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In 2019, a serious dengue virus (DENV) infection broke out in the Xishuangbanna Dai Autonomous Prefecture, China. Therefore, we conducted a molecular epidemiological analysis in people that contracted DENV serotype 1 (DENV-1) during this year. We analyzed the molecular epidemiology of six DENV-1 epidemic strains in 2019 by full-length genome sequencing, amino acid mutation site analysis, evolutionary tree analysis, and recombination site comparison analysis. Through the analysis of amino acid mutation sites, it was found that DENV-1 strain (MW386867) was different from the other five epidemic DENV-1 strains in Xishuangbanna in 2019. MW386867 had unique mutation sites at six loci. The six epidemic DENV-1 strains in Xishuangbanna in 2019 were divided into two clusters. MW386867 was highly similar to the MG679800 (Myanmar 2017), MG679801 (Myanmar 2017), and KC172834 (Laos 2008), and the other five strains were highly similar to JQ045660 (Vietnam 2011), FJ176780 (GuangDong 2006). Genetic recombination analysis revealed that there was no recombination signal in the six epidemic DENV-1 strains in Xishuangbanna in 2019. We speculate that the DENV-1 epidemic in 2019 has a co-epidemic of local strains and cross-border strains., (© 2022 Wiley Periodicals LLC.)

Dengue fever, as a mosquito-borne viral disease widely spread in tropical and subtropical regions, remarkably threatens public health, while the mechanism involved in host-DENV interaction has not been fully elucidated. Firstly, we analyzed the expression levels of long non-coding RNAs (lncRNAs) in THP-1 cells after DENV-3 infection and Antibody- Dependent Enhancement of viral infection (ADE-VI) by RNA-Seq. Secondly, through the RT-qPCR to confirm those differentially expressed (DE) lncRNAs. Then, we also analyzed the competitive endogenous RNA (CeRNA) regulatory network of DE lncRNAs. Finally, we predicted the encode ability of DE lncRNAs. It was found that on the X and Y chromosomes, the expression levels of lncRNAs in THP-1 cells after ADE-VI were significantly different from those in the negative control and the DENV-3 infection groups. There were 71 DE lncRNAs after DENV-3 infection, including 42 up-regulated and 29 down-regulated lncRNAs. A total of 70 DE lncRNAs after ADE-VI were detected, including 38 up-regulated and 32 down- regulated lncRNAs. After ADE-VI and DENV-3 infection, there were 35 DE lncRNAs, including 11 up-regulated and 24 down-regulated lncRNAs. The analysis of the CeRNA regulatory network of DE lncRNAs revealed that, TRIM29, STC2, and IGFBP5 were correlated with the ADE-VI. Additionally, it was found that lncRNAs not only participated in the CeRNA regulatory network, but also maybe encoded small peptides. Our findings provided clues for further investigation into the lncRNAs associated antiviral mechanism of ADE-VI and DENV-3 infection., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Dengue virus infection mainly causes dengue hemorrhagic fever (DHF) and/or dengue shock syndrome (DSS). However, ADE (antibody-dependent enhancement) is one of the main pathogenic factors, and its pathogenic mechanism has not been fully elucidated. Recently, with the development of high-throughput sequencing, an increased number of RNAs have been confirmed to play a vital regulatory role in the process of virus infection. However, there is a lack of research on dengue virus infection and ADE. In this study, we used RNA-Seq to detect differentially expressed RNAs (DE RNAs) profiles in mock-infected, DENV-3-infected, and ADE-infected THP-1 cells. Firstly, we found 69 circRNAs, 259 miRNAs, and 18 mRNAs were differentially expressed in THP-1 vs DENV-3. In THP-1 vs ADE, 94 circRNAs, 263 miRNAs, and 111 mRNAs were differentially expressed. In DENV-3 vs ADE, 68 circRNAs, 105 miRNAs, and 94 mRNAs were differentially expressed. Functional enrichment analysis of these DE RNAs mainly focused on immune system, viral infectious diseases, cytokine-cytokine receptor interactions, and NOD/RIG-I-like receptor signaling pathways. In DENV-3 vs ADE, notably, the expression of HBB was up-regulated, which was a Fcγ Receptor-mediated phagocytosis protein. Additionally, we predicted the encoding ability of DE circRNAs, and it was found that a small peptide was encoded by novel_circ_001562 and that its amino acid sequence was consistent with that of DDX60L, which is a class of interferon-stimulated genes. Finally, we constructed the ceRNA regulatory network pathway. Therefore, our study provides a new strategy for further investigation on DENV-host interactions.

In 2018, a small-scale dengue epidemic broke out in Hunan Province, an inland province in central South China, with 172 people infected. To verify the causative agent, complete genome information was obtained by PCR and sequencing based on the viral RNAs extracted from patient serum samples. Mutation and evolutionary analysis were performed by MEGA7.0 software. The online softwares "Predict protein" and "Mfold" were used to predict the secondary structure of proteins and untranslated regions, respectively. Phylogenetic analysis showed that all five isolates in this study were DENV type 2, which is most closely related to the Zhejiang strain (2017-MH110588). Compared with the DENV-2 standard strain, 773 nucleotide mutations occurred in the isolated strain, of which 666 were nonsense mutations. Of the 80 mutated amino acids, 22 occurred in the structural protein region (2 in C region, 8 in PrM/M region, 12 in E region), and 58 in the non-structural (NS) protein region (9 in NS1 region, 10 in NS2 region, 12 in NS3 region, 7 in NS4 region, 20 in NS5 region). The prM/M region had the highest AA mutation rate, while NS4B was conservative. Three amino acid mutations (E: N390 th S, and NS5: S676 th N, K800 th T) may important for the replication and virulence of the DENV. Secondary structure prediction observed 28 changes in polynucleotide binding regions and 110 changes in protein binding sites of coding sequence. 2 and 4 base substitutions resulted in 2 and 6 significant changes in the RNA secondary structure of 5' UTR and 3' UTR, respectively. Two significant positive selection sites were observed in NS5. To our knowledge, this research is the first complete genome analysis of the epidemic strain of the 2018 dengue outbreak in Hunan and will benefit further research for virus traceability and vaccine development., (Copyright © 2021 Elsevier B.V. All rights reserved.)

A dengue virus serotype 1 (DENV-1) epidemic occurred from October to December 2018 in Xishuangbanna, Yunnan, Southwest China, neighboring Myanmar, Laos, and Vietnam. In this study, we investigated the molecular characteristics, evolution, and potential source of DENV from Xishuangbanna. The C (capsid), prM (premembrane), and E (envelope) genes of DENV isolated from 87 serum samples obtained from local patients were amplified and sequenced, and the sequences were evaluated by identification of mutations, phylogenetic and homologous recombination analysis, and secondary structure prediction. Phylogenetic analysis showed that all of the epidemic DENV strains from Xishuangbanna could be grouped in a branch with DENV-1 isolates, and were most similar to the Fujian 2005 (China, DQ193572) and Singapore 2016 (MF314188) strains. When compared with DENV-1SS (the standard strain), there were 31 non-synonymous mutations, but no obvious homologous recombination signal was found. Secondary structure prediction showed that some changes had occurred in a helical region in proteins of the MN123849 and MN123854 strains, but there were few changes in the disordered region. This study reveals the molecular characteristics of the structural genes of the Xishuangbanna epidemic strains in 2018 and provides a reference for molecular epidemiology, infection, and pathogenicity research and vaccine development.

Background: An unexpected dengue outbreak occurred in Hunan Province in 2018. This was the first dengue outbreak in this area of inland China, and 172 cases were reported., Methods: To verify the causative agent of this outbreak and characterise the viral genes, the genes encoding the structural proteins C/prM/E of viruses isolated from local residents were sequenced followed by mutation and phylogenetic analysis. Recombination, selection pressure, potential secondary structure and three-dimensional structure analyses were also performed., Results: Phylogenetic analysis revealed that all epidemic strains were of the cosmopolitan DENV-2 genotype and were most closely related to the Zhejiang strain (MH010629, 2017) and then the Malaysia strain (KJ806803, 2013). Compared with the sequence of DENV-2SS, 151 base substitutions were found in the sequences of 89 isolates; these substitutions resulted in 20 non-synonymous mutations, of which 17 mutations existed in all samples (two in the capsid protein, six in the prM/M proteins, and nine in the envelope proteins). Moreover, amino acid substitutions at the 602nd (E322:Q → H) and 670th (E390: N → S) amino acids may have enhanced the virulence of the epidemic strains. One new DNA binding site and five new protein binding sites were observed. Two polynucleotide binding sites and seven protein binding sites were lost in the epidemic strains compared with DENV-2SS. Meanwhile, five changes were found in helical regions. Minor changes were observed in helical transmembrane and disordered regions. The 429th amino acid of the E protein switched from a histamine (positively charged) to an asparagine (neutral) in all 89 isolated strains. No recombination events or positive selection pressure sites were observed. To our knowledge, this study is the first to analyse the genetic characteristics of epidemic strains in the first dengue outbreak in Hunan Province in inland China., Conclusions: The causative agent is likely to come from Zhejiang Province, a neighbouring province where dengue fever broke out in 2017. This study may help clarify the intrinsic geographical relatedness of DENV-2 and contribute to further research on pathogenicity and vaccine development.

Background: Xishuangbanna, a border area of China, Burma and Laos, had its first major DENV-1 outbreak in 2017. This study aims to explore the genetic characterization, potential source and evolution of the viruses in outbreak., Methods: The structural protein C/prM/E genes of viruses isolated from local residents or Burmese travelers were sequenced followed by mutation, phylogenetic, homologous recombination, molecular clock and demographic reconstruction analysis., Results: Phylogenetic analysis revealed that all of the strains were classified as three cluster of DENV-1. Cluster 1, 2 and 3 were most similar to China Guangzhou 2011, China Hubei 2014 and Laos 2008 strain, respectively. Among 236 base mutations, 31 caused nonsynonymous mutations when compared with the DENV-1SS. No homologous recombination signal was discovered. The structural protein of these strains had similar three-dimensional structure. Only site 434 showed differences among five predicted protein binding sites. Molecular clock phylogenetic and demographic reconstruction analysis showed that DENV-1 became highly diversified in 1972 followed by a slightly decreased period until 2017., Conclusions: Dengue isolated strains show diversification between Burma and China. Amino acid substitution (I440T) may lead to weakened virulence of the epidemic strains. DENV-1 became highly diversified in 1972 followed by a slightly decreased period., (Copyright © 2018 The Authors. Published by Elsevier Ltd.. All rights reserved.)

In 2015, a dengue outbreak with 1,067 reported cases occurred in Xishuangbanna, a city in China that borders Burma and Laos. To characterize the virus, the complete genome sequence was obtained and phylogenetic, mutation, substitution and recombinant analyses were performed. DENV-NS1 positive serum samples were collected from dengue fever patients, and complete genome sequences were obtained through RT-qPCR from these serum samples. Phylogenetic trees were then constructed by maximum likelihood phylogeny test (MEGA7.0), followed by analysis of nucleotide mutation and amino acid substitution. The recombination events among DENVs were also analyzed by RDP4 package. The diversity analysis of secondary structure for translated viral proteins was also performed. The complete genome sequences of four amplified viruses (YNXJ10, YNXJ12, YNXJ13, and YNXJ16) were 10,742, 10,742, 10,741, and 10,734 nucleotides in length, and phylogenetic analysis classified the viruses as cosmopolitan genotype of DENV-2. All viruses were close to DENV Singapore 2013 (KX380828.1) and the DENV China 2013 (KF479233.1). In comparison to DENV-2SS (M29095), the total numbers of base substitutions were 712 nt (YNXJ10), 809 nt (YNXJ12), 772 nt (YNXJ13), and 841 nt (YNXJ16), resulting in 109, 171, 130, and 180 amino acid substitutions in translated regions, respectively. In addition, compared with KX380828.1, there were 44, 105, 64, and 116 amino acid substitutions in translated regions, respectively. The highest mutation rate occurred in the prM region, and the lowest mutation rate occurred in the NS4B region. Most of the recombination events occurred in the prM, E and NS2B/3 regions, which corresponded with the mutation frequency of the related portion. Secondary structure prediction within the 3,391 amino acids of DENV structural proteins showed there were 7 new possible nucleotide-binding sites and 6 lost sites compared to DENV-2SS. In addition, 41 distinct amino acid changes were found in the helix regions, although the distribution of the exposed and buried regions changed only slightly. Our findings may help to understand the intrinsic geographical relatedness of DENV-2 and contributes to the understanding of viral evolution and its impact on the epidemic potential and pathogenicity of DENV.

A dengue outbreak abruptly occurred at the border of China, Myanmar, and Laos in June 2017. By November 3rd 2017, 1184 infected individuals were confirmed as NS1-positivein Xishuangbanna, a city located at the border. To verify the causative agent, complete genome information was obtained through PCR and sequencing based on the viral RNAs extracted from patient samples. Phylogenetic trees were constructed by the maximum likelihood method (MEGA 6.0). Nucleotide and amino acid substitutions were analyzed by BioEdit, followed by RNA secondary structure prediction of untranslated regions (UTRs) and protein secondary structure prediction in coding sequences (CDSs). Strains YN2, YN17741, and YN176272 were isolated from local residents. Stains MY21 and MY22 were isolated from Burmese travelers. The complete genome sequences of the five isolates were 10,735 nucleotides in length. Phylogenetic analysis classified all five isolates as genotype I of DENV-1, while isolates of local residents and Burmese travelers belonged to different branches. The three locally isolates were most similar to the Dongguan strain in 2011, and the other two isolates from Burmese travelers were most similar to the Laos strain in 2008. Twenty-four amino acid substitutions were important in eight evolutionary tree branches. Comparison with DENV-1SS revealed 658 base substitutions in the local isolates, except for two mutations exclusive to YN17741, resulting in 87 synonymous mutations. Compared with the local isolates, 52 amino acid mutations occurred in the CDS of two isolates from Burmese travelers. Comparing MY21 with MY22, 17 amino acid mutations were observed, all these mutations occurred in the CDS of non-structured proteins (two in NS1, 10 in NS2, two in NS3, three in NS5). Secondary structure prediction revealed 46 changes in the potential nucleotide and protein binding sites of the CDSs in local isolates. RNA secondary structure prediction also showed base changes in the 3'UTR of local isolates, leading to two significant changes in the RNA secondary structure. To our knowledge, this study is the first complete genome analysis of isolates from the 2017 dengue outbreak that occurred at the border areas of China, Burma, and Laos.

In the past few decades, dengue has spread rapidly and is an emerging disease in China. An unexpected dengue outbreak occurred in Xishuangbanna, Yunnan, China, resulting in 1331 patients in 2013. In order to obtain the complete genome information and perform mutation and evolutionary analysis of causative agent related to this largest outbreak of dengue fever. The viruses were isolated by cell culture and evaluated by genome sequence analysis. Phylogenetic trees were then constructed by Neighbor-Joining methods (MEGA6.0), followed by analysis of nucleotide mutation and amino acid substitution. The analysis of the diversity of secondary structure for E and NS1 protein were also performed. Then selection pressures acting on the coding sequences were estimated by PAML software. The complete genome sequences of two isolated strains (YNSW1, YNSW2) were 10,710 and 10,702 nucleotides in length, respectively. Phylogenetic analysis revealed both strain were classified as genotype II of DENV-3. The results indicated that both isolated strains of Xishuangbanna in 2013 and Laos 2013 stains (KF816161.1, KF816158.1, LC147061.1, LC147059.1, KF816162.1) were most similar to Bangladesh (AY496873.2) in 2002. After comparing with the DENV-3SS (H87) 62 amino acid substitutions were identified in translated regions, and 38 amino acid substitutions were identified in translated regions compared with DENV-3 genotype II stains Bangladesh (AY496873.2). 27(YNSW1) or 28(YNSW2) single nucleotide changes were observed in structural protein sequences with 7(YNSW1) or 8(YNSW2) non-synonymous mutations compared with AY496873.2. Of them, 4 non-synonymous mutations were identified in E protein sequences with (2 in the β-sheet, 2 in the coil). Meanwhile, 117(YNSW1) or 115 (YNSW2) single nucleotide changes were observed in non-structural protein sequences with 31(YNSW1) or 30 (YNSW2) non-synonymous mutations. Particularly, 14 single nucleotide changes were observed in NS1 sequences with 4/14 non-synonymous substitutions (4 in the coil). Selection pressure analysis revealed no positive selection in the amino acid sites of the genes encoding for structural and non-structural proteins. This study may help understand the intrinsic geographical relatedness of dengue virus 3 and contributes further to research on their infectivity, pathogenicity and vaccine development., (Copyright © 2017 Elsevier B.V. All rights reserved.)

Antibody dependent enhancement (ADE) of dengue virus (DENV) infection is identified as the main risk factor of severe Dengue diseases. Through opsonization by subneutralizing or non-neutralizing antibodies, DENV infection suppresses innate cell immunity to facilitate viral replication. However, it is largely unknown whether suppression of type-I IFN is necessary for a successful ADE infection. Here, we report that both DENV and DENV-ADE infection induce an early ISG (NOS2) expression through RLR-MAVS signalling axis independent of the IFNs signaling. Besides, DENV-ADE suppress this early antiviral response through increased autophagy formation rather than induction of IL-10 secretion. The early induced autophagic proteins ATG5-ATG12 participate in suppression of MAVS mediated ISGs induction. Our findings suggest a mechanism for DENV to evade the early antiviral response before IFN signalling activation. Altogether, these results add knowledge about the complexity of ADE infection and contribute further to research on therapeutic strategies.

Background: Dengue poses a large burden on the public health systems worldwide. severe dengue (SD) could lead to more serious clinical symptoms and even death. This study aimed to identify the cause of SD in a clinical trial during the dengue outbreak in Xishuangbanna in 2019, and could provide new insights into the pathogenic mechanisms of SD. Methods: Mosquito-borne viral (DENV, JEV, and CHIKV) infections were identified. The epidemiological factors and clinical symptoms of inpatients in Xishuangbanna were recorded. The IgG and IgM levels in the serum of dengue inpatients were evaluated, and secondary infections were identified. Then, the structural proteins (C/PrM/E) were sequenced and compared with those of the same type of DENV in the same area as before, and their structures were predicted by the SWISS-MODEL (expasy.org). The full-length viral genomes were sequenced and aligned with representative strains by BioEidt or MEGA 5.0. Results: In this outbreak, the clinical symptoms were more serious in SD. The proportion of SD inpatients of male and Han nationality was larger than that of dengue fever (DF) inpatients (p < 0.05). DENV-2 infection was the majority in DF, with 45 inpatients. However, DENV-1 infection was the most common SD, with 54 inpatients. There were 3 DENV-3-positive inpatients in the DF group and 6 ZIKV-positive inpatients in the SD group. A secondary infection accounted for 76.47% (78 cases) of SD inpatients, but secondary infections were only in 20% (17 cases) of DF inpatients. In the three-dimensional structure of protein analysis, the C/PrM/E of DENV-1 and DENV-2 showed more stability than previous epidemic strains, while DENV-3 in 2019 showed a looser spatial structure. After a complete genome sequencing and analysis, all six DENV-2 strains belonged to cosmopolitan, five of which clustered into one branch. The GC/AT of the five strains decreased from 2014 to 2018. Compared with DF strains, SD strains had no mutations of commonness. Conclusions: SD may related to secondary heteromorphic dengue in Xishuangbanna in 2019. The coinfection of ZIKV could be another related factor for SD. The currently datas were very limited and only suggestive. [ABSTRACT FROM AUTHOR]

Keywords: severe dengue fever; IgG; IgM; dengue inpatients; dengue gene sequence EN severe dengue fever IgG IgM dengue inpatients dengue gene sequence 1 3 3 06/23/22 20220620 NES 220620 In the original article, the reference for " B The genome encodes a polyprotein, which is processed into three structural proteins [the capsid (C), premembrane (prM), and envelope (E) protein] and seven non-structural proteins (NS1-NS5) (Langerak et al., 2019) b " was incorrectly written as "Langerak, T., Mumtaz, N., Tolk, V. I., van Gorp, E. C. M., Martina, B. E., Rockx, B., et al. (2019). IgG, IgM, severe dengue fever, dengue inpatients, dengue gene sequence Phenotypic and genotypic characterization of dengue virus isolates differentiates dengue fever and dengue hemorrhagic fever from dengue shock syndrome. 10.1038/nrmicro303023652323 10 Tuiskunen, A, Monteil, V, Plumet, S, Boubis, L, Wahlstr?m, M, Duong, V, Phenotypic and genotypic characterization of dengue virus isolates differentiates dengue fever and dengue hemorrhagic fever from dengue shock syndrome. [Extracted from the article]

• A large-scale dengue fever outbreak occurred in the China, Myanmar, and Laos border area in 2019. • Three dengue virus (DENV) serotypes were found to be co-circulating, which is very rare in this area. • DENV-1 and DENV-2 were the main pathogenic agents of this outbreak. • The co-circulation may have led to more serious clinical symptoms and a larger scale epidemic. Dengue fever was included in the top 10 global health threats announced by the World Health Organization (WHO) in early 2019. In some southern provinces of China, autochthonous outbreaks have also been reported over the last decade. An unexpected large outbreak of dengue fever was reported in Xishuangbanna, a border area of China, Myanmar, and Laos, in 2019. Among the 226 hospitalized cases, 90 were diagnosed as severe dengue according to the 2009 WHO guidelines. Serotyping and phylogenetic analyses of envelope gene sequences from 246 randomly selected samples showed that three serotypes of dengue virus were co-circulating in this outbreak, which is very rare in this area. Dengue virus serotype 1 (DENV-1, genotype I) and serotype 2 (DENV-2, Cosmopolitan genotype and Asian genotype) were the main pathogenic agents of this outbreak. Dengue virus serotype 3 (DENV-3) epidemic strains were classified as genotype III and formed a close cluster with the Thailand 2015 epidemic strain. The co-circulation may have led to more serious clinical symptoms and a larger scale epidemic. This finding is of great importance in understanding the circulation of DENV and to strengthen the detection and management of dengue fever in border areas. [ABSTRACT FROM AUTHOR]

To elucidate the differences in microRNAs during dengue virus infection between Vero cell-adapted strain (DENV-2-Vero) and its source, the clinical C6/36 isolated strain (DENV-2-C6/36), a comparison analysis was performed in Vero cells by high throughput sequencing. The results showed that the expression of 16 known and 3 novel miRNAs exhibited marked differences. 5 known miRNAs were up-regulated in DENV-2-C6/36 group, while 11 known microRNAs were down-regulated in DENV-2-Vero group. The GO enrichment and KEGG pathway analysis showed that there was a distinct difference in regulating viral replication between two strains. In DENV-2-Vero infection group, significantly enriched GO terms included virion attachment to host cells, viral structural protein/genome processing and packaging. Meanwhile, the regulation of cell death and apoptosis between two groups were different in the early stage of infection. KEGG enrichment analysis showed that DENV-2-C6/36 infection induced more intense regulation of immune-related pathways, including Fc gamma R-mediated phagocytosis, etc. DENV-2-Vero infection could partially alleviate the immune defense of Vero cells compared with DENV-2-C6/36. The results indicated that the distinct microRNA changes induced by two DENV-2 strains may be partly related to their infective abilities. Our data provide useful insights that help elucidate the host-pathogen interactions following DENV infection. [ABSTRACT FROM AUTHOR]