMDV, labeled with colloidal gold was used as the detector, the staphylococcal protein A (SPA) and swine anti-foot-and-mouth disease virus (FMDV) antibody were blotted on the nitrocellulose membrane for the test and control lines, respectively. 296 swine serum samples were collected to evaluate the characteristics of the strip in comparison with existing commercial liquid-phage blocking ELISA (LPB ELISA) kit and peptide ELISA kit. The strip was shown to be of high specificity and sensitivity. Furthermore, the dipstick assay based on the strip is rapid (5min) and easy to perform with no requirement of professional skills, reagents nor equipment. This suggests that the immunochromatographic strip is an acceptable alternative for use in clinical laboratories lacking specialized equipment and for field diagnosis. [Copyright &y& Elsevier]

The use of sulfonamides, such as sulfamethazine (SM2), in pig production is recognized as a public health risk as it inevitably results in sulfamethazine residues in pork. This study is aimed at establishing rapid, simple, reliable methods, with both sensitivity and specificity, for detecting sulfamethazine residues. For this purpose, monoclonal antibodies against sulfamethazine were prepared and characterized. No cross-reaction of the monoclonal antibodies was identified with other sulfonamides or analytes. Based on the competitive immunoassay principle, an indirect competitive ELISA kit (SM2 kit) and a rapid detection strip for detecting sulfamethazine residues were developed using monoclonal antibodies and the colloidal gold technique. The indirect competitive ELISA kit and the strip assay could be performed within 2 h and 5-10 min, respectively. The results showed that the detection limits were 1 ng ml-1 for the indirect competitive ELISA kit and 8 ng ml-1 with the unaided eye and 1 ng ml-1 with the strip reader for the rapid strip assay. Comparing the HPLC method with the SM2-kit or the strip in pig urine spiked with standards of SM2, the difference was <4.6% for SM2-kit and 4.3% for the strip. The two methods are suitable for the rapid screening of sulfamethazine residues in swine urine. Experimental data revealed that the two methods, especially the strip, proved to be sensitive, specific, rapid and easy to use for the quantitative, semi-quantitative or qualitative detection of SM2 residues in swine urine. [ABSTRACT FROM AUTHOR]

• PRRSV infection induces vimentin rearrangement during post-entry stage. • Vimentin enwraps PRRSV replication complex at the perinuclear site. • Vimentin rearrangement is beneficial for PRRSV replication. • Vimentin (Ser38) phosphorylation by CaMKIIγ is responsible for rearrangement. Vimentin, a member of intermediate filaments, has been documented to be involved in viral infections. Despite several studies focusing on its involvement in porcine reproductive and respiratory syndrome virus (PRRSV) infection, the detailed mechanisms by which vimentin takes effect remain to be fully elucidated. In the present study, we identified a previously unrecognized role of vimentin rearrangement in PRRSV replication. We monitored that PRRSV infection induced vimentin reorganization during post-entry stage, which was beneficial for viral replication. In detail, the serine residue of vimentin was phosphorylated at position 38 (Ser38) by calcium calmodulin-dependent protein kinase II gamma (CaMKIIγ), and vimentin filaments reorganized into cage-like structures enwrapping PRRSV replication complex (RC) at the perinuclear location. Taken together, these results expand the knowledge of PRRSV replication, and provide novel targets for prevention and control of PRRSV. [ABSTRACT FROM AUTHOR]

• Elastase is highly expressed in HP-PRRSV-infected lung tissues. • Elastase facilitates HP-PRRSV entry independent of low pH and CME. • Elastase mediates HP-PRRSV entry via membrane fusion at the cell surface. • HP-PRRSV GP5 is cleaved by elastase. Infection by enveloped viruses includes endocytosis and/or membrane fusion at the plasma membrane, where host cell proteases play an essential role. Among them, elastase-mediated infection has been documented for several enveloped viruses. Porcine reproductive and respiratory syndrome virus (PRRSV), an economically critical factor in global swine industry, is previously reported to infect host cells via low pH-dependent clathrin-mediated endocytosis (CME) and undergo membrane fusion in recycling endosomes. In the current study, we identified that elastase was significantly elevated in the lung tissues of highly pathogenic PRRSV (HP-PRRSV)-infected pigs compared to the mock-infected ones. We subsequently demonstrated that elastase contributed to HP-PRRSV infection in both MARC-145 cells and porcine alveolar macrophages (PAMs). Mechanistically, HP-PRRSV entered host cells at the cell surface via elastase-mediated membrane fusion, independent of low pH and CME, and its glycoprotein 5 (GP5) was cleaved by the protease during this process. All these findings deepen our understanding of HP-PRRSV infection, and are beneficial for prevention and control of the disease. [ABSTRACT FROM AUTHOR]

• A highly sensitive monoclonal antibody (mAb) against bacitracin was produced. • An immunoassay strip was developed for monitoring BAC in milk. • This strip was accurate and reliable based on the results of recovery test. In this study, a murine monoclonal antibody (mAb) of 6D2-G10 against bacitracin zinc (BAC) was produced and applied to an immunochromatographic strip (ICS) for the initial detection of BAC in milk. The ICS with a cut-off value of 25 ng/mL could be perceived by the naked eye within 10 min. With the assist of the strip reader, the limit of detection (LOD) was measured as 0.82 ng/mL, the half-maximal inhibitory concentration (IC 50) was recorded as 3.16 ng/mL, and the linear detection range was from 0.97 to 10.30 ng/mL. The recoveries ranged from 87.7% to 96.0% with the highest coefficient of variation (CV) of 9.1% in the intra-assay and from 84.3% to 90.2% with the highest CV of 10.7% in the inter-assay. In short, the established ICS provided a serviceable analytical tool for qualitatively and quantitatively monitoring BAC in milk. [ABSTRACT FROM AUTHOR]

• This assay can determine zeranol and its five family analogues in milk. • The cutoff values for zeranol and its analogues were all 10 ng/mL coincidentally. • The lower detection limits for zeranol and its analogues were all below 0.3 ng/mL. • The detection time of this assay is less than 15 min. Based on colloidal gold and broad-spectrum monoclonal antibody that binds to zeranol and its five analogues with high sensitivity, a lateral flow immunochromatographic assay (LFIA) in a competitive format was developed to specifically determine residues of zeranol, an illegal growth promoter in livestock. In this study, the assay had high sensitivity and was broad-spectrum only for zeranol and its five analogues, and the results were obtained within 10 min without needing sophisticated procedures. The cutoff values for zeranol and its five analogues were 10 ng/mL, and the IC 50 values for zeranol, β-zearalanol, zearalanone, α-zearalenol, β-zearalenol and zearalenone were 1.250, 1.800, 1.775, 1.225, 1.709 and 1.319 ng/mL, respectively. The recovery rates were ranged from 85.6 to 93.9%, with the coefficient of variations less than 12.4%. The results demonstrated that the LFIA could be used for rapid, simultaneous, semi-quantitative and quantitative detection of residues of zeranol and its five analogous in milk. [ABSTRACT FROM AUTHOR]

• The recombinant fiber protein of EDSV is expressed in Escherichia coli with highly solubility. • The recombinant protein retains the native trimeric structure, with high HA-titer. • A single inoculation with the structure-stabilized fiber protein could protect chickens from virus challenge. Egg-drop syndrome virus (EDSV) is an avian adenovirus that causes markedly decrease in egg production and in the quality of the eggs when it infects chickens. In this report, we engineered truncated fiber protein containing the entire knob domain and part of the shaft region as a vaccine candidate. The protein was obtained in the soluble fraction in Escherichia coli (E. coli), and expression level after nickel-affinity purification was 126 mg/L. By means of multiple characterization methods, it is demonstrated that the recombinant protein retains the native trimeric structure. A single inoculation with the structure-stabilized recombinant protein, even at the lowest dose of 2 μg, stimulated hemagglutination inhibition (HI) antibody responses in chickens, for at least 16 weeks. Neutralizing titers in sera from the protein immunized groups was similar to that of inactivated vaccine immunized group. The lymphocyte proliferation response and cytokine secretion were also induced in immunized SPF chickens. In addition, immunization with the fiber protein also significantly reduced the viral load in the liver. Taken together, these results suggest the truncated fiber protein as an effective single dose, long lasting and rapidly effective vaccine to protect against EDSV. [ABSTRACT FROM AUTHOR]

Maduramicin (MD) is a type of monoglycoside polyether ionophore antibiotic that can effectively treat coccidiosis and facilitate animal growth. However, its extensive and excessive use brings potential risk to human health. Herein, an electrochemical immunosensor based on indirect competitive format was fabricated for analysis of MD residue in eggs by a multiple signal amplification system. Initially, Au nanoparticles were deposited onto glassy carbon electrode surface to load the coating antigen MD-BSA and to improve conductivity. Then the signal amplification platform was constructed by encapsulating hemin into Fe-MIL-88 NH 2 metal-organic frameworks (hemin@MOFs), and then the obtained composites were decorated with AuPt nanoparticles. The synthesized hemin@MOFs/AuPt was not only used as a signal amplification mediator, but also utilized as a carrier for immobilization of horseradish peroxidase-conjugated affinipure goat anti-mouse antibody (Ab 2 -HRP) and horseradish peroxidase (HRP). The constructed hemin@MOFs/AuPt-Ab 2 -HRP bioconjugates could effectively amplify the current signal since hemin@MOFs, AuPt and HRP all exhibited high catalytic activity towards the hydrogen peroxide. Moreover, the established immunosensor showed high sensitivity and stability during the detection procedure. With the synergistic catalytic effect of hemin@MOFs, AuPt and HRP, a wide detection range of 0.1–50 ng mL−1 and a low detection limit of 0.045 ng mL−1 were achieved (S/N = 3), respectively. Ultimately, the developed method displayed excellent performance in practical applications, providing a promising probability to detect other veterinary drug residues to guarantee food safety. • An ultrasensitive biosensor for MD detection was constructed by using hemin@MOFs/AuPt-Ab 2 -HRP/HRP as bioprobes. • A linear calibration curve of MD ranged from 0.1 ng mL−1 to 50 ng mL−1 was obtained with LOD as low as 0.045 ng mL−1. • A multiple signal amplification strategy was triggered by the co-synergistic effect of hemin@MOFs, AuPt and HRP. • The proposed immunosensor shows a great potential for sensitive detection of small molecule drug residues. [ABSTRACT FROM AUTHOR]