BHK-21 cells, cytopathic effects (CPE) were detected, and flavivirus primers were amplified as positive. After the complete sequence of the virus was determined and spliced, a 10 840 nucleotide long sequence was obtained, encoding 3 432 amino acids. Phylogenetic analysis based on the whole gene sequence and E gene sequence showed that: The newly isolated SJM23-22 was most closely related to the GⅠa strain (C081) in Cambodia, with 98.5% nucleotide homology and 99.8% amino acid homology, while the homology with other genotypes was below 90% for nucleotides and below 98% for amino acids. The results of site analysis revealed 22 amino acid difference sites on the E gene compared to the live attenuated vaccine strain SA14-14-2, with 7 differences at 8 neurovirulence-related key amino acid sites. The results of important epitopes analysis indicated an exact match in three important epitopes in domain Ⅲ between the Shuangjiang isolates and the live attenuated vaccine strains. The results of secondary structure and tertiary structure prediction showed that the strain was characterized by random curling. Conclusions One strain of GⅠa-type JEV was isolated from Culex tritaeniorhynchus in Shuangjiang County, with no significant changes in the key amino acid sites related to antigenic epitopes. This study enriches the virus-carrying situation of Culex tritaeniorhynchus in Shuangjiang County, Yunnan Province, providing a reference for the prevention and control of the insect-borne epidemic in the province.Objective　To investigate the status and molecular characteristics of Japanese encephalitis virus (JEV) carried by mosquitoes in Shuangjiang County, Lincang City, Yunnan Province. Methods Mosquito specimens were collected from cattle pens using mosquito traps in Shuangjiang County, Lincang City in August 2023. After mosquito species identification, BHK-21 cells and C6/36 cells were used in one group of 25 mosquitoes each. Positive isolates were identified by flavivirus primers. Subsequently, the full-length GⅠ-type JEV was amplified using 15 pairs of primers with RT-PCR, sequenced, and spliced, and sequence analysis was performed using bioinformatics software such as MEGA X, DNAstar, GeneDoc, SOPMA, and SWISS-MODEL. Results A total of 1 300 Culex tritaeniorhynchus were collected and divided into 52 groups for virus isolation, leading to the identification of one positive isolate (SJM23-22). After inoculation with C6/36 and

Objective To isolate the Japanese encephalitis virus carried by Culex tritaeniorhynchus in Dongchuan District of Yunnan Province and analyze its molecular characteristics, so as to provide insights into the prevention and control of Japanese encephalitis in Yunnan Province. Methods Mosquito specimens were collected using mosquito-trapping lamps from pig farms in Batang Village and Xiaoxin Village, Dongchuan District, Kunming City, Yunnan Province in July 2016, and the mosquito species was identified according to the mosquito morphology. Then, 60 to 100 mosquitoes of each species served as a group and were ground. Baby hamster kidney-21 (BHK-21) cells and Aedes albopictus clone C6/36 cells were used for virus isolation, and positive isolates were identified using flavivirus primers. The positive isolates were amplified using reverse transcription polymerase chain reaction (RT-PCR) assay with 15 pairs of specific primers covering the full length of the genotype I Japanese encephalitis virus, and DNA sequence assembly was performed using the software SeqMan in the DNASTAR package. The obtained sequences were aligned with the complete sequences of 38 Japanese encephalitis virus downloaded from the GenBank with the software MegAlign, and the nucleotide and amino acid homology analyses of the obtained sequences were performed. The difference in amino acid sites was analyzed with the software GeneDoc, and phylogenetic trees were created based on the sequences of the coding region and E protein of the isolated Japanese encephalitis virus with the software Mega X. In addition, the secondary and tertiary structures of the E protein of the Japanese encephalitis virus were predicted using the online tool SOPMA and the software Swiss-Model. Results A total of 5 820 mosquitoes were collected and 3 843 Cx. tritaeniorhynchus (66.03%) were identified according to the mosquito morphology. A positive virus isolate, termed YNDC55-33, was isolated from Cx. tritaeniorhynchoides following batches of virus isolation from mosquito specimens, and cytopathic effect was observed following inoculation into BHK-21 and C6/36 cells. The YNDC55-33 virus isolate was successfully amplified with the flavivirus primes, and a long sequence containing 300 nucleotides was obtained. Following sequence alignment using the BLAST tool, the sequence of the YNDC55-33 virus isolate had high homology with that of the genotype I Japanese encephalitis virus. A long sequence with 10 845 nucleotides in length, which encoded 3 432 amino acids, was obtained by splicing the full sequence of the YNDC55-33 virus isolate. Phylogenetic analysis based on the whole-genome sequence and E gene sequence of the YNDC55-33 virus isolate showed that the new YNDC55-33 virus isolate was most closely related to the genotype I Guizhou isolate (GenBank accession number: HM366552), with nucleotide homology of 98.5% and amino acid homology of 99.4%, and the YNDC55-33 virus isolate shared 97.96% ± 0.33% nucleotide homology and 99.35% ± 0.08% amino acid homology with other genotype I Japanese encephalitis virus isolates, and < 90% nucleotide homology and < 98% amino acid homology with other genotypes of Japanese encephalitis virus. The YNDC55-33 virus isolate and the live attenuated virus vaccine candidate SA14-14-2 isolate differed at 16 amino acid sites on E gene, and 7 out of 8 key amino acid sites related to neurovirulence. The secondary and tertiary structures of the E protein of the YNDC55-33 virus isolate were predicted to be characterized by random coils. Conclusions A genotype I Japanese encephalitis virus was isolated from Cx. tritaeniorhynchus in Dongchuan District, Kunming City. This virus isolate and the live attenuated virus vaccine candidate SA14-14-2 isolate does not differ at antigenic epitopes-related key amino acid sites, and the major protein structure of the virus isolate is random coils. This study adds new data for the epidemiological distribution of Japanese encephalitis virus in Yunnan Province, which may provide insights into the prevention and control of Japanese encephalitis in the province.

Objective To evaluate the curative effect of relaxing music applied in complex wisdom teeth extraction.Methods A total of 200 patients with complex wisdom teeth were selected and divided into A,B groups,relaxing music was applied in group A 30 minutes before and during teeth extraction,while group B underwent teeth extraction directly.Beck Anxiety Inventory (BAI) and Visual Analogue Scale (VAS) were selected as the measuring tools of anxiety and pain intensity.Parameters including blood pressure (BP),heart rate (HR) and respiratory rate (RR) were measured before relaxing music application and after.Results After informed their disease condition,the score of BAI of group A and B was (52.18 ± 10.75),(52.41 ± 14.08),respectively,and no significant difference was found (t =0.13,P ＞ 0.05).While 30 minutes after they listened to relaxing music,the score of BAI of group A and B was (38.24 ±6.59) and (54.12 ±9.95),and group A was better than group B,and the difference was statistically significant (t =13.31,P ＜ 0.05).After the operation,the score of VAS in group A was significantly better than that in group B [(1.96 ± 0.84) vs (3.42 ±0.91) ;t=11.79,P＜0.05)].Conclusions Relaxing music could effectively relieve the anxiety and uncomfortable from complex wisdom teeth extraction.