Chronic liver disease is a general term for a variety of liver diseases， and its prevalence rate is increasing year by year. With the progression of the disease， patients may experience a variety of serious complications and even progress to liver failure. In recent years， a number of studies have revealed the association between ectopically colonized oral bacteria and chronic liver disease and explored their potential value in the diagnosis and treatment of chronic liver disease. This article systematically reviews the association of ectopically colonized oral bacteria with metabolic associated fatty liver disease， liver cirrhosis， and liver cancer and analyzes the mechanism for the influence of ectopically colonized oral bacteria on chronic liver disease， so as to provide a reference for the diagnosis and treatment of chronic liver disease using ectopically colonized oral bacteria.

ObjectiveTo evaluate the clinical efficacy of Zuogui Pill （左归丸，ZGP） decoction on neurological function， motor function， and activities of daily living in patients with ischemic stroke （IS） of liver-kidney yin deficiency syndrome， and to assess its potential mechanisms from the perspectives of cerebral perfusion and corticospinal tract （CST） remodeling. MethodsA total of 108 patients with IS of liver-kidney yin deficiency syndrome and onset time ≤14 days were enrolled and randomly divided into a control group and a treatment group， with 54 cases in each group. The control group received conventional western medicine， while the treatment group received ZGP decoction （one dose per day） in addition to the conventional treatment. Both groups were treated for 90 days. The National Institutes of Health Stroke Scale （NIHSS）， modified Rankin Scale （mRS）， modified Barthel Index （MBI）， Fugl-Meyer Assessment （FMA）， and Chinese medicine syndrome score （CMS） were evaluated before treatment and after （90±3） days of treatment. Three-dimensional arterial spin labeling （3D-ASL） and diffusion tensor imaging （DTI） were performed to obtain ipsilesional cerebral blood flow （CBF） in the unaffected and affected hemispheres， relative CBF （rCBF）， and fractional anisotropy （FA） of the CST. Nodal analysis of FA values along the CST was also conducted. Correlation analysis was used to assess the relationships between post-treatment cerebral perfusion parameters （CBF and rCBF）， and FA values with clinical neurological function scores. ResultsAfter treatment， both groups showed significantly decreased NIHSS， mRS and CMS scores， together with significantly increased FMA and MBI scores （P＜0.01）. Intergroup comparison showed that NIHSS， mRS， and CMS scores in the treatment group were significantly lower than those in the control group ， while MBI and FMA scores were significantly higher （P＜0.01）. The 3D-ASL results revealed that CBF and rCBF values in the affected hemisphere significantly increased in both groups after treatment， and the treatment group had significantly higher values （P＜0.01）. DTI showed that the FA value of the CST on the affected side in the treatment group was significantly higher than that in the control group after treatment （P＜0.05）. Nodal analysis along the CST further demonstrated that the continuity of FA values at lesion-related nodes 45~99 in the treatment group was significantly better than that in the control group （P＜0.01）， with significant increases at nodes 65~71 and 76~81 compared with baseline （P＜0.05 or P＜0.01）， whereas the control group showed a decreasing trend at nodes 80~93 （P＜0.05）. Correlation analysis indicated that CBF and rCBF values in the affected hemisphere were negatively correlated with NIHSS and mRS scores and positively correlated with MBI score （P＜0.05）. FA value of the CST on the affected side， both before and after treatment， were negatively correlated with NIHSS score and positively correlated with FMA score （P＜0.05 or P＜0.01）. ConclusionZGP decoction can effectively improve neurological function， motor function， and activities of daily living in IS patients with liver-kidney yin deficiency syndrome， and its mechanism may be related to enhanced cerebral perfusion in the affected hemisphere and corticospinal tract structural remodeling.

Stresses induced by the deficiency or excess of trace mineral elements, such as manganese (Mn), represent a common limiting factor for the production of crops like Brassica napus. To identify key genes involved in Mn allocation in B. napus and elucidate the underlying mechanisms, a member of the metal tolerance protein (MTP) family obtained in the previous screening of cDNA library of B. napus under Mn stress was selected as the research subject. Based on the sequence information and phylogenetic analysis, it was named as BnMTP10. It belongs to the Mn-cation diffusion facilitator (CDF) subfamily. Expression of BnMTP10 in yeast significantly improved the tolerance of transformants to excessive Mn and iron (Fe) and reduced the accumulation of Mn and Fe. However, the yeast transformants exhibited no significant changes in tolerance to excess cadmium, boron, aluminum, zinc, or copper. The qRT-PCR results demonstrated that the flowers of B. napus had the highest expression of BnMTP10, followed by roots and leaves. Subcellular localization studies revealed that BnMTP10 was localized in the endoplasmic reticulum (ER). Compared with wild-type plants, transgenic Arabidopsis overexpressing BnMTP10 exhibited enhanced tolerance to excessive Mn stress but showed no significant difference under Fe stress. Correspondingly, under excessive Mn stress, the Mn content in the roots of transgenic Arabidopsis increased significantly. However, under excessive Fe stress, the Fe content in transgenic Arabidopsis did not alter significantly. According to the results, we hypothesize that BnMTP10 may alleviate excessive Mn stress in plants by mediating Mn transport to the ER. This study facilitated our understanding of efficient mineral nutrients, and provided theoretical foundations and gene resources for breeding B. napus.
Brassica napus/genetics* ; Manganese/metabolism* ; Plants, Genetically Modified/genetics* ; Plant Proteins/physiology* ; Arabidopsis/metabolism* ; Gene Expression Regulation, Plant ; Phylogeny ; Cation Transport Proteins/metabolism* ; Stress, Physiological

Objective:To explore the high-risk factors for parenteral nutrition associated cholestasis（PNAC）in extremely/ultra-low birth weight infants，and establish a risk Alignment Diagram prediction model.Methods:We retrospectivly analyzed the clinical data of hospitalized extremely/ultra-low birth weight infants admitted to Neonatology Department at Quanzhou Children's Hospital from January 2019 to December 2020，using multivariate Logistic regression analysis to screen for independent risk factors for the occurrence of PNAC.An Alignment Diagram model prediction model for PNAC was constructed by using R software，and the performance of the model was evaluated through receiver operating characteristic curves.Results:A total of 203 extremely/ultra-low birth weight infants were included，with a median gestational age of 29.14（28.00，30.86）weeks and a median birth weight of 1　170（1　000,1　300）g.Among them，26（12.81%）cases developed PNAC.Multivariate Logistic regression analysis showed that the duration of parenteral nutrition（ OR=1.015 ，95% CI 1.003-1.034），the cumulative amount of glucose（ OR=1.014 ，95% CI 1.001-1.028），small for gestational age（ OR=3.455 ，95% CI 1.127-10.589），and neonatal sepsis（ OR=3.142 ，95% CI 1.039-9.503）were independent risk factors for PNAC（ P＜0.05）；The four independent risk factors mentioned above were introduced into R software to construct an Alignment Diagram model，the area under the receiver operating characteristic curve was 0.835（95% CI 0.842-0.731），and the results of the Hosmer Limeshow goodness of fit test show that：χ 2=5.34，degree of freedom=8， P=0.72.A calibration curve indicated good consistency between the predicted probability of the model and the actual occurrence rate，with good accuracy. Conclusion:The Alignment Diagram model constructed based on four independent risk factors of the duration of parenteral nutrition，glucose accumulation，small for gestational age infants，and neonatal sepsis exhibits high predictive ability，and is expected to provide an intuitive and convenient visualization tool for preventing or reducing the occurrence of PNAC in extremely/ultra-low birth weight infants

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.

Primary central nervous system lymphoma (PCNSL) is a highly aggressive malignant lymphoma. As most chemotherapy drugs have difficulty in crossing the blood-brain barrier, PCNSL shows a difficulty in clinical treatment, a high recurrence rate and a poor prognosis. Early identification of relapsed patients and prompt initiation of salvage therapy play a critical role in the improvement of patients' prognosis. Brain biopsy is the gold standard to identify recurrence, while the risk of operation failure and complications is still high. Non-invasive imaging techniques are beneficial for early identification of recurrence in PCNSL and can provide an important basis for guiding relapsed patients to adjust treatment plans in time. However, there is no unified evaluation standard for imaging methods of monitoring the relapsed lesions of PCNSL. With the further research of the pathophysiological mechanism of PCNSL, biomarker detection has become a new method to identify recurrence and more clinical evidence is still needed in the future.

Objective:To understand the prevalence of arboviruses in mosquito samples in Xichang City, Sichuan Province, and enrich the data of arbovirus activity and genetic characteristics in southwestern Sichuan Province.Methods:In June 2018, the nucleic acid was extracted from Culex tritaeniorhynchus mosquitoes collected from different pigsties in three villages and suburbs of Xichang City. The specific primers of Yunnan orbivirus, Banna virus, Tibet orbivirus (S7, S10), Flavivirus and alphavirus were used for quantitative polymerase chain reaction examination, and the positive product was cloned for sequencing analysis. Results:A total of 9 012 mosquitoes were collected, of which Cx. tritaeniorhynchus was the dominant species. A number of 88 batches of these mosquitoes were amplified, and 2 strains of Japanese encephalitis virus (JEV), 7 strains of Banna virus (BAV), 7 strains of Tibet orbivirus (TIBOV) and 1 strain of Yunnan orbivirus virus (YOUV) were detected, respectively. By the results of cluster analysis and evolutionary tree analysis, the 17 newly found virus strains were close to the Yunnan isolates, and 2 JEV strains were located in the GI-b clade. The other 7 strains of BAV were A2 evolutionary clades. Of the 7 TIBOV plants, 6 were located in the same clade. One TOUV was in the same clade as the Yunnan strain. Conclusions:Culex tritaeniorhynchus mosquitoes in Xichang city might carry JEV, BAV, YOUV and TIBOV, among them JEV was GI-b type and BAV was A2 type. The results provide data supporting the detection and analysis of arboviruses in Xichang city.

Objective To preliminarily understand the infection of Chikungunya virus (CHIKV) in Cangyuan County, a southern border area of Yunnan Province, and provide a reference basis for the prevention and control of Chikungunya fever. Methods In April 2020, a total of 400 serum samples from individuals seeking medical care at the People's Hospital of Cangyuan County in Yunnan Province were collected. Among these, 121 samples were from healthy individuals undergoing physical examinations, and 279 samples were from patients with fever. The serum samples collected underwent CHIKV neutralizing antibody testing using a serum micro-neutralization assay. Real-time fluorescence quantitative RT-PCR was used to detect CHIKV nucleic acid in the samples, followed by analysis of the test results. Results The results of neutralizing antibodies showed that 18 of the 400 human serum samples were positive for neutralizing antibodies against CHIKV, with an overall positivity rate for serum samples of 4.5% (18/400). Among the 279 serum samples collected from patients with fever, 18 were positive for neutralizing antibodies against CHIKV, with a positive rate of 6.45% (18/279), and the neutralizing antibody titers ranged from 1∶10 to 1∶320. The results of 121 healthy human serum samples were negative for neutralizing antibodies against CHIKV. The results of real-time fluorescence quantitative RT-PCR showed that 3 of the 400 human serum samples were positive for CHIKV nucleic acid, and the positive rate was 0.75% (3/400). Among the 279 serum samples collected from patients with fever, 3 samples were positive for CHIKV nucleic acid, with a positive rate of 1.08% (3/279), and Ct values ranged from 36.58 to 37.74. While all healthy human serum samples were negative for CHIKV nucleic acid. Conclusions The findings indicate that infection of CHIKV exists in the population of Cangyuan County, a southern border area of Yunnan Province, and an outbreak of the disease is occurring. Therefore, it is necessary to strengthen the monitoring, prevention, and control of CHIKV in this area.

Objective This study aims to comprehensively investigate the molecular characteristics of the predominant circulating Dengue virus type 1 (DENV-1) during the 2019 Dengue fever outbreak in Xishuangbanna, providing an essential insight to support the prevention and control of dengue fever in the local area. Methods A Dengue virus type 1 (DENV-1) strain, designated as JHS45, isolated from the blood of a febrile patient in Xishuangbanna in 2019, underwent a process of inoculation and cultivation in C6/36 cells. Second-generation sequencing was employed to capture the viral genetic sequence. Bioinformatics software, including CLC, was used for assembling the sequencing data. Sequentially, sequence alignment, construction of a phylogenetic tree, and analysis of amino acid sites were conducted using software such as Lasergene and MEGA6.1. Results Cytopathic effects of JHS45 appeared in C6/36 cells after 6 days. After sequencing and assembly, a 10 687-nucleotide (nt) long sequence of the JHS45 virus was obtained (GenBank accession number: OR593353). Genetic evolutionary analysis revealed that the JHS45 virus formed an evolutionary branch with DENV-1 genotype I viruses prevalent in Xishuangbanna, Guangzhou, Henan, and Zhejiang in China during 2019, as well as the DENV-1 genotype I virus prevalent in Thailand in 2013, with nucleotide homology of 97.6% to 99.9% and amino acid homology of 99.1% to 100%. Further analysis revealed that the JHS45 strain shared a smaller evolutionary branch with the DENV-1 genotype I viruses prevalent in Xishuangbanna (MW386863) and Guangzhou (MW261839) in 2019, showing the highest homology with nucleotide and amino acid homology of 99.9% and 100%, respectively. Amino acid differential site analysis between the JHS45 strain and the DENV-1 prevalent in Xishuangbanna since 2015 revealed 40 amino acid differential sites in the coding region of the JHS45 virus, primarily concentrated in the NS3 and NS5 regions of non-structural proteins. Conclusion The comprehensive analysis of the JHS45 strain's whole genome sequence indicates it is a DENV-1 genotype I virus. The genetic evolutionary relationship between this Xishuangbanna dengue fever outbreak is closely related to the prevalent virus strains in Xishuangbanna, Guangzhou, Henan, and Zhejiang. These findings provide a robust scientific foundation for monitoring dengue fever outbreaks, conducting virus evolution studies, and shaping effective prevention and control strategies, not only within Yunnan Province but also on a broader scale throughout China.

Objective:To study the prevalence of Akabane virus (AKAV) in Yunnan province.Methods:A group of sentinel animals including 5 goats and 10 cattles which were sero-negative for bluetongue virus and AKAV were located in Mangshi, Yunnan province, to monitor arbovirus activity from April to October 2015. The heparin-anticoagulated blood of the animals was collected weekly for arbovirus isolation. Erythrocytes were lysed in distilied water and inoculated onto BHK-21 monolayer for virus isolation. After the cells showed cytopathic effect (CPE), AKAV in the cell cuture were identified by RT-PCR amplification with AKAV S gene segment specific primers and gene sequencing.Results:The result indicated that BHK-21 cell inoculated with the blood from one sentinel goat showed CPE 48 h post inoculation. One day old suckling mice intracerebrally inoculated with the supernatant of the cell culture, they started to become sick and died after 3 days. RT-PCR identification and S gene sequencing showed that the isolate was AKAV (numbered as 16415). The full length of S segment gene of 16415 is 856 nt, encoding 233 amino acids of protein N, and 91 amino acids of protein Ns. Phylogenetic analysis showed that, among the AKAV strains isolated from China and abroad, the newly isolated 16415 was in the same branch with the other viruses in different geographical regions, and their S segment nucleotide sequence have a high homology of 83.8%-97.7%. The further study show that 16415 and the AKAV isolated from the domestic bamboo rat in Guangxi in 2013 have a closest realtionship in evolution, the nucleotide sequence homology was 97.7%, and amino acid homology is 99.6%. Compared with the Japanese OBE-1 strain, the amino acids of protein N of 16415 and the AKAVs, isolated from banboo rat in Guangxi and Anopheles vagas in Mangshi of Yunnan province of China, have two common diverse amino acids loci located at the 115th and 206th sites respectively. Conclusions:It is concluded that AKAV was newly isolated from goat in Mangshi of Yunnan province, which may have important epidemiological significance.