Specialized cis-Acting RNA Elements Balance Genome Cyclization to Ensure Efficient Replication of Yellow Fever Virus.

Genome cyclization is essential for viral RNA (vRNA) replication of the vertebrate-infecting flaviviruses, and yet its regulatory mechanisms are not fully understood. Yellow fever virus (YFV) is a notorious pathogenic flavivirus. Here, we demonstrated that a group of cis-acting RNA elements in YFV balance genome cyclization to govern efficient vRNA replication. It was shown that the downstream of the 5'-cyclization sequence hairpin (DCS-HP) is conserved in the YFV clade and is important for efficient YFV propagation. By using two different replicon systems, we found that the function of the DCS-HP is determined primarily by its secondary structure and, to a lesser extent, by its base-pair composition. By combining in vitro RNA binding and chemical probing assays, we found that the DCS-HP orchestrates the balance of genome cyclization through two different mechanisms, as follows: the DCS-HP assists the correct folding of the 5' end in a linear vRNA to promote genome cyclization, and it also limits the overstabilization of the circular form through a potential crowding effect, which is influenced by the size and shape of the DCS-HP structure. We also provided evidence that an A-rich sequence downstream of the DCS-HP enhances vRNA replication and contributes to the regulation of genome cyclization. Interestingly, diversified regulatory mechanisms of genome cyclization, involving both the downstream of the 5'-cyclization sequence (CS) and the upstream of the 3'-CS elements, were identified among different subgroups of the mosquito-borne flaviviruses. In summary, our work highlighted how YFV precisely controls the balance of genome cyclization to ensure viral replication. IMPORTANCE Yellow fever virus (YFV), the prototype of the Flavivirus genus, can cause devastating yellow fever disease. Although it is preventable by vaccination, there are still tens of thousands of yellow fever cases per year, and no approved antiviral medicine is available. However, the understandings about the regulatory mechanisms of YFV replication are obscure. In this study, by a combination of bioinformatics, reverse genetics, and biochemical approaches, it was shown that the downstream of the 5'-cyclization sequence hairpin (DCS-HP) promotes efficient YFV replication by modulating the conformational balance of viral RNA. Interestingly, we found specialized combinations for the downstream of the 5'-cyclization sequence (CS) and upstream of the 3'-CS elements in different groups of the mosquito-borne flaviviruses. Moreover, possible evolutionary relationships among the various downstream of the 5'-CS elements were implied. This work highlighted the complexity of RNA-based regulatory mechanisms in the flaviviruses and will facilitate the design of RNA structure-targeted antiviral therapies.

Key Residue in the Precursor Region of M Protein Contributes to the Neurovirulence and Neuroinvasiveness of the African Lineage of Zika Virus.

The Zika virus (ZIKV) represents an important global health threat due to its unusual association with congenital Zika syndrome. ZIKV strains are phylogenetically grouped into the African and Asian lineages. However, the viral determinants underlying the phenotypic differences between the lineages remain unknown. Here, multiple sequence alignment revealed a highly conserved residue at position 21 of the premembrane (prM) protein, which is glutamic acid and lysine in the Asian and African lineages, respectively. Using reverse genetics, we generated a recombinant virus carrying an E21K mutation based on the genomic backbone of the Asian lineage strain FSS13025 (termed E21K). The E21K mutation significantly increased viral replication in multiple neural cell lines with a higher ratio of M to prM production. Animal studies showed E21K exhibited increased neurovirulence in suckling mice, leading to more severe defects in mouse brains by causing more neural cell death and destruction of hippocampus integrity. Moreover, the E21K substitution enhanced neuroinvasiveness in interferon alpha/beta (IFN-α/ß) receptor knockout mice, as indicated by the increased mortality, and enhanced replication in mouse brains. The global transcriptional analysis showed E21K infection profoundly altered neuron development networks and induced stronger antiviral immune response than wild type (WT) in both neural cells and mouse brains. More importantly, the reverse K21E mutation based on the genomic backbone of the African strain MR766 caused less mouse neurovirulence. Overall, our findings support the 21st residue of prM functions as a determinant for neurovirulence and neuroinvasiveness of the African lineage of ZIKV. IMPORTANCE The suspected link of Zika virus (ZIKV) to birth defects led the World Health Organization to declare ZIKV a Public Health Emergency of International Concern. ZIKV has been identified to have two dominant phylogenetic lineages, African and Asian. Significant differences exist between the two lineages in terms of neurovirulence and neuroinvasiveness in mice. However, the viral determinants underlying the phenotypic differences are still unknown. Here, combining reverse genetics, animal studies, and global transcriptional analysis, we provide evidence that a single E21K mutation of prM confers to the Asian lineage strain FSS130125 significantly enhanced replication in neural cell lines and more neurovirulent and neuroinvasiveness phenotypes in mice. Our findings support that the highly conserved residue at position 21 of prM functions as a determinant of neurovirulence and neuroinvasiveness of the African lineage of ZIKV in mice.

Short Direct Repeats in the 3' Untranslated Region Are Involved in Subgenomic Flaviviral RNA Production.

Mosquito-borne flaviviruses consist of a positive-sense genome RNA flanked by the untranslated regions (UTRs). There is a panel of highly complex RNA structures in the UTRs with critical functions. For instance, Xrn1-resistant RNAs (xrRNAs) halt Xrn1 digestion, leading to the production of subgenomic flaviviral RNA (sfRNA). Conserved short direct repeats (DRs), also known as conserved sequences (CS) and repeated conserved sequences (RCS), have been identified as being among the RNA elements locating downstream of xrRNAs, but their biological function remains unknown. In this study, we revealed that the specific DRs are involved in the production of specific sfRNAs in both mammalian and mosquito cells. Biochemical assays and structural remodeling demonstrate that the base pairings in the stem of these DRs control sfRNA formation by maintaining the binding affinity of the corresponding xrRNAs to Xrn1. On the basis of these findings, we propose that DRs functions like a bracket holding the Xrn1-xrRNA complex for sfRNA formation.IMPORTANCE Flaviviruses include many important human pathogens. The production of subgenomic flaviviral RNAs (sfRNAs) is important for viral pathogenicity as a common feature of flaviviruses. sfRNAs are formed through the incomplete degradation of viral genomic RNA by the cytoplasmic 5'-3' exoribonuclease Xrn1 halted at the Xrn1-resistant RNA (xrRNA) structures within the 3'-UTR. The 3'-UTRs of the flavivirus genome also contain distinct short direct repeats (DRs), such as RCS3, CS3, RCS2, and CS2. However, the biological functions of these ancient primary DR sequences remain largely unknown. Here, we found that DR sequences are involved in sfRNA formation and viral virulence and provide novel targets for the rational design of live attenuated flavivirus vaccine.

The long-term impact of a chronic total occlusion in a non-infarct-related artery on acute ST-segment elevation myocardial infarction after primary coronary intervention.

OBJECTIVES: To investigate the long-term outcome of patients with acute ST-segment elevation myocardial infarction (STEMI) and a chronic total occlusion (CTO) in a non-infarct-related artery (IRA) and the risk factors for mortality. METHODS: The enrolled cohort comprised 323 patients with STEMI and multivessel diseases (MVD) that received a primary percutaneous coronary intervention between January 2008 and November 2013. The patients were divided into two groups: the CTO group (n = 97) and the non-CTO group (n = 236). The long-term major adverse cardiovascular and cerebrovascular events (MACCE) experienced by each group were compared. RESULTS: The rates of all-cause mortality and MACCE were significantly higher in the CTO group than they were in the non-CTO group. Cox regression analysis showed that an age ≥ 65 years (OR = 3.94, 95% CI: 1.47-10.56, P = 0.01), a CTO in a non-IRA(OR = 5.09, 95% CI: 1.79 ~ 14.54, P < 0.01), an in-hospital Killip class ≥ 3 (OR = 4.32, 95% CI: 1.71 ~ 10.95, P < 0.01), and the presence of renal insufficiency (OR = 5.32, 95% CI: 1.49 ~ 19.01, P = 0.01), stress ulcer with gastraintestinal bleeding (SUB) (OR = 6.36, 95% CI: (1.45 ~ 28.01, P = 0.01) were significantly related the 10-year mortality of patients with STEMI and MVD; an in-hospital Killip class ≥ 3 (OR = 2.97,95% CI:1.46 ~ 6.03, P < 0.01) and the presence of renal insufficiency (OR = 5.61, 95% CI: 1.19 ~ 26.39, P = 0.03) were significantly related to the 10-year mortality of patients with STEMI and a CTO. CONCLUSIONS: The presence of a CTO in a non-IRA, an age ≥ 65 years, an in-hospital Killip class ≥ 3, and the presence of renal insufficiency, and SUB were independent risk predictors for the long-term mortality of patients with STEMI and MVD; an in-hospital Killip class ≥ 3 and renal insufficiency were independent risk predictors for the long-term mortality of patients with STEMI and a CTO.

Effects of antidiabetic drugs on left ventricular function/dysfunction: a systematic review and network meta-analysis.

BACKGROUND: Although a variety of antidiabetic drugs have significant protective action on the cardiovascular system, it is still unclear which antidiabetic drugs can improve ventricular remodeling and fundamentally delay the process of heart failure. The purpose of this network meta-analysis is to compare the efficacy of sodium glucose cotransporter type 2 (SGLT-2) inhibitors, dipeptidyl peptidase-4 (DPP-4) inhibitors, glucagon-like peptide-1 (GLP-1) agonists, metformin (MET), sulfonylurea (SU) and thiazolidinediones (TZDs) in improving left ventricular (LV) remodeling in patients with type 2 diabetes (T2DM) and/or cardiovascular disease (CVD). METHODS: We searched articles published before October 18, 2019, regardless of language or data, in 4 electronic databases: PubMed, EMBASE, Cochrane Library and Web of Science. We included randomized controlled trials in this network meta-analysis, as well as a small number of cohort studies. The differences in the mean changes in left ventricular echocardiographic parameters between the treatment group and control group were evaluated. RESULTS: The difference in the mean change in LV ejection fraction (LVEF) between GLP-1 agonists and placebo in treatment effect was greater than zero (MD = 2.04% [0.64%, 3.43%]); similar results were observed for the difference in the mean change in LV end-diastolic diameter (LVEDD) between SGLT-2 inhibitors and placebo (MD = - 3.3 mm [5.31, - 5.29]), the difference in the mean change in LV end-systolic volume (LVESV) between GLP-1 agonists and placebo (MD = - 4.39 ml [- 8.09, - 0.7]); the difference in the mean change in E/e' between GLP-1 agonists and placebo (MD = - 1.05[- 1.78, - 0.32]); and the difference in the mean change in E/e' between SGLT-2 inhibitors and placebo (MD = - 1.91[- 3.39, - 0.43]). CONCLUSIONS: GLP-1 agonists are more significantly associated with improved LVEF, LVESV and E/e', SGLT-2 inhibitors are more significantly associated with improved LVEDD and E/e', and DPP-4 inhibitors are more strongly associated with a negative impact on LV end-diastolic volume (LVEDV) than are placebos. SGLT-2 inhibitors are superior to other drugs in pairwise comparisons.

The Emerging Duck Flavivirus Is Not Pathogenic for Primates and Is Highly Sensitive to Mammalian Interferon Antiviral Signaling.

UNLABELLED: Flaviviruses pose a significant threat to both animals and humans. Recently, a novel flavivirus, duck Tembusu virus (DTMUV), was identified to be the causative agent of a serious duck viral disease in Asia. Its rapid spread, expanding host range, and uncertain transmission routes have raised substantial concerns regarding its potential threats to nonavian hosts, including humans. Here, we demonstrate that DTMUV is not pathogenic for nonhuman primates and is highly sensitive to mammal type I interferon (IFN) signaling. In vitro assays demonstrated that DTMUV infected and replicated efficiently in various mammalian cell lines. Further tests in mice demonstrated high neurovirulence and the age-dependent neuroinvasiveness of the virus. In particular, the inoculation of DTMUV into rhesus monkeys did not result in either viremia or apparent clinical symptoms, although DTMUV-specific humoral immune responses were detected. Furthermore, we revealed that although avian IFN failed to inhibit DTMUV in avian cells, DTMUV was more sensitive to the antiviral effects of type I interferon than other known human-pathogenic flaviviruses. Knockout of the type I IFN receptor in mice caused apparent viremia, viscerotropic disease, and mortality, indicating a vital role of IFN signaling in protection against DTMUV infection. Collectively, we provide direct experimental evidence that this novel avian-origin DTMUV possesses a limited capability to establish infection in immunocompetent primates due to its decreased antagonistic activity in the mammal IFN system. Furthermore, our findings highlight the potential risk of DTMUV infection in immunocompromised individuals and warrant studies on the cross-species transmission and pathogenesis of this novel flavivirus. IMPORTANCE: Mosquito-borne flaviviruses comprise a large group of pathogenic and nonpathogenic members. The pathogenic flaviviruses include dengue, West Nile, and Japanese encephalitis viruses, and the nonpathogenic flaviviruses normally persist in a natural cycle and rarely cause disease in humans. A novel flavivirus, DTMUV (also known as duck egg drop syndrome flavivirus [DEDSV]) was identified in 2012 in ducks and then rapidly spread to several Asian countries. This new flavivirus was then shown to infect multiple avian species, resulting in neurological symptoms with unknown routes of transmission. There is public concern regarding its potential transmission from birds to humans and other nonavian hosts. Our present study shows that the mammalian IFN system can efficiently eliminate DTMUV infection and that the emergence of severe DTMUV-associated disease in mammals, especially humans, is unlikely. Currently, DTMUV infection mostly affects avian species.

In vitro and in vivo characterization of chimeric duck Tembusu virus based on Japanese encephalitis live vaccine strain SA14-14-2.

Duck Tembusu virus (DTMUV), a newly identified flavivirus, has rapidly spread to China, Malaysia and Thailand. The potential threats to public health have been well-highlighted; however its virulence and pathogenesis remain largely unknown. Here, by using reverse genetics, a recombinant chimeric DTMUV based on Japanese encephalitis live vaccine strain SA14-14-2 was obtained by substituting the corresponding prM and E genes (named ChinDTMUV). In vitro characterization demonstrated that ChinDTMUV replicated efficiently in mammalian cells with small-plaque phenotype in comparison with its parental viruses. Mouse tests showed ChinDTMUV exhibited avirulent phenotype in terms of neuroinvasiveness, while it retained neurovirulence from its parental virus DTMUV. Furthermore, immunization with ChinDTMUV was evidenced to elicit robust IgG and neutralizing antibody responses in mice. Overall, we successfully developed a viable chimeric DTMUV, and these results provide a useful platform for further investigation of the pathogenesis of DTMUV and development of a live attenuated DTMUV vaccine candidate.

Genotype-specific neutralization determinants in envelope protein: implications for the improvement of Japanese encephalitis vaccine.

Japanese encephalitis remains the leading cause of viral encephalitis in children in Asia and is expanding its geographical range to larger areas in Asia and Australasia. Five genotypes of Japanese encephalitis virus (JEV) co-circulate in the geographically affected areas. In particular, the emergence of genotype I (GI) JEV has displaced genotype III (GIII) as the dominant circulating genotype in many Asian regions. However, all approved vaccine products are derived from GIII strains. In the present study, bioinformatic analysis revealed that GI and GIII JEV strains shared two distinct amino acid residues within the envelope (E) protein (E222 and E327). By using reverse genetics approaches, A222S and S327T mutations were demonstrated to decrease live-attenuated vaccine (LAV) SA14-14-2-induced neutralizing antibodies in humans, without altering viral replication. A222S or S327T mutations were then rationally engineered into the infectious clone of SA14-14-2, and the resulting mutant strains retained the same genetic stability and attenuation characteristics as the parent strain. More importantly, immunization of mice with LAV-A222S or LAV-S327T elicited increased neutralizing antibodies against GI strains. Together, these results demonstrated that E222 and E327 are potential genotype-related neutralization determinants and are critical in determining the protective efficacy of live Japanese encephalitis vaccine SA14-14-2 against circulating GI strains. Our findings will aid in the rational design of the next generation of Japanese encephalitis LAVs capable of providing broad protection against all JEV strains belonging to different genotypes.

Novel cis-acting element within the capsid-coding region enhances flavivirus viral-RNA replication by regulating genome cyclization.

cis-Acting elements in the viral genome RNA (vRNA) are essential for the translation, replication, and/or encapsidation of RNA viruses. In this study, a novel conserved cis-acting element was identified in the capsid-coding region of mosquito-borne flavivirus. The downstream of 5' cyclization sequence (5'CS) pseudoknot (DCS-PK) element has a three-stem pseudoknot structure, as demonstrated by structure prediction and biochemical analysis. Using dengue virus as a model, we show that DCS-PK enhances vRNA replication and that its function depends on its secondary structure and specific primary sequence. Mutagenesis revealed that the highly conserved stem 1 and loop 2, which are involved in potential loop-helix interactions, are crucial for DCS-PK function. A predicted loop 1-stem 3 base triple interaction is important for the structural stability and function of DCS-PK. Moreover, the function of DCS-PK depends on its position relative to the 5'CS, and the presence of DCS-PK facilitates the formation of 5'-3' RNA complexes. Taken together, our results reveal that the cis-acting element DCS-PK enhances vRNA replication by regulating genome cyclization, and DCS-PK might interplay with other cis-acting elements to form a functional vRNA cyclization domain, thus playing critical roles during the flavivirus life cycle and evolution.

Rational design of a flavivirus vaccine by abolishing viral RNA 2'-O methylation.

Viruses that replicate in the cytoplasm cannot access the host nuclear capping machinery. These viruses have evolved viral methyltransferase(s) to methylate N-7 and 2'-O cap of their RNA; alternatively, they "snatch" host mRNA cap to form the 5' end of viral RNA. The function of 2'-O methylation of viral RNA cap is to mimic cellular mRNA and to evade host innate immune restriction. A cytoplasmic virus defective in 2'-O methylation is replicative, but its viral RNA lacks 2'-O methylation and is recognized and eliminated by the host immune response. Such a mutant virus could be rationally designed as a live attenuated vaccine. Here, we use Japanese encephalitis virus (JEV), an important mosquito-borne flavivirus, to prove this novel vaccine concept. We show that JEV methyltransferase is responsible for both N-7 and 2'-O cap methylations as well as evasion of host innate immune response. Recombinant virus completely defective in 2'-O methylation was stable in cell culture after being passaged for >30 days. The mutant virus was attenuated in mice, elicited robust humoral and cellular immune responses, and retained the engineered mutation in vivo. A single dose of immunization induced full protection against lethal challenge with JEV strains in mice. Mechanistically, the attenuation phenotype was attributed to the enhanced sensitivity of the mutant virus to the antiviral effects of interferon and IFIT proteins. Collectively, the results demonstrate the feasibility of using 2'-O methylation-defective virus as a vaccine approach; this vaccine approach should be applicable to other flaviviruses and nonflaviviruses that encode their own viral 2'-O methyltransferases.

A chimeric dengue virus vaccine using Japanese encephalitis virus vaccine strain SA14-14-2 as backbone is immunogenic and protective against either parental virus in mice and nonhuman primates.

The development of a safe and efficient dengue vaccine represents a global challenge in public health. Chimeric dengue viruses (DENV) based on an attenuated flavivirus have been well developed as vaccine candidates by using reverse genetics. In this study, based on the full-length infectious cDNA clone of the well-known Japanese encephalitis virus live vaccine strain SA14-14-2 as a backbone, a novel chimeric dengue virus (named ChinDENV) was rationally designed and constructed by replacement with the premembrane and envelope genes of dengue 2 virus. The recovered chimeric virus showed growth and plaque properties similar to those of the parental DENV in mammalian and mosquito cells. ChinDENV was highly attenuated in mice, and no viremia was induced in rhesus monkeys upon subcutaneous inoculation. ChinDENV retained its genetic stability and attenuation phenotype after serial 15 passages in cultured cells. A single immunization with various doses of ChinDENV elicited strong neutralizing antibodies in a dose-dependent manner. When vaccinated monkeys were challenged with wild-type DENV, all animals except one that received the lower dose were protected against the development of viremia. Furthermore, immunization with ChinDENV conferred efficient cross protection against lethal JEV challenge in mice in association with robust cellular immunity induced by the replicating nonstructural proteins. Taken together, the results of this preclinical study well demonstrate the great potential of ChinDENV for further development as a dengue vaccine candidate, and this kind of chimeric flavivirus based on JE vaccine virus represents a powerful tool to deliver foreign antigens.

miR-125b regulates side population in breast cancer and confers a chemoresistant phenotype.

Resistance to chemotherapy is a major obstacle for the effective treatment of breast cancer and is partially due to the presence of drug resistant stem cell-like side population (SP). Previous studies have shown elevated miR-125b is associated with chemoresistance and metastasis; however, the relationship between miR-125b and SP cells remains unknown. In this study, we isolated and characterized SP cells in a panel of breast cancer cell lines and primary cancer cells from breast cancer patients. SP cells showed cancer stem cells (CSCs) properties, including self-renewal, resistance to chemotherapy and high expression of stem cell markers. The percentage of SP cells was higher in chemotherapy resistant patients compared to that in chemotherapy responsive patients (5.8 ± 2.4% in non-responsive patients vs. 1.2 ± 0.5% in responsive patients, P = 0.012). Importantly, SP cells had higher level of miR-125b than NSP cells and the elevated miR-125b expression in chemoresistant cancer cells were due to high percentage of SP cells. Overexpression of miR-125b correlated with an increase in tumor SP and CSC property, whereas knockdown of miR-125b correlated with decreased incidence of SP. In addition, miR-125b overexpression in breast cancer cells induced epithelial-mesenchymal transition (EMT)-like cellular marker alteration, suggesting a potential mechanism of miR-125b in the regulation of cancer stem-like SP cells. Taken together, these results suggest an important role for miR-125b in breast cancer chemoresistance by maintaining cancer stem-like SP fraction, and raise the possibility that miR-125b may be a significant prognostic response marker for cancer therapy.

Identification and characterization of a linearized B-cell epitope on the pr protein of dengue virus.

The four serotypes of dengue virus (DENV) represent one of the major mosquito-borne pathogens globally; so far no vaccine or specific antiviral is available. During virion maturation, the pr protein is cleaved from its precursor form the prM protein on the surface of immature DENV by host protease. Recent findings have demonstrated that the pr protein not only played critical roles in virion assembly and maturation, but was also involved in antibody-dependent enhancement of DENV infection. However, the B-cell epitopes on the pr protein of DENV have not been well characterized. In this study, a set of 11 partially overlapping peptides spanning the entire pr protein of DENV-2 were fused with glutathione S-transferase and expressed in Escherichia coli. ELISA screening with murine hyperimmune antiserum against immature DENV identified the P8 peptide (57KQNEPEDIDCWCNST7¹) in the pr protein as the major immunodominant epitope. Fine mapping by truncated protein assays confirmed the 8-e peptide 57KQNEPEDI64 was the smallest unit capable of antibody binding. Importantly, the 8-e epitope reacted with sera from dengue fever patients. Site-directed mutagenesis revealed the asparagine residue at position 59 was important for epitope recognition. The 8-e epitope coincided well with the B-cell epitopes predicted by Immune Epitope Database analysis, and 3D structural modelling mapped the 8-e peptide on the surface of prM-E heterodimers. Overall, our findings characterized a linearized B-cell epitope on the pr protein of DENV, which will help to understand the life cycle of DENV and pathogenesis of dengue infections in human.

Development of chimaeric West Nile virus attenuated vaccine candidate based on the Japanese encephalitis vaccine strain SA14-14-2.

Mosquito-borne flaviviruses include a large group of important human medical pathogens. Several chimaeric flaviviruses have been constructed, and show potential for vaccine development. Although Japanese encephalitis virus (JEV) live vaccine SA14-14-2 has been widely used with ideal safety and efficacy profiles, no chimaeric flavivirus based on the JEV vaccine has been described to date. Based on the reverse genetic system of the JEV vaccine SA14-14-2, a novel live chimaeric flavivirus carrying the protective antigens of West Nile virus (WNV) was constructed and recovered in this study. The resulting chimaera (ChinWNV) replicated efficiently in both mammalian and mosquito cells and possessed genetic stability after in vitro serial passaging. ChinWNV exhibited a small-plaque phenotype, and its replication was significantly restricted in mouse peripheral blood and brain compared with parental WNV. Importantly, ChinWNV was highly attenuated with regard to both neurovirulence and neuroinvasiveness in mice. Furthermore, a single ChinWNV immunization stimulated robust WNV-specific adaptive immune responses in mice, conferring significant protection against lethal WNV infection. Our results demonstrate that chimaeric flaviviruses based on the JEV vaccine can serve as a powerful platform for vaccine development, and that ChinWNV represents a potential WNV vaccine candidate that merits further development.

Complete genome sequence of a chikungunya virus isolated in Guangdong, China.

Chikungunya virus belongs to the genus Alphavirus in the family Togaviridae. Here we report the complete genome sequence of a chikungunya virus strain, GD05/2010, isolated in 2010 from a patient with chikungunya fever in Guangdong, China. The sequence information is important for surveillance of this emerging arboviral infection in China.

Noninvasive bioluminescence imaging of dengue virus infection in the brain of A129 mice.

Dengue virus (DENV) infection is one of the most important public health threats globally; however, no vaccines or effective antivirals are currently available. The bioluminescence imaging technique has emerged as a powerful tool for studies on viral pathogenesis in vitro and in vivo. In this study, using a recombinant DENV that stably expressed Renilla luciferase (Rluc-DENV), we used bioluminescence for imaging of DENV infection in the brain of A129 mice that lacked type I interferon receptors. Upon intracranial inoculation with Rluc-DENV, A129 mice developed typical neurological symptoms and rapidly succumbed to viral infection. Real-time bioluminescence intensity analysis revealed the replication kinetics of Rluc-DENV in the brain of A129 mice. Linear regression analyses showed a good correlation between photon flux and viral titers (R(2) = 0.9923). Finally, the bioluminescence model was validated using a known mouse monoclonal antibody, 2A10G6, and the therapeutic effects of this neutralizing antibody were readily monitored by live imaging in the same animal. The noninvasive bioluminescence imaging of DENV infection as described here shows distinct advantages over traditional animal models and provides a powerful tool for potential antiviral or vaccine assays against DENV infection in vivo.

Comparisons of drug-eluting balloon versus drug-eluting stent for the treatment of cancer patients presenting with acute myocardial infarction.

BACKGROUND: Treatment for cancer patients presenting with acute myocardial infarction (AMI) remains challenging. The objective of the study was to investigate the safety and efficiency of drug eluting balloon (DEB) versus drug eluting stent (DES) in this high-risk group. METHODS: Between 1st January 2017 and 1st January 2022, cancer patients admitted to Beijing Chaoyang Hospital with AMI were retrospectively enrolled. The primary endpoint was major adverse cardiovascular event (MACE). The secondary endpoints included major bleeding events, heart failure and cardiac complications. RESULTS: A total of 164 cancer patients presenting with AMI were included in the final analysis. Patients treated with DEB had a numerically lower rate of MACE than those treated with DES during a median follow-up of 21.8 months (22.9% vs. 37.1%, p = 0.23). Patients treated with DEB had a trend towards lower rate of major bleeding events than patients treated with DES (6.3% vs. 18.1%, HR 2.96, 95% CI [0.88, 9.92], p = 0.08). There were no significant differences between the two groups with regards to the rate of heart failure (4.2% vs. 9.5%, p = 0.32) and cardiac complications (0.0% vs. 2.6%, p = 0.56). CONCLUSIONS: The present study demonstrated that in cancer patients with AMI, DEB had a trend towards lower rate of major bleeding events and a numerically lower rate of MACE compared with DES.

Comparisons of different new-generation transcatheter aortic valve implantation devices for patients with severe aortic stenosis: a systematic review and network meta-analysis.

BACKGROUND: Whether there are differences among the new-generation transcatheter aortic valve implantation (TAVI) devices for patients with aortic stenosis remains unclear. The aim of the study was to compare the efficiency and safety of different new-generation TAVI devices for patients with aortic stenosis. MATERIALS AND METHODS: A comprehensive search of PubMed, Embase and Web of Science from their inception to 1 February 2022. Randomized clinical trials and observational studies that compared two or more different TAVI devices were enroled. Pairwise meta-analysis and frequentist network meta-analysis were conducted to pool the outcome estimates of interest. RESULTS: A total of 79 studies were finally included. According to the surface under the cumulative ranking, the top two ranked valves for lower rates of events were as follows: direct flow medical (DFM) (4.6%) and Lotus (48.8%) for lower rate of device success; Sapien 3 (16.8%) and DFM (19.7%) for lower mortality; DFM (8.6%) and Sapien 3 (25.5%) for lower rates of stroke; Evolut (27.6%) and DFM (35.8%) for lower rates of major and life-threatening bleeding; Portico (22.6%) and Sapien 3 (41.9%) for lower rates of acute kidney injury; Acurate (8.6%) and DFM (13.2%) for lower rates of permanent pacemaker implantation; Lotus (0.3%) and Sapien 3 (22.7%) for lower rates of paravalvular leak; Evolut (1.4%) and Portico (29.1%) for lower rates of mean aortic valve gradients. CONCLUSIONS: The findings of the present study suggested that the device success rates were comparable among these new-generation valves except for DFM. After excluding DFM, Sapien 3 might be the best effective for decreased mortality and stroke; Lotus might be the best effective for decreased paravalvular leak; Evolut might be the best effective for decreased major and life-threatening bleeding and mean aortic valve gradients; Acurate and Portico might be the best effective for decreased permanent pacemaker implantation and acute kidney injury, respectively.

Comparisons of Drug-Eluting Balloon versus Drug-Eluting Stent in the Treatment of Young Patients with Acute Myocardial Infarction.

BACKGROUND: The incidence of acute myocardial infarction (AMI) in the younger population has been increasing gradually in recent years. The objective of the present study is to investigate the safety and effectiveness of drug-eluting balloons (DEBs) in young patients with AMI. METHODS: All consecutive patients with AMI aged ≤ 45 years were retrospectively enrolled. The primary endpoint was a device-oriented composite endpoint (DOCE) of cardiac death, target vessel myocardial infarction (MI), or target lesion revascularization (TLR). The secondary study endpoints included heart failure and major bleeding events. RESULTS: A total of 276 young patients presenting with AMI were finally included. The median follow-up period was 1155 days. Patients treated with DEBs had a trend toward a lower incidence of DOCEs (3.0% vs. 11.0%, p = 0.12) mainly driven by the need for TLR (3.0% vs. 9.1%, p = 0.19) than those treated with DESs. No significant differences between the two groups were detected in the occurrence of cardiac death (0.0% vs. 0.5%, p = 0.69), MI (0.0% vs. 1.4%, p = 0.40), heart failure (0.0% vs. 1.9%, p = 0.39), or major bleeding events (1.5% vs 4.8%, p = 0.30). Multivariate regression analysis showed that DEBs were associated with a trend toward a lower risk of DOCEs (HR 0.13, 95% CI [0.02, 1.05], p = 0.06). CONCLUSIONS: The findings of the present study suggested that DEBs might be a potential treatment option in young patients with AMI. A larger scale, randomized, multicenter study is required to investigate the safety and effectiveness of DEBs in this setting.

A single nucleotide mutation in NS2A of Japanese encephalitis-live vaccine virus (SA14-14-2) ablates NS1' formation and contributes to attenuation.

Japanese encephalitis (JE) remains the leading cause of viral encephalitis in the Asia-Pacific region, and the live vaccine SA14-14-2 is currently recommended by WHO and widely used in Asian countries with a good safety and efficacy profile. In this study, we demonstrated that SA14-14-2 failed to produce NS1', the larger NS1-related protein, compared with its parental strain SA14 in various cells. Sequence analysis and secondary structure prediction identified a single silent mutation G66A in the NS2A-coding region of SA14-14-2 destabilized the conserved pseudoknot structure, which was associated with a -1 ribosomal frame shift event. Using reverse genetic technology and animal study, we provided solid evidence that this single silent mutation G66A in the NS2A gene abolished the production of NS1' in vitro and reduced neurovirulence and neuroinvasiveness in mice. These findings provide critical information in understanding the molecular mechanism of JE vaccine attenuation and is critical for JE vaccine quality control.