BF/C2 is a crucial molecule in the coagulation complement cascade pathway and plays a significant role in the immune response of grass carp through the classical, alternative, and lectin pathways during GCRV infection. In vivo experiments demonstrated that the mRNA expression levels of BF/C2 (A, B) in grass carp positively correlated with GCRV viral replication at various stages of infection. Excessive inflammation leading to death coincided with peak levels of BF/C2 (A, B) mRNA expression and GCRV viral replication. Correspondingly, BF/C2 (A, B) recombinant protein, CIK cells and GCRV co-incubation experiments yielded similar findings. Therefore, 3 h (incubation period) and 9 h (death period) were selected as critical points for this study. Transcriptome sequencing analysis revealed significant differences in the expression of BF/C2A and BF/C2B during different stages of CIK infection with GCRV and compared to the blank control group (PBS). Specifically, the BF/C2A_3 and BF/C2A_9 groups exhibited 2729 and 2228 differentially expressed genes (DEGs), respectively, with 1436 upregulated and 1293 downregulated in the former, and 1324 upregulated and 904 downregulated in the latter. The BF/C2B_3 and BF/C2B_9 groups showed 2303 and 1547 DEGs, respectively, with 1368 upregulated and 935 downregulated in the former, and 818 upregulated and 729 downregulated in the latter. KEGG functional enrichment analysis of these DEGs identified shared pathways between BF/C2A and PBS groups at 3 and 9 h, including the C-type lectin receptor signaling pathway, protein processing in the endoplasmic reticulum, Toll-like receptor signaling pathway, Salmonella infection, apoptosis, tight junction, and adipocytokine signaling pathway. Additionally, the BF/C2B groups at 3 and 9 h shared pathways related to protein processing in the endoplasmic reticulum, glycolysis/gluconeogenesis, and biosynthesis of amino acids. The mRNA levels of these DEGs were validated in cellular models, confirming consistency with the sequencing results. In addition, the mRNA expression levels of these candidate genes (mapk1, il1b, rela, nfkbiab, akt3a, hyou1, hsp90b1, dnajc3a et al.) in the head kidney, kidney, liver and spleen of grass carp immune tissue were significantly different from those of the control group by BF/C2 (A, B) protein injection in vivo. These candidate genes play an important role in the response of BF/C2 (A, B) to GCRV infection and it also further confirmed that BF/C2 (A, B) of grass carp plays an important role in coping with GCRV infection.

Rice black-streaked dwarf disease caused by Rice black-streaked dwarf virus (RBSDV) is one of the most destructive viral diseases of rice. Thus, it is imperative that resistant rice germplasms are screened for novel RBSDV-resistant genes. RBSDV resistance of a diverse global collection comprising 1,953 rice accessions was evaluated under natural conditions across 3 years. The average disease incidences of the Xian/indica (XI) subgroup were significantly lower than those of the Geng/japonica (GJ) subgroup. Interestingly, most XI-1A accessions in the Xian subgroup were significantly more susceptible than XI-1B accessions, even though XI-1A and XI-1B have a close phylogenetic relationship. Four Xian accessions stably and highly resistant to RBSDV were consistently identified in 2 years. Ten genomic regions (GRs) with 147 single nucleotide polymorphisms associated with RBSDV resistance were detected by a single-locus genome-wide association study (GWAS), of which five were repeatedly identified in a multilocus GWAS. Two previously reported GRs, grRBSDV-6.1 and grRBSDV-6.3, which were repeatedly detected as stably and highly associated with RBSDV resistance, contained 17 and seven genes, respectively, with significant differences of resistance among haplotypes. Haplotype analyses of the candidate genes LOC_Os06g03150 in grRBSDV-6.1 and LOC_Os06g31190 in grRBSDV-6.3 suggested that the former gene is mainly associated with the differentiation of resistance within the Xian subgroup and the latter gene mainly explains the difference in the resistance between Xian and Geng. Another three novel resistance GRs (grRBSDV-1.1, grRBSDV-7.1, and grRBSDV-9.1) were identified. Our findings may enhance the application of disease-resistant rice germplasms for breeding RBSDV-resistant varieties.

Grass carp reovirus II (GCRV II) causes severe hemorrhagic disease with high mortality in grass carp, Cyenopharyngodon idellus. DNA vaccination has been proven to be a very effective method in conferring protection against fish viruses. However, DNA vaccines for GCRV II have not yet been conducted on grass carp. In the current work, we vaccinated grass carp with a DNA vaccine consisting of the segment 6 (pC-S6; encoding VP4) or 10 (pC-S10; encoding NS38) of GCRV II and comparatively analyzed the immune responses induced by these two vaccines. The protective efficacy of pC-S6 and pC-S10, in terms of relative percentage survival (RPS), was 59.9% and 23.1% respectively. This suggests that pC-S6 and pC-S10 DNA vaccines could increase the survival rate of grass carp against GCRV, albeit with variations in immunoprotective effect. Immunological analyses indicated the following. First, post-vaccination (pv), both pC-S6 and pC-S10 up-regulated the expression of interferon (IFN-1), Mx1, IL-1β, and TNF-α. However, CD4 and CD8α were up-regulated in the case of pC-S6 but not pC-S10. Second, comparing non-vaccinated and pC-S10-vaccinated fish, the T cell response related genes, such as CD4, CD8α, and GATA3, were elevated in pC-S6-vaccinated fish at 48 h post-challenge (pc). Third, pC-S6 and pC-S10 induced similar patterns of specific antibody response pv. However, only anti-VP4 IgM in the sera of surviving fish infected with GCRV was significantly increased pc compared with that pre-challenge. Taken together, these results indicate that pC-S6 promotes both innate (IFN-1 and Mx1 induction) and adaptive (T cell and specific antibody response) immunity pv and that the induction of a memory state promptly primes the immune response upon later encounters with the virus, whereas pC-S10 only induces the type I IFN-related response pv and a lower inflammatory response pc.

Oncolytic virotherapy is a novel treatment involving replication-competent virus in the elimination of cancer. We have previously reported the oncolytic effects of reovirus in various canine cancer cell lines. This study aims to establish the safety profile of reovirus in dogs with spontaneously occurring tumours and to determine a recommended dosing regimen. Nineteen dogs with various tumours, mostly of advanced stages, were treated with reovirus, ranging from 1.0 × 108 to 5.0 × 109 TCID50 given as intratumour injection (IT) or intravenous infusion (IV) daily for up to 5 consecutive days in 1 or multiple treatment cycles. Adverse events (AEs) were graded according to the Veterinary Cooperative Oncology Group- Common Terminology Criteria for Adverse Events (VCOG-CTCAE) v1.1 guidelines. Viral shedding, neutralizing anti-reovirus antibody (NARA) production and immunohistochemical (IHC) detection of reovirus protein in the tumours were also assessed. AE was not observed in most dogs and events were limited to Grade I or II fever, vomiting, diarrhoea and inflammation of the injected tumour. No infectious virus was shed and all dogs had elevated NARA levels post-treatment. Although IHC results were only available in 6 dogs, 4 were detected positive for reovirus protein. In conclusion, reovirus is well-tolerated and can be given safely to tumour-bearing dogs according to the dosing regimen used in this study without significant concerns of viral shedding. Reovirus is also potentially effective in various types of canine tumours.

To assess whether the addition of oncolytic reovirus (Reolysin®) to weekly paclitaxel prolonged progression-free survival (PFS) in the treatment of women with recurrent or persistent ovarian, tubal or primary peritoneal cancer.
Patients with recurrent or persistent epithelial ovarian, tubal, or peritoneal carcinoma, measurable or detectable disease, and three or fewer prior regimens were randomly assigned to paclitaxel (80mg/m2 intravenously days 1, 8, and 15 every 4weeks) or the combination of paclitaxel (80mg/m2 intravenously days 1, 8, and 15) plus reovirus 3×1010TCID50/day intravenously on days 1-5, both every 4weeks until disease progression or toxicity. The primary end point was PFS. The study was designed with 80% power for a one-sided alternative at a 10% level of significance to detect a reduction in the hazard by 37.5%.
The study accrued 108 patients, 100 of whom were evaluable for toxicity. Median PFS was 4.3months for paclitaxel and 4.4months for paclitaxel plus reovirus (hazard ratio, 1.11; 90% two-sided CI, 0.78 to 1.59; one-sided P=0.687). The proportion responding (overall response rate) to paclitaxel was 20% among 45 patients with measurable disease receiving paclitaxel alone, and 17.4% among the 46 patients treated with the combination. The asymptotic relative probability of responding was 0.87 (90% CI, 0.42 to 1.79). Severe adverse events were more common in the combination regimen than in paclitaxel arm for severe neutropenia (grade≥4, 12% versus 0%), and severe respiratory adverse events (grade≥3, 25% versus 2%). No deaths were considered treatment related.
The addition of reovirus to weekly paclitaxel in the treatment of women with recurrent or persistent ovarian, tubal or peritoneal cancer did not sufficiently reduce the hazard of progression or death to warrant further investigation.

Arctic charr (Salvelinus alpinus) are the northernmost distributed freshwater fish and can grow at water temperatures as low as 0.2 °C. Other teleost species have impaired immune function at temperatures that Arctic charr thrive in, and thus, charr may maintain immune function at these temperatures. In this study, a fibroblastic cell line, named ACBA, derived from the bulbus arteriosus (BA) of Arctic charr was developed for use in immune studies at various temperatures. ACBA has undergone more than forty passages at 18 °C over 3 years, while showing no signs of senescence-associated β-galactosidase activity and producing nitric oxide. Remarkably, ACBA cells survived and maintained some mitotic activity even at 1 °C for over 3 months. At these low temperatures, ACBA also continued to produce MH class I proteins. After challenge with poly I:C, only antiviral Mx proteins were induced while MH proteins remained constant. When exposed to live viruses, ACBA was shown to permit viral infection and replication of IPNV, VHSV IVa and CSV at 14 °C. Yet at the preferred temperature of 4 °C, only VHSV IVa was shown to replicate within ACBA. This study provides evidence that Arctic charr cells can maintain immune function while also resisting infection with intracellular pathogens at low temperatures.

Naturally occurring reoviruses are live replication-proficient viruses specifically infecting human cancer cells while sparing the normal counterparts. Stem cells can be highly susceptible to viral infection due to their innate high proliferation potential and other active signaling pathways of cells that might be involved in viral tropism. In the previous study, we showed that reoviruses could adversely affect murine embryonic stem cells\' integrity in vitro and in vivo. Oncolytic viruses, delivered systemically face many hurdles that also impede their localization and infection of, metastatic tumors, due to a variety of immune and physical barriers. To overcome such hurdles to systemic delivery, several studies supported the idea that certain types of cells, including mesenchymal stem cells, might play a role as cell carriers for oncolytic viruses. Thus, it would be interesting to examine whether human adult stem cells such as human adipose-derived mesenchymal stem cells could be saved by the reoviral challenge. In this study, we report that biological activities such as proliferation and multipotency of human adipose-derived stem cells are not affected by wild-type reovirus challenge as evidenced by survival, osteogenic and adipogenic differentiation potential assays following treatment with reoviruses. Therefore, unlike murine embryonic stem cells, our study strongly suggests that human adipose-derived adult stem cells could be spared in vivo during wild-type reoviral anti-cancer therapeutics in a clinical setting. Furthermore, the results support the possible clinical use of human adipose-derived stem cells as an effective cell carrier of oncolytic reovirus to maximize their tumor tropism and anti-tumor activity.

To study the efficacy and safety of Saccharomyces boulardii (SB) in acute childhood rotavirus diarrhea.
Children (3 months to 5 years) with WHO-defined acute watery diarrhea and stool rotavirus positive (n  =  60) were randomized into intervention (n  =  30) and control (n  =  30) groups. The intervention group received SB (500 mg/day) for 5 days.
The median duration (hours) of diarrhea was significantly shorter in the intervention group (60 vs. 89; 95% CI: -41.2 to - 16.8). A significantly shorter duration of hospitalization (74 vs. 91; 95% CI: -33.46 to - 0.54) was also seen in the intervention group, but no significant difference was seen for fever and vomiting. There was also no difference between the two groups in the proportion of children requiring parenteral rehydration and persistence of diarrhea lasting beyond day 7. There was no report of any adverse events.
The present trial showed that SB is effective and safe in acute rotavirus diarrhea.

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Antheraea mylitta cytoplasmic polyhedrosis virus (AmCPV) contains 11 double stranded RNA genome segments and infects tasar silkworm A. mylitta. RNA-dependent RNA polymerase (RdRp) is reported as a key enzyme responsible for propagation of the virus in the host cell but its structure function relationship still remains elusive. Here a computational approach has been taken to compare sequence and secondary structure of AmCPV RdRp with other viral RdRps to identify consensus motifs. Then a reliable pairwise sequence alignment of AmCPV RdRp with its closest sequence structure homologue λ3 RdRp is done to predict three dimensional structure of AmCPV RdRp. After comparing with other structurally known viral RdRps, important sequence and/or structural features involved in substrate entry or binding, polymerase reaction and the product release events have been identified. A conserved RNA pentanucleotide (5\'-AGAGC-3\') at the 3\'-end of virus genome is predicted as cis-acting signal for RNA synthesis and its docking and simulation study along with the model of AmCPV RdRp has allowed to predict mode of template binding by the viral polymerase. It is found that template RNA enters into the catalytic center through nine sequence-independent and two sequence-dependent interactions with the specific amino acid residues. However, number of sequence dependent interactions remains almost same during 10 nano-second simulation time while total number of interactions decreases. Further, docking of N(7)-methyl-GpppG (mRNA cap) on the model as well as prediction of RNA secondary structure has shown the template entry process in the active site. These findings have led to postulate the mechanism of RNA-dependent RNA polymerization process by AmCPV RdRp. To our knowledge, this is the first report to evaluate structure function relationship of a cypoviral RdRp.