niryuu/sni-each-converted huggingface dataset:
The IFITM3 polymorphism rs12252-C, which encodes an IFITM3 isoform (D21 IFITM3) lacking 21 amino acids at the amino terminus, has been controversially associated with poor clinical outcomes in patients with H1N1 influenza A virus (IAV) infections. In vitro studies have shown that D21 IFITM3 loses its ability to restrict H1N1 IAV. Subsequent research has also revealed that tyrosine 20 is the key determinant for IFITM3 endocytic trafficking, which is essential for the efficient anti-viral activity of IFITM3. In contrast to previous studies, we demonstrated that both D21 IFITM3 and an IFITM3 variant (Y20A IFITM3), in which tyrosine 20 is substituted with alanine, strongly restricted entry mediated by IAV H1, H3, H5, and H7 proteins. D21 IFITM3 also efficiently suppressed replication of H1N1 and, to a lesser extent, H3N2 IAV. D21 IFITM3 and Y20A IFITM3 had broader subcellular distributions than full-length IFITM3 but an abundant amount of both IFITM3 variants still localized to late endosomes and lysosomes. Our data indicate that tyrosine 20 partially regulates the subcellular localization of IFITM3 but is not functionally essential for IFITM3-mediated IAV restriction. They also suggested that mechanisms, other than viral entry restriction, might contribute to variations in clinical outcomes of H1N1 influenza associated with rs12252-C.
Output: IFITM3 Polymorphism rs12252-C Restricts Influenza A Viruses
The baculovirus group of insect viruses is widely used for foreign gene introduction into mammalian cells for gene expression and protein production; however, the efficiency of baculovirus entry into mammalian cells is in general still low. In this study, two recombinant baculoviruses were engineered and their ability to improve viral entry was examined: (1) cytoplasmic transduction peptide (CTP) was fused with baculovirus envelope protein, GP64, to produce a cytoplasmic membrane penetrating baculovirus (vE-CTP); and (2) the protein transduction domain (PTD) of HIV TAT protein was fused with the baculovirus capsid protein VP39 to form a nuclear membrane penetrating baculovirus (vE-PTD). Transduction experiments showed that both viruses had better transduction efficiency than vE, a control virus that only expresses EGFP in mammalian cells. Interestingly, vE-CTP and vE-PTD were also able to improve the transduction efficiency of a co-transduced baculovirus, resulting in higher levels of gene expression. Our results have described new routes to further enhance the development of baculovirus as a tool for gene delivery into mammalian cells.
Output: Membrane penetrating peptides greatly enhance baculovirus transduction efficiency into mammalian cells
Background: Influenza is a severe contagious disease especially in children, elderly and immunocompromised patients. Beside vaccination, the discovery of new anti-viral agents represents an important strategy to encounter seasonal and pandemic influenza A virus (IAV) strains. The bacterial extra-cellular ribonuclease binase is a well-studied RNase from Bacillus pumilus. Treatment with binase was shown to improve survival of laboratory animals infected with different RNA viruses. Although binase reduced IAV titer in vitro and in vivo, the mode of action (MOA) of binase against IAV at the molecular level has yet not been studied in depth and remains elusive. Methods: To analyze whether binase impairs virus replication by direct interaction with the viral particle we applied a hemagglutination inhibition assay and monitored the integrity of the viral RNA within the virus particle by RT-PCR. Furthermore, we used Western blot and confocal microscopy analysis to study whether binase can internalize into MDCK-II cells. By primer extension we examined the effect of binase on the integrity of viral RNAs within the cells and using a mini-genome system we explored the effect of binase on the viral expression. Results: We show that (i) binase does not to attack IAV particle-protected viral RNA, (ii) internalized binase could be detected within the cytosol of MDCK-II cells and that (iii) binase impairs IAV replication by specifically degrading viral RNA species within the infected MDCK-II cells without obvious effect on cellular mRNAs. Conclusion: Our data provide novel evidence suggesting that binase is a potential anti-viral agent with specific intra-cellular MOA.
Output: Bacterial ribonuclease binase exerts an intra-cellular anti-viral mode of action targeting viral RNAs in influenza a virus- infected MDCK-II cells
The Collaborative Cross (CC) is a multiparent panel of recombinant inbred (RI) mouse strains derived from eight founder laboratory strains. RI panels are popular because of their long-term genetic stability, which enhances reproducibility and integration of data collected across time and conditions. Characterization of their genomes can be a community effort, reducing the burden on individual users. Here we present the genomes of the CC strains using two complementary approaches as a resource to improve power and interpretation of genetic experiments. Our study also provides a cautionary tale regarding the limitations imposed by such basic biological processes as mutation and selection. A distinct advantage of inbred panels is that genotyping only needs to be performed on the panel, not on each individual mouse. The initial CC genome data were haplotype reconstructions based on dense genotyping of the most recent common ancestors (MRCAs) of each strain followed by imputation from the genome sequence of the corresponding founder inbred strain. The MRCA resource captured segregating regions in strains that were not fully inbred, but it had limited resolution in the transition regions between founder haplotypes, and there was uncertainty about founder assignment in regions of limited diversity. Here we report the whole genome sequence of 69 CC strains generated by paired-end short reads at 303 coverage of a single male per strain. Sequencing leads to a substantial improvement in the fine structure and completeness of the genomes of the CC. Both MRCAs and sequenced samples show a significant reduction in the genome-wide haplotype frequencies from two wild-derived strains, CAST/EiJ and PWK/PhJ. In addition, analysis of the evolution of the patterns of heterozygosity indicates that selection against three wild-derived founder strains played a significant role in shaping the genomes of the CC. The sequencing resource provides the first description of tens of thousands of new genetic variants introduced by mutation and drift in the CC genomes. We estimate that new SNP mutations are accumulating in each CC strain at a rate of 2.4 6 0.4 per gigabase per generation. The fixation of new mutations by genetic drift has introduced thousands of new variants into the CC strains. The majority of these mutations are novel compared to currently sequenced laboratory stocks and wild mice, and some are predicted to alter gene function. Approximately one-third of the CC inbred strains have acquired large deletions (.10 kb) many of which overlap known coding genes and functional elements. The sequence of these mice is a critical resource to CC users, increases threefold the number of mouse inbred strain genomes available publicly, and provides insight into the effect of mutation and drift on common resources.
Output: Genomes of the Mouse Collaborative Cross
In memory of Federica Novelli. 1-Aryl-4,6-diamino-1,2-dihydrotriazine derivatives Anti-influenza A and B viruses activity Anti-RSV activity Host (human) DHFR inhibition a b s t r a c t We have identified a series of 1-aryl-4,6-diamino-1,2-dihydrotriazines, structurally related to the antimalarial drug cycloguanil, as new inhibitors of influenza A and B virus and respiratory syncytial virus (RSV) via targeting of the host dihydrofolate reductase (DHFR) enzyme. Most analogues proved active against influenza B virus in the low micromolar range, and the best compounds (11, 13, 14 and 16) even reached the sub-micromolar potency of zanamivir (EC 50 ¼ 0.060 mM), and markedly exceeded (up to 327 times) the antiviral efficacy of ribavirin. Activity was also observed for two influenza A strains, including a virus with the S31N mutant form of M2 proton channel, which is the most prevalent resistance mutation for amantadine. Importantly, the compounds displayed nanomolar activity against RSV and a superior selectivity index, since the ratio of cytotoxic to antiviral concentration was >10,000 for the three most active compounds 11, 14 and 16 (EC 50~0 .008 mM), far surpassing the potency and safety profile of the licensed drug ribavirin (EC 50 ¼ 5.8 mM, SI > 43).
Output: Host dihydrofolate reductase (DHFR)-directed cycloguanil analogues endowed with activity against influenza virus and respiratory syncytial virus
Patients undergoing solid organ transplantation (SOT) may acquire infections from the transplanted organ. Routine screening for common infections are an established part of the pretransplant evaluation of donors and recipients. Likewise, strategies exist for prophylaxis and surveillance for common donorassociated infections including hepatitis B, CMV and EBV. However, despite advances in diagnostic testing to evaluate the infectious risk of donors, unanticipated transmission of pathogens occurs, particularly when donors are asymptomatic or have subtle or unusual manifestations of a transmissible Infection. Infectious diseases (ID) providers play an integral role in donor and recipient risk assessment and can advise transplant centers on organ utilization and guide evaluation and management of the SOT recipient. Consideration of the donor cause of death and preceding clinical syndromes are important for characterizing the potential risk for recipient infection. This allows a more accurate analysis of the risk: benefit of accepting a life-saving organ and risk of infection. ID providers and transplant teams should work closely with organ procurement organizations (OPOs) to solicit additional donor information when a donor-derived infection is suspected so that reporting can be facilitated to ensure communication with the care-teams of other organ recipients from the same donors. National advisory committees work closely with federal agencies to provide oversight, guide policy development, and assess outcomes to assist with the prevention and management of donor-transmitted disease through organ transplantation. The clinical vignettes in this review highlight some of the complexities in the evaluation of potential donor transmission.
Output: Journal of the Pediatric Infectious Diseases Society Clinical Vignettes: Donor-Derived Infections
Influenza A virus RNA replication requires an intricate regulatory network involving viral and cellular proteins. In this study, we examined the roles of cellular ubiquitinating/deubiquitinating enzymes (DUBs). We observed that downregulation of a cellular deubiquitinating enzyme USP11 resulted in enhanced virus production, suggesting that USP11 could inhibit influenza virus replication. Conversely, overexpression of USP11 specifically inhibited viral genomic RNA replication, and this inhibition required the deubiquitinase activity. Furthermore, we showed that USP11 interacted with PB2, PA, and NP of viral RNA replication complex, and that NP is a monoubiquitinated protein and can be deubiquitinated by USP11 in vivo. Finally, we identified K184 as the ubiquitination site on NP and this residue is crucial for virus RNA replication. We propose that ubiquitination/deubiquitination of NP can be manipulated for antiviral therapeutic purposes. The EMBO Journal VOL 29 | NO 22 | 2010 EMBO THE EMBO JOURNAL THE EMBO JOURNAL USP11 and influenza virus RNA replication T-L Liao et al
Output: Ubiquitination and deubiquitination of NP protein regulates influenza A virus RNA replication
Several mammalian arenaviruses (mammarenaviruses) cause hemorrhagic fevers in humans and pose serious public health concerns in their endemic regions. Additionally, mounting evidence indicates that the worldwide-distributed, prototypic mammarenavirus, lymphocytic choriomeningitis virus (LCMV), is a neglected human pathogen of clinical significance. Concerns about human-pathogenic mammarenaviruses are exacerbated by of the lack of licensed vaccines, and current anti-mammarenavirus therapy is limited to off-label use of ribavirin that is only partially effective. Detailed understanding of virus/host-cell interactions may facilitate the development of novel anti-mammarenavirus strategies by targeting components of the host-cell machinery that are required for efficient virus multiplication. Here we document the generation of a recombinant LCMV encoding a nucleoprotein (NP) containing an affinity tag (rLCMV/Strep-NP) and its use to capture the NP-interactome in infected cells. Our proteomic approach combined with genetics and pharmacological validation assays identified ATPase Na + /K + transporting subunit alpha 1 (ATP1A1) and prohibitin (PHB) as pro-viral factors. Cell-based assays revealed that ATP1A1 and PHB are involved in different steps of the virus life cycle. Accordingly, we observed a synergistic inhibitory effect on LCMV multiplication with a combination of ATP1A1 and PHB inhibitors. We show that ATP1A1 inhibitors suppress multiplication of Lassa virus and Candid#1, a live-attenuated vaccine strain of Junín virus, suggesting that the requirement of ATP1A1 in virus multiplication is conserved among genetically distantly related mammarenaviruses. Our findings suggest that clinically approved inhibitors of ATP1A1, like digoxin, could be repurposed to treat infections by mammarenaviruses pathogenic for humans.
Output: Interactome analysis of the lymphocytic choriomeningitis virus nucleoprotein in infected cells reveals ATPase Na + /K + transporting subunit Alpha 1 and prohibitin as host-cell factors involved in the life cycle of mammarenaviruses
As a city famous for tourism, the public healthcare system of Macau SAR has been under great pressure during the outbreak of the Coronavirus Disease 2019 . In this study, we report clinical and microbiological features of ten COVID-19 patients enrolled in the Centro Hospitalar Conde de São Januário (CHCSJ) between January 21 to February 16, 2020. Clinical samples from all patients including nasopharyngeal swab (NPS)/sputum, urine, and feces were collected for serial virus RNA testing by standard qRT-PCR assay. In total, seven were imported cases and three were local cases. The median duration from Macau arrival to admission in imported cases was 3 days. Four patients required oxygen therapy but none of them needed machinal ventilation. No fatal cases were noted. The most common symptoms were fever (80%) and diarrhea (80%). In the "Severe" group, there was significantly more elderly patients (p=0.045), higher lactate dehydrogenase levels (p=0.002), and elevated C-Reactive protein levels compared to the "Mild to Moderate" group (p<0.001). There were positive SARS-CoV-2 RNA signals in all patients' NPS and stool specimens but negative in all urine specimens. Based on our data on SARS-CoV-2 RNA shedding in stool and the possibility of a lag in viral detection in NPS specimens, the assessment of both fecal and respiratory specimen is recommended to enhance diagnostic sensitivity, and also to aid discharge decision before the role of viral RNA shedding in stool is clarified.
Output: Evaluation of SARS-CoV-2 RNA shedding in clinical specimens and clinical characteristics of 10 patients with COVID-19 in Macau
In order to estimate the diversity, clinical involvement and zoonotic potential of parasites in pigs submitted for diagnosis to the PathoPig project of the Swiss Federal Food Safety and Veterinary Office, faeces (n = 125) from suckling piglets (n = 39), weaners (n = 60) and piglets beginning fattening (n = 26) from 74 Swiss farms were examined by 3 coproscopical methods (i.e. sedimentation/zinc chloride-flotation; SAFC and Ziehl-Neelsen staining). Samples microscopically positive for Cryptosporidium were further tested by PCR/sequencing for species assessment. The most frequently detected parasite was Balantidium coli, a facultative pathogenic ciliate with zoonotic potential, in 5.1, 36.7 and 50.0% of suckling, weaners and fatteners and 43.2% of farms; however, no association with disease was observed. Isospora (syn. Cystoisospora) suis infections were detected in 13.3 and 11.1% of suckling piglets with and without diarrhoea, and in 10.0 and 13.3% of weaners and fatteners with diarrhoea, respectively, and were significant associated with emaciation. Cryptosporidium infections were detected in 10.3, 15.0 and 19.2% of sucklings, weaners and fatteners, respectively, and in 18.9% of the farms. Interestingly, two age-related species were identified: C. suis in younger piglets (2 to 6 weeks) and C. scrofarum in older ones (6 to 17 weeks). None of the pigs infected with C. scrofarum (n = 8), but 3 of 4 piglets infected with C. suis (coinfection with I. suis in 2 cases) had diarrhoea. The zoonotic species C. parvum was not detected, nevertheless, sporadic cases of human infection with the porcine-adapted species have been reported. Ascaris suum, Trichuris suis and Strongylida were rarely detected (b 4%) in all age categories.
Output: Occurrence, clinical involvement and zoonotic potential of endoparasites infecting Swiss pigs
Coronavirus (CoV) envelope (E) protein ion channel activity was determined in channels formed in planar lipid bilayers by peptides representing either the transmembrane domain of severe acute respiratory syndrome CoV (SARS-CoV) E protein, or the full-length E protein. Both of them formed a voltage independent ion conductive pore with symmetric ion transport properties. Mutations N15A and V25F located in the transmembrane domain, prevented the ion conductivity. E protein derived channels showed no cation preference in non-charged lipid membranes, whereas they behaved as pores with mild cation selectivity in negatively-charged lipid membranes. The ion conductance was also controlled by the lipid composition of the membrane. Lipid charge also regulated the selectivity of a HCoV-229E E protein derived peptide. These results suggested that the lipids are functionally involved in E protein ion channel activity, forming a protein-lipid pore, a novel concept for CoV E protein ion channel entity.
Output: CORONAVIRUS E PROTEIN FORMS ION CHANNELS WITH FUNCTIONALLY AND STRUCTURALLY-INVOLVED MEMBRANE LIPIDS
In the older adult the benefits of vaccination to prevent infectious disease are limited, mainly due to the adaptive immune system's inability to generate protective immunity. Age-dependent decline in immune competence, often referred to as immunosenescence, results from progressive deterioration of innate and adaptive immune responses and most of the insights into mechanisms of immune aging have derived from studies in murine models. In this Review, we explore how well these models are applicable to understand the aging process throughout the 80-100 years of human life and discuss recent advances in uncovering and characterizing the mechanisms underlying age-associated defective adaptive immunity in humans.
Output: Understanding immune senescence to improve vaccine responses
Zinc may be a new risk factor for pneumonia in the elderly. In this special article, we reviewed the magnitude of the problem of pneumonia (its prevalence, morbidity and mortality) in the elderly, its etiology, and the dysregulation of the immune system associated with increasing age. In addition, we presented evidence from the literature, including work we did recently, that low zinc status (commonly reported in the elderly) impairs immune function, decreases resistance to pathogens, and is associated not only with increased incidence and duration of pneumonia, increased use and duration of antimicrobial treatment, but also with increased overall mortality in the elderly. Inadequate stores of zinc might therefore be a risk factor for pneumonia in the elderly. Randomized, double blind, controlled studies are needed to determine the efficacy of zinc supplementation as a potential low cost intervention to reduce morbidity and mortality due to pneumonia in this vulnerable population. Please provide between three and five key words or phrases
Output: Zinc: a new risk factor for pneumonia in the elderly? NIH Public Access
Antibodies in human serum against an enteropathogenic bovine coronavirus were detected by double immunodiffusion (DID), neutralization of infectivity, indirect immunofluorescence, and immune electron microscopy. Human sera reacting in the DID test neutralized the infectivity of the bovine coronaviruses to indices of 2.5 to ~> 5. Nineteen of 40 DID-negative, heat-inactivated sera had neutralizing indices of 1 to 3.0. Human serum with neutralizing and DID antibodies produced juxtanuclear and cytoplasmic fluorescence identical to that of bovine immune serum in cells infected with the bovine coronavirus. Antibodies in human and bovine sera interacted with the peplomeres of the bovine coronavirus, matting and bridging them, when present in excess, and facilitated formation of large viral aggregates when present in equivalent concentrations. Complement added to the virus-antibody complexes did not alter specifically the morphology of single, antibody-laden viral particles or viral particles in aggregates. Evidence of the transmission of coronavirus from experimentally inoculated calves to man, with ensuing gastroenteritis, was found by electron microscopic tracing of the coronavirus and its virus-antibody complexes.
Output: Medical Microbiology and Immunology Reactivity of Antibodies in Human Serum with Antigens of an Enteropathogenic Bovine Coronavirus
The descriptive distribution and phylogeny of feline coronaviruses (FCoVs) were studied in cats suspected of having feline infectious peritonitis (FIP) in Malaysia. Ascitic fluids and/or biopsy samples were subjected to a reverse transcription polymerase chain reaction (RT-PCR) targeted for a conserved region of 3'untranslated region (3'UTR) of the FCoV genome. Eighty nine percent of the sampled animals were positive for the presence of FCoV. Among the FCoV positive cats, 80% of cats were males and 64% were below 2 years of age. The FCoV positive cases included 56% domestic short hair (DSH), 40% Persian, and 4% Siamese cats. The nucleotide sequences of 10 selected amplified products from FIP cases were determined. The sequence comparison revealed that the field isolates had 96% homology with a few point mutations. The extent of homology decreased to 93% when compared with reference strains. The overall branching pattern of phylogenetic tree showed two distinct clusters, where all Malaysian isolates fall into one main genetic cluster. These findings provided the first genetic information of FCoV in Malaysia.
Output: Descriptive distribution and phylogenetic analysis of feline infectious peritonitis virus isolates of Malaysia
Background: Middle East respiratory syndrome coronavirus (MERS-CoV) consists of a positive-sense, single-stranded RNA genome and four structural proteins: the spike, envelope, membrane, and nucleocapsid protein. The assembly of the viral genome into virus particles involves viral structural proteins and is believed to be mediated through recognition of specific sequences and RNA structures of the viral genome. Methods and Results: A culture system for the production of MERS coronavirus-like particles (MERS VLPs) was determined and established by electron microscopy and the detection of coexpressed viral structural proteins. Using the VLP system, a 258-nucleotide RNA fragment, which spans nucleotides 19,712 to 19,969 of the MERS-CoV genome (designated PS258(19712-19969) ME ), was identified to function as a packaging signal. Assembly of the RNA packaging signal into MERS VLPs is dependent on the viral nucleocapsid protein. In addition, a 45-nucleotide stable stem-loop substructure of the PS258(19712-19969) ME interacted with both the N-terminal domain and the Cterminal domain of the viral nucleocapsid protein. Furthermore, a functional SARS-CoV RNA packaging signal failed to assemble into the MERS VLPs, which indicated virus-specific assembly of the RNA genome. Conclusions: A MERS-oV RNA packaging signal was identified by the detection of GFP expression following an incubation of MERS VLPs carrying the heterologous mRNA GFP-PS258(19712-19969) ME with virus permissive Huh7 cells. The MERS VLP system could help us in understanding virus infection and morphogenesis.
Output: Nucleocapsid protein-dependent assembly of the RNA packaging signal of Middle East respiratory syndrome coronavirus
This paper, based on relevant literature articles and the authors' clinical experience, presents a goal-oriented respiratory management for critically ill patients with acute respiratory distress syndrome (ARDS) that can help improve clinicians' ability to care for these patients. Early recognition of ARDS modified risk factors and avoidance of aggravating factors during hospital stay such as nonprotective mechanical ventilation, multiple blood products transfusions, positive fluid balance, ventilator-associated pneumonia, and gastric aspiration can help decrease its incidence. An early extensive clinical, laboratory, and imaging evaluation of "at risk patients" allows a correct diagnosis of ARDS, assessment of comorbidities, and calculation of prognostic indices, so that a careful treatment can be planned. Rapid administration of antibiotics and resuscitative measures in case of sepsis and septic shock associated with protective ventilatory strategies and early short-term paralysis associated with differential ventilatory techniques (recruitment maneuvers with adequate positive end-expiratory pressure titration, prone position, and new extracorporeal membrane oxygenation techniques) in severe ARDS can help improve its prognosis. Revaluation of ARDS patients on the third day of evolution (Sequential Organ Failure Assessment (SOFA), biomarkers and response to infection therapy) allows changes in the initial treatment plans and can help decrease ARDS mortality.
Output: Goal-Oriented Respiratory Management for Critically Ill Patients with Acute Respiratory Distress Syndrome
Duchenne muscular dystrophy (DMD) is an X-linked disorder characterized by progressive muscle degeneration, caused by the absence of dystrophin. Exon skipping by antisense oligonucleotides (ASOs) has recently gained recognition as therapeutic approach in DMD. Conjugation of a peptide to the phosphorodiamidate morpholino backbone (PMO) of ASOs generated the peptide-conjugated PMOs (PPMOs) that exhibit a dramatically improved pharmacokinetic profile. When tested in animal models, PPMOs demonstrate effective exon skipping in target muscles and prolonged duration of dystrophin restoration after a treatment regime. Herein we summarize the main pathophysiological features of DMD and the emergence of PPMOs as promising exon skipping agents aiming to rescue defective gene expression in DMD and other neuromuscular diseases. The listed PPMO laboratory findings correspond to latest trends in the field and highlight the obstacles that must be overcome prior to translating the animal-based research into clinical trials tailored to the needs of patients suffering from neuromuscular diseases.
Output: Peptide-conjugate antisense based splice-correction for Duchenne muscular dystrophy and other neuromuscular diseases
Pathological characterization of autopsied tissues from patients with SARS revealed severe damage in restricted tissues, such as lung, with no apparent cell damage in other tissues, such as intestine and brain. Here, we examined the susceptibility of neural cell lines of human (OL) and rat (C6) origins to SARS-associated coronavirus. Both of the neural cell lines showed no apparent cytopathic effects (CPE) by infection but produced virus with infectivity of 10 2-5 per ml, in sharp contrast to the production by infected Vero E6 cells of >10 9 per ml that showed a lytic infection with characteristic rounding CPE. Interestingly, the infection of intestinal cell line CaCo-2 also induced no apparent CPE, with production of the virus at a slightly lower level as that of the Vero E6 cell culture. Notably, the cellular receptor for the virus, angiotensin-converting enzyme 2 was expressed at similar levels on Vero E6 and CaCo-2 cells, but at undetectable levels on OL and C6 cells.
Output: Susceptibility of human and rat neural cell lines to infection by SARS-coronavirus
Real-time quantitative reverse transcriptase polymerase chain reaction (rRT-PCR) is now widely used to detect viral pathogens in various human specimens. The application of internal controls to validate the entire process of these assays is necessary to prevent false-negative results caused by unexpected inhibition or inefficient extraction. In the present study, we describe a strategy to produce a stable internal control for rRT-PCR by packaging foreign RNA into influenza virions using plasmid-based reverse genetics technology. The envelope structure of influenza virus can effectively protect RNA segments from RNase digestion, which provides an advantage for its routine use as an internal control. Utilizing this approach, we successfully generated a recombinant influenza virus (rPR8-HCV) containing the 5′ untranslated region (5′ UTR) of the hepatitis C virus (HCV) RNA genome. After inactivation and purification, the rPR8-HCV particles were demonstrated to be RNase resistant and stable at 4°C for at least 252 days in human plasma, with no degradation even after being frozen and thawed multiple times. These results were reproducible in the COBAS TaqMan HCV test for 164 days. Moreover, the chimeric influenza virus particles could be easily produced in embryonated eggs and were noninfectious after inactivation treatment. Additionally, this strategy could also be adapted for real-time clinical applications of other RNA targets, providing a universal approach with broad clinical applications in rRT-PCR assays.
Output: Use of chimeric influenza viruses as a novel internal control for diagnostic rRT-PCR assays
Clinical patient profiles with H5N1 or SARS infection. The two cases of H5N1 infection have been reported previously (Uiprasertkul et al., 2005) . Briefly, the first case was a previously healthy 6year old boy who developed fever and shortness of breath. Despite antibiotic treatment he deteriorated and Chest X-Ray showed right lower lobe opacification. Nasopharyngeal aspirate was positive for influenza A and PCR showed positive result for H5 and negative for H1 and H3. He was started on oseltamivir 2 days before death. The second case was a 33 year old man with previous good health who was admitted with a 4-day history of fever, malaise, sore throat and cough with blood-stained sputum. Chest X-Ray showed right lower-lobe consolidation. Despite treatment with oseltamivir 2 days after admission his condition deteriorated and he developed ARDS. PCR for H5 was positive on day 6 of illness -the day on which he died. The clinical profiles of the nine SARS patients used in our study have been previously reported (Nicholls et al., 2006) . Human lung obtained from a patient who died of extra-pulmonary disease served as a normal control and was kindly provided by Prof. D. Kerjaschki, Department of Pathology, Medical University of Vienna, according to ethical guidelines. For mouse H5N1 infection experiment, female C57BL/6N mice (6-8 week of age) were used. Mice were anesthetized with isoflurane and infected intranasally with live H5N1 avian influenza virus (strain A/HK/483/97) at an equivalent TCID 50 dose of 10 2 and 10 1 . Mice were observed daily and euthanized for tissue sampling on day 4 after challenge. Lung tissues were collected and fixed in 10% formalin. The experiment was conducted in a biosafety level III laboratory. Lethal lung infections with Anthrax, Y. pestis, and monkey pox virus, as well as infections with Smallpox virus, Marburg virus, Western Equine Encephalitis virus (WEE), and Eastern Equine Encephalitis virus (EEE) and lung sampling were performed at the US Army Medical Research Institute of Infectious Diseases, Fort Detrick, according to institutional guidelines. The samples were obtained from USAMRIID's pathology archival collection.
Output: Supplemental Data Identification of Oxidative Stress and Toll-like Receptor 4 Signaling as a Key Pathway of Acute Lung Injury SUPPLEMENTAL EXPERIMENTAL PROCEDURES
The envelope spike (S) proteins of MERS-CoV and SARS-CoV determine the virus host tropism and entry into host cells, and constitute a promising target for the development of prophylactics and therapeutics. Here, we present high-resolution structures of the trimeric MERS-CoV and SARS-CoV S proteins in its pre-fusion conformation by single particle cryo-electron microscopy. The overall structures resemble that from other coronaviruses including HKU1, MHV and NL63 reported recently, with the exception of the receptor binding domain (RBD). We captured two states of the RBD with receptor binding region either buried (lying state) or exposed (standing state), demonstrating an inherently flexible RBD readily recognized by the receptor. Further sequence conservation analysis of six human-infecting coronaviruses revealed that the fusion peptide, HR1 region and the central helix are potential targets for eliciting broadly neutralizing antibodies.
Output: ARTICLE Cryo-EM structures of MERS-CoV and SARS-CoV spike glycoproteins reveal the dynamic receptor binding domains
Porcine epidemic diarrhea virus (PEDV) outbreaks on pig farms have caused significant economic loss in the swine industry since it was first reported in Thailand a decade ago. Anecdotal evidence suggests that PEDV is now endemic in this region, therefore genome information of circulating PEDV is important for molecular surveillance and evaluation of potential benefits of field vaccination. Here, we characterized PEDV infection on commercial Thai swine farms by screening 769 samples of feces and small intestinal contents from pigs with diarrhea between 2011 and 2016. Using reverse-transcription polymerase chain reaction targeting the spike (S) gene, 153 PEDV-positive samples were further subjected to analysis of the open reading frame 3 and nucleocapsid (N) genes. Comparison of 95 samples in which nucleotide sequencing was successfully obtained for all three genes revealed evolutionary diversity among the Thai PEDV strains. Phylogenetic analyses suggest that although some Thai strains changed little from years past, others resembled more closely to the recent strains reported in China. Interestingly, eight Thai PEDV strains possessed amino acid deletions in the N protein. The PEDV sequence divergence may be responsible for driving periodic outbreaks and continued persistence of PEDV on commercial swine farms. Our findings provide important insight into regional PEDV strains in circulation, which may assist future inclusions of suitable strains for future PEDV vaccines.
Output: Analysis of the spike, ORF3, and nucleocapsid genes of porcine epidemic diarrhea virus circulating on Thai swine farms, 2011-2016 Distributed under Creative Commons CC-BY 4.0
The mosquito-borne chikungunya virus (CHIKV) has become a major global health problem. Upon infection, chikungunya fever (CHIKF) can result in long-term joint pain and arthritis, and despite intense research, no licensed vaccine for CHIKV is available. We have developed two recombinant chimpanzee adenovirus-vectored vaccines (ChAdOx1) that induce swift and robust anti-CHIKV immune responses with a single dose, without the need for adjuvants or booster vaccines. Here, we report the vaccines' protective efficacies against CHIKV infection in a lethal A129 mouse model. Our results indicate that a single, un-adjuvanted ChAdOx1 Chik or ChAdOx1 Chik ∆Cap dose provided complete protection against a lethal virus challenge and prevented CHIKV-associated severe inflammation. These candidate vaccines supported survival equal to the attenuated 181/25 CHIKV reference vaccine but without the vaccine-related side effects, such as weight loss. Vaccination with either ChAdOx1 Chik or ChAdOx1 Chik ∆Cap resulted in high titers of neutralizing antibodies that are associated with protection, indicating that the presence of the capsid within the vaccine construct may not be essential to afford protection under the conditions tested. We conclude that both replication-deficient ChAdOx1 Chik vaccines are safe even when used in A129 mice and afford complete protection from a lethal challenge.
Output: A Single and Un-Adjuvanted Dose of a Chimpanzee Adenovirus-Vectored Vaccine against Chikungunya Virus Fully Protects Mice from Lethal Disease
Strong evidences (Table 2) supporting the role of NIPPV in the treatment of different conditions leading to acute or acute-onchronic respiratory failure include four conditions as follows: (a) Prevention of intubation in patients with acute exacerbations of chronic obstructive pulmonary disease (AECOPD) (b) Acute cardiogenic pulmonary oedema (ACPO) ABSTRACT Non-invasive positive pressure ventilation (NIPPV) is the technique of delivering mechanical ventilation without endotracheal intubation or tracheostomy. This is increasingly being utilised in both acute and chronic conditions. Strong evidence supports the use of NIPPV for acute respiratory failure (ARF) to prevent endotracheal intubation (ETI) and to facilitate extubation in patients with acute exacerbations of chronic obstructive pulmonary disease, to avoid ETI in acute cardiogenic pulmonary oedema (ACPO), and in immunocompromised patients. Weaker evidence supports the use of NIPPV for patients with ARF due to asthma exacerbations, with postoperative ARF, pneumonia and acute lung injury/acute respiratory distress syndrome. NIPPV should be applied under close monitoring for signs of treatment failure and, in such cases, ETI should be promptly available. A trained team, at an appropriate location, with careful patient selection and optimal choice of devices can optimise the outcome of NIPPV.
Output: Recent advances in the role of non-invasive ventilation in acute respiratory failure
Enveloped viruses require viral fusion proteins to promote fusion of the viral envelope with a target cell membrane. To drive fusion, these proteins undergo large conformational changes that must occur at the right place and at the right time. Understanding the elements which control the stability of the prefusion state and the initiation of conformational changes is key to understanding the function of these important proteins. The construction of mutations in the fusion protein transmembrane domains (TMDs) or the replacement of these domains with lipid anchors has implicated the TMD in the fusion process. However, the structural and molecular details of the role of the TMD in these fusion events remain unclear. Previously, we demonstrated that isolated paramyxovirus fusion protein TMDs associate in a monomer-trimer equilibrium, using sedimentation equilibrium analytical ultracentrifugation. Using a similar approach, the work presented here indicates that trimeric interactions also occur between the fusion protein TMDs of Ebola virus, influenza virus, severe acute respiratory syndrome coronavirus (SARS CoV), and rabies virus. Our results suggest that TM-TM interactions are important in the fusion protein function of diverse viral families. IMPORTANCE Many important human pathogens are enveloped viruses that utilize membrane-bound glycoproteins to mediate viral entry. Factors that contribute to the stability of these glycoproteins have been identified in the ectodomain of several viral fusion proteins, including residues within the soluble ectodomain. Although it is often thought to simply act as an anchor, the transmembrane domain of viral fusion proteins has been implicated in protein stability and function as well. Here, using a biophysical approach, we demonstrated that the fusion protein transmembrane domains of several deadly pathogens-Ebola virus, influenza virus, SARS CoV, and rabies virus-self-associate. This observation across various viral families suggests that transmembrane domain interactions may be broadly relevant and serve as a new target for therapeutic development. Citation Webb SR, Smith SE, Fried MG, Dutch RE. 2018. Transmembrane domains of highly pathogenic viral fusion proteins exhibit trimeric association in vitro. mSphere 3:e00047-18.
Output: Transmembrane Domains of Highly Pathogenic Viral Fusion Proteins Exhibit Trimeric Association In Vitro
Inflammasomes have emerged as critical innate sensors of host immune that defense against pathogen infection, metabolism syndrome, cellular stress and cancer metastasis in the liver. The assembly of inflammasome activates caspase-1, which promotes the maturation of interleukin-1β (IL-1β) and interleukin-18 (IL-18), and initiates pyroptotic cell death (pyroptosis). IL-18 exerts pleiotropic effects on hepatic NK cells, priming FasL-mediated cytotoxicity, and interferon-γ (IFN-γ)-dependent responses to prevent the development of liver diseases. However, considerable attention has been attracted to the pathogenic role of inflammasomes in various acute and chronic liver diseases, including viral hepatitis, nanoparticle-induced liver injury, alcoholic and non-alcoholic steatohepatitis. In this review, we summarize the latest advances on the physiological and pathological roles of inflammasomes for further development of inflammasome-based therapeutic strategies for human liver diseases.
Output: MINI REVIEW Inflammasome: A Double-Edged Sword in Liver Diseases
Zika virus (ZIKV) infection can be the cause of congenital malformations, including microcephaly in infants and can cause other disorders such as Guillain-Barré syndrome, meningoencephalitis, and myelitis, which can also occur in some infected adults. However, at this time, there is no drug approved to treat ZIKV infection. Drug repurposing is the promptest way to obtain an effective drug during a global public health emergency such as the spread of Zika virus. In this study, we report a US Food and Drug Admistration-approved drug that is safe for pediatric use. Nitazoxanide and its bioactive metabolite, tizoxanide, have anti-ZIKV potential in vitro, and we identified that they exerts antiviral effect possibly by targeting the viral postattachment step.
Output: Pediatric Drug Nitazoxanide: A Potential Choice for Control of Zika
The spectrum of human pathogens and the infectious diseases they cause is continuously changing through evolution and changes in the way human populations interact with their environment and each other. New human pathogens most often emerge from an animal reservoir, emphasizing the central role that non-human reservoirs play in human infectious diseases. Pathogens may also re-emerge with new characteristics, such as multidrug-resistance, or in different places, such as West Nile virus in the USA in 1999, to cause new epidemics. Most human pathogens have a history of evolution in which they first emerge and cause epidemics, become unstably adapted, re-emerge periodically, and eventually become endemic with the potential for future outbreaks.
Output: Emerging infectious diseases
We investigate the effects of social distancing in controlling the impact of the COVID-19 epidemic using a simple susceptible-infected-removed epidemic model. We show that an alternative or complementary approach based on targeted isolation of the vulnerable subpopulation may provide a more efficient and robust strategy at a lower economic and social cost within a shorter timeframe resulting in a collectively immune population. . CC-BY-NC-ND 4.0 International license It is made available under a is the author/funder, who has granted medRxiv a license to display the preprint in perpetuity. The copyright holder for this preprint .
Output: Targeted adaptive isolation strategy for COVID-19 pandemic
We describe the unusual structure of a vaccinia virus late mRNA. In these molecules, the protein-coding sequences of a major late structural polypeptide are preceded by long leader RNAa, which In some cases are thousands of nucleotides long. These sequences map to different regions of the viral genome and in one instance are separated from the late gene by more than 100 kb of DNA. Moreover, the leader sequences map either upstream or downstream of the late gene, are transcribed from either DNA strand, and are fused to the late gene coding sequence via a poly(A) stretch. This demonstrates that vaccinia virus produces late mRNAs by tagging the protein-coding sequences onto the 3' end of other RNAa.
Output: Vaccinia Virus Produces Late mRNAs by Discontinuous Synthesis
Major incidents during the recent past have reinforced the value that the NHS and other agencies have invested into the comprehensive Emergency, Preparedness, Resilience and Response framework. This gives a detailed structure of the role of the NHS in any type of major incident from man-made disaster to pandemic flu. This has required preparation of communication, transport, security, military and healthcare systems. Also included is how the response to the incident is handled at a local level and for different levels of response. Examples of how this has played out are described. Specialist training at the higher and advanced level for trainees is established so that victims are triaged at the scene and received by consultants with appropriate training. Hospitals, ambulance services and intensive care units across the country can use networks to ensure not only rapid access to Major Trauma Centres but also to highly sophisticated skills when advanced life support is required. The NHS response to major incidents has been shown to be effective and successful.
Output: Preparation for and organization during a major incident
Epizootic outbreaks of diarrhoeas have emerged and disseminated in different rabbit farms in Mexico causing great economical losses, during the past years. Seven, 5-weeks-old New Zealand White (NZW) rabbits chosen at random from 35 ill animals that were remitted for postmortem, histopathology, and ultrastructural examinations were studied. Bacteriological and parasitological studies were carried out in three additional ill rabbits of same age. In a field trail 45, 5-weeks-old apparently healthy NZW rabbits were observed daily for sanitary status for a 5-week period. Some of the rabbits did not response to the preventive drug treatment and were therefore, used to study the development of the disease. Clinical signs, gross lesions, and mortality throughout the fattening period were recorded. Eight, 8-weeks-old NZW rabbits who survived an outbreak were assessed for gamma-globulins in serum of the total protein fraction during a 3week period. Gamma-globulins were also measured in eight free-disease healthy rabbits of same breed and age. Lesions of the small intestine consisted of mucoid enteropathy, lymphocytic plasmocytic enteritis with atrophy and fusion of villi, and hyperplasia of globet cells. Serosal edema was present. Ultrastructural examinations of jejunum and ileum from 3/7 diseased rabbits, revealed enterocytes in apoptosis, mixed with degenerative and/or necrotic changes together with infiltration of lymphocytes, macrophages, neutrophils, and loss of microvillus. There were electron dense structures suggestive of virus particles inside the nuclei and cytoplasm of some enterocytes. There was lymphoid spleen atrophy and proliferation of reticuloendothelial cells in 7/7 rabbits. Interstitial pneumonia in 4/7 rabbits was found. Encephalitozoon cuniculi was detected in the brain of 1/7 rabbits. Escherichia coli were detected in 3/3 cases and Eimeria spp. in 2/3 cases. Mortality rate in the field study was 51.1% and the spread of the disease occur in 9/9 cages. The proportion of gamma-globulins in rabbits who survive an outbreak was much lower (P = 0.0001) than free-disease healthy rabbits (8.1 ± 1.0 and 14.0 ± 1.0, respectively). The disease was multifactorial and consisted of sub-acute mucoid enteropathy probably induced by viral infection and aggravated by the proliferation of opportunistic pathogens common to rabbits. This may explain the severe degenerative and necrotic changes observed in the small intestine of diarrhoeic rabbits.
Output: Studies on the evolution, pathology, and immunity of commercial fattening rabbits affected with epizootic outbreaks of diarrhoeas in Mexico: A case report