Search

Over the past two decades, numerous non-coding RNAs (ncRNAs) have been identified in different biological systems including virology, especially in large DNA viruses such as herpesviruses. As a representative oncogenic alphaherpesvirus, Marek's disease virus (MDV) causes an important immunosuppressive and rapid-onset neoplastic disease of poultry, namely Marek's disease (MD). Vaccinations can efficiently prevent the onset of MD lymphomas and other clinical disease, often heralded as the first successful example of vaccination-based control of cancer. MDV infection is also an excellent model for research into virally-induced tumorigenesis. Recently, great progress has been made in understanding the functions of ncRNAs in MD biology. Herein, we give a review of the discovery and identification of MDV-encoded viral miRNAs, focusing on the genomics, expression profiles, and emerging critical roles of MDV-1 miRNAs as oncogenic miRNAs (oncomiRs) or tumor suppressor genes involved in the induction of MD lymphomas. We also described the involvements of host cellular miRNAs, lincRNAs, and circRNAs participating in MDV life cycle, pathogenesis, and/or tumorigenesis. The prospects, strategies, and new techniques such as the CRISPR/Cas9-based gene editing applicable for further investigation into the ncRNA-mediated regulatory mechanisms in MDV pathogenesis/oncogenesis were also discussed, together with the possibilities of future studies on antiviral therapy and the development of new efficient MD vaccines. [ABSTRACT FROM AUTHOR]

Marek's disease virus (MDV) induces severe immunosuppression and lymphomagenesis in the chicken, its natural host, and results in a condition that investigated the pathogenesis of MDV and have begun to focus on the expression profiling of circular RNAs (circRNAs). However, little is known about how the expression of circRNAs is referred to as Marek's disease. Previous reports have is regulated during MDV replication. Here, we carried out a comprehensive profiling analysis of N6-methyladenosine (m6A) modification on the circRNA transcriptome in infected and uninfected chicken embryonic fibroblast (CEF) cells. Methylated RNA immunoprecipitation sequencing (MeRIP-Seq) revealed that m6A modification was highly conserved in circRNAs. Comparing to the uninfected group, the number of peaks and conserved motifs were not significantly different in cells that were infected with MDV, although reduced abundance of circRNA m6A modifications. However, gene ontology and Kyoto encyclopedia of genes and genomes (KEGG) pathway analyses revealed that the insulin signaling pathway was associated with the regulation of m6A modified circRNAs in MDV infection. This is the first report to describe alterations in the transcriptome-wide profiling of m6A modified circRNAs in MDV-infected CEF cells. [ABSTRACT FROM AUTHOR]

Infectious bursal disease virus (IBDV) is an important immunosuppressive virus in chickens. Surface immunoglobulin M (sIgM)-bearing B lymphocytes act as the major targets of IBDV in the bursa of Fabricius, and sIgM may function as one of the membrane binding sites responsible for IBDV infection. Recently, using the virus overlay protein binding assay, the chicken λ light chain of sIgM was identified to specifically interact with IBDV in a virulence-independent manner in vitro. To further investigate sIgM λ light chain-mediated IBDV binding and infection in pre-B cells, the cell line DT40, which is susceptible to both pathogenic and attenuated IBDV, was used. Based on the RNA interference strategy, the DT40 cell line whose λ light chain of sIgM was stably knocked down, herein termed DT40LKD, was generated by the genomic integration of a specific small hairpin RNA and a green fluorescence protein co-expression construct. Flow cytometry analysis indicated that the binding of IBDV to DT40LKD cells was significantly reduced due to the loss of sIgM λ light chain. In particular, reduced viral replication was observed in IBDV-incubated DT40LKD cells, and no viral release into cell culture medium was detected by the IBDV rapid diagnostic strips. In addition, the rescue of sIgM λ light chain expression restored viral binding and replication in DT40LKD cells. These results show that sIgM λ light chain appears to be beneficial for IBDV attachment and infection, suggesting that sIgM acts as a binding site involved in IBDV infection. [ABSTRACT FROM AUTHOR]

The authors describe an ultrasensitive method for simultaneous detection of neomycin (NEO) and quinolones antibiotics (QNS). It is based on the use of (a) two immuno-nanoprobes (a probe for NEO and a probe for QNS), (b) surface-enhanced Raman scattering (SERS) detection, and (c), a portable lateral flow assay (LFA). The two probes consist of gold nanoparticles (AuNPs) conjugated to the Raman active molecule 4-aminothiophenol (PATP), and to monoclonal antibody against NEO (NEO mAb) or against NOR (NOR mAb). Quantitative detection of NEO and QNS was realized via SERS of the PATP-coated AuNPs captured in the test line of a LFA. Under optimized condition, the visual limits of LFA are 10 ng·mL for NEO and 200 ng·mL for NOR, and with LODs down to 0.37 pg·mL and 0.55 pg·mL by using SERS. The NEO test line is not interfered by the NEO analogues gentamycin, streptomycin and tobramycin, but the NOR test line suffers from different degrees of cross-reactivity (CR) to 12 common other QNS, the CRs ranging from 1.5% to 136%. The recoveries of NEO and NOR from spiked milk samples ranged between 86% and 121%, with relative standard deviations (RSD) from 3% to 6%. The method is highly sensitive, accurate and effective. It may be applied to simultaneous detection of NEO and 8 QNS, including NOR, enoxacin, ciprofloxacin, ofloxacin, fleroxacin, marbofloxacin, enrofloxacin, and pefloxacin. [ABSTRACT FROM AUTHOR]

In the past decade, a large number of microRNAs (miRNAs) have been identified in the viral genome of Gallid herpesvirus 2 (GaHV-2), which is historically known as Marek's disease virus type 1. The biological role of most GaHV-2 miRNAs remains unclear. In the present study, we have performed an overall gene expression profile of GaHV-2 miRNAs during the virus life cycle at each phase of the developing disease, a highly contagious, lymphoproliferative disorder, and neoplastic immunosuppressive disease of poultry known as the Marek's disease. According to their distinct in vivo expression patterns, the GaHV-2 miRNAs can be divided into three groups: 12 miRNAs in group I, including miR-M4-5p, displayed a typical expression pattern potentially correlated to the latent, late cytolytic, and/or the proliferative phases in the cycle of GaHV-2 pathogenesis; group II consisting of another 12 miRNAs with expression correlated to the early cytolytic and/or latent phases in GaHV-2's life cycle; while the other two miRNAs in group III showed no identical expression features. Our findings may provide meaningful clues in the search for further potential functions of viral miRNAs in GaHV-2 biology. [ABSTRACT FROM AUTHOR]

Marek's disease is a highly contagious, oncogenic, and immunosuppressive avian viral disease. Surveillance of newly registered Marek's disease virus (MDV) isolates is meaningful for revealing the potential factors involved in increased virulence. Presently, we have focused on the molecular characteristics of all available MDVs from China, including 17 new Henan isolates. Based on Meq, gE, and gI genes, we found that most Chinese isolates contain conserved amino acid point mutations in Meq, such as E, A, A, R, and A, compared to USA virulent MDVs. However, the 59-aa or 60-aa insertions are only found in a few mild MDVs rather than virulent MDVs in China. Further phylogenetic analysis has demonstrated that a different genotype of MDV has been prevalent in China, and for virulent MDVs, their recent evolution has possibly been geographically restricted. Our study has provided more detailed information regarding the field MDVs circulating in China. [ABSTRACT FROM AUTHOR]

In order to assess the influence of long-term irrigation of polluted river water on the ecological health of the rice fields in the low reach of Liaohe River Plain, a series of environmental samples and biological samples in rice fields, including sediments, paddy soils, mitten-handed crabs ( Eriocheir sinensis), loaches ( Misgurnus anguillicaudatu) and frog ( Hoplobatrachus tigerinus), were collected from the area far away from industrial zones, and polychlorinated biphenyls (PCBs) and organochlorine pesticides (HCH, DDT, HCB and Mirex) were analyzed by high resolution gas chromatography coupled with high resolution mass spectrometry (HRGC/HRMS). The comparison of pollutant concentrations and profiles verified that the significant contribution of river water irrigation to PCBs contaminations in paddy soils and the mutual influence of organochlorine pesticide contamination between river sediments and paddy soils. Bioaccumulations of PCBs and organochlorine pesticides in mitten-handed crabs and loaches were site-specific, suggested that they were both the good indicators of ecosystem health. Mitten-handed crabs had the highest capacity to accumulate PCBs and organochlorine pesticides, and also they had the strong ability to degrade these pollutants in their bodies, especially for DDT and HCH. The biota-soil accumulation factors (BSAF) of ΣHCH, ΣDDT, Mirex, HCB and ΣPCB in mitten-handed crab were calculated to be 24.5, 15.3, 122.2, 28.1 and 54.6, respectively. In view of the strong bioaccumulation ability, the health risk should be concerned for the cultivation of mitten-handed crabs in paddy fields in the low reach of Liaohe River Plain. [ABSTRACT FROM AUTHOR]

A method for the selective separation of polychlorinated naphthalene (PCNs), hexabromocyclododecanes (HBCDs) and tetrabromobisphenol A (TBBPA) in complex matrices was developed. In the present study, the type of sorbents and the volume and composition of eluting solvents were investigated in detail. Under the optimal conditions, the recoveries of PCNs, HBCDs and TBBPA in different fractions were determined with liquid/gas chromatography tandem mass spectrometry (LC or GC-MS/MS), which was nearly close to 100%. To investigate the feasibility of the developed method, it was applied to isolate target compounds from spiked soil extracts. PCNs, HBCDs and TBBPA could be subsequently eluted with hexane, hexane/dichloromethane (1:1), and hexane/dichloromethane (1:3), and the recoveries of the analysts from three fractions were satisfactory (70.9%-107%) with the exception of γ-HBCD at low levels. The results indicated that the method exhibited good performance for the selective separation of PCNs, HBCDs and TBBPA and showed great potential in sample pretreatment in terms of alleviating influence from different class of impurities to some extent. [ABSTRACT FROM AUTHOR]

Since the first Chinese case report of Japanese encephalitis, Japanese encephalitis virus (JEV) has circulated in China for at least 60 years. Even though pigs play a critical role in the JEV transmission cycle information on the prevalence of JEV in pigs has not been investigated in China. As the central Chinese province of Henan has the largest human population in China, a history of serious JEV and is the largest pig producing province it was chosen for this study. We have found that currently natural infection with JEV in pigs and mosquitoes is prevalent and both genotypes 1 and 3 co-circulate in pigs and mosquitoes in central China. Phylogenetic analysis showed that all of the newly obtained pig-derived JEV isolates are more closely related to isolates from the 1950s to 1960s than to those recently isolated from humans and mosquitoes. Further analyses based on all the previous reported Chinese isolates indicates that presently genotype 3 JEV is the predominant genotype in pigs but genotype 1 JEV is emerging and spreading rapidly in recent years. Our study provides information for understanding the current epidemiology of JEV in China and suggests possible measures applicable to the further control of JEV. [ABSTRACT FROM AUTHOR]

Japanese encephalitis virus (JEV) causes severe viral encephalitis in humans and some other mammalian animals. Highly efficient culture of the virus is critical for antigen preparation, vaccine production and other basic researches. We have investigated the influence of a number of variables such as the strain virulence, the state of the host cells, medium composition, infection method and others on the proliferation of distinct JEV strains in BHK-21 cells. The results showed that two distinct strategies are needed for the propagation of virulent or attenuated JEV strains. The most critical variables were the method of infection, and especially the density of the host cell. Our studies indicate the general approaches to the in vitro culture of other JEV strains using BHK-21 cell line. [ABSTRACT FROM AUTHOR]

MicroRNAs (miRNAs) are the recently discovered small non-coding RNA molecules that have post-transcriptional regulatory functions in many important biological processes. A large number of miRNAs have been found to be encoded by viral genomes, especially in herpesviruses. Previous research regarding miRNAs encoded by herpesviruses, including Marek's disease virus (MDV), has demonstrated their involvement in lytic replication, latent infection, T-lymphocyte transformation and tumorigenesis. MDV is an oncogenic alphaherpesvirus, with the ability to induce tumors in natural hosts; however, formation of these tumors can be prevented by immunization with attenuated or nonpathogenic forms of the virus. Marek's disease is considered to be a good biomedical model for investigating the biology, genetics, and immunology of tumorigenesis. In this paper, we review the discovery and identification of MDV-encoded miRNAs, along with their genomics, expression profiles, and currently known functions. We also discuss the prospects and techniques possibly applicable to the further investigation of the biological roles of MDV-encoded miRNAs. [ABSTRACT FROM AUTHOR]

An efficient vector, designated as pCAGX, was designed for direct cloning and enhanced expression of PCR-amplified ORFs in mammalian cells. It relied on the well-known TA-cloning principle, and utilized the CMV enhancer/chicken β-actin/rabbit β-globin (CAG) hybrid promoter instead of the classical CMV promoter to drive more efficient transgene expression in wider host cells. The specially designed cassette under CAG hybrid promoter contained two tandemly arrayed XcmI sites which were spaced by an additional EcoRV site. For direct cloning and expressing PCR-amplified ORFs, the T-vector was prepared by further digesting the EcoRV-linearized pCAGX with XcmI to produce T tails on both 3′-ends, which could efficiently minimize the non-recombinant background of T-vector and eliminate the necessity of selective marker genes such as LacZ that allowed blue/white screening. Various PCR fragments in length were prepared to verify the cloning efficiency by ligation with this vector, and GFP gene expression under control of the CAG hybrid promoter in different host cells was assayed by flow cytometry. The results indicated that this vector was higher efficient, especially suitable for cloning and expressing a number of interesting ORFs in parallel, and higher-level transgene expression in different mammalian cells was obtained than the reported vectors using the CMV promoter. [ABSTRACT FROM AUTHOR]