Hepatitis G virus genomic RNA is pathogenic to Macaca mulatta

AIM: To explore the pathogenicity and infectivity of hepatitis G virus (HGV) by observing replication and expression of the virus, as well as the serological and histological changes of Macaca mulatta infected with HGV genomic RNA or HGV RNA-positive serum.METHODS: Full-length HGV cDNA clone (HGVqz) was constructed and proved to be infectious, from which HGV genomic RNA was transcribed in vitro. Macaca mulatta BY1 was intra-hepatically inoculated with HGV genomic RNA, HGV RNA-positive serum from BY1 was intravenously inoculated into Macaca mulatta BM1, and then BB1 was infected with serum from BM1. Serum and liver tissue were taken regularly, and checked with RT-PCR, in situ hybridization and other immunological, serological,histological assays.RESULTS: Serum HGV RNA was detectable in all the 3Macaca mulattas, serological and histological examinations showed the experimental animals had slightly elevated alanine transaminase (ALT) and developed HGV viremia during the infectious period. The histology, immunohistochemistry, and in situ hybridization in liver tissues of the inoculated animals demonstrated a very mild hepatitis with HGV antigen expression in cytoplasm of hepatocytes.RT-PCR and quantitative PCR results showed that HGV could replicate in liver.CONCLUSION: The genomic RNA from full-length HGV cDNA is infectious to the Macaca mulatta and can cause mild hepatitis. HGV RNA-positive serum, from HGV RNA inoculated Macaca mulatta, is infectious to other Macaca mulattas. Macaca mulatta is susceptible to the inoculated HGV, and therefore can be used as an experimental animal model for the studies of HGV infection and pathogenesis.

Clinical application value of long non-coding RNAs signatures of genomic instability in predicting prognosis of hepatocellular carcinoma

Hepatocellular carcinoma(HCC)presents challenges due to its high recurrence and metastasis rates and poor prognosis.While current clinical diagnostic and prognostic indicators exist,their accuracy remains imperfect due to their biol-ogical complexity.Therefore,there is a quest to identify improved biomarkers for HCC diagnosis and prognosis.By combining long non-coding RNA(lncRNA)expression and somatic mutations,Duan et al identified five representative lncRNAs from 88 lncRNAs related to genomic instability(GI),forming a GI-derived lncRNA signature(LncSig).This signature outperforms previously re-ported LncSig and TP53 mutations in predicting HCC prognosis.In this editorial,we comprehensively evaluate the clinical application value of such prognostic evaluation model based on sequencing technology in terms of cost,time,and practicability.Additionally,we provide an overview of various prognostic models for HCC,aiding in a comprehensive understanding of research progress in pro-gnostic evaluation methods.

Genomics of pancreatic ductal adenocarcinoma

Pancreatic cancer is one of the worst prognostic cancers because of the late diagnosis and the absence of effective treatment. Within all subtypes of this disease, ductal adenocarcinoma has the shortest survival time. In recent years,global genomics profiling allowed the identification of hundreds of genes that are perturbed in pancreatic cancer. The integration of different omics sources in the study of pancreatic cancer has revealed several molecular mechanisms, indicating the complex history of its development. However, validation of these genes as biomarkers for early diagnosis, prognosis or treatment efficacy is still incomplete but should lead to new approaches for the treatment of the disease in the future.

The long non-coding RNA expression profile of Coxsackievirus A16 infected RD cells identified by RNA-seq

Coxsackievirus A16(CVA16) is one of major pathogens of hand, foot and mouth disease(HFMD) in children. Long non-coding RNAs(Inc RNAs) have been implicated in various biological processes,but they have not been associated with CVA16 infection. In this study, we comprehensively characterized the landscape of Inc RNAs of normal and CVA16 infected rhabdomyosarcoma(RD)cells using RNA-Seq to investigate the functional relevance of Inc RNAs. We showed that a total of 760 Inc RNAs were upregulated and 1210 Inc RNAs were downregulated. Out of these dysregulated Inc RNAs, 43.64% were intergenic, 22.31% were sense, 15.89% were intronic, 8.67% were bidirectional, 5.59% were antisense, 3.85% were s RNA host Inc RNAs and 0.05% were enhancer. Six dysregulated Inc RNAs were validated by quantitative PCR assays and the secondary structures of these Inc RNAs were projected. Moreover, we conducted a bioinformatics analysis of an Inc RNAs(ENST00000602478) to elucidate the diversity of modification and functions of Inc RNAs. In summary, the current study compared the dysregulated Inc RNAs profile upon CVA16 challenge and illustrated the intricate relationship between coding and Inc RNAs transcripts. These results may not only provide a complete picture of transcription in CVA16 infected cells but also provide novel molecular targets for treatments of HFMD.

Stability and infectivity of enteroviruses on dry surfaces:Potential for indirect transmission control

Hand,foot,and mouth disease(HFMD)is a contagious disease mainly occurring in young children,and outbreaks commonly occur among young children in the Asia–Pacific region including Thailand.Moreover,the World Health Organization(WHO)monitors HFMD in the Western Pacific region to detect outbreaks and other significant events by the Regional Event Based Surveillance System.HFMD is mainly caused by a group of enteroviruses(EVs)transmitted through direct contact(person to person)and indirect contact with contaminated objects(surface-to-hand).However,few studies have examined the surface stability of EVs.In this study,we investigated the stability of enterovirus A71(EV-A71)and coxsackievirus A16(CVA16)on three different dry surfaces(wood,plastic,and stainless steel)using the endpoint titration using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide(MTT)staining of viable cells and real-time polymerase chain reaction(viral genome detection).The results revealed that virus infectivity dramatically decreased within a few hours on dry surfaces.However,viral RNA could be detected on dry surfaces for up to 28 days.Concerning heat inactivation,both EV-A71 and CVA16 were inactivated after exposure to 60°C for 15 min.Information on virus stability on different dry surfaces will provide useful information for HFMD transmission control.

Genome Editing with CRISPR-Cas9:Can It Get Any Better?

The CRISPR-Cas revolution is taking place in virtually all fields of life sciences.Harnessing DNA cleavage with the CRISPR-Cas9 system of Streptococcus pyogenes has proven to be extraordinarily simple and efficient,relying only on the design of a synthetic single guide RNA（sgRNA） and its co-expression with Cas9.Here,we review the progress in the design of sgRNA from the original dual RNA guide for S.pyogenes and Staphylococcus aureus Cas9（SpCas9 and SaCas9）.New assays for genome-wide identification of offtargets have provided important insights into the issue of cleavage specificity in vivo.At the same time,the on-target activity of thousands of guides has been determined.These data have led to numerous online tools that facilitate the selection of guide RNAs in target sequences.It appears that for most basic research applications,cleavage activity can be maximized and off-targets minimized by carefully choosing guide RNAs based on computational predictions.Moreover,recent studies of Cas proteins have further improved the flexibility and precision of the CRISPR-Cas toolkit for genome editing.Inspired by the crystal structure of the complex of sgRNA-SpCas9 bound to target DNA,several variants of SpCas9 have recently been engineered,either with novel protospacer adjacent motifs（PAMs） or with drastically reduced off-targets.Novel Cas9 and Cas9-like proteins called Cpf 1 have also been characterized from other bacteria and will benefit from die insights obtained from SpCas9.Genome editing with CRISPR-Cas9 may also progress with better understanding and control of cellular DNA repair pathways activated after Cas9-induced DNA cleavage.

Secondary Structural Elements of the HCV X-region Involved in Viral Replication

Background and Aims:The noncoding regions in the 3'-untranslated region (UTR) of the hepatitis C virus (HCV)genome contain secondary structures that are important for replication.The aim of this study was to identify detailed conformational elements of the X-region involved in HCV replication.Methods:Ribonucleic acid (RNA) structural analogs X94,X12,and X12c were constructed to have identical conformation but 94%,12%,and 0% sequence identity,respectively,to the X region of HCV genotype 2a.Effects of structural analogs on replication of HCV genotypes 1b and 2a HCV RNA were studied by quantitative reverse transcriptase polymerase chain reaction.Results:In replicon BB7 cells,a constitutive replication model,HCV RNA levels decreased to 55%,52%,53%,and 54% after transfection with expression plasmids generating RNA structural analogs 5B-46,X-94,X-12,and X-12c,respectively (p<0.001 for all).In an HCV genotype 2a infection model,RNA analogs 5B-46,X-94,and X-12 in hepatic cells inhibited replication to 11%,9%,and 12%,respectively.Because the X-12 analog was only 12% identical to the corresponding sequence of HCV genotype 2a,the sequence per se,or antisense effects were unlikely to be involved.Conclusions:The data suggest that conformation of secondary structures in 3'-UTR of HCV RNA genome is required for HCV replication.Stable expression of RNA analogs predicted to have identical stem-loop structures might inhibit HCV infection of hepatocytes in liver and may represent a novel approach to design anti-HCV agents.