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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. [ABSTRACT FROM AUTHOR]

• This is the first complete-genome analysis of dengue epidemic strains from Hunan. • Epidemic DENV-2 strains were closest to the Zhejiang strain in 2017. • The PrM/M region had the highest AA mutation rate, while NS4B was conservative. • Three amino acid mutations may important for replication and virulence of DENV-2. 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: N390thS, and NS5: S676thN, K800thT) 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. [ABSTRACT FROM AUTHOR]