Singapore grouper iridovirus (SGIV) is a large double-stranded DNA virus that is a major threat to grouper aquaculture. The pathogenesis of SGIV is not well understood so far. Previous studies have revealed that ICP18, an immediate early protein encoded by SGIV ORF086R gene, promotes viral replication by regulating cell proliferation and virus assembly. In the present study, the potential functions of ICP18 were further explored by probing into its interactors using a proximity-dependent BioID method. Since our in-house grouper embryonic cells (a natural host cell of SGIV) could not be efficiently transfected with the plasmid DNA, and the grouper genome data for mass spectrometry-based protein identification is not currently available, we chosen a non-permissive cell (HEK293 T) as a substitute for this study. A total of 112 cellular proteins that potentially bind to ICP18 were identified by mass spectrometry analysis. Homology analysis showed that among these identified proteins, 110 candidate ICP18-interactors had homologous proteins in zebrafish (a host of SGIV), and shared high sequence identity. Further analysis revealed that the identified ICP18-interacting proteins modulate various cellular processes such as cell cycle and cell adhesion. In addition, the interaction between ICP18 and its candidate interactor, i.e., cyclin-dependent kinase1 (CDK1), was confirmed using Co-immunoprecipitation (Co-IP) and Pull-down assays. Collectively, our present data provides additional insight into the biological functions of ICP18 during viral infection, which could help in further unraveling the pathogenesis of SGIV.

Epstein-Barr virus (EBV) infection is highly prevalent in the population and is known to be associated with a variety of human tumors, such as nasopharyngeal carcinoma, gastric cancer, and lymphoma; however, the mechanisms of EBV carcinogenesis remain unclear. Recent studies have revealed that many non-coding RNAs participate in the regulation of proliferation, migration, invasion, and other processes in EBV-associated tumor, and the interaction between ncRNAs and the potential target genes has gradually become a research hotspot. Therefore, here, we discuss the expression and roles of ncRNAs in EBV-associated epithelial tumors.

Infectious diseases related to viruses, as well as bacterial pathogens, abound in all parts of the world, burdening health and economy. Thus, there is a dire need to find new prevention and treatment strategies to improve clinical practices related to viral infections. Human gut contains trillions of bacteria which have regulatory roles in immune development, homeostasis, and body metabolism. Today, it is difficult to find any prominent viral infection that hasn't had any link with the human gut microbiota. In this opinion-based review article, I argued the significance of manipulating human gut microbiota as novel therapeutics through probiotics or FMT in alleviating complexities related to viral infections, and pinpointed bottlenecks involved in this research.

Circular RNAs (circRNAs) are a class of non-coding RNAs with a special covalently closed circular structure, which is formed by precursor mRNA (pre-mRNA) through "back-splicing". CircRNAs are more stable than linear RNAs because they are resistant to exoribonucleases. Viral infections often cause abnormal expression of circRNAs, which could serve as novel biomarkers for the diagnosis of viral infections by detecting specific circRNAs in cells, body fluids, or tissues. CircRNAs also play a critical role in regulating host immune response and virus replication. Here, we reviewed the production and function of circRNAs, mainly focusing on their regulation on virus infection, to provide novel insights into the potential role of circRNAs as diagnostic marker or treatment targets for viral infection.

The endogenous free radical nitric oxide (NO) plays a pivotal role in the immunological system. NO has already been reported as a potential candidate for use in the treatment of human coronavirus infections, including COVID-19. In fact, inhaled NO has been used in clinical settings for its antiviral respiratory action, and in the regulation of blood pressure to avoid clot formation. In this mini-review, we discuss recent progress concerning the antivirus activity of NO in clinical, pre-clinical and research settings, and its beneficial effects in the treatment of clinical complications in patients infected with coronaviruses and other respiratory viral diseases, including COVID-19. We also highlight promising therapeutic effects of NO donors allied to nanomaterials to combat COVID-19 and other human coronavirus infections. Nanomaterials can be designed to deliver sustained, localized NO release directly at the desired application site, enhancing the beneficial effects of NO and minimizing the side effects. Challenges and perspectives are presented to open new fields of research.

Here a bioinformatic pipeline VVV has been developed to analyse viral populations in a given sample from Next Generation Sequencing (NGS) data. To date, handling large amounts of data from NGS requires the expertise of bioinformaticians, both for data processing and result analysis. Consequently, VVV was designed to help non-bioinformaticians to perform these tasks. By providing only the NGS data file, the developed pipeline generated consensus sequences and determined the composition of the viral population for an avian Metapneumovirus (AMPV) and three different animal coronaviruses (Porcine Epidemic Diarrhea Virus (PEDV), Turkey Coronavirus (TCoV) and Infectious Bronchitis Virus (IBV)). In all cases, the pipeline produced viral consensus genomes corresponding to known consensus sequence and made it possible to highlight the presence of viral genetic variants through a single graphic representation. The method was validated by comparing the viral populations of an AMPV field sample, and of a copy of this virus produced from a DNA clone. VVV demonstrated that the cloned virus population was homogeneous (as designed) at position 2934 where the wild-type virus demonstrated two variant populations at a ratio of almost 50:50. A total of 18, 10, 3 and 28, viral genetic variants were detected for AMPV, PEDV, TCoV and IBV respectively. The simplicity of this pipeline makes the study of viral genetic variants more accessible to a wide variety of biologists, which should ultimately increase the rate of understanding of the mechanisms of viral genetic evolution.

Cyclic GMP-AMP synthase (cGAS) is a main sensor used to detect microbial DNA in the cytoplasm, which subsequently induces the production of interferon (IFN) via the cGAS/STING/IRF3 signaling pathway, leading to an antiviral response. However, some viruses have evolved multiple strategies to escape this process. Pseudorabies virus (PRV) is a double-stranded DNA virus belonging to the Alphaherpesvirinae subfamily, which can cause serious damage to the porcine industry. Many herpesvirus components have been reported to counteract IFN production, whereas little is known of PRV. In the present study, we found that PRV glycoprotein E (gE) was involved in counteracting cGAS/STING-mediated IFN production. Ectopic expression of gE decreased cGAS/STING-mediated IFN-β promoter activity and the level of mRNA expression. Moreover, gE targeted at or downstream of IRF3 was found to inhibit IFN-β production. However, gE did not affect the phosphorylation, dimerization and nuclear translocation of IRF3. Furthermore, gE is located on the nuclear membrane and could subsequently degrade CREB-binding protein (CBP). MG132, a proteasome inhibitor, decreased CBP degradation and restored the IFN-β production induced by gE. Finally, gE-deleted PRV induced a higher level of IFN-β production and reduced CBP degradation compared to wild-type PRV. Together, these results demonstrate that PRV gE can inhibit cGAS/STING-mediated IFN-β production by degrading CBP to interrupt the enhanced assembly of IRF3 and CBP.

Characterization of host adaptation markers among human isolates is important for recognizing the potential for cross-species transmission in avian influenza A viruses. Here, we studied two new potential adaptive mutations, V292I and D740A, in the PB2 protein that were identified by a multi-factor regression model. The study shows that the prevalence of the PB2-V292I mutation is increased in H7N9 influenza viruses isolated from both humans and birds over the past 6 years. The phylogenetic tree showed that influenza A/H7N9 has a lineage based on the strains containing PB2-292I. Polymerase complexes containing PB2-292I/627K derived from H7N9 exhibit increased polymerase activity. PB2-292I coupled with 627K also enhances viral transcription and replication in cells, whereas PB2-292I alone did not show the same effect in the H7N9 virus. However, PB2-740A only had a limited prevalence in 2013, and the change from D to A in PB2-740A may have a negative effect on the replication of the H7N9 virus in cells.

RNA helicase A (RHA) is a ubiquitously expressed DExH-box helicase enzyme that is involved in a wide range of biological processes including transcription, translation, and RNA processing. A number of RNA viruses recruit RHA to the viral RNA to facilitate virus replication. DNA viruses contain a DNA genome and replicate using a DNA-dependent DNA polymerase. RHA has also been reported to associate with some DNA viruses during replication, in which the enzyme acts on the viral RNA or protein products. As shown for Epstein-Barr virus and Kaposi's sarcoma-associated herpesvirus, RHA has potential to allow the virus to control a switch in cellular gene expression to modulate the antiviral response. While the study of the interaction of RHA with DNA viruses is still at an early stage, preliminary evidence indicates that the underlying molecular mechanisms are diverse. We now review the current status of this emerging field.

The main objective of this narrative review is to describe the available evidence on the possible antiviral activity of ozone in patients with COVID-19 and its therapeutic applicability through hospital protocols. Amongst different possible therapies for SARS-CoV-2 pneumonia, ozone therapy seems to have an immunological role because of the modulation of cytokines and interferons, including the induction of gamma interferon. Some data suggest the possible role of ozone therapy in SARS, either as a monotherapy or, more realistically, as an adjunct to standard treatment regimens; therefore, there is increasing interest in the role of ozone therapy in COVID-19 treatment The PubMed and Scopus databases and the Italian Scientific Society of Oxygen Ozone Therapy website were used to identify articles focused on ozone therapy. The search was limited to articles published from January 2011 to July 2020. Of 280 articles found on ozone therapy, 13 were selected and narratively reviewed. Ozone exerts antiviral activity through the inhibition of viral replication and direct inactivation of viruses. Ozone is an antiviral drug enhancer and is not an alternative to antiviral drugs. Combined treatment with involving ozone and antivirals demonstrated a reduction in inflammation and lung damage. The routes of ozone administration are direct intravenous, major autohaemotherapy and extravascular blood oxygenation-ozonation. Systemic ozone therapy seems useful in controlling inflammation, stimulating immunity and as antiviral activity and providing protection from acute coronary syndromes and ischaemia reperfusion damage, thus suggesting a new methodology of immune therapy. Systemic ozone therapy in combination with antivirals in COVID-19-positive patients may be justified, helpful and synergic.

The family Togaviridae comprises several significant human and veterinary mosquito-borne pathogens. Two togaviruses (genus Alphavirus) have been previously identified in association with marine mammals, the southern elephant seal virus (SESV) and Eastern equine encephalitis virus (EEEV) from a fatal captive harbor seal infection. Herein we report the ultrastructural and phylogenomic characterization of a novel marine togavirus, the first isolated from a cetacean, an Alaskan harbor porpoise (Phocoena phocoena) displaying ulcerative dermatitis. A skin sample was processed for virus isolation on Vero.DogSLAMtag cells and cytopathic effects (CPE) were observed on primary isolation approximately 20 days post-infection. Transmission electron microscopy of the infected Vero.DogSLAMtag cells revealed typical alphavirus particles budding from both plasma and vacuolar membranes of infected cells. A next-generation sequencing approach was used to determine the near complete genome of the Alaskan harbor porpoise alphavirus (AHPV). Phylogenetic analysis supported the AHPV as the sister species to the SESV, forming a marine mammal alphavirus clade separate from the recognized alphavirus antigenic complexes. Genetic comparison of the protein coding sequence of the AHPV to other alphaviruses demonstrated amino acid identities ranging from 42.1-67.1%, with the highest identity to the SESV. Based on its genetic divergence, we propose the AHPV represents a novel alphavirus species, pending formal proposal to and ratification by the International Committee on Taxonomy of Viruses. The ecological and genetic characteristics of the AHPV and the SESV also suggest they represent a novel antigenic complex within the genus Alphavirus, which we propose to be named the Marine Mammal Virus Complex. The role of the AHPV in the associated harbor porpoise cutaneous pathology, if any, remains unclear. Further research is needed to determine AHPV's route(s) of transmission and potential vectors, host range, prevalence, and pathogenicity in cetaceans including harbour porpoises.

Geminiviruses cause devastating diseases in solanaceous crops, with the bipartite begomoviruses tomato yellow leaf curl Kanchanaburi virus (TYLCKaV) and pepper yellow leaf curl Thailand virus (PYLCThV) major threats in Southeast Asia. To determine the molecular mechanism of geminivirus infection, we constructed infectious clones of TYLCKaV and PYLCThV. Both constructs infected Nicotiana benthamiana, but only TYLCKaV could infect Solanum lycopersicum 'A39'. A genome-swapping of TYLCKaV with PYLCThV revealed the TYLCKaV-B genome segment as the determinant of TYLCKaV infectivity in tomato. We constructed five geminivirus clones with chimeric TYLCKaV-B and PYLCThV-B genome segments to narrow down the region determining TYLCKaV infectivity in tomato. Only chimeric clones carrying the TYLCKaV intergenic region (IR) showed infectivity in S. lycopersicum 'A39', indicating that the IR of TYLCKaV-B is essential for TYLCKaV infectivity in tomato. Our results provide a foundation for elucidating the molecular mechanism of geminivirus infection in plants.

Rotavirus (RV) replication occurs in cytoplasmic compartments, known as viroplasms, that are composed of viral and cellular proteins. Viroplasm formation requires RV nonstructural proteins NSP2 and NSP5 and cellular lipid droplets (LDs); however, the mechanisms required for viroplasm assembly remain largely unknown. We previously identified two conformationally-distinct forms of NSP2 (dNSP2, vNSP2) found in RV-infected cells that interact differentially with hypo- and hyperphosphorylated NSP5, respectively, and indicate a coordinated phosphorylation-dependent mechanism regulating viroplasm assembly. We also reported that phosphorylation of dNSP2 on serine 313 by the cellular kinase CK1α triggers the localization of vNSP2 to sites of viroplasm assembly and its association with hyperphosphorylated NSP5. To directly evaluate the role of CK1α-mediated NSP2 phosphorylation on viroplasm formation, we used a recently published plasmid-based reverse genetics method to generate a recombinant rotavirus (rRV) with a phosphomimetic NSP2 mutation (rRV NSP2 S313D). The rRV NSP2 S313D virus is significantly delayed in viroplasm formation, virus replication, and interferes with wild type RV replication during co-infection. The rRV NSP2 S313A virus was not rescued. Taking advantage of the delay in viroplasm formation, the NSP2 S313D phosphomimetic mutant was used as a tool to observe very early events in viroplasm assembly. We show that (1) viroplasm assembly correlates with NSP5 hyperphosphorylation, and (2) that vNSP2 S313D co-localizes with RV-induced LDs without NSP5, suggesting that vNSP2 phospho-S313 is sufficient for interacting with LDs and may be the virus factor required for RV-induced LD formation. Further studies with the rRV NSP2 S313D virus are expected to reveal new aspects of viroplasm and LD initiation and assembly.

Rotavirus species A (RVA) is the etiological agent of acute gastroenteritis in young individuals of various animal species, including humans. Vaccination has helped to reduce the impact of these viruses on humans and some species of domestic mammals, but they do not confer complete immunity, so antirotavirus agents are another important control option. In this study, millimolar concentrations of benzimidazole inhibited the replication of the Rhesus rotavirus (RRV) strain of RVA. Two mutants partially resistant to the inhibitory effect of benzimidazole were independently selected, and their genomes and those of their parental strains were fully sequenced. Most (7/11) mutations occurred in the gene that encodes the VP2 protein, and similarly most of the missense mutations (5/9), including the only one shared by the two mutants (G2,414 → R[G/A], D800 N), occurred in the VP2 gene. Our results identify the VP2 gene as the primary target affected by benzimidazole.

The envelope glycoprotein (E) is the smallest structural component of SARS-CoVs; plays an essential role in the viral replication starting from envelope formation to assembly. The in silico analysis of 2086 whole genome sequences from India performed in this study provides the first observation on the extensive deletion of amino acid residues in the C-terminal region of the envelope glycoprotein in 34 Indian SARS-CoV-2 genomes. These amino acid deletions map to the homopentameric interface and PDZ binding motif (PBM) present in the C-terminal region of E protein as well as immediately after the reverse primer binding region as per Charité protocol in 26 of these genomes, hence, their detection through RT-qPCR may not be hampered and therefore E gene-based RT-qPCR would still detect these isolates. Eight genomes from the State of Odisha had deletion even in the primer binding site. It is possible that the deletions in the C-terminal region of E protein of these genomes are a result of adapting to a newer geographical area and host. The information on the clinical status was available only for 9 out of 34 cases and these were asymptomatic. However, further studies are indispensable to understand the functional consequences of amino acid deletion in the C terminal region of SARS-CoV-2 envelope protein in the viral pathogenesis and host adaptation.

Erratum in Virus Res. 2022 Jan 2;307:198620.

Escherichia coli is an opportunistic bacterial pathogen that causes a wide range of nosocomial infections. The emergence of multidrug resistance in E. coli poses a severe threat to global health. Phage therapies are an alternative method to control multidrug-resistant pathogens, which have been attracting increasing attention. Owing to their ability to lyse bacteria specifically and efficiently, bacteriophages are considered novel antimicrobial agents. In this study, we used multidrug-resistant E. coli as an indicator and isolated, characterized, and compared two new phages of the Siphoviridae family referred to as vB_EcoS_XF and vB_EcoS_XY2. These phages were able to infect several pathogenic multidrug-resistant E. coli strains. A short latent period and large burst size ensured their rapidly reproduction in host cells. Their tolerance of high temperatures and high pH levels meant that remained stable when used to control pathogenic E. coli strains. No obvious cytotoxicity was observed when either HEK293 T or A549 cells were incubated with these two phages. Mass spectrometry analysis allowed us to identify several phage-encoded proteins. Genomic analysis revealed that no toxic proteins or antibiotic proteins were encoded. Genome comparison and phylogenetic analysis indicated that the phages identified show high similarity with E. coli phages of the genus Kagunavirus. The desirable characteristics of the novel phages identified make them good potential therapeutic candidates, and components of phage cocktails to treat multidrug-resistant E. coli in the future.

Red sea bream iridovirus (RSIV) is the causative agent of the iridoviral disease with high mortality rates in cultured fish. Our laboratory reported the first case of RSIV infection in India which resulted in mass mortalities of Asian seabass, Lates calcarifer. The RSIV-LC strain isolated from infected fish was subjected to complete genome sequencing and analysis. The complete genome of RSIV-LC was found to be of 111,557 bp in size having a G + C content of 53 %. The complete genome has 114 open reading frames (ORFs) of which 38 ORFs were predicted as functional proteins while the rest were hypothetical proteins. Among the ORFs 26 were found to be core genes reported earlier to be homologous in iridovirus complete genomes. Phylogenetic tree constructed based on the 26 core gene sequences, major capsid protein and ATPase genes revealed RSIV-LC in this study to belong to the genus Megalocytivirus of the RSIV-Genotype II. The present study provides the first report of the complete genome sequence and annotation of the RSIV strain isolated from India.

A molecular chaperone heat shock protein 90 (Hsp90) is required for efficient infection by several viruses. Hsp90 has been recently implicated in baculovirus infection, but its exact role remains obscure. This study investigated the effect of 17-N-allylamino-17-demethoxygeldanamycin (17-AAG), an Hsp90-specific inhibitor, on Bombyx mori nucleopolyhedrovirus (BmNPV) infection. The 17-AAG treatment significantly decreased the production of budded viruses and occlusion bodies in BmNPV-infected Bombyx mori cultured cells. Immunoblot and SDS-PAGE analyses showed that the expression of early and delayed early gene products, DBP and BRO, was delayed and dysregulated, and the very late gene product POLH was almost completely diminished. RT-qPCR experiments revealed that 17-AAG treatment did not affect initiation of the immediate early gene ie-1 expression, but the expression decreased by ∼50 % during the late stage of infection. 17-AAG treatment also decreased ie-1 promoter activity by ∼50 %. In addition, the expression of delayed early and late genes was dysregulated and inhibited, respectively. These results indicated that Hsp90 function is required for stable ie-1 transcription. Inhibiting Hsp90 function negatively affects ie-1 expression, resulting in dysregulation of delayed early genes and a severe decrease in late and very late gene expression.

The Bombyx mori nucleopolyhedrovirus (BmNPV)-based baculoviral expression vector system is among the most efficient expression vector systems for eukaryotic proteins especially when used in combination with silkworms as a host. We newly isolated a novel BmNPV strain (BmNPV H4) in Hokkaido, Japan that outperforms the type strain T3 in terms of both proliferation and expression of polyhedrin protein in silkworm larvae; however, it proliferates poorly in the BmN cell line. We inferred the gene responsible for the differences in proliferation between viral strains by quantifying amino acid similarity distances in protein functional domains and identifying highly divergent alleles between the H4 and T3 strains. Among proteins that differ markedly in functional domain sequence between H4 and T3, we identified the F gene, which encodes the F protein, as a putative cause of proliferative differences between the two strains. Using recombinant viruses with the F protein-coding sequence exchanged between H4 and T3, we determined that the T3 F protein increases H4 proliferation in BmN while the H4 F protein does not improve T3 proliferation in silkworm larvae. Our results suggest that the BmNPV F protein can strongly affect viral proliferation in a genetic background-specific manner and may be an important target for manipulating the proliferation characteristics of BmNPV-based expression vectors.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiologic agent of coronavirus disease 19 (COVID-19), and is genetically related to the 2003 SARS and Middle East respiratory syndrome (MERS-CoV) coronaviruses. Recent studies have reported that similar to SARS-CoV, this strain expresses a spike protein (S) with a receptor binding domain (RBD) that binds to angiotensin-converting enzyme 2 (ACE2) - an enzyme expressed mostly in the endothelium, kidneys, heart, gastrointestinal tract and lungs - to facilitate viral entry and intracellular replication. Incidentally, the renin-angiotensin-aldosterone system (RAAS) is integral to physiologic control of both ACE and ACE2 expression, and is an essential system utilized by SARS-CoV-2, albeit with varying schools of thought on how it can affect viral entry. In this paper, we will review current knowledge on the RAAS and how it can be affected by non-steroidal anti-inflammatory drugs (NSAIDs) and corticosteroid use at the organ and cellular levels. We will then discuss the relevance of these interactions on organ-specific ACE2 expression, and provide scientific insights on how this mechanism can potentially affect SARS-CoV-2 infection in the early phases of disease. From the standpoint of other known viruses, we will then aim to discuss the potential uses or restrictions of these drugs in viral infection, and provide an update on relevant studies about COVID-19.

Seneca Valley virus (SVV) is a novel Picornaviridae that is closely associated with porcine idiopathic vesicular disease (PIVD). Here, a novel SVV strain (CH-GX-01-2019) was detected and isolated from swine in Guangxi Province, China. The complete genomic sequence of CH-GX-01-2019 exhibited 93.3-98.9 % identify with other SVV isolates at the nucleotide level. CH-GX-01-2019 showed the highest level of similarity (98.9 %) with Vietnamese strains. And CH-GX-01-2019 exhibited two consecutive amino acid mutations in VP1 gene. Phylogenetic analysis based on the complete genome and the VP1 gene showed that Chinese SVV isolates can be divided into three clusters. We analyzed the geographical distributions of SVV strains in China and found that the epidemiology of SVV in China is complicated; most strains are distributed predominantly in south and central China. Between 2015 and 2019, the dominant epidemic SVV isolates in China have changed from clusters 1 and 3 to cluster 2. CH-GX-01-2019 (cluster 3) is a recombinant strain from Colombia-2016 (cluster 2) and HB-CH-2016 (cluster 1). Our findings will enhance our understanding of the prevalence and genetic variation of SVV in the swine herds of China and provide important insights into the molecular epidemiology of SVV.

The SD12-F120 is a live-attenuated genotype I strain of Japanese encephalitis virus (JEV) and was obtained by serial passage of wild-type strain SD12 on BHK-21 cells combined with multiple plaque purification and virulence selection in mice. The large scale production and vast clinical trials always demand ideal safety and efficacy profile of live-attenuated vaccines. In the present study, SD12-F120VC has undergone serial passaging of P1-P30 in WHO qualified Vero cells to assess the potential effect of adaptation to growth on Vero cells. The series of experiments showed that vaccine SD12-F120VC (Vero cell adapted) variants have consistently increased in peak virus titer compared to early passages and have good adaptation to growth in Vero cells. The animal experiments showed that Vero cell adapted SD12-F120VC variants have attenuation phenotype in suckling mice and the plaque morphology for all SD12-F120VC variants was small. Vaccination of mice with SD12-F120VC vaccine produced complete protection for homologous SD12 genotype I strain, but failed to give the complete protection of vaccinated mice against the challenge of heterologous N28 genotype III strain. In response to immunization of SD12-F120VC in mice, the neutralizing antibodies titer against homologous SD12-F120VC and SD12 (GI) was higher than heterologous N28 (GIII) strain. The prM protein has 6 amino acid substitutions, of which 5 amino acid changes were confined at the start of the pr domain in the ∼40 amino acids, and some mutations in the pr domain of prM might contribute to Vero cell adaptation. Our findings in this study are important for validation, evaluation and quality control study of live attenuated flaviviruses vaccines and show that Vero cells are a suitable substrate for the production of a safe and stable live-attenuated JEV vaccine.

The resistance of hepatitis C virus (HCV) to direct-acting antiviral agents, used in chronic hepatitis C treatment, consists of a natural process resulting from resistance-associated substitutions (RASs) at specific amino acid regions. To identify and establish the natural prevalence of RASs in the NS3 gene in patients with chronic hepatitis C in the state of Pará, northern Brazil. Molecular analysis was performed on a total of 35 patients infected with HCV genotype 1, who were treatment-naive to protease inhibitors. HCV RNA was extracted from plasma and the NS3 region was amplified and submitted to DNA sequencing (Sanger). The general natural prevalence of RASs in the NS3 gene was 37.5 % (Y56F and S122T). The substitutions Y56F (34.3 %), S122T (3.1 %), V132I (15.6 %) and V170I (9.3 %) were identified. Y56F and S122T provide resistance to the protease inhibitors grazoprevir and simeprevir, respectively. All amino acid substitutions in the NS3 gene, including RASs, identified in patients from the state of Pará were present in other Brazilian studies. The natural presence of RASs in this study reflects the elevated genetic variability of HCV.

Bombyx mori cypovirus 1 (BmCPV1) is a member of the Reoviridae family which is characterized by its single-layered capsid. Similar with other turreted viruses in the Reoviridae, transcription of BmCPV1 occurs inside the capsid, and the nascent mRNA is released to the turret which consists of five turret proteins (TPs) and located at the 5-fold axis of the outer capsid, then the capping enzyme TP will guanylate and methylate the nascent viral mRNA to produce a matured mRNA. However, during these processes, how the BmCPV1 draws other cellular proteins to facilitate its replication is still lesser-known. Here we used an ELISA to investigate the interaction between ALP and BmCPV1. A co-immunoprecipitation technique was employed to detect the interaction of ALP with the Methylase domain of TP. We further studied whether ALP affects the replication of BmCPV1 inside the cell, results show that reducing the expression of ALP through RNAi reduced the transcription level of the BmCPV1 VP1 gene, which was increased by overexpression of ALP. In summary, our data demonstrate an interaction between ALP and BmCPV1 and that ALP promoted the replication of BmCPV1, and support our hypothesis of the ALP is an RTPase to facilitate the capping process of BmCPV1.

Multidrug resistant bacterial infections are difficult to treat and contribute to high morbidity and mortality. The phage vB PaeP-SaPL was isolated from a sewage drain (Lahore, Pakistan) against Pseudomonas aeruginosa PA-1 (NCBI Accession number MG763232). SaPL produced circular, transparent plaques, 4-5 mm in diameter and showed broad host range infecting 57 % of tested MDR P. aeruginosa clinical isolates (N = 38), while no infectivity was observed against any tested strains of other genera. SaPL inhibited PA-1 growth until 24 h post infection at MOI of 1. The SaPL showed stability at varying temperature and pH, with optimum stability at pH 7 and 45 °C. The latent period of SaPL was 20 min with burst size of 155 virions. The genome of SaPL was double stranded DNA of 45,796 bps having 63 CDS (13 for known proteins and 50 for hypothetical proteins) with a GC content of 52 %. The termini analysis revealed that SaPL genome ends are redundant and permuted. The packaging strategy used by SaPL was a headful (pac) strategy like P1 phage. Survivability of PA-1 challenged mice, treated with SaPL (100 %) was statistically significant (P < 0.05) than in untreated challenged mice (0%). Based on its efficacy in reducing bacterial growth, selective infectivity against majority of P. aeruginosa strains and its ability to increase survivability in PA-1 challenged mice, SaPL is proposed to be a potential candidate for bacteriophage therapy against difficult to treat MDR P. aeruginosa infections.

Human parvovirus 4 (PARV4) is a novel tetraparvovirus that was isolated from intravenous drug users in 2005. Recombinant PARV4 capsid protein VP2 can form stable virus-like particles (VLPs) in yeast. These VLPs could act as antigen carriers during vaccine development. Therefore, the information about PARV4 VP2 VLP antigenic sites could advance further research in this area. In this work, human parvovirus 4 VLPs obtained from yeast were used to generate monoclonal antibodies (mAbs) in mice. Epitope mapping of the obtained mAbs showed at least three distinct antigenic sites of the VP2 protein. On top of that, molecular cloning was used to replace PARV4 VP2 antigenic sites with heterologous peptides. The chimeric PARV4 VLPs bearing polyhistidine inserts obtained from yeast were observed using electron microscopy while polyhistidine-specific antibodies detected heterologous peptides of the chimeric VP2 proteins.

A deterministic compartmental model of the corona virus diseases has been introduced to investigate the current outbreak and epidemic peaks of the 2019-nCoV in South Asian countries. We have done details analysis of the above mentioning model and demonstrated its application using publicly reported data. Based on the reported data, we have determined the new infective rate, β = 0.0017, β = 0.00069, β = 0.0011 and β = 0.00047 for India, Pakistan, Bangladesh and Afghanistan, respectively and these values are not found for other countries due to insufficient data. According to present model, the epidemic under the most restrictive measures was observed of peak on around June 29 in Bangladesh with a peak size of 4100 infectious individuals which was closed to real size of 4014. In the same way, the model results also showed that the epidemic peaks were found on around September 16 in India, June 13 in Pakistan and on June 5 in Afghanistan with the peak sizes of 97,500, 6950 and 935, respectively those were closed to the real values. In addition, we have derived a model-implied basic reproduction number for each day of currently infected cases so that the mitigation and defeat strategies can be imposed to control the size of the epidemic.

Given the dual life cycle of arboviruses in insect and animal hosts and the importance of serum factors as a first line antiviral defense, we have examined the outcome of interactions between the arbovirus La Crosse Virus (LACV) and human serum. To mimic the life cycle between species, we used LACV derived from insect (I-LACV) and human keratinocyte (HaCaT) cells. Incubation of I-LACV with normal human serum did not result in neutralization, but instead stabilized I-LACV virions and enhanced the amount of infectious virus. Enhanced infectivity was also seen with heat-inactivated serum devoid of complement activity and with serum from a range of animals including mouse, ferret, and non-human primates. Depletion of antibodies from serum resulted in loss of enhancement of infectivity and sucrose gradient sedimentation assays showed IgG co-sedimenting with I-LACV particles. In agreement with our results with I-LACV, HaCaT-derived LACV was not neutralized by complement or antibodies in normal human serum. However, in contrast to I-LACV, HaCaT-derived LACV infectivity was stable when incubated alone and treatment with serum did not enhance infectivity. Our results indicate that LACV derived from insect cells differs substantially from virus derived from human cells, with I-LACV being dependent on serum factors to enhance infectivity. These findings suggest that understanding differential composition of insect versus animal cell-derived LACV may form the foundation for potential new antiviral approaches.

Newcastle disease is a severe clinical manifestation of avian species caused by Newcastle disease virus (NDV). Although several vaccination strategies are available to protect poultry against NDV infection, even then, outbreaks have been reported in the vaccinated birds. The lack of therapeutics against NDV makes the need for effective anti-viral drugs is of utmost importance. Lithium Chloride (LiCl) is a widely prescribed drug for the treatment of bipolar disorder, acute brain injuries, and chronic neurodegenerative diseases. Also, LiCl has been repurposed as an effective anti-viral drug for some viral infections. In the present work, we have investigated the efficacy of LiCl to inhibit NDV replication using in vitro, in ovo, and in vivo models. Our results collectively showed the modulation of NDV replication after the LiCl treatment. We also demonstrated that NDV induces endoplasmic reticulum stress (ER-stress), and a stress-inducible ER chaperone, glucose-regulating protein 78 (GRP78), was found to be over-expressed after NDV infection. Subsequently, the treatment of NDV infected cells with LiCl significantly reduced the transcript and protein levels of GRP78. Finally, we concluded that LiCl treatment protects the cells from ER-stress induced by the NDV infection.

Tembusu virus (TMUV) causes disease in poultry, especially in ducks, resulting in abnormality in egg production and with high morbidity and mortality, resulting in great loss in duck farming industry in China and Southeast Asia. Previous studies on the pathogenesis of TMUV infection have been mostly conducted in poultry, with a few studies being undertaken in mice. While TMUV does not cause disease in humans, it has been reported that antibodies against TMUV have been found in serum samples from duck farmers, and thus data on TMUV infection in humans is limited, and the pathogenesis is unclear. In this study we investigated the cell tropism and potential susceptibility of humans to TMUV using several human cell lines. The results showed that human nerve and liver cell lines were both highly susceptible and permissive, while human kidney cells were susceptible and permissive, albeit to a lower degree. In addition, human muscle cells, lung epithelial cells, B-cells, T-cells and monocytic cells were largely refractory to TMUV infection. This data suggests that liver, neuron and kidney are potential target organs during TMUV infection in humans, consistent with what has been found in animal studies.

The channel catfish virus (CCV) can cause lethal hemorrhagic infection in channel catfish, resulting in significant economic losses in the fish industry. Effective drugs for the virus are still lacking. Acyclovir is known as a potent antiviral agent against human herpes viruses and some animal DNA viruses. The present study was undertaken to explore the antiviral response and mechanism of acyclovir against CCV in channel catfish ovary (CCO) cells. Acyclovir was able to significantly inhibit the expression of viral genes related to CCV viral DNA synthesis and suppress viral replication at a safe concentration. Furthermore, acyclovir blocked the cytopathic effects and apoptosis induced by CCV, thereby maintaining the normal cellular morphological structure, as shown by the protection of CCO cells from the formation of apoptotic bodies or nuclear fragmentation. Moreover, reverse transcript quantitative polymerase chain reaction (RT-qPCR) demonstrated that acyclovir suppressed the expression of caspase 3, caspase 8 and caspase 9, while there was no significant impact on the expression of the apoptosis-inhibiting gene bcl-2 in CCV-infected cells. In addition, acyclovir did not promote the expression of immune-related genes such as MyD88, Mx1, IRF3, IRF7, IFN-I, NF-kB and IL-1β, suggesting that the antiviral activity of acyclovir to CCV infection is not achieved by facilitating the expression of immune-related genes in CCO cells. Taken together, the results from this study suggest that acyclovir could effectively regulate CCV-induced infection, and thus is a promising therapeutic agent against CCV. Our results will aid our understanding of the pharmacological mechanisms of antiviral agents.

Enterovirus A71 (EV-A71) emerged as a leading cause of virus derived infant encephalitis in most Asian countries. Some recent studies point out the critical role of microRNA (miRNA) in the regulation of pyroptosis. However, the role of miRNAs in the regulation of EV-A71 infection-induced pyroptosis was not previously explored. In this study, we utilized microRNA array and real-time PCR to verify that miR-195 significantly down-regulate in EV-A71-infected SH-SY5Y human neuroblastoma cells. An inverse correlation of NLRX1 with miR-195 expression in EV-A71-infected SH-SY5Y cells was found. Target prediction of miR-195 showed that NLRX1 could directly interact with miR-195. Results from luciferase reporter assays, qRT-PCR and western blotting demonstrated the negative regulation between miR-195 and NLRX1. Silencing NLRX1 expression with small interfering RNAs (siRNAs-NLRX1) and over-expression of miR-195 also attenuate the EV-A71 associated pyroptosis. Our findings provided evidence showed that miR-195 can regulate EV-A71 infection-induced pyroptosis, by directly targeting NLRX1.

Coronavirus Disease 2019 (COVID-19) is a global public health threat. Genomic surveillance of SARS-CoV-2 was implemented in March of 2020 at a major diagnostic hub in Bangkok, Thailand. Several virus lineages supposedly originated in many countries were found, and a Thai-specific lineage, designated A/Thai-1, has expanded to be predominant in Thailand. A virus sample in the SARS-CoV-2 A/Thai-1 lineage contains a frame-shift deletion at ORF7a, encoding a putative host antagonizing factor of the virus.

Infectious bronchitis virus (IBV) of GI-19 (QX), GI-7 (TW), GI-13 (4/91) and GI-1 (Mass) lineages have been frequently detected in China in recent years. Here, An IBV strain, referred as GD17/04, was isolated from the dead yellow feather chicken vaccinated with H52 and 4/91 vaccines, whose genome sequence was obtained through high-throughput sequencing. Then it has been confirmed by the RDP and SimPlot analysis that GD17/04 is a recombinant strain deriving from YX10, 4/91, TW 2575/98 and H52 strains. Therein S1 gene of GD17/04 consists of sequences of TW2575/98 and 4/91, the former for the region of 20,371 to 21,072 nt and 21,847 to 21,975 nt, the latter for the sandwiched region of 21,073 to 21,846 nt. Moreover, as a nephropathogenic variant which caused high morbidity of 100 % and mortality of 60 %, unlike most other IBV strains, GD17/04 can cause obvious cell lesion in primary CEK cell, and even in DF-1 cells, without the process of continuous passage. As the few IBV strain can infect avian passage cell line, GD17/04 provides a material basis for further study of the interaction mechanism between IBV and avian host. Collectively, the findings highlight the significance that biological characteristics of novel strain should be studied, in addition to constant epidemiologic and molecular surveillance for IBV.

Suspect has been directed towards some direct acting antivirals (DAAs) due to their reported association with hepatocellular carcinoma (HCC) development in chronic hepatitis C (CHC) patients. The mechanisms behind HCC development, following CHC treatment, were not well understood and may be linked to genetic variabilities in different patients which affect several cytokine productions involved in angiogenesis and inflammation. Of these variabilities, is the genetic polymorphisms in the interleukin-17 (IL-17) A receptor gene. Being an important pleiotropic cytokine, this study aimed to investigate the association between haplotypes in IL-17A receptor rs2275913 and rs3819024 and development of HCC in CHC patients treated with either triple therapy (sofosbuvir (SOF), pegylated interferon-alpha-2a (Peg-IFNα-2a) & ribavirin(RBV)) or with dual therapy (Peg-IFNα-2a&RBV). A cohort of 100 CHC patients was recruited in this study. Samples were tested for single nucleotide polymorphism (SNPs) in IL-17A receptor (rs2275913 and rs3819024) using TaqMan Genotyping assay. Our results showed that the presence of G-G haplotype in IL-17A (rs2275913& rs3819024) is inversely associated with HCC development in patients receiving triple therapy. While, high serum AFP levels are directly associated with HCC development in patients receiving triple therapy. However, in patients receiving dual therapy, HCC development was only associated with high serum alpha fetoprotein (AFP) levels and was not correlated to any specific allele in our studied SNPs. Such results highlight the importance of IL17A receptor gene haplotyping in the prediction of HCC development in patients receiving triple therapy. These results will aid in performing tailored, personalized strategy for CHC treatment.

New therapies against hepatitis B virus (HBV) require the elimination of covalently closed circular DNA (cccDNA), the episomal HBV genome. HBV plasmids containing an overlength 1.3-mer genome and bacterial backbone (pHBV1.3) are used in many different models, but do not replicate the unique features of cccDNA. Since the stable cccDNA pool is a barrier to HBV eradication in patients, we developed a recombinant circular HBV genome (rcccDNA) to mimic the cccDNA using Cre/LoxP technology. We validated four LoxP insertion sites into the HBV genome using hydrodynamic tail vein injection into murine liver, demonstrating high levels of HBV surface antigen (HBsAg) and HBV DNA expression with rcccDNA formation. HBsAg expression from rcccDNA was >30,000 ng/mL over 78 days, while HBsAg-expression from pHBV1.3 plasmid DNA declined from 2753 ng/mL to 131 ng/mL over that time in immunodeficient mice (P < 0.001), reflective of plasmid DNA silencing. We then cloned Cre-recombinase in cis on the LoxP-HBV plasmids, achieving plasmid stability in bacteria with intron insertion into Cre and demonstrating rcccDNA formation after transfection in vitro and in vivo. These cis-Cre/LoxP-HBV plasmids were then used to create HBx-mutant and GFP reporter plasmids to further probe cccDNA biology and antiviral strategies against cccDNA. Overall, we believe these auto-generating rcccDNA plasmids will be of great value to model cccDNA for testing new therapies against HBV infection.

BACKGROUND: Human immunodeficiency virus (HIV) increasing molecular diversity and emergence of drug resistant mutants remain a major concern in China. Enfuvirtide (ENF/T-20) is the first entry inhibitor used in patients failing highly active antiretroviral therapy (HAART). However, data on HIV-1 gp41genetic diversity and primary ENF resistance-associated mutations among treatment-naïve patients in China is limited. The objective was to identify molecular diversity and ENF resistance patterns of HIV-1 in southern China, using envelope (env) gp41 sequences and bioinformatics tools, which may help optimize antiretroviral therapy. METHODS: From November 2018 to January 2019, 439 blood plasma samples from ENF-naïve patients were collected from Shenzhen, Wuhan and Chongqing, of which 396 HIV env regions were sequenced and subtyped, and were performed the analysis of drug resistance-associated mutations (DRMs). RESULTS: The main subtypes were circulating recombinant form (CRF) 01_AE (30.6 %) and CRF07_BC (48.7 %). CRF55_01B had been the fourth subtype in the study, and many rare CRFs were observed. Notably, CRF02_AG and CRF_BF strains typically found in Africa and US respectively were identified amongst Chinese patients. Known DRMs were detected in 27.5 % (109/396) of ENF treatment-naïve patients. One major DRM (L44 M), many secondary DRMs (including N126 K, E137 K, S138A), and lots of polymorphisms were found in the study, which have been proved to elevate resistance to ENF. CONCLUSIONS: HIV-1 molecular diversity was observed in the study, which indicating that HIV-1 variability is becoming increasingly complex in southern China. A diverse set of primary DRMs discovered in this study described the serious threat to ART, which reminds us the urgent need of timely surveillance of HIV-1 viral diversity and drug resistance in China.

crAssphages are a broad group of diverse bacteriophages in the order Caudovirales that have been found to be highly abundant in the human gastrointestinal tract. Despite their high prevalence, we have an incomplete understanding of how crAssphages shape and respond to ecological and evolutionary dynamics in the gut. Here, we report genomes of crAssphages from feces of one South African woman and three infants. Across the complete genome sequences of the South African crAssphages described here, we identify particularly elevated positive selection in RNA polymerase and phage tail protein encoding genes, contrasted against purifying selection, genome-wide. We further validate these findings against a crAssphage genome from previous studies. Together, our results suggest hotspots of selection within crAssphage RNA polymerase and phage tail protein encoding genes are potentially mediated by interactions between crAssphages and their bacterial partners.

Foot-and-mouth disease (FMD) virus 3A protein regulates viral replication and virulence; thus, we generated BHK-Flp-In cell line expressing 3A protein because it can serve as helper cell line for infecting a replication defective FMDV to produce a live disabled vaccine. FMDV Asia1 3A was amplified, cloned in pcDNA vector and confirmed by sequencing. The 3A gene was subcloned in pcEF/FRT vector and transfected in BHK-Flp-In cells and transformed cells were selected by resistance to hygromycin and susceptibility to zeocin antibiotics. The BHK-Flp-In cells expressing 3A protein was designated as Flp-In3A. Western blot and immunofluorescence confirmed that Flp-In3A cells expressed FMDV3A protein. Absolute quantitation of 3A transcripts showed peak expression at 6 h in Flp-In3A cells followed by a sharp decrease and the cells showed growth retardation for 2 h post-seeding with cytoplasmic vacuolations with advancing time. Response to infection with FMDV Asia1 virus revealed smaller plaques in Flp-In3A cells. Then, we investigated the effect of FMDV3A expression on autophagy related genes by real time PCR. Most autophagy genes were upregulated by 9 h post-seeding of which, autophagosome marker LC3B-II was demonstrated by western blot. Transient expression of 3A in PK-15 cells upregulated both Th1 and Th2 genes. The study suggested that the expressed 3A protein of FMDV cannot be used for 3A trans-supplementation in helper cells; however, it acts as an endogenously processed antigen that has the potential to elicit immune response in vivo.

Update of bioRxiv. 2020 Aug 05;:

To evaluate and quantify the evolutionary dynamics of the bipartite begomovirus tomato severe rugose virus (ToSRV) in a cultivated and a non-cultivated host, plants of tomato and Nicandra physaloides were biolistically inoculated with an infectious clone and systemically infected leaves were sampled at 30, 75 and 120 days after inoculation. Total DNA was extracted and sequenced in the Illumina HiSeq 2000 platform. The datasets were trimmed with the quality score limit set to 0.01, and the assembly was performed using the infectious clone sequence as reference. SNPs were filtered using a minimum p-value of 0.001 and the sum frequencies were used to calculate the deviation from the original clone sequence. Nucleotide substitution rates were calculated for the two DNA components in both hosts: 1.73 × 10-3 and 3.07 × 10-4 sub/site/year for the DNA-A and DNA-B, respectively, in N. physaloides, and 8.05 × 10-4 and 7.02 × 10-5 sub/site/year the for DNA-A and DNA-B, respectively, in tomato. These values are in the same range of those estimated for viruses with single-stranded RNA genomes and for other begomoviruses. Strikingly, the number of substitutions decreased over time, suggesting the presence of bottlenecks during systemic infection. Determination of Shannon's entropy indicated different patterns of variation in the DNA-A and the DNA-B, suggesting distinct evolutionary forces acting upon each component.

The first incidence of COVID-19 was reported in the Wuhan city of Hubei province in China in late December 2019. Because of failure in timely closing of borders of the affected region, COVID-19 spread across like a wildfire through air travel initiating a pandemic. It is a serious lower respiratory track viral infection caused by highly contagious, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2). Coronavirus including COVID-19 causing SARS-CoV-2 causes zoonotic diseases and thought to be originated from bats. Since its first incidence, the virus has spread all across the world, causing serious human casualties, economic losses, and disrupting global supply chains. As with SARS-CoV, COVID-19 causing SARS-CoV-2 follows a similar path of airborne infection, but is less lethal and more infectious than SARS and MERS. This review focusses on the pathogenesis of SARS-CoV-2, especially on the dysfunctional immune responses following a cytokine storm in severely affected persons. The mode of entry of SARS-CoV-2 is via the angiotensin converting enzyme 2 (ACE-2) receptors present on the epithelial lining of lungs, gastrointestinal tract, and mucus membranes. Older persons with weaker immune system and associated co-morbidities are more vulnerable to have dysfunctional immune responses, as most of them concomitantly have severe hypovitaminosis D. Consequently, causing severe damage to key organs of the body including lungs and the cardiovascular system. Since, vast majority of persons enters to the intensive care units and died, had severe vitamin D deficiency, thus, this area must be investigated seriously. In addition, this article assesses the role of vitamin D in reducing the risk of COVID-19. Vitamin D is a key regulator of the renin-angiotensin system that is exploited by SARS-CoV-2 for entry into the host cells. Further, vitamin D modulates multiple mechanisms of the immune system to contain the virus that includes dampening the entry and replication of SARS-CoV-2, reduces concentration of pro-inflammatory cytokines and increases levels of anti-inflammatory cytokines, enhances the production of natural antimicrobial peptide and activates defensive cells such as macrophages that could destroy SARS-CoV-2. Thus, this article provides the urgency of needed evidences through large population based randomized controlled trials and ecological studies to evaluate the potential role of vitamin D in COVID-19.

Programmed death ligand 1 (PD-L1) up-regulation on antigen presenting cells induces T cell dysfunction, strongly impairing immune response. Human Herpesviruses (HHV) 6B is a β-herpesvirus that, although displays a higher tropism for T cells, can infect other immune cells including monocytes and dendritic cells (DCs) and neuronal cells. We have previously shown that HHV-6B infection of primary monocytes reduced autophagy and induced Endoplasmic Reticulum (ER) stress/ Unfolded Protein Response (UPR), impairing their survival and differentiation into DCs. In this study, we found that PD-L1 expression was up-regulated by HHV-6B on the surface of infected monocytes and that its extracellular release also increased, effects known to lead to an impairment of anti-viral immune response. At molecular level, PD-L1 up-regulation correlated with the activation of a positive regulatory circuit between the increase of intracellular ROS and the activation of STAT1 and STAT3 induced by HHV-6B, accompanied by a high release of pro-inflammatory/immune suppressive cytokines. In conclusion, this study unveils new strategies put in place by HHV-6B to induce immune dysfunction and the underlying molecular pathways that could be targeted to counteract such immune suppressive effects.

Senecavirus A (SVA), previously known as Seneca Valley virus, is classified into the genus Senecavirus in the family Picornaviridae. SVA is not pathogenic to normal human cells, but has potent oncolytic activity in some tumor cells with neuroendocrine feature, such as small cell lung cancer (SCLC) NCI-H446 cell line. In this study, we rescued and characterized a recombinant SVA that could efficiently express a novel luciferase, NanoLuc® luciferase (NLuc), which was smaller and "brighter" than others. This NLuc-tagged recombinant SVA (rSVA-NLuc) exhibited high capacity for viral replication, but genetic instability of NLuc during serial virus passages. The NLuc as a reporter facilitated oncolytic analysis of rSVA-NLuc in H446 cells. The rSVA-NLuc-infected H446 cells exhibited an oncolytic phenotype characterized by cell rounding, swelling, detachment and lysis at 48 h post infection. Kinetic curve showed that the NLuc was rapidly expressed in H446 cells during an exponential phase of viral growth. Because the NLuc offered several advantages over fluorescent proteins for assay scalability in vivo, the rSVA-NLuc would play a potential role in facilitating in vivo imaging studies of oncolytic virotherapy.

Reported here is the complete genome sequence of Mourilyan virus (MoV) that infects giant tiger (Penaeus monodon) and kuruma prawns (P. japonicas) in Australia. Its genome was determined using various PCR strategies based on the sequences of 3 randomly-amplified cDNA clones to its L and M RNA segments discovered in a library generated to determine the genome sequence of gill-associated ronivirus. The sequences of PCR products and clones obtained showed the MoV genome to comprise 4 ssRNA segments (L, M, S1 and S2), as confirmed by Northern blotting using RNA from naïve and MoV-infected prawns, and by Illumina sequence analysis of semi-purified MoV. BLASTn searches identified the MoV L, M and S1 RNA segments to be homologous to Wēnzhōu shrimp virus 1 (WzSV1) segments discovered recently in a P. monodon RNA-Seq library (SRR1745808). Mapping this read library to the MoV S2 RNA segment identified WzSV1 to also possess an equivalent segment. BLASTp searches identified the putative non-structural protein (NSs2; 393-394 aa) encoded in their S2 RNA segments to have no homologs in GenBank. Possibly due to NSs2 being encoded in a discrete RNA segment rather than in ambisense relative to the N protein as in the S RNA segments of other phenuiviruses, each of 6 MoV S1 RNA segment clones sequenced possessed a variable-length (≤ 645 nt) imperfect GA-repeat extending from the N protein stop codon to the more variable ∼90 nt segment terminal sequence. Read mapping of RNA-Seq library SRR1745808 showed the WzSV1 S1 RNA segment to possess a similar GA-repeat. However, paired-read variations hindered definitive assembly of a consensus sequence. All 4 MoV and WzSV1 RNA segments terminated with a 10 nt inverted repeat sequence (5'-ACACAAAGAC.) identical to the RNA segment termini of uukuviruses. Phylogenetic analyses of MoV/WzSV1 RNA-dependant RNA polymerase (L RNA), G1G2 precursor glycoprotein (M RNA) and nucleocapsid (N) protein (S1 RNA) sequences generally clustered them with as yet unassigned crustacean/diptera bunya-like viruses on branches positioned closely to others containing tick-transmitted phenuiviruses. As genome sequences of most phenuiviruses discovered recently have originated from meta-transcriptomics studies, the data presented here showing the MoV and WzSV1 genomes to comprise more than 3 RNA segments, like the plant tenuiviruses, suggests a need to investigate the genomes of these unassigned viruses more closely.

Viruses are highly diverse and are the sole agents that can infect organisms in all domains of life. Viruses are defined as capsid-encoding organisms as opposed to ribosome-encoding cellular organisms. However, recent advances in virology indicate the existence of unique viruses that do not meet this basic definition, such as capsidless viruses. During virome analysis of the soft tick Argas japonicus, we identified virus-like sequences closely related to the members of genus Nyavirus (family Nyamiviridae). Further analysis revealed sequences derived from a novel nyavirus that lacks two structural protein genes, matrix (M) and glycoprotein (G). This unique nyavirus is tentatively named Sekira virus (SEKRV). To our knowledge, this is the first study to report a nyavirus deficient in M and G genes in nature. The mechanism of infection, replication, and persistence of SEKRV remain unknown, yet this finding provides new insight into virus evolution and the diverse way of viral life in nature.

The preferential use of a specific codon, out of a group of synonymous codons encoding the same amino acid, in a gene transcript results from the bias in codon choice. Various evolutionary forces namely mutation pressure and natural selection influence the pattern of codon usage i.e. distinct for each gene/genome. We investigated the pattern of codon usage of eight human herpesvirus genomes and compared them with two other herpesvirus genomes namely murine herpesvirus 68 and bovine herpesvirus type 1.1 to elucidate its compositional features, pattern of codon usage across the genomes and report the differences of codon usage pattern of human herpesviruses from that of other two other viruses. We also identified the similarity of the codon usage of human herpesviruses with its host (human). The genes were found to be CG rich in HHV2, HHV3, HHV4, HHV6, HHV7 and BH genomes while TA rich in HHV1, HHV5, HHV8 and MH genomes. The codon usage bias (CUB) of genes was low. A highly significant correlation was found among compositional contents depicting the role of mutational pressure along with natural selection in framing CUB. Several more frequently used codons as well as less frequently used codons were identified to be similar between each human virus and its host (human), while murine herpesvirus 68 and bovine herpesvirus type 1.1 genomes did not possess similar adaptation strategy as human herpesviruses to human (host), thus we could conclude that viral CUB might have been shaped as per their host's nature for better surveillance. Neutrality plot revealed mutational pressure mostly influenced the CUB of HHV1, HHV8 and MH viruses, while natural selection had a major impact in the CUB of HHV2, HHV3, HHV4, HHV5, HHV6, HHV7 and BH genomes.

The objective of this study was to investigate safety and efficacy using a low-temperature immunization protocol with NNV in sevenband grouper, Epinephelus septemfasciatus. Further, NNV specific antibody post immunization and intramuscularly challenge was also evaluated. Immunization at low temperature resulted in a low titer virus infection in brain tissues without any clinical symptoms of infection such as sluggish behavior and/or spinning, rotating swimming being observed, and no mortality was observed. Post challenge, NNV titer NNV giving an RPS of 100 %, increased in brain tissues of naïve (non-immunized) sevenband grouper NNV giving an RPS of 100 %, with a cumulative mortality of 100 % at 25 days post-infection. No mortality or disease symptoms NNV giving an RPS of 100 %, as NNV giving and of 100 %, observed in the groups immunized at low temperature with live NNV giving an RPS of 100 %. NNV giving an RPS of 100 %. NNV specific antibody was not detected in live NNV vaccinated sevenband grouper. This is the first study that confirms that field-scale NNV immersion vaccine can protect sevenband grouper against lethal infection with NNV at natural seawater temperature under the gradually increased from 14.3-24.8 °C.