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Emerging coronaviruses (CoVs) can cause severe diseases in humans and animals, and, as of yet, none of the currently available broad-spectrum drugs or vaccines can effectively control these diseases. Host antiviral proteins play an important role in inhibiting viral proliferation. One of the isoforms of cytoplasmic poly(A)-binding protein (PABP), PABPC4, is an RNA-processing protein, which plays an important role in promoting gene expression by enhancing translation and mRNA stability. However, its function in viruses remains poorly understood. Here, we report that the host protein, PABPC4, could be regulated by transcription factor SP1 and broadly inhibits the replication of CoVs, covering four genera (Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus) of the Coronaviridae family by targeting the nucleocapsid (N) protein through the autophagosomes for degradation. PABPC4 recruited the E3 ubiquitin ligase MARCH8/MARCHF8 to the N protein for ubiquitination. Ubiquitinated N protein was recognized by the cargo receptor NDP52/CALCOCO2, which delivered it to the autolysosomes for degradation, resulting in impaired viral proliferation. In addition to regulating gene expression, these data demonstrate a novel antiviral function of PABPC4, which broadly suppresses CoVs by degrading the N protein via the selective autophagy pathway. This study will shed light on the development of broad anticoronaviral therapies. IMPORTANCE Emerging coronaviruses (CoVs) can cause severe diseases in humans and animals, but none of the currently available drugs or vaccines can effectively control these diseases. During viral infection, the host will activate the interferon (IFN) signaling pathways and host restriction factors in maintaining the innate antiviral responses and suppressing viral replication. This study demonstrated that the host protein, PABPC4, interacts with the nucleocapsid (N) proteins from eight CoVs covering four genera (Alphacoronavirus, Betacoronavirus, Gammacoronavirus, and Deltacoronavirus) of the Coronaviridae family. PABPC4 could be regulated by SP1 and broadly inhibits the replication of CoVs by targeting the nucleocapsid (N) protein through the autophagosomes for degradation. This study significantly increases our understanding of the novel host restriction factor PABPC4 against CoV replication and will help develop novel antiviral strategies.

Porcine epidemic diarrhea virus (PEDV) is a globally distributed alphacoronavirus that has reemerged lately, resulting in large economic losses. During viral infection, type I interferon (IFN-I) plays a vital role in the antiviral innate immunity. However, PEDV has evolved strategies to limit IFN-I production. To suppress virus replication, the host must activate IFN-stimulated genes and some host restriction factors to circumvent viral replication. This study observed that PEDV infection induced early growth response gene 1 (EGR1) expression in PEDV-permissive cells. EGR1 overexpression remarkably suppressed PEDV replication. In contrast, depletion of EGR1 led to a significant increase in viral replication. EGR1 suppressed PEDV replication by directly binding to the IFN-regulated antiviral (IRAV) promoter and upregulating IRAV expression. A detailed analysis revealed that IRAV interacts and colocalizes with the PEDV nucleocapsid (N) protein, inducing N protein degradation via the E3 ubiquitin ligase MARCH8 to catalyze N protein ubiquitination. Knockdown of endogenous MARCH8 significantly reversed IRAV-mediated N protein degradation. The collective findings demonstrate a new mechanism of EGR1-mediated viral restriction, in which EGR1 upregulates the expression of IRAV to degrade PEDV N protein through MARCH8. IMPORTANCE PEDV is a highly contagious enteric coronavirus that has rapidly emerged worldwide and has caused severe economic losses. No currently available drugs or vaccines can effectively control PEDV. PEDV has evolved many strategies to limit IFN-I production. We identified EGR1 as a novel host restriction factor and demonstrated that EGR1 suppresses PEDV replication by directly binding to the IRAV promoter and upregulating the expression of IRAV, which interacts with and degrades the PEDV N protein via the E3 ubiquitin ligase MARCH8 to catalyze nucleocapsid protein ubiquitination, which adds another layer of complexity to the innate antiviral immunity of this newly identified restriction factor. A better understanding of the innate immune response to PEDV infection will aid the development of novel therapeutic targets and more effective vaccines against virus infection.

Porcine reproductive and respiratory syndrome virus (PRRSV) causes great economic losses to the swine industry globally; however, effective control measures for this virus are limited. Here, we screened a porcine alveolar macrophage (PAM) cDNA library with a yeast two-hybrid system to reveal that galectin-1 (Gal-1), an endogenous innate immune protein encoded by LGALS1, interacts with nonstructural protein 11 (Nsp11) of PRRSV. Western blotting and viral titer assays indicated that Gal-1 overexpression suppressed replication in multiple PRRSV strains (P < 0.001), whereas Gal-1 knockdown or knockout increased viral titer and nucleocapsid protein expression. The Gal-1-specific anti-PRRSV effect was associated with the endoribonuclease domain of Nsp11 through inactivation of interferon-antagonist function and stimulation of interferon-stimulated gene expression. Additionally, Gal-1 interacted with PRRSV E protein but not with PRRSV glycoproteins, and recombinant Gal-1 treatment inhibited PRRSV in PAMs and MARC-145 cells. Furthermore, Gal-1 inhibited replication in multiple viruses, including equine arteritis virus, porcine epidemic diarrhea virus, pseudorabies virus, Japanese encephalitis virus, and classical swine fever virus, suggesting its potential broad application for antiviral strategies. Our findings provide insight into the important role of Gal-1 in PRRSV pathogenesis and its potential use as a novel therapeutic target against PRRSV infection.

Currently, various porcine reproductive and respiratory syndrome virus (PRRSV) variants emerged worldwide with different genetic characteristics and pathogenicity, increasing the difficulty of PRRS control. In this study, a PRRSV strain named HBap4-2018 was isolated from swine herds suffering severe respiratory disease with high morbidity in Hebei Province of China in 2018. The genome of HBap4-2018 is 15,003 nucleotides in length, and compared with NADC30-like PRRSV, nsp2 of HBap4-2018 has an additional continuous deletion of five amino acids. Phylogenetic analysis based on complete genome and ORF5 showed that HBap4-2018 belonged to lineage 8 of PRRSV-2, which was characterized by highly variable genome. However, HBap4-2018 was classified into lineage 1 based on phylogenetic analysis of nsp2, sharing higher amino acid homology (85.3%–85.5%) with NADC30-like PRRSV. Further analysis suggested that HBap4-2018 was a novel natural recombinant PRRSV with three recombinant fragments in the genome, of which highly pathogenic PRRSV (HP-PRRSV) served as the major parental strains, while NADC30-like PRRSV served as the minor parental strains. Five recombination break points were identified in nsp2, nsp3, nsp5, nsp9 and ORF6, respectively, presenting a novel recombinant pattern in the genome. Piglets inoculated with HBap4-2018 presented typical clinical signs with a mortality rate of 60%. High levels of viremia and obvious macroscopic and histopathological lesions in the lungs were observed, revealing the high pathogenicity of HBap4-2018 in piglets. SUPPLEMENTARY INFORMATION: The online version contains supplementary material available at 10.1007/s12250-021-00453-0.

Porcine reproductive and respiratory syndrome (PRRS) is one of the major drivers of economic loss in the swine industry worldwide. In commercial pig production, vaccination is the first option in an attempt to control infectious diseases. Pigs are therefore often immunized with different vaccines, and almost all of them are delivered via the intramuscular (IM) route. However, the IM injection may result in physical damage, stress reactions, and is labor demanding. An alternative route is urgently needed to reduce the disadvantages of conventional vaccination. In this study, a needle-free intradermal (ID) delivery system was evaluated for delivering a live PRRS vaccine as compared with the traditional needle-syringe method. Fifty-two 4-week-old piglets were divided into six groups: piglets in groups A-C were immunized using ID delivery system with 104, 105 and 106 TCID50 of PRRS candidate vaccine strain rHN-NP49, respectively; piglets in group D were immunized IM with 105 TCID50 of rHN-NP49; and group E and F were used as challenge and control groups, respectively. At 28 days post vaccination, piglets in group A to E were challenged with a lethal dose of highly-pathogenic PRRSV. Similar results were found in viremia and antibody response among the ID and IM groups during the immunization stage. After challenge, similar results were found in average body weight gain, viral shedding, serum viral load, and clinical score among the immunization groups, with a higher protection ratio in the ID group compared with IM group with the same immunization dose. These results demonstrated that the ID delivery system could provide similar or even better protection compared with IM route, and could be an effective route for PRRS vaccination.

Meishan pigs tend to have higher disease resistance than commercial breeds, although more studies are needed to confirm this difference. This study compared intestinal barrier function between Meishan and crossbred neonatal piglets to provide guidance for both the breeding and nutritional regulation of pigs. Six Meishan piglets and 6 Duroc × (Landrace × Yorkshire) crossbred neonatal piglets (all with normal birth weights) were obtained and allocated into the MEIS and CROSS groups, respectively. Intestinal morphology, goblet cell numbers, antioxidant enzyme activity, and cytokine gene and tight junction protein expression were assessed. The results showed that BW was lower in the MEIS group than in the CROSS group (P < 0.01). The relative lengths of the duodenum (P < 0.05), jejunum (P < 0.01) and ileum (P < 0.01) in the MEIS group were higher than those in the CROSS group. Compared with the CROSS group, the MEIS group exhibited shorter villus lengths in the duodenum and jejunum (P < 0.01), a shallower crypt depth in the ileum (P < 0.001) and denser and longer microvilli in the intestine. The numbers of GCs in the duodenum (P < 0.01) and jejunum (P < 0.001) and the activity levels of glutathione peroxidase (P < 0.05) in the jejunum and of catalase (P < 0.01) and superoxide dismutase (P < 0.01) in the ileum were higher in the MEIS group than in the CROSS group. Compared with the CROSS group, the MEIS group exhibited higher gene expression levels of interleukin (IL) 4 and interferon γ (IFNγ) in the jejunum (P < 0.05); IL2 (P < 0.05), IL4 (P < 0.01) and IFNγ (P < 0.001) in the ileum; and mucin 2 (P < 0.01) and occludin (P < 0.05) in the duodenum. In conclusion, Meishan neonatal piglets showed lower birth weights but higher intestinal barrier function than crossbred piglets.

Porcine reproductive and respiratory syndrome virus (PRRSV), which seriously endangers the world pig industry, invades host cells through receptor-mediated endocytosis involving clathrin. CD163 is an essential receptor for PRRSV during its infection of cells. The scavenger receptor cysteine-rich 5 (SRCR5) domain of the CD163 molecule is necessary for PRRSV infection, and interacts with glycoproteins GP2a and GP4 of PRRSV, allowing the virus to infect the host cells. In this study, a monoclonal antibody (mAb) against the SRCR5-6 region of porcine CD163 was developed, and the target epitope of the mAb was determined as 497TWGTVCDSDF506, which is directly adjacent to the ligand-binding pocket (LBP) domain (487-495aa) of CD163. Further study indicated that the mAb could partially block PRRSV infection of its target cells, pulmonary alveolar macrophages. The mAb developed in the study may provide a foundation of antiviral therapy for PRRSV.

Selenised glucose (SeGlu) is a newly invented organic selenium compound being synthesised through the selenisation reaction of glucose with NaHSe. We hypothesised that glucose could be used as a carrier for the stable low-valent organoselenium to enhance the selenium concentrations of eggs. To probe the effects of SeGlu on production performances of laying hens, egg selenium concentration, egg quality, and antioxidant indexes, 360 Hy-Line Brown laying hens were randomly assigned to three treatment groups fed with a basal diet alone or the diet supplemented with 5 or 10 mg/kg of Se from SeGlu. The results showed that SeGlu treatment not only enhanced (P < 0.001) the Se concentration in albumen and yolks, glutathione peroxidase activity, and total antioxidant capacity of eggs but also increased (P = 0.032) the Haugh unit of eggs being stored for 2 weeks, while the production performances and egg qualities of fresh eggs were not affected. Moreover, SeGlu supplementation linearly (P < 0.001) increased the scavenging ability of superoxide radicals in eggs. Briefly, SeGlu can enhance the selenium deposition and antioxidant activity of eggs, thereby meeting the nutritional requirement for Se-deficient humans.

Porcine reproductive and respiratory syndrome virus (PRRSV) is one of the most important pathogens in the swine industry worldwide. Our previous study had indicated that proprotein convertase subtilisin/kexin type 9 (PCSK9) was a responsive gene in porcine alveolar macrophages (PAMs) upon PRRSV infection. However, whether PCSK9 impacts the PRRSV replication and how the PRRSV modulates host PCSK9 remains elusive. Here, we demonstrated that PCSK9 protein suppressed the replication of both type-1 and type-2 PRRSV species. More specifically, the C-terminal domain of PCSK9 was responsible for the antiviral activity. Besides, we showed that PCSK9 inhibited PRRSV replication by targeting the virus receptor CD163 for degradation through the lysosome. In turn, PRRSV could down-regulate the expression of PCSK9 in both PAMs and MARC-145 cells. By screening the nonstructural proteins (nsps) of PRRSV, we showed that nsp11 could antagonize PCSK9&rsquo, s antiviral activity. Furthermore, mutagenic analyses of PRRSV nsp11 revealed that the endoribonuclease activity of nsp11 was critical for antagonizing the antiviral effect of PCSK9. Collectively, our data provide further insights into the interaction between PRRSV and the cell host and offer a new potential target for the antiviral therapy of PRRSV.

The widespread changes in forest cover caused by climatological and anthropogenic factors can influence the forest ecosystem and climate system to a great extent. With the increasing availability of remote sensing data, monitoring of forest changes at high temporal resolution and on various scales is becoming more realistic. Though several methods based on time series data have been used to detect forest disturbance, there are few studies paying attention to boreal areas where the forest is significant in regulating the global carbon cycle and biogeophysical processes. In this paper, we present a robust method of Breaks Detection Based On Polynomial Model (BDPM) to track boreal (e.g. Lesser Khingan Mountains) deforestation and forest fires based on the MODIS and Landsat TM time series data. Compared with the previous methods, the BDPM offers the following advantages: (1) Fitting of the polynomial model using the seasonal variation of forests in the whole region instead of a single pixel to avoid error accumulation; (2) to avoid confusion between vegetation change due to climate changes and abrupt forest disturbances, we segmented the long-time NDVI series data into 12 seasonal cycles and simulated the temporal variations in each seasonal cycle.

Numerous studies have documented the effects of irrigation on local, regional, and global climate. However, most studies focused on the cooling effect of irrigated dryland in semiarid or arid regions. In our study, we focused on irrigated paddy fields in humid regions at mid to high latitudes and estimated the effects of paddy field expansion from rain-fed farmland on local temperatures based on remote sensing and observational data. Our results revealed much significant near-surface cooling in spring (May and June) rather than summer (July and August) and autumn (September), which was −2.03 K, −0.73 K and −1.08 K respectively. Non-radiative mechanisms dominated the local temperature response to paddy field expansion from rain-fed farmland in the Sanjiang Plain. The contributions from the changes to the combined effects of the non-radiative process were 123.6%, 95.5%, and 66.9% for spring (May and June), summer (July and August), and autumn (September), respectively. Due to the seasonal changes of the biogeophysical properties for rain-fed farmland and paddy fields during the growing season, the local surface temperature responses, as well as their contributions, showed great seasonal variability. Our results showed that the cooling effect was particularly obvious during the dry spring instead of the warm, wet summer, and indicated that more attention should be paid to the seasonal differences of these effects, especially in a region with a relatively humid climate and distinct seasonal variations.

The purified whole-virus proteins derived from A/swine/Shanghai/1/2014 (H1N1) (SH1) were chosen to immunize BALB/c mice to prepare the monoclonal antibody (MAb) against hemagglutinin (HA) protein of an European avian-like (EA) H1N1 swine influenza virus (SIV). After cloning three times by limiting dilution, one strain of hybridoma cells named 3C7 secreting anti-HA protein MAb was obtained by hybridoma technique. The results of indirect immunofluorescence assay and western blot analyses showed that the MAb 3C7 specifically reacted with the HA protein of EA H1N1 SIV. This work indicated that the MAb 3C7 would be a valuable tool as a specific diagnostic reagent for SIV epidemiological surveys and identification of HA protein epitopes of the EA H1N1 SIVs in the future.

Highly pathogenic porcine reproductive and respiratory syndrome (HP-PRRS), which emerged in China in 2006, was characterized by high fever, high morbidity and high mortality. The causative agent of the disease was a highly pathogenic variant of porcine reproductive and respiratory syndrome virus (also called HP-PRRSV), which has a discontinuous deletion of 1 + 29 amino acids (aa) in the Nsp2 coding region, compared to classical PRRSV. In 2014, fattened pigs on a pig farm in Jiangxi Province suffered from clinical symptoms of high fever, dyspnoea and death. A PRRSV, termed JX2014T2, was isolated from samples of the dead pigs. Genomic analysis of the isolated PRRSV indicated that the genome of the virus was 14,960 bp in length and belonged to the North American genotype. In the Nsp2-coding region, there was a discontinuous deletion of 1 + 29 aa, similar to HP-PRRSV; however, an additional continuous deletion of 120 amino acids between aa 628 and 747 was found. Further analysis of the pathogenicity of PRRSV JX2014T2 was performed in piglets, and the results indicated that all infected piglets suffered from typical clinical symptoms of PRRS, such as high fever, cough, mental depression, anorexia, dyspnoea and palpebral swelling and died within 15 days postinfection (dpi). This demonstrated that the newly isolated PRRSV JX2014T2 strain containing an additional deletion of 120 aa is highly pathogenic to piglets, suggesting that a highly pathogenic variant with new genetic features is circulating in China.

Significant differences exist between the highly pathogenic (HP) porcine reproductive and respiratory syndrome virus (PRRSV) and its attenuated pathogenic (AP) strain in the ability to infect host cells. The mechanisms by which different virulent strains invade host cells remain relatively unknown. In this study, pulmonary alveolar macrophages (PAMs) were infected with HP-PRRSV (HuN4) and AP-PRRSV (HuN4-F112) for 24 h, then harvested and subjected to label-free quantitative mass spectrometry. A total of 2849 proteins were identified, including 95 that were differentially expressed. Among them, 26 proteins were located on the membrane. The most differentially expressed proteins were involved in response to stimulus, metabolic process and immune system process, which mainly had the function of binding and catalytic activity. CD163, Vimentin and nmII as well as detected proteins were assessed together by string analysis, which elucidated a potentially different infection mechanism. According to the function annotations, PRRSV with different virulence may mainly differ in immunology, inflammation, immune evasion as well as cell apoptosis. This is the first attempt to explore the differential characteristics between HP-PRRSV and its attenuated PRRSV infected PAMs focusing on membrane proteins which will be of great help to further understand the different infective mechanisms of HP-PRRSV and AP-PRRSV. This article is protected by copyright. All rights reserved

SAMHD1 is a type I interferon (IFN) inducible host innate immunity restriction factor that inhibits an early step of the viral life cycle. The underlying mechanisms of SAMHD1 transcriptional regulation remains elusive. Here, we report that inducing SAMHD1 upregulation is part of an early intrinsic immune response via TLR3 and RIG-I/MDA5 agonists that ultimately induce the nuclear translocation of the interferon regulation factor 3 (IRF3) protein. Further studies show that IRF3 plays a major role in upregulating endogenous SAMHD1 expression in a mechanism that is independent of the classical IFN-induced JAK-STAT pathway. Both overexpression and activation of IRF3 enhanced the SAMHD1 promoter luciferase activity, and activated IRF3 was necessary for upregulating SAMHD1 expression in a type I IFN cascade. We also show that the SAMHD1 promoter is a direct target of IRF3 and an IRF3 binding site is sufficient to render this promoter responsive to stimulation. Collectively, these findings indicate that upregulation of endogenous SAMHD1 expression is attributed to the phosphorylation and nuclear translocation of IRF3 and we suggest that type I IFN induction and induced SAMHD1 expression are coordinated.

The sterile alpha motif and HD domain 1 (SAMHD1) protein has been identified as a novel innate immunity restriction factor that participates in processes crucial to the viral life cycle. In the present study, we describe a procedure to generate monoclonal antibodies (MAbs) against porcine SAMHD1 and investigate its characteristics to analyze the expression of endogenous SAMHD1. The open reading frame of porcine SAMHD1 was cloned into the prokaryotic expression vector pCold-TF DNA to construct a recombinant plasmid pcold-pSAMHD1 and induce expression of recombinant porcine SAMHD1 protein by IPTG in Escherichia coli Rosetta. The purified recombinant porcine SAMHD1 protein was used to prepare MAbs of SAMHD1. After subcloning five times hybridoma cell clones expressing SAMHD1, MAbs were generated. Western blot analysis and indirect immunofluorescence assay showed that the overexpressed porcine SAMHD1 in 293T cells and endogenous SAMHD1 protein in porcine cell lines could be specifically recognized by the MAbs produced in this study. In conclusion, specific MAbs of porcine SAMHD1 are reported, and these MAbs provide a valuable tool for further studies of SAMHD1-mediated signaling in virus-infected cells to elucidate the underlying antiviral mechanism.

Highlights • Over-expression of the miR-26 family strongly inhibited PRRSV replication in two PRRSV genotypes in a dose-dependent manner. • Luciferase reporter gene assay showed miR-26a does not target PRRSV genome. • Over-expression of miR-26a increases the expression of type I IFN and the IFN-stimulated genes MX1 and ISG15 during PRRSV infection., MicroRNAs (miRNAs) play important roles in viral infections, especially by modulating the expression of cellular factors essential to viral replication or the host innate immune response to infection. To identify host miRNAs important to controlling porcine reproductive and respiratory syndrome virus (PRRSV) infection, we screened 15 miRNAs that were previously implicated in innate immunity or antiviral functions. Over-expression of the miR-26 family strongly inhibited PRRSV replication in vitro, as shown by virus titer assays, Western blotting, and qRT-PCR assays. MiR-26a inhibited the replication of both type 1 and type 2 PRRSV strains. Mutating the seed region of miR-26 restored viral titers. Luciferase reporters showed that miR-26a does not target the PRRSV genome directly but instead affects the expression of type I interferon and the IFN-stimulated genes MX1 and ISG15 during PRRSV infection. These results demonstrate the important role of miR-26a in modulating PRRSV infection and also support the possibility of using host miR-26a to achieve RNAi-mediated antiviral therapeutic strategies.

The objective of this study was to determine the effects of dietary n-6:n-3 polyunsaturated fatty acid (PUFA) ratios on heterophil:lymphocyte (H:L) ratios and T cell subsets in the peripheral blood of the growing Yangzhou goose. A total of 160 healthy Yangzhou geese (0.407 ± 0.023 kg), aged 21 d, were randomly divided into 4 groups consisting of diets with different n-6:n-3 PUFA ratios: 12:1, 9:1, 6:1, and 3:1. White blood cell counts and H:L ratios were measured at 42, 56, and 70 d of age; T lymphocyte subsets were also analyzed using monoclonal antibodies and flow cytometry simultaneously. The results showed that differences were found in white blood cells between groups (P0.05) but not within groups (P0.05), and white blood cells of 70-d-old geese were significantly higher than those of other ages within the same group. The H:L ratios of all groups decreased as age increased, and differences were found in 6:1 and 3:1 groups between ages (P0.05). As for the comparison of H:L ratio between treatments, 6:1 and 3:1 were lower than the other 2 groups at 42 or 70 d of age (P0.05), and 3:1 was lower than the other 3 groups at 56 d of age (P0.05). Moreover, H:L ratios had a tendency to decrease with the decrease of n-6:n-3 PUFA ratios. Furthermore, cluster of differentiation (CD) 3(+) and CD8(+) increased gradually, whereas CD4(+) and CD4(+):CD8(+) ratios first increased and then decreased as age increased. Differences were detected in T cell subsets among ages (P0.05). In addition, CD4(+) and CD4(+):CD8(+) ratios were affected remarkably by n-6:n-3 PUFA ratios but CD3(+) and CD8(+) were not, and 6:1 had much higher CD4(+) percentage and CD4(+):CD8(+) ratio compared with the other groups (P0.05). Taken together, diets containing low n-6:n-3 PUFA ratios could decrease H:L ratios in the peripheral blood. Additionally, CD4(+) percentage and CD4(+):CD8(+) ratio were much higher in the 6:1 group, which could be relevant for improving positive immune responses for Yangzhou goslings from 42 to 70 d of age.

The highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) is still a serious threat to the swine industry. However, the pathogenic mechanism of HP-PRRSV remains unclear. We infected host porcine alveolar macrophages (PAMs) with the virulent HuN4 strain and the attenuated HuN4-F112 strain and then utilized fluorescent two-dimensional difference gel electrophoresis (2D-DIGE) to screen for intracellular proteins that were differentially expressed in host cells infected with the two strains. There were 153 proteins with significant different expression (P