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Zika virus (ZIKV) is an emerging mosquito-borne flavivirus that poses significant threats to global public health. Macrophages and dendritic cells are both key sentinel cells in the host immune response and play critical roles in the pathogenesis of flavivirus infections. Recent studies showed that ZIKV could productively infect monocyte-derived dendritic cells (moDCs), but the role of macrophages in ZIKV infection remains incompletely understood. In this study, we first compared ZIKV infection in monocyte-derived macrophages (MDMs) and moDCs derived from the same donors. We demonstrated that while both MDMs and moDCs were susceptible to epidemic (Puerto Rico) and pre-epidemic (Uganda) strains of ZIKV, virus replication was largely restricted in MDMs but not in moDCs. ZIKV induced significant apoptosis in moDCs but not MDMs. The restricted virus replication in MDMs was not due to inefficient virus entry but was related to post-entry events in the viral replication cycle. In stark contrast with moDCs, ZIKV failed to inhibit STAT1 and STAT2 phosphorylation in MDMs. This resulted in the lack of efficient antagonism of the host type I interferon-mediated antiviral responses. Importantly, depletion of STAT2 but not STAT1 in MDMs significantly rescued the replication of ZIKV and the prototype flavivirus yellow fever virus. Overall, our findings revealed a differential interplay between macrophages and dendritic cells with ZIKV. While dendritic cells may be exploited by ZIKV to facilitate virus replication, macrophages restricted ZIKV infection.

Background Among various cooking methods, steaming is favored by many because it can cause less damage to nutrient components in muscle, retain the inherent food flavor, and reduce the generation of harmful substances. Steaming conditions are closely related to fish flavor, fat and protein oxidation, and digestibility. Results The black carp steamed for 4 to 14 min was studied in this article based on sensory assessment, electronic tongue, free amino acids, adenosine triphosphate (ATP)-related compounds, total nitrogen and non-protein nitrogen to explore the effect of steaming time on the taste substances and protein degradation of pickled black carp. The experimental result showed that the meat steamed within 8 min tasted better, showing high tastiness. The sensory assessment score increased significantly to the maximum value of 82.33 at 6 min. The content of umami and sweet amino acids increased significantly to the maximum value of 1.6801 g kg-1 at 6 min. In the meantime, the IMP (inosine monophosphate) content was 1.9128 g kg-1 , with its taste activity value (TAV) reaching 7.65, which proved that IMP affected the taste most. Furthermore, the total nitrogen content was 30.77 g kg-1 , which meant protein degraded a great deal. Based on equivalent umami concentration (EUC) and its TAV, the meat tasted best at 6-8 min. The longer the steaming time, the faster the protein degradation and the more the flavor precursors. Conclusion The black carp pickled products (with a weight of 20 g, with the size of 3 cm × 3 cm × 2 cm) is suggested to be steamed for 6 to 8 min. This conclusion provides a theoretical basis for its better taste quality. © 2020 Society of Chemical Industry.

PIM2 (proviral integration site for Moloney murine leukemia virus 2) kinase plays an important role as an oncogene in multiple cancers, such as leukemia, liver, lung, myeloma, prostate and breast cancers. PIM2 is largely expressed in both leukemia and solid tumors, and it promotes the transcriptional activation of genes involved in cell survival, cell proliferation, and cell-cycle progression. Many tumorigenic signaling molecules have been identified as substrates for PIM2 kinase, and a variety of inhibitors have been developed for its kinase activity, including SMI-4a, SMI-16a, SGI-1776, JP11646 and DHPCC-9. Here, we summarize the signaling pathways involved in PIM2 kinase regulation and PIM2 mechanisms in various neoplastic diseases. We also discuss the current status and future perspectives for the development of PIM2 kinase inhibitors to combat human cancer, and PIM2 will become a therapeutic target in cancers in the future.

Zika virus (ZIKV) is an emerging flavivirus that may be associated with congenital anomalies in infected fetuses and severe neurological and genital tract complications in infected adults. Currently, antiviral treatments to revert these ZIKV-induced complications are lacking. ZIKV infection has recently been suggested to upregulate the host unfolded protein response, which may contribute to the congenital neurological anomalies. As an extension from these findings, we thoroughly investigated the ZIKV-induced unfolded protein response using a combination of the neuronal cell line, induced pluripotent stem cell-derived human neuronal stem and progenitor cells, and an interferon receptor-deficient A129 mouse model. Our results revealed a critical contribution of the inositol-requiring enzyme-1 (IRE1) arm of the unfolded protein response to ZIKV-induced neurological and testicular complications. Importantly, the inhibition of the IRE1 signaling pathway activation with KIRA6 (kinase-inhibiting RNAse attenuator 6), a selective small molecule IRE1 inhibitor that promotes cell survival, potently reverted the ZIKV-induced perturbations of the key gene expressions associated with neurogenesis and spermatogenesis in vitro and in vivo, highlighting the potential of IRE1 inhibition as a novel host-targeting antiviral strategy in combating against ZIKV-induced neurological and testicular pathologies.

BackgroundSevere acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is an emerging coronavirus that has resulted in more than 2 000 000 laboratory-confirmed cases including over 145 000 deaths. Although SARS-CoV-2 and SARS-CoV share a number of common clinical manifestations, SARS-CoV-2 appears to be highly efficient in person-to-person transmission and frequently causes asymptomatic or presymptomatic infections. However, the underlying mechanisms that confer these viral characteristics of high transmissibility and asymptomatic infection remain incompletely understood.MethodsWe comprehensively investigated the replication, cell tropism, and immune activation profile of SARS-CoV-2 infection in human lung tissues with SARS-CoV included as a comparison.ResultsSARS-CoV-2 infected and replicated in human lung tissues more efficiently than SARS-CoV. Within the 48-hour interval, SARS-CoV-2 generated 3.20-fold more infectious virus particles than did SARS-CoV from the infected lung tissues (P < .024). SARS-CoV-2 and SARS-CoV were similar in cell tropism, with both targeting types I and II pneumocytes and alveolar macrophages. Importantly, despite the more efficient virus replication, SARS-CoV-2 did not significantly induce types I, II, or III interferons in the infected human lung tissues. In addition, while SARS-CoV infection upregulated the expression of 11 out of 13 (84.62%) representative proinflammatory cytokines/chemokines, SARS-CoV-2 infection only upregulated 5 of these 13 (38.46%) key inflammatory mediators despite replicating more efficiently.ConclusionsOur study provides the first quantitative data on the comparative replication capacity and immune activation profile of SARS-CoV-2 and SARS-CoV infection in human lung tissues. Our results provide important insights into the pathogenesis, high transmissibility, and asymptomatic infection of SARS-CoV-2.

Down-regulation of CaMET1-like1 in pepper leads to DNA hypomethylation and premature fruit ripening, and interactions between DNA methylation and phytohormones appear to fine-tune the ripening process., There is growing evidence to suggest that epigenetic tags, especially DNA methylation, are critical regulators of fruit ripening. To examine whether this is the case in sweet pepper (Capsicum annuum) we conducted experiments at the transcriptional, epigenetic, and physiological levels. McrBC PCR, bisulfite sequencing, and real-time PCR demonstrated that DNA hypomethylation occurred in the upstream region of the transcription start site of some genes related to pepper ripening at the turning stage, which may be attributed to up-regulation of CaDML2-like and down-regulation of CaMET1-like1, CaMET1-like2, CaCMT2-like, and CaCMT4-like. Silencing of CaMET1-like1 by virus-induced gene silencing led to DNA hypomethylation, increased content of soluble solids, and accumulation of carotenoids in the fruit, which was accompanied by changes in expression of genes involved in capsanthin/capsorubin biosynthesis, cell wall degradation, and phytohormone metabolism and signaling. Endogenous ABA increased during fruit ripening, whereas endogenous IAA showed an opposite trend. No ethylene signal was detected during ripening. DNA hypomethylation repressed the expression of auxin and gibberellin biosynthesis genes as well as cytokinin degradation genes, but induced the expression of ABA biosynthesis genes. In mature-green pericarp, exogenous ABA induced expression of CaDML2-like but repressed that of CaCMT4-like. IAA treatment promoted the transcription of CaMET1-like1 and CaCMT3-like. Ethephon significantly up-regulated the expression of CaDML2-like. Treatment with GA3 and 6-BA showed indistinct effects on DNA methylation at the transcriptional level. On the basis of the results, a model is proposed that suggests a high likelihood of a role for DNA methylation in the regulation of ripening in the non-climacteric pepper fruit.

Grass carp is the most important freshwater fish in China, and salting is the main processing method. This study investigated the effect of brine injection on flavor of grass carp muscle compared with brining at low temperature (4°C) and room temperature (20°C) respectively. Salt content, free amino acids, nucleotides, equivalent umami concentration (EUC) and texture were evaluated as flavor indicators during salting. The results showed that brine injection groups reached equilibrium in 3 h and 5 h at 20°C and 4°C, respectively. Brining groups reached equilibrium in 5 h and 7 h at 20°C and 4°C, respectively. The equilibrium salt content was about 1.76%, and there was no significant deterioration in terms of the other indicators mentioned above by two different methods. The results of this research indicate the feasibility and the great economic potential of brine injection, and also provide the theoretical basis to promote this method in processing of aquatic products.

Avian hepatitis E virus (HEV) is the major causative pathogen of the big liver and spleen disease, hepatitis-splenomegaly syndrome, and hepatic rupture hemorrhage syndrome. Until now, there are 6 different avian HEV genotypes that infect chickens have been reported worldwide. Epidemiologic investigations of the avian HEV demonstrated that avian HEV has been widely spread in China in recent years. In this study, an avian HEV named YT-aHEV was obtained from white-feathered broilers using LMH cells by virus isolation assay in Shandong province, China. The complete genome consists of 6656-nt excluding the poly(A) tail. The isolate was highly similar to the CaHEV strain and segregated into the same branch belonging to avian HEV genotype 3. Indirect immunofluorescence using capsid protein-specific polyclonal antibodies confirmed that YT-aHEV could establish productive infection and replicate stably in LMH cells. Furthermore, an in vivo avian HEV infection model was established successfully in specific pathogen-free chicken embryos by intravenous experiments. In the present study, we demonstrate an avian HEV infection associated with liver lesions of hemorrhage and swelling by LMH cells for the first time in a white-feather broiler flock in China. This research also provides a new diagnosis method for detection of avian HEV, which laid a foundation for the understanding of pathogenicity and molecular biology of this virus for further study.

Background Wildtype mice are not susceptible to SARS-CoV-2 infection. Emerging SARS-CoV-2 variants, including B.1.1.7, B.1.351, P.1, and P.3, contain mutations in spike that has been suggested to associate with an increased recognition of mouse ACE2, raising the postulation that these SARS-CoV-2 variants may have evolved to expand species tropism to wildtype mouse and potentially other murines. Our study evaluated this possibility with substantial public health importance. Methods We investigated the capacity of wildtype (WT) SARS-CoV-2 and SARS-CoV-2 variants in infecting mice (Mus musculus) and rats (Rattus norvegicus) under in vitro and in vivo settings. Susceptibility to infection was evaluated with RT-qPCR, plaque assays, immunohistological stainings, and neutralization assays. Findings Our results reveal that B.1.1.7 and other N501Y-carrying variants but not WT SARS-CoV-2 can infect wildtype mice. High viral genome copies and high infectious virus particle titres are recovered from the nasal turbinate and lung of B.1.1.7-inocluated mice for 4-to-7 days post infection. In agreement with these observations, robust expression of viral nucleocapsid protein and histopathological changes are detected from the nasal turbinate and lung of B.1.1.7-inocluated mice but not that of the WT SARS-CoV-2-inoculated mice. Similarly, B.1.1.7 readily infects wildtype rats with production of infectious virus particles. Interpretation Our study provides direct evidence that the SARS-CoV-2 variant, B.1.1.7, as well as other N501Y-carrying variants including B.1.351 and P.3, has gained the capability to expand species tropism to murines and public health measures including stringent murine control should be implemented to facilitate the control of the ongoing pandemic. Funding A full list of funding bodies that contributed to this study can be found in the Acknowledgements section.

Chicken infectious anemia virus (CIAV) can be transmitted by contaminated live vaccines, and causes huge economic losses. This study evaluated the contamination status of CIAV in 24 batches of vaccines by recombinase-aided amplification assay (RAA), fluorescence quantitative PCR and dot blot assay, and then determined a contaminated avian pox attenuated vaccine. The whole genome of the CIAV contaminant was then sequenced and named as JS2020-PFV (Genbank accession number: MW234428, 2296bp). It showed 94.5–99.9% identities with reference strains, and shared the closest evolution relationship with AB1K which was isolated from a chicken farm in Turkey. All these suggested that the use of CIAV contaminated live vaccine may be the reason for its epidemic in poultry.

BackgroundLive attenuated vaccines have been extensively used to prevent infectious disease in poultry flocks. However, exogenous virus contamination in attenuated vaccines had been reported several times in the past, which brought enormous threat to poultry production and diseases prevention and control. Recently, an attenuated vaccine against Newcastle Disease produced in China was detected contamination with chicken infectious anemia virus (CIAV) in a routine inspection for exogenous virus. To understand the multiple routes of transmission of CIAV, and to better formulate correct prevention and control, it is necessary to find out the possible source of this contamination.Methodslood samples of SPF chickens that generated vaccines were collected to investigate CIAV antibody titers by ELISA test. Then, 14- to 18-day-old SPF chicken embryos (n=40) were randomly selected, DNA was extracted and detected by quantitative real-time PCR and nucleic acid dot hybridization assays. To further investigate the molecular features of the CIAV isolate, the complete genome of CIAV was amplified and analyzed.ResultsThe results showed both SPF chickens and embryos for vaccine preparation were CIAV-positive. In addition, the full-length genome sequences of CIAV from vaccines and SPF chicken embryos were consisted of 2,298 nucleotides (nt) with 100% homology, named as SDSPF2020 (Genbank accession number: MW660821). It demonstrated 95.7%-99.6% homology with the complete nucleotide sequences of reference strains, and shared the closest evolution relationship with the Chinese strain HLJ15125. Conclusion This study illustrated that vertical transmission of CIAV from SPF chickens and embryos was an important way for exogenous virus contamination in vaccine production. As such, vaccine quality monitoring and health control are significant in the poultry industry from an environmental safety point of view.

Leaf size and flatness directly affect photosynthesis and are closely related to agricultural yield. The final leaf size and shape are coordinately determined by cell proliferation, differentiation, and expansion during leaf development. Lettuce (Lactuca sativa L.) is one of the most important leafy vegetables worldwide, and lettuce leaves vary in shape and size. However, the molecular mechanisms of leaf development in lettuce are largely unknown. In this study, we showed that the lettuce APETALA2 (LsAP2) gene regulates leaf morphology. LsAP2 encodes a transcriptional repressor that contains the conserved EAR motif, which mediates interactions with the TOPLESS/TOPLESS-RELATED (TPL/TPR) corepressors. Overexpression of LsAP2 led to small and crinkly leaves, and many bulges were seen on the surface of the leaf blade. LsAP2 physically interacted with the CINCINNATA (CIN)-like TEOSINTE BRANCHED1/CYCLOIDEA/PROLIFERATING CELL FACTOR (TCP) transcription factors and inhibited their transcriptional activation activity. RNA sequencing analysis showed that LsAP2 affected the expression of auxin- and polarity-related genes. In addition, LsAP2 directly repressed the abaxial identity gene KANADI2 (LsKAN2). Together, these results indicate that LsAP2 regulates leaf morphology by inhibiting CIN-like TCP transcription factors and repressing LsKAN2, and our work provides insights into the regulatory mechanisms of leaf development in lettuce.

Background: The use of Reticuloendotheliosis virus (REV) from contaminated live virus vaccine is suspected to be one of the most important causes of massive outbreaks of Reticuloendotheliosis in China. Methods: In this study, we established a droplet digital PCR (ddPCR) detection method for REV and compared its sensitivity to different methods to detect REV contamination in a vaccine. Results: The results indicated that both quantitative PCR and dot-blot methods could detect REV contamination at a dose of 1 TCID50/1,000 feathers, whereas ddPCR could detect REV contamination at a dose of 0.1 TCID50/1,000 feathers, which is 1,000-fold more sensitive than conventional polymerase chain reaction detection (102 TCID50/1000 feathers). ddPCR not only exhibited the highest sensitivity but also proved extremely intuitive, especially to detect REV contamination in vaccines.Conclusions: The ddPCR method established in this study to detect REV contamination in vaccines can effectively detect and quantify low-dose REV contamination. This provides a new method for the rapid detection of REV contamination in various samples, especially vaccines.

Koi herpesvirus (KHV) is highly contagious and lethal to cyprinid fish, causing significant economic losses to the carp aquaculture industry, particularly to koi carp breeders. Vaccines delivered through intramuscular needle injection or gene gun are not suitable for mass vaccination of carp. So, the development of cost-effective oral vaccines that are easily applicable at a farm level is highly desirable. In this study, we utilized chitosan-alginate capsules as an oral delivery system for a live probiotic (Lactobacillus rhamnosus) vaccine, pYG-KHV-ORF81/LR CIQ249, expressing KHV ORF81 protein. The tolerance of the encapsulated recombinant Lactobacillus to various digestive environments and the ability of the probiotic strain to colonize the intestine of carp was tested. The immunogenicity and the protective efficacy of the encapsulated probiotic vaccine was evaluated by determining IgM levels, lymphocyte proliferation, expression of immune-related genes, and viral challenge to vaccinated fish. It was clear that the chitosan-alginate capsules protected the probiotic vaccine effectively against extreme digestive environments, and a significant level (P < 0.01) of antigen-specific IgM with KHV-neutralizing activity was detected, which provided a protection rate of ca. 85% for koi carp against KHV challenge. The strategy of using chitosan-alginate capsules to deliver probiotic vaccines is easily applicable for mass oral vaccination of fish. IMPORTANCE An oral probiotic vaccine, pYG-KHV-ORF81/LR CIQ249, encapsulated by chitosan-alginate capsules as an oral delivery system was developed for koi carp against koi herpesvirus (KHV) infection. This encapsulated probiotic vaccine can be protected from various digestive environments and maintain effectively high viability, showing a good tolerance to digestive environments. This encapsulated probiotic vaccine has a good immunogenicity in koi carp via oral vaccination, and a significant level of antigen-specific IgM was effectively induced after oral vaccination, displaying effective KHV-neutralizing activity. This encapsulated probiotic vaccine can provide effective protection for koi carp against KHV challenge, which is handling-stress free for the fish, cost effective, and suitable for the mass oral vaccination of koi carp at a farm level, suggesting a promising vaccine strategy for fish.

Coronaviruses have repeatedly crossed species barriers to cause epidemics1. “Pan-coronavirus” antivirals targeting conserved viral components involved in coronavirus replication, such as the extensively glycosylated spike protein, can be designed. Here we show that the rationally engineered H84T-banana lectin (H84T-BanLec), which specifically recognizes high-mannose found on viral proteins but seldom on healthy human cells2, potently inhibits the highly virulent MERS-CoV, pandemic SARS-CoV-2 and its variants, and other human-pathogenic coronaviruses at nanomolar concentrations. MERS-CoV-infected human DPP4-transgenic mice treated by H84T-BanLec have significantly higher survival, lower viral burden, and reduced pulmonary damage. Similarly, prophylactic or therapeutic H84T-BanLec is effective against SARS-CoV-2 in hamsters. Importantly, intranasally and intraperitoneally administered H84T-BanLec are comparably effective. Time-of-drug-addition assay shows that H84T-BanLec targets virus entry. Real-time structural analysis with high-speed atomic force microscopy depicts multi-molecular associations of H84T-BanLec dimers with the SARS-CoV-2 spike trimer. Single-molecule force spectroscopy demonstrates binding of H84T-BanLec to multiple SARS-CoV-2 spike mannose sites with high affinity, and that H84T-BanLec competes with SARS-CoV-2 spike for binding to cellular ACE2. Modelling experiments identify distinct high-mannose glycans in spike recognized by H84T-BanLec. The multiple H84T-BanLec binding sites on spike likely account for the activity against SARS-CoV-2 variants and the lack of resistant mutants. The broad-spectrum H84T-BanLec should be clinically evaluated in respiratory viral infections including COVID-19.

Bacteria are one of the main groups of organisms, which dynamically and closely participate in human health and disease development. With the integration of chemical biotechnology, bacteria have been utilized as an emerging delivery system for various biomedical applications. Given the unique features of bacteria such as their intrinsic biocompatibility and motility, bacteria-based delivery systems have drawn wide interest in the diagnosis and treatment of various diseases, including cancer, infectious diseases, kidney failure, and hyperammonemia. Notably, at the interface of chemical biotechnology and bacteria, many research opportunities have been initiated, opening a promising frontier in biomedical application. Herein, the current synergy of chemical biotechnology and bacteria, the design principles for bacteria-based delivery systems, the microbial modulation, and the clinical translation are reviewed, with a special focus on the emerging advances in diagnosis and therapy.

Norovirus (NoV), a major food-borne virus, causes non-bacterial acute gastroenteritis in humans. Berries are generally harvested from low-growing bushes by hand and are minimally processed before being sold to consumers. Therefore, the consumption of berries has been linked to numerous food-borne gastroenteritis outbreaks caused by NoV in many countries. We performed a survey of NoV contamination in commercial fresh/frozen berry fruits collected from 2016 to 2017 in the Heilongjiang Province, the main berry-producing area in China, using a TaqMan-based real-time reverse transcription-PCR assay. Among 900 frozen and 900 fresh domestic retail berry samples, the prevalence of NoV was 9% (81/900) and 12.11% (109/900), including 35.80% (29/81) and 29.36% (32/109) of genotype GI alone, 54.32% (44/81) and 60.55% (66/109) of GII alone, and 9.88% (8/81) and 10.09% (11/109) of both GI and GII, respectively. No NoV was detected among the 677 frozen berry samples for export. Thus, the occurrence of NoV contamination was significantly higher in domestic berries than in exported berries and higher in fresh berries than in frozen berries. This study highlights the need for further risk surveillance for NoV contamination in berries produced in the Heilongjiang Province and recommends region-extended monitoring of retail berries for NoV.

Background Pigeon crop has the unique ability to produce a nutrient rich substance termed pigeon ‘milk’ (PM), which has functional resemblance with the mammalian milk. Previous researches have demonstrated that a large number of exosomes and exosomal miRNAs exist in mammalian milk, and many of them are associated with immunity, growth and development. However, to date, little is known about the exosomes and exosomal miRNAs in PM. Results In this study, we isolated the exosomes from PM and used small RNA sequencing to investigate the distribution and expression profiles of exosomal miRNAs. A total of 301 mature miRNAs including 248 conserved and 53 novel miRNAs were identified in five lactation stages i.e. 1d, 5d, 10d, 15d, and 20d. From these, four top 10 conserved miRNAs (cli-miR-21-5p, cli-miR-148a-3p, cli-miR-10a-5p and cli-miR-26a-5p) were co-expressed in all five stages. We speculate that these miRNAs may have important role in the biosynthesis and metabolism of PM. Moreover, similar to the mammalian milk, a significant proportion of immune and growth-related miRNAs were also present and enriched in PM exosomes. Furthermore, we also identified 41 orthologous miRNAs group (giving rise to 81 mature miRNA) commonly shared with PM, human, bovine and porcine breast milk. Additionally, functional enrichment analysis revealed the role of exosomal miRNAs in organ development and in growth-related pathways including the MAPK, Wnt and insulin pathways. Conclusions To sum-up, this comprehensive analysis will contribute to a better understanding of the underlying functions and regulatory mechanisms of PM in squabs. Electronic supplementary material The online version of this article (10.1186/s12864-018-5201-0) contains supplementary material, which is available to authorized users.

Highly pathogenic coronaviruses including severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)1,2, Middle East respiratory syndrome coronavirus (MERS-CoV)3,4, and SARS-CoV-15 vary in their transmissibility and pathogenicity. However, infection by all three viruses result in substantial apoptosis in cell culture6-8 and in patient samples9-11, suggesting a potential link between apoptosis and the pathogenesis of coronaviruses. To date, the underlying mechanism of how apoptosis modulates coronavirus pathogenesis is unknown. Here we show that a cysteine-aspartic protease of the apoptosis cascade, caspase-6, serves as an essential host factor for efficient coronavirus replication. We demonstrate that caspase-6 cleaves coronavirus nucleocapsid (N) proteins, generating N fragments that serve as interferon (IFN) antagonists, thus facilitating virus replication. Inhibition of caspase-6 substantially attenuates the lung pathology and body weight loss of SARS-CoV-2-infected golden Syrian hamsters and improves the survival of mouse-adapted MERS-CoV (MERS-CoVMA)-infected human DPP4 knock-in (hDPP4 KI) mice. Overall, our study reveals how coronaviruses exploit a component of the host apoptosis cascade to facilitate their replication. These results further suggest caspase-6 as a potential target of intervention for the treatment of highly pathogenic coronavirus infections including COVID-19 and MERS.

Cigarette smoke extract (CSE), a complex mixture of compounds, contributes to a range of eye diseases; however, the underlying pathophysiological responses to tobacco smoke remain ambiguous. The purpose of the present study was to evaluate the cigarette smoke-induced phenotypic and transcriptomic changes in the corneal epithelium with a view to elucidating the likely underlying mechanism. Accordingly, for the first time, we characterized the genome-wide effects of CSE on the corneal epithelium. The ocular surface of the mice in the experimental groups was exposed to CSE for 1 h per day for a period of one week, while mice in the control group were exposed to preservative-free artificial tears. Corneal fluorescein staining, in vivo confocal microscopy and scanning electron microscopy were performed to examine the corneal ultrastructure. Transcriptome sequencing and bioinformatics analysis were performed followed by RT-qPCR to validate gene expression changes. The results indicate that CSE exposure disrupted the structural integrity of the superficial epithelium, decreased the density of microvilli, and compromised the corneal epithelial barrier intactness. RNA-seq revealed 667 differentially expressed genes, and functional analysis highlighted the enhancement of several biological processes such as antioxidant activity and the response to oxidative stress. Moreover, the Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis showed that glutathione metabolism and drug metabolism cytochrome P450 were the most relevant pathways contributing to the effects of CSE on the corneal epithelium. Protein–protein interaction (PPI) network analysis illustrated that GCLC, NQO1, and HMOX1 were the most relevant nodes. In conclusion, the present study indicates that CSE exposure induces changes in the phenotype and genotype of the corneal epithelium. The antioxidant response element is essential for counteracting the effects of cigarette smoke on this tissue layer. These results shed novel insights into how cigarette smoke damages this ocular surface.

BackgroundEndometriosis is a chronic hormonal inflammatory disease characterized by the presence of endometrial tissue (glands and stroma) outside the uterus. Endometriosis seriously affects the physical health of women of childbearing age, often causes infertility, and affects the body and mind of patients and their families.MethodsWe examined the effect of HSF1 on endometriosis through cell count, scratch and clone formation experiments. We used real-time quantitative PCR and western blotting to detect the effect of HSF1 on mRNA and protein of endometriosis cells. Collect the cell culture medium and Glucose levels and lactate levels were determined using a glucose (GO) assay kit and a lactate assay kit. Furthermore, we established a mouse model of endometriosis, and the effect of HSF1 on endometriosis was observed by inhibiting HSF1 with KRIBB11 in the mice.ResultsHSF1 is highly expressed in endometriosis and plays an indispensable role in endometriosis development in both cell and animal studies. We found that HSF1 promotes endometriosis development and glucose consumption and lactic acid production. Further research showed that HSF1 functions in endometriosis by up-regulating PFKFB3, a key enzyme in glycolysis. And the HSF1 inhibitor KRIBB11 can abrogate all of the above experimental effects both in vivo and in vitro. ConclusionsOur study shows that HSF1 plays a significant role in the occurrence and development of endometriosis, which may become a new target for the treatment of endometriosis and provide a new idea for the clinical treatment of endometriosis.

Infection of highly pathogenic coronaviruses triggers apoptosis that can be targeted to reduce disease severity., Infection by highly pathogenic coronaviruses results in substantial apoptosis. However, the physiological relevance of apoptosis in the pathogenesis of coronavirus infections is unknown. Here, with a combination of in vitro, ex vivo, and in vivo models, we demonstrated that protein kinase R–like endoplasmic reticulum kinase (PERK) signaling mediated the proapoptotic signals in Middle East respiratory syndrome coronavirus (MERS-CoV) infection, which converged in the intrinsic apoptosis pathway. Inhibiting PERK signaling or intrinsic apoptosis both alleviated MERS pathogenesis in vivo. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and SARS-CoV induced apoptosis through distinct mechanisms but inhibition of intrinsic apoptosis similarly limited SARS-CoV-2– and SARS-CoV–induced apoptosis in vitro and markedly ameliorated the lung damage of SARS-CoV-2–inoculated human angiotensin-converting enzyme 2 (hACE2) mice. Collectively, our study provides the first evidence that virus-induced apoptosis is an important disease determinant of highly pathogenic coronaviruses and demonstrates that this process can be targeted to attenuate disease severity.

Seeds are major vehicles of propagation and dispersal in plants. A number of transcription factors, including APETALA2 (AP2), play crucial roles during the seed development process in various plant species. However, genes essential for seed development and the regulatory networks during seed development remain unclear in lettuce. Here, we identified a lettuceAP2(LsAP2) gene that was highly expressed at the early stages of seed development.LsAP2knockout plants obtained by the CRISPR/Cas9 system were used to explore the biological function ofLsAP2. Compared with wild-type, the seeds of theLsap2mutant plants had increased length and decreased width, and developed an extended tip at the seed top. After further investigating the seed structural characteristics ofLsap2mutant plants, we proposed a new function ofLsAP2in seed dispersal. Moreover, we identified several interactors of LsAP2. Our results showed that LsAP2 directly interacted with the lettuce homolog of BREVIPEDICELLUS (LsBP) and promoted the expression ofLsBP. Transcriptome analysis revealed that LsAP2 might also be involved in brassinosteroid biosynthesis and signaling pathways. Taken together, our data indicate that LsAP2 has a significant function in regulating seed shape in lettuce.

Avian leukemia virus (ALV) is a retrovirus that infects a variety of poultry, and is a vertical transmission of poultry-borne disease. The harm of ALV gradually magnifies with the increase of generations. siRNA with high efficiency, high safety, silence specificity, and unlimited targets have aroused increasing attention from the medical profession, However, there is no suitable vehicle to assist ALV-targeted siRNA for cell internalization. In this paper, ALV-targeted siRNA was first loaded by 13 nm Au NPs layer by layer to synthesize Au@PRP NPs with good stability (78 nm, ζ= +38), which provided an opportunity for the safe diffusion of Au@PRP NPs into cells and specific binding with ALV. Au @PRP NPs could effectively deliver siRNA to DF-1 cells (92.1%) and contain 20 pmol Cy3-siRNA/per million. The strong polyelectrolyte on the surface of Au@PRP NPs can effectively capture cells and co-infect with ALV as high as 88-100%. Moreover, Au@PRP NPs can locate C/E chicken embryo fibroblasts (DF-1 cells) with ALV, and have a significant inhibitory effect on the release of virus particles. Au@PRP NPs will bring new opportunity to research how the multifunctional load design and tracking path of gold nanoparticles influence the uptake/inhibition mechanisms of ALV and the other poultry diseases and guide different target genes to exert their effects.

Background And Aims Besides prominent respiratory involvement, gastrointestinal manifestations are commonly reported in Coronavirus Disease 2019 (COVID-19) patients. We compared infection of ex vivo human intestinal tissues by SARS-CoV-2 and SARS-CoV with respect to their replication kinetics and immune activation profile. Methods Human intestinal tissues were obtained from patients while undergoing surgical operations at the Queen Mary Hospital, Hong Kong. Upon surgical removal, the tissues were immediately processed and infected with SARS-CoV-2 or SARS-CoV. Replication kinetics were determined with immunohistochemistry, qRT-PCR, and plaque assays. Immune activation in the infected intestinal tissues was assessed by detecting the gene expression of interferons and representative pro-inflammatory cytokines and chemokines. Results SARS-CoV-2 could infect and productively replicate in the ex vivo human intestinal tissues with the release of infectious virus particles, but not in ex vivo human liver and kidney tissues. Importantly, SARS-CoV-2 replicated less efficiently than SARS-CoV, induced less cytopathology in the human intestinal epithelium, and induced a more robust innate immune response including the activation of both type I and type III interferons, than SARS-CoV in human intestinal tissues. Conclusion Using the ex vivo human intestinal tissues as a physiologically relevant model, our data indicated that SARS-CoV-2 could productively replicate in the human guts, suggesting the gastrointestinal tract might serve as an alternative route of virus dissemination. SARS-CoV-2 replicated less efficiently and induced less cytopathology than SARS-CoV in keeping with the clinical observations reported for SARS-2003 and COVID-19, which might be a result of the more robust immune activation by SARS-CoV-2 than SARS-CoV in the human intestine., SARS-CoV-2 induces a higher innate immune response while replicates less efficiently and induces less cytopathology than SARS-CoV in the human intestinal tissues. These findings potentially explained the differential gastrointestinal manifestations observed in COVID-19 and SARS-2003.

Understanding the factors that contribute to efficient SARS-CoV-2 infection of human cells may provide insights on SARS-CoV-2 transmissibility and pathogenesis, and reveal targets of intervention. Here, we analyze host and viral determinants essential for efficient SARS-CoV-2 infection in both human lung epithelial cells and ex vivo human lung tissues. We identify heparan sulfate as an important attachment factor for SARS-CoV-2 infection. Next, we show that sialic acids present on ACE2 prevent efficient spike/ACE2-interaction. While SARS-CoV infection is substantially limited by the sialic acid-mediated restriction in both human lung epithelial cells and ex vivo human lung tissues, infection by SARS-CoV-2 is limited to a lesser extent. We further demonstrate that the furin-like cleavage site in SARS-CoV-2 spike is required for efficient virus replication in human lung but not intestinal tissues. These findings provide insights on the efficient SARS-CoV-2 infection of human lungs., Here, using lung epithelial cells and ex vivo tissue explants, the authors show that, in addition to ACE2, host heparan sulfate is directly involved in SARS-CoV-2 attachment and entry and provide data suggesting that host sialic acids may act as viral restriction factor in lung tissues.

Heme Oxygenase-1 (HO-1) is a ubiquitously expressed enzyme which plays important functions in antioxidant, anti-inflammatory and anti-apoptosis. Recent studies have demonstrated that HO-1 also has significant antiviral properties, inhibiting the replication of some kinds of viruses such as human immunodeficiency virus (HIV), hepatitis C virus (HCV), and dengue fever virus (DFV). In this study, we evaluated the role of HO-1 in Duck Tembusu virus (DTMUV) replication in vitro. The results showed that, the mRNA expression level of HO-1 was transient up-regulated and then significantly decreased in duck embryo fibroblast (DEF) infected with DTMUV. HO-1 induction by transfection of HO-1 over-expression plasmid or treatment with cobalt protoporphyrin (CoPP), a potent HO-1 inducer, could inhibit DTMUV replication effectively. In contrast, HO-1 siRNA knockdown in DEF increased DTMUV replication, implied that HO-1 was an important cellular factor against DTMUV replication. Furthermore, we found that ferric ion (Fe3+) but not biliverdin and carbon monoxide, products of heme degradation by HO-1, mediated the HO-1-induced anti-DTMUV effect. Overall, these finding revealed that a drug induced the HO-1 signal pathway was a promising strategy for treating DTMUV infection.

Clinical manifestations of coronavirus disease 2019 (COVID-19) vary from asymptomatic virus shedding, nonspecific pharyngitis, to pneumonia with silent hypoxia and respiratory failure. Dendritic cells and macrophages are sentinel cells for innate and adaptive immunity that affect the pathogenesis of severe acute respiratory syndrome (SARS) and Middle East respiratory syndrome (MERS). The interplay between SARS-CoV-2 and these cell types remains unknown. We investigated infection and host responses of monocyte-derived dendritic cells (moDCs) and macrophages (MDMs) infected by SARS-CoV-2. MoDCs and MDMs were permissive to SARS-CoV-2 infection and protein expression but did not support productive virus replication. Importantly, SARS-CoV-2 launched an attenuated interferon response in both cell types and triggered significant proinflammatory cytokine/chemokine expression in MDMs but not moDCs. Investigations suggested that this attenuated immune response to SARS-CoV-2 in moDCs was associated with viral antagonism of STAT1 phosphorylation. These findings may explain the mild and insidious course of COVID-19 until late deterioration.

Highlights • SARS-CoV-2 infection was more robust than SARS-CoV in Calu3. In contrast, SARS-CoV infected intestinal epithelial cells more efficiently. • SARS-CoV-2 infection launched an attenuated interferon and pro-inflammatory cytokines/chemokines response in both Calu3 and Caco2 cells, despite robust virus infection and propagation. • SARS-CoV-2 was more sensitive to IFNβ and poly(I:C) pretreatment than that of SARS-CoV., Objectives Respiratory and intestinal tract were two primary target organs of SARS-CoV-2 infection. However, detailed characterization of the host-virus interplay in infected human lung and intestinal epithelial cells is lacking. Methods We utilized immunofluorescence assays, flow cytometry, and RT-qPCR to delineate the virological features and the innate immune response of the host cells against SARS-CoV-2 infection in two prototype human cell lines representing the human pulmonary (Calu3) and intestinal (Caco2) epithelium when compared with SARS-CoV. Results Lung epithelial cells were significantly more susceptible to SARS-CoV-2 compared to SARS-CoV. However, SARS-CoV-2 infection induced an attenuated pro-inflammatory cytokines/chemokines induction and type I and type II IFN responses. A single dose of 10 U/mL IFN-β (IFNβ) pretreatment potently protected both Calu3 and Caco2 against SARS-CoV-2 infection. Interestingly, SARS-CoV-2 was more sensitive to the pretreatment with IFNβ and IFN inducer than SARS-CoV in Calu3. Conclusions Despite robust infection efficiency in both the human lung and intestinal epithelial cells, SARS-CoV-2 could attenuate the virus-induced pro-inflammatory response and IFN response. Pre-activation of the type I IFN signaling pathway primed a highly efficient antiviral response in the host against SARS-CoV-2 infection, which could serve as a potential therapeutic and prophylactic maneuver to COVID-19 patients.

Avian leukosis (AL) is one of the most pandemic immunosuppressive diseases and has been widely spread between 2006 and 2009 in China. The contamination of avian leukosis virus (ALV) in attenuated vaccine is considered as one of the possible transmission routes of this disease. Based on a retrospective survey of 918 batches of attenuated vaccine produced before 2010, three of them were identified as ALV-positive and corresponding ALV strains were successfully isolated from a live Fowlpox virus vaccine, a live Newcastle disease virus vaccine and a live Infectious Bursal Disease virus vaccine, respectively, and whole-genome sequencing showed that these three isolates shared the highest homology with ALV-A wild strains isolated in China (97.7%) over the same period, and the phylogenetic analysis based on their gp85 genes further confirmed that they belong to subgroup A. Meanwhile, although these three ALV-A strains isolated from contaminated vaccines shared a close genetic relationship, their U3 region of genome have a relatively low identity, suggesting that these three strains may have different sources. This study reminds us once again that the possibility of ALV infecting chickens through contaminated live vaccines, requiring us to carry out stricter exogenous virus monitoring in vaccines.

Proliferative diabetic retinopathy (PDR) is the leading cause of blindness and accounts for approximately 12% of all new cases of blindness. Prostaglandin E2 (PGE2) and nucleotide-binding domain and Leucine-rich repeat Receptor containing a Pyrin domain 3 (NLRP3) inflammasomes have been long considered to be associated with PDR. Levels of pro-inflammatory mediators were examined by Enzyme-linked immunosorbent assay (ELISA) measurements. PGE2 levels were analyzed by using the Prostaglandin E2 Monoclonal EIA Kit. Human retinal microvascular endothelial cells (HRMECs) were stained with Annexin V/propidium iodide and analyzed by flow cytometry for testing the apoptosis. Expression levels of NLRP3 inflammasome components under various conditions were detected by Western blot and real-time PCR. Inflammasome markers and PGE2 were highly expressed in the vitreous of PDR patients. PGE2 and NLRP3 induced apoptosis of HRMECs. PGE2 upregulated the expression of NLRP3 inflammasome components and inflammatory chemokines. Knockdown of NLRP3 attenuated the expression of NLRP3 inflammasome components and inhibited the effect of PGE2. Our results suggest that PGE2 levels and NLRP3 inflammasome activation are closely related to the pathogenesis of PDR and nonsteroidal anti-inflammatory drugs may have a potential therapeutic effect on PDR.

Coronavirus tropism is predominantly determined by the interaction between coronavirus spikes and the host receptors. In this regard, coronaviruses have evolved a complicated receptor-recognition system through their spike proteins. Spikes from highly related coronaviruses can recognize distinct receptors, whereas spikes of distant coronaviruses can employ the same cell-surface molecule for entry. Moreover, coronavirus spikes can recognize a broad range of cell-surface molecules in addition to the receptors and thereby can augment coronavirus attachment or entry. The receptor of Middle East respiratory syndrome coronavirus (MERS-CoV) is dipeptidyl peptidase 4 (DPP4). In this study, we identified membrane-associated 78-kDa glucose-regulated protein (GRP78) as an additional binding target of the MERS-CoV spike. Further analyses indicated that GRP78 could not independently render nonpermissive cells susceptible to MERS-CoV infection but could facilitate MERS-CoV entry into permissive cells by augmenting virus attachment. More importantly, by exploring potential interactions between GRP78 and spikes of other coronaviruses, we discovered that the highly conserved human GRP78 could interact with the spike protein of bat coronavirus HKU9 (bCoV-HKU9) and facilitate its attachment to the host cell surface. Taken together, our study has identified GRP78 as a host factor that can interact with the spike proteins of two Betacoronaviruses, the lineage C MERS-CoV and the lineage D bCoV-HKU9. The capacity of GRP78 to facilitate surface attachment of both a human coronavirus and a phylogenetically related bat coronavirus exemplifies the need for continuous surveillance of the evolution of animal coronaviruses to monitor their potential for human adaptations.

Background Reticuloendotheliosis is an immunosuppressive disease caused by avian reticuloendotheliosis virus (REV). It is commonly found in poultry farms and has caused a notable economic loss worldwide. Despite this, there is currently no effective vaccine available to protect against REV infection. Method In this study, gp90 protein derived from an REV isolated from a contaminated vaccine was co-administered with cytosine-phosphate-guanine oligodeoxynucleotide (CpG-ODN) adjuvant to hens to determine if it protects their chicks against REV infection. To synthesize the gp90 protein, the gp90 gene was amplified using polymerase chain reaction, expressed in Escherichia coli, and purified. The resulting recombinant protein was injected intramuscularly into breeder hens along with CpG-ODN adjuvant and then serum antibody levels were regularly evaluated. After the fertilized eggs from these vaccinated hens had hatched, the resulting chicks were challenged with a 102.7 50% tissue culture infectious dose (TCID50) of REV at 1 day old and the REV antibody levels in these hatched chickens were evaluated before and after the challenge. Viremia and growth rate were measured weekly and statistically analyzed. Results The results suggest that the gp90 recombinant protein was successfully prepared and, when used with CpG-ODN adjuvant to immunize breeder hens, induced serological antibody production against REV in both hens and their hatched chicks. In addition, the maternal antibodies induced by the gp90 protein vaccine effectively protected majority of the chicks from REV infection. Conclusions Overall, we found the gp90 protein obtained in this study may be a potential vaccine candidate that had good immunogenicity and could be an auxiliary measure to accelerate the eradication of REV.

Bacteria-based therapy has shown great promise in treating various diseases by regulating intestinal flora balance in pre-clinical and clinical studies. However, weak resistance against stress in the gastrointestinal (GI) tract and limited retention time in the intestine result in low bacteria availability and persistence, restricting further clinical application. Here, we report a double-layer coating strategy employing tannic acid (TA) and enteric L100 for probiotics encapsulation to address the delivery challenges after oral administration. The Escherichia coli Nissle 1917 (EcN) sequentially encapsulated by TA and L100 layers display robust resistances against the harsh external environment of the GI tract. Furthermore, the pH-responsive degradation of the outer L100 layer leads to the selective delivery of TA-EcN to the intestine, where the strong mucoadhesive capability of the TA layer prolongs the retention time of EcN without compromising the viability and proliferation capabilities of EcN, resulting in superior efficacy of prophylactic and treatment efficacy against colitis. To mitigate the potential side effects caused by long-term mucoadhesion of TA, the TA layer can be further removed by addition of ethylenediaminetetraacetic acid (EDTA). The spatiotemporal delivery of probiotics through double-layer encapsulation can augment the therapeutic efficacy of bacteria-based therapy against GI tract-related diseases.

Background: Staphylococcus aureus is a common human pathogen capable of causing diverse illnesses with possible recurrent infections. Although recent studies have highlighted the role of cellular immunity in recurrent infections, the mechanism by which S. aureus evades host responses remains largely unexplored. Methods: This study utilizes in vitro and in vivo infection experiments to investigate difference of pro-inflammatory responses and subsequent adaptive immune responses between adsA mutant and WT S. aureus strain infection. Findings: We demonstrated that adenosine synthase A (AdsA), a potent S. aureus virulence factor, can alter Th17 responses by interfering with NLRP3 inflammasome-mediated IL-1β production. Specifically, S. aureus virulence factor AdsA dampens Th1/Th17 immunity by limiting the release of IL-1β and other Th polarizing cytokines. In particular, AdsA obstructs the release of IL-1β via the adenosine/A2aR/NLRP3 axis. Using a murine infection model, pharmacological inhibition of A2a receptor enhanced S. aureus-specific Th17 responses, whereas inhibition of NLRP3 and caspase-1 downregulated these responses. Our results showed that AdsA contributes to recurrent S. aureus infection by restraining protective Th1/Th17 responses. Interpretation: Our study provides important mechanistic insights for therapeutic and vaccination strategies against S. aureus infections. Funding This work was supported by grants from Shenzhen Peacock project ( KQTD2015033-117210153 ), and Guangdong Science and Technology Department ( 2020B1212030004 ) to J.H. and China Postdoctoral Science Foundation ( 2019M663167 ) to BZZ . We also thank the L & T Charitable Foundation , the Guangdong Science and Technology Department ( 2020B1212030004 ), and the Program for Guangdong Introducing Innovative and Entrepreneurial Teams ( 2019BT02Y198 ) for their support.

Objective Hemicastration is a unilateral orchiectomy to remove an injured testis, which can induce hormonal changes and compensatory hypertrophy of the remaining testis, and may influence spermatogenesis. However, the underlying molecular mechanisms are poorly understood. Here, we investigated the impact of hemicastration on remaining testicular function. Methods Prepubertal mice (age 24 days) were hemicastrated, and their growth was monitored until they reached physical maturity (age 72 days). Subsequently, we determined testis DNA methylation patterns using reduced representation bisulfite sequencing of normal and hemicastrated mice. Moreover, we profiled the testicular gene expression patterns by RNA sequencing (RNA-seq) to examine whether methylation changes affected gene expression in hemicastrated mice. Results Hemicastration did not significantly affect growth or testosterone (p>0.05) compared with control. The genome-wide DNA methylation pattern of remaining testis suggested that substantial genes harbored differentially methylated regions (1,139) in gene bodies, which were enriched in process of protein binding and cell adhesion. Moreover, RNA-seq results indicated that 46 differentially expressed genes (DEGs) involved in meiotic cell cycle, synaptonemal complex assembly and spermatogenesis were upregulated in the hemicastration group, while 197 DEGs were downregulated, which were related to arachidonic acid metabolism. Integrative analysis revealed that proteasome 26S subunit ATPase 3 interacting protein gene, which encodes a protein crucial for homologous recombination in spermatocytes, exhibited promoter hypomethylation and higher expression level in hemicastrated mice. Conclusion Global profiling of DNA methylation and gene expression demonstrated that hemicastration-induced compensatory response maintained normal growth and testicular morphological structure in mice.

Background Avian leukosis virus (ALV) is one of the main causes of tumour development within the poultry industry in China. The subgroup J avian leukosis viruses (ALV-J), which induce erythroblastosis and myelocytomatosis, have the greatest pathogenicity and transmission ability within this class of viruses. ALV can be transmitted both horizontally and vertically; however, the effects of ALV infection in chickens—especially roosters—during the propagation, on future generations is not clear. Knowing the role of the cock in the transmission of ALV from generation to generation might contribute to the eradication programs for ALV. Results The results showed that two hens inseminated with ALV-J-positive semen developed temporary antibody responses to ALV-J at 4–5 weeks post insemination. The p27 antigen was detected in cloacal swabs of six hens, and in 3 of 26 egg albumens at 1–6 weeks after insemination. Moreover, no viremia was detected at 6 weeks after insemination even when virus isolation had been conducted six times at weekly intervals for each of the 12 females. However, ALV-J was isolated from 1 of their 34 progeny chicks at 1 week of age, and its gp85 had 98.4%–99.2% sequence identity with the gp85 of ALV-J isolated from semen samples of the six cocks. Conclusions Our findings indicated that females that were late horizontally infected with ALV-J by artificial insemination might transmit the virus to progeny through eggs, which amounts to vertical transmission.

To investigate the possible causes of the massive spread of fowl adenovirus (FAdV) infection among chickens in recent years in China, 32 batches of live-virus vaccines were tested for contamination with FAdV by PCR. Among these, 1 live Newcastle disease virus (NDV) vaccine of the LaSota strain was demonstrated to be positive for contamination. The amplified hexon gene exhibited 99.8% identity with a recent Chinese field isolate (JSJ13) of FAdV-4. The positive LaSota vaccine was first neutralized with anti-NDV serum and then inoculated into specific pathogen-free embryos at embryonic day 5 through the yolk sac for isolation of the contaminated FAdV. The same hexon gene bands were amplified from extracted DNA of the liver tissues and chicken embryo allantoic fluid of the inoculated embryos, indicating the replication and isolation of the FAdV-4 virus strain that had contaminated the vaccine. This represents the first report of FAdV-4 contamination in a live vaccine for poultry in China. These findings suggest that contamination of live vaccine might represent one of the most important causes of massive outbreaks of FAdV infection among chickens and indicate that FAdV should therefore be included in the regular monitoring list for the detection of exogenous viral contamination of attenuated vaccines for poultry.

Background Recurrent pregnancy loss (RPL) refers to two or more spontaneous abortions that occur consecutively with the same spouse. RPL severely affects human reproduction health, and causes extreme physical and mental suffering to patients and their families. Methods We used isobaric tags for relative and absolute quantitation (iTRAQ), which was coupled with liquid chromatography mass spectrometry (LC-MS) proteomic analysis, in order to identify differentially expressed proteins. Moreover, we used western blot to analyze differentially expressed proteins. Results Of the 2350 non-redundant proteins identified, 38 proteins were significantly altered and were identified as potential biomarkers for RPL. The protein-protein interaction network constructed using GeneMANIA revealed that 35.55% displayed similar co-expression, 30.87% were predicted, and 20.95% had physical interaction characteristics. Based on Gene ontology classification and KEGG pathway enrichment analyses, the majority of these differentially expressed proteins were found to be related to biological regulation, metabolic and cellular processes, protein binding and different enzymes activities, as well as disorder of fat and glucose metabolic pathways. It is noteworthy that three metabolism related biomarkers (HK1, ACLY, and FASN) were further confirmed through western blot analysis. Conclusions These results suggest that these differentially expressed proteins may be used as biomarkers for RPL, and related signaling pathways may play crucial roles in male induced RPL. Electronic supplementary material The online version of this article (10.1186/s12958-019-0496-5) contains supplementary material, which is available to authorized users.

Lung metastasis is a common and deadly occurrence in many types of solid tumors. Chemokine receptor CXCR4 and transcription factor signal transducer and activator of transcription 3 (STAT3) are among potential therapeutic targets in lung metastatic cancer. Both CXCR4 and STAT3 play important roles in the proliferation, angiogenesis, and metastasis of cancer cells. Here, we report on the development of a pulmonary delivery (p.d.) system based on perfluorocarbon (PFC) nanoemulsions for combined delivery of a partially fluorinated polymeric CXCR4 antagonist (FM) and anti-STAT3 small interfering RNA (siRNA). We have prepared FM-stabilized PFC (FM@PFC) as a delivery system of therapeutic siRNA adsorbed on the surface of the emulsion. These FM@PFC/siRNA nanoemulsions inhibited both CXCR4 and STAT3, as demonstrated by effective anti-invasive ability in vitro and related antimetastatic activity in vivo. The combined nanoemulsions provided a comprehensive anticancer effect in the model of established lung metastasis of breast carcinoma, which was dependent on induction of cancer cell apoptosis, anti-angiogenic effect, anti-invasive activity, and overcoming of the immunosuppressive tumor microenvironment. Direct comparison with intravenous (i.v.) injection showed superior activity of pulmonary administration as indicated by significantly increased animal survival. Overall, this work established the suitability of the PFC nanoemulsions for p.d. of combination anticancer treatments and as a promising method to treat lung metastasis.

Growth retardation reduces the incomes of livestock farming. However, effective nutritional interventions to promote compensatory growth and the mechanisms involving digestive tract microbiomes and transcripts have yet to be elucidated. In this study, Qinghai plateau yaks, which frequently suffer from growth retardation due to malnutrition, were used as an experimental model. Young growth-retarded yaks were pastured (GRP), fed basal ration (GRB), fed basal ration addition cysteamine hydrochloride (CSH; GRBC) or active dry yeast (ADY; GRBY). Another group of growth normal yak was pastured as a positive control (GNP). After 60-day nutritional interventions, the results showed that the average daily gain (ADG) of GRB was similar to the level of GNP, and the growth rates of GRBC and GRBY were significantly higher than the level of GNP (P < 0.05). Basal rations addition of CSH or ADY either improved the serum biochemical indexes, decreased serum LPS concentration, facilitated ruminal epithelium development and volatile fatty acids (VFA) fermentation of growth-retarded yaks. Comparative transcriptome in rumen epithelium between growth-retarded and normal yaks identified the differentially expressed genes mainly enriched in immune system, digestive system, extracellular matrix and cell adhesion pathways. CSH addition and ADY addition in basal rations upregulated ruminal VFA absorption (SLC26A3, PAT1, MCT1) and cell junction (CLDN1, CDH1, OCLN) gene expression, and downregulated complement system (C2, C7) gene expression in the growth-retarded yaks. 16S rDNA results showed that CSH addition and ADY addition in basal rations increased the rumen beneficial bacterial populations (Prevotella_1, Butyrivibrio_2, Fibrobacter) of growth-retarded yaks. The correlation analysis identified that ruminal VFAs and beneficial bacteria abundance were significantly positively correlated with cell junction and VFA absorption gene expressions and negatively correlated with complement system gene expressions on the ruminal epithelium. Therefore, CSH addition and ADY addition in basal rations promoted rumen health and body growth of growth-retarded yaks, of which basal ration addition of ADY had the optimal growth-promoting effects. These results suggested that improving nutrition and probiotics addition is a more effective method to improve growth retardation caused by gastrointestinal function deficiencies.

Background Cross protection is a promising alternative to control plant viral diseases. One critical factor limiting the application of cross protection is the availability of attenuated mutants or mild strains. Potato virus X (PVX) infects many crops and induces huge economic losses to agricultural production. However, researches on the variability and mechanism of PVX virulence are scarce. Methods The mutants were obtained by introducing mutations into the RNA dependent RNA polymerase (RdRp) gene of PVX via site-directed mutagenesis. Attenuated mutants were screen according to their symptoms in Nicotiana benthamiana plants. The protection efficacy against severe infection were evaluated with interval of 5, 10 and 15 days. Results Among the 40 mutants obtained, four mutants carrying substitutions of either Glu46, Asn863, Asn968 or Glu1001 to Ala in PVX RdRp showed drastically attenuated symptom, accompanying with reduced accumulation levels of coat protein, plus- and minus-sense RNAs. When the interval between protective and challenging inoculations was 15 days, mutant E1001A (with substitution of Glu1001 to Ala in RdRp) provided complete protection against severe infection in both Nicotiana benthamiana and tomato, while E46A (Glu46 mutated to Ala) provided incomplete protection. To reduce the risk of reverse mutation, we constructed mutant dM which carries double mutations of both Glu46 and Glu1001 to Ala in RdRp. The mutant dM could provide effective protection against severe PVX infection. Conclusion Mutations of Glu46, Asn863, Asn968 or Glu1001 to Ala in PVX RdRp significantly reduced the viral symptoms. Mutants E1001A and E46A could provide effective protection against wild type PVX in both Nicotiana benthamiana and tomato. These results provide theoretical and practical bases for the control of PVX via cross protection.

Nervous necrosis virus (NNV) infects more than 120 fish species worldwide and causes high mortality and significant economic losses to the aquaculture industry. Among different genotypes of NNV, the red-grouper nervous necrosis virus (RGNNV) genotype is the most widely reported with the highest number of susceptible species. Interferon (IFN) is a powerful cytokine driving host to establish an antiviral response. RGNNV may have an elaborate strategy to overcome the host's IFN-based antiviral response. In this study, we demonstrated that the B2 protein of RGNNV blocked IFN-mediated antiviral response by inhibiting host transcription directed by RNA polymerase II (RNAP II) complex. We firstly found that the upregulation of IFNφ1 promoter activity stimulated by poly(I:C) was suppressed by RGNNV infection. By systematically screening the structural and nonstructural proteins of RGNNV, we identified B2 protein as the virus's most potent antihost protein, which inhibited the IFNφ1 promoter activities stimulated by MDA5, MAVS, TBK1, IRF3, and IRF7 and decrease their protein expression. Importantly, B2 also suppressed the constitutively active cytomegalovirus (CMV) promoter, which indicated that B2 might be a general inhibitor of gene transcription. Moreover, western blotting showed that B2 could decrease RNAP II expression and RNAP II phosphorylation. All these results showed RGNNV blocked host's IFN response by B2 protein directly dysregulating RNAP II expression. To the best of our knowledge, this is the first report about the mechanism of RGNNV for anti-host IFN response. These findings will provide a new insight for further research on the viral infection mechanism and effective strategies to control RGNNV.

In our previous study, six subgroup J strains of avian leukosis virus (ALV-J)-associated acutely transforming viruses carrying different lengths of the v-fps oncogene, designated as Fu-J and Fu-J1-5, were isolated and characterized from fibrosarcomas in ALV-J-infected chickens. In the present study, the oncogenic potential of Fu-J and Fu-J1-5 was investigated using a reverse genetics technique. Six replication-defective viruses, named rFu-J and rFu-J1-5, were rescued with the replication-competent rescued ALV-J strain rSDAU1005 as a helper virus by co-transfection of chicken embryo fibroblast monolayers with infectious clone plasmids. Experimental bird studies were performed, demonstrating that only the rescued rFu-J virus carrying the complete v-fps oncogene with rSDAU1005 as the helper virus could induce acute fibrosarcoma after inoculation in specific-pathogen-free (SPF) chickens. These results provide direct evidence that the replication-defective acutely transforming Fu-J virus, with the complete v-fps oncogene, was associated with acute fibrosarcoma in chickens infected with ALV-J in the field, as reported previously.

Transduction of oncogenes by ALVs and generation of acute transforming viruses is common in natural viral infections. In order to understand the molecular basis for the rapid oncogenicity of Fu-J, an acutely transforming avian leukosis virus isolated from fibrosarcomas in crossbreed broilers infected with subgroup J avian leukosis virus (ALV-J) in China, complete genomic structure of Fu-J virus was determined by PCR amplification and compared with those of Fu-J1, Fu-J2, Fu-J3, Fu-J4, and Fu-J5 reported previously. The results showed that the genome of Fu-J was defective, with parts of gag gene replaced by the complete v-fps oncogene and encoded a 137 kDa Gag-fps fusion protein. Sequence analysis revealed that Fu-J and Fu-J1 to Fu-J5 were related quasi-species variants carrying different lengths of v-fps oncogenes generated from recombination between helper virus and c-fps gene. Comparison of virus carrying v-fps oncogene also gave us a glimpse of the molecular characterization and evolution process of the acutely transforming ALV.

Bisphenol A (BPA), one of the most prevalent chemicals for daily use, was recently reported to disturb the homeostasis of energy metabolism and insulin signaling pathways, which might contribute to the increasing prevalence rate of mild cognitive impairment (MCI). However, the underlying mechanisms are remained poorly understood. Here we studied the effects of low dose BPA on glucose transport and the IR/IRS/AKT/GSK3β axis in adult male mice to delineate the association between insulin signaling disruption and neurotoxicity mediated by BPA. Mice were treated with subcutaneous injection of 100μg/kg/d BPA or vehicle for 30 days, then the insulin signaling and glucose transporters in the hippocampus and prefrontal cortex were detected by western blot. Our results showed that mice treated with BPA displayed significant decrease of insulin sensitivity, and in glucose transporter 1, 3 (GLUT1, 3) protein levels in mouse brain. Meanwhile, hyperactivation of IR/IRS/AKT/GSK3β axis was detected in the brain of BPA treated mice. Noteworthily, significant increases of phosphorylated tau and β-APP were observed in BPA treated mice. These results strongly suggest that BPA exposure significantly disrupts brain insulin signaling and might be considered as a potential risk factor for neurodegenerative diseases.

To understand the cytogenetic characteristics of acute fibrosarcoma in chickens infected with the subgroup J avian leukosis virus associated with the v-src oncogene, we performed a karyotype analysis of fibrosarcoma cell cultures. Twenty-nine of 50 qualified cell culture spreads demonstrated polyploidy of some macrochromosomes, 21 of which were trisomic for chromosome 7, and others were trisomic for chromosomes 3, 4, 5 (sex chromosome w), and 10. In addition, one of them was trisomic for both chromosome 7 and the sex chromosome 5 (w). In contrast, no aneuploidy was found for 10 macrochromosomes of 12 spreads of normal chicken embryo fibroblast cells, although aneuploidy for some microchromosomes was demonstrated in five of the 12 spreads. The cytogenetic mosaicism or polymorphism of the aneuploidy in the acute fibrosarcoma described in this study suggests that the analysed cells are polyclonal.