Objective To investigate T cell subtypes and their functions in liver immune microenvironments among patients with alveolar echinococcosis (AE) using single-cell RNA sequencing (scRNA-seq). Methods Four AE patients that were admitted to Qinghai Provincial People’s Hospital in 2023 for hepatic surgery for the first time were enrolled, and liver specimens were sampled 1 cm (peri-lesion, PL group) and > 5 cm from AE lesions (distal lesion, DL group) among each patient. Finally, a total of eight liver specimens were sampled from four AE patients for scRNA-seq analysis. Genome and transcriptome data of liver specimens were processed using the software Cell Ranger and R package. Differentially expressed genes (DEGs) and their biological functions were analyzed using gene ontology (GO) enrichment analysis and Kyoto encyclopedia of genes and genomes (KEGG) pathway analysis, and the primary intercellular communication patterns and interaction mechanisms were identified among T cell subtypes in liver specimens using the CellChat package. In addition, the developmental stages of T cells were subjected to trajectory analysis with the monocle package to investigate the expression of genes associated with cell growth and tumor transformation, and to predict the developmental trajectories of T cells. Results All four AE patients were female, with a mean age of (25.00 ± 9.06) years, and there were three cases from Jiuzhi County, Golog Tibetan Autonomous Prefecture and one case from Chengduo County, Yushu Tibetan Autonomous Prefecture, Qinghai Province. The viability of single-cell samples from eight liver specimens was 90.41% to 96.33%, and a total of 81 763 cells were analyzed, with 19 cell types annotated. Of these cell types, 13 were immune cells (87.60%), and T cells (33.13%), neutrophils (15.40%), and natural killer cells (11.92%) were the three most common cell types. Re-clustering of 27 752 T cells and proliferative T cells identified 10 distinct T cell subtypes, with CD8⁺ cytotoxic T cells (23.43%), CD8⁺ naive T cells (12.80%), and CD4⁺ effector memory T cells (17.73%) as dominant cell types. The proportions of T helper 2 (Th2) cells (5.19% vs. 3.63%; χ² = 38.35, P < 0.01) and CD4⁺ effector memory T cells (21.59% vs. 13.67%; χ² = 244.70, P < 0.01) were significantly higher in liver specimens in the PL group than in the DL group, and the proportion of CD4⁺ helper T cells was significantly lower in the PL group than in the DL group (7.50% vs. 14.75%; χ² = 330.52, P < 0.01). KEGG pathway analysis revealed that Th2 cells were significantly enriched in cell apoptosis and multiple cancer-associated pathways, and CD4⁺ effector memory T cells were significantly enriched in the regulation of cytokines and chronic inflammation, while CD4⁺ helper T cells were significantly enriched in immune responses regulation. Trajectory analysis of T cells showed that CD4⁺ helper T cells were at an earlier developmental stage relative to Th2 cells and CD4⁺ effector memory T cells, and the expression of inhibitor of DNA binding 3 (ID3), thioredoxin interacting protein (TXNIP), Bcl2-associated athanogene 3 (BAG3) and heat shock protein family B (small) member 1 (HSPB1) genes appeared a tendency towards a decline over time. Conclusions CD4⁺ effector memory T cells and CD8⁺ cytotoxic T cells are primary interacting cells in the liver specimens of AE patients. Reduced expression of Th2 cells and CD4⁺ helper T cells contributes to an inhibitory immune microenvironment, which promotes immune evasion by Echinococcus multilocularis, and Th2 cells are significantly enriched in multiple cancer-associated pathways, which may be linked to the invasive growth of E. multilocularis.

Objective:To analyze the genus, drug resistance/virulence and phylogenetic characteristics of Brucella strains isolated from brucellosis surveillance sentinels in Henan Province from 2013 to 2022, and provide baseline data for the surveillance, early warning and outbreak tracing of brucellosis. Methods:Blood samples were collected from patients with Brucella infection for strain isolation, culture and species identification, drug susceptibility test, whole genome sequencing, splicing and assembly, functional/virulence/resistance gene prediction analysis and phylogenetic tree drawing based on single nucleotide polymorphism (SNP). Results:In 36 brucellosis patients, the majority were men (86.11%, 31/36), young adults aged 18-50 (88.89%, 32/36) and farmers/herdsmen (72.22%, 26/36). A total of 36 strains of Brucella melitensis were isolated, and average 1 305 functional proteins of 21 categories were predicted by strain genome; all the strains carried four main virulence factors (pmm, VirB group, BtpA/BtpB, BvrS/BvrR). The drug sensitivity rate was 100.00% to six types of antibiotics including levofloxacin, rifampicin, doxycycline, streptomycin, tetracycline and gentamicin, they showed different resistances to three antibiotics including compound trimethoprim-sulfamethoxazole, ciprofloxacin and ampicillin. The strains carried four types of resistance genes and two clusters of resistance genes, with four combinations of genotypes, the resistance mechanisms included antibiotic degradation/modification enzymes, resistant nodular cell differentiation (RND) efflux pumps, 16S/23S ribosomal rRNA binding site mutations, etc. The number of SNP differed in the genomes of 36 Brucellamelitensis strains ranged from 0 to 454 and phylogenetic tree was divided into three major branches, with relative branch distances between 0.000 0 and 0.498 6 for each strain. Conclusions:Human Brucellamelitensis strains isolated from surveillance sentinels in Henan from 2013 to 2022 carried multiple virulence and antibiotic resistance genes and had different drug resistance phenotypes. Single nucleotide polymorphism analysis and phylogenetic tree analysis showed significant differences in phylogenetic relationships among different strains.

Objective:To analyze the genetic evolution and molecular characteristics of H7N9 avian influenza virus (AIV) isolated in a live poultry market.Methods:Samples such as poultry feces, sewage, and hair removal machine and chopping board swabs were collected. Real-time fluorescent quantitative PCR was used to detect influenza A virus and H7N9 AIV in the samples. The whole genome of H7N9 AIV was amplified with influenza A virus universal primers and sequenced. BLAST and MEGA X were used for sequence alignment, phylogenetic analysis and molecular characterization.Results:Seven poultry-related environment samples were collected in the live poultry market in Xuchang city in February 2023, and four were positive for H7N9 AIV. The whole genome sequences of three H7N9 AIV isolates were successfully obtained, and the isolates shared high nucleotide identity in different genes (98.37%-100.00%). BLAST analysis showed they were highly identical to H7N9 strains isolated from domestic poultry in China from 2020 to 2021. Genetic evolution analysis showed that the three isolates clustered in the same branch and were closer to the recent environmental isolates than to the recent strains isolated from human or avian. Through comparison with the sequences of the representative strains in different periods, it was found that the isolated strains in this study showed high avian pathogenicity with four amino acids KRAA inserted at the cleavage site; the hemagglutinin receptor-binding site was QSG, which was an avian binding receptor; there was a G186I mutation in hemagglutinin. Mammalian-adaptive mutation E627K was not detected in polymerase basic protein 2. Mutations (R292K and I38T) associated with drug resistance to neuraminidase inhibitor (oseltamivir) and polymerase acidic protein inhibitor (baloshavir) were not detected, suggesting that these isolates remained susceptible to these drugs. A S31N mutation was found in M2 protein, indicating they were resistant to alkamines.Conclusions:The three H7N9 AIV strains isolated in the live poultry market have high avian pathogenicity, but there are no significant increase in mutations related to the binding ability to human receptors, mammalian pathogenicity, viral transmissibility, or drug resistance as compared with previous representative strains causing human or avian infection.

Objective:To analyze the whole genome of Omicron variants causing the first local Omicron outbreak in Henan Province and to investigate the mutations in the SARS-CoV-2 genome for source tracing.Methods:Respiratory tract samples from COVID-19 cases in the Omicron outbreak in Henan Province from January 7 to 29, 2022 were subjected to whole-genome sequencing and sequence alignment analysis. Whole-genome identity, variations and evolution of the Omicron variants were analyzed.Results:Through high-throughput sequencing, the whole-genome sequences of SARS-CoV-2 were obtained from 120 cases, which accounted for 25.64% (120/468) of all COVID-19 cases in Anyang during the same period. Compared with the genome of Wuhan reference strain (NC_045512.2), there were 57-59 nucleotide mutation sites in the 120 whole genome sequences, and one or two nucleotide mutation sites were added to the shared 57 nucleotide sites. All of the 120 strains were VOC/Omicron (BA.1.1) variants and shared high homology. The whole-genome sequence obtained from the first case A contained 57 nucleotide mutation sites, while apart from the 57 identical nucleotide mutation sites, one specific mutation site (C1594T) was found in the whole-genome sequence obtained from the first case B, suggesting that the two cases were in the same transmission chain. After comparing with the database of domestic and imported cases by the Chinese Center for Disease Control and Prevention and the Henan Provincial Center for Disease Control and Prevention, it was found that the current outbreak was linked with the same transmission chain as the existing local epidemics in other provinces. Moreover, epidemiological investigation showed that on January 2, case A had come into contact with her cousin and his family who returned from an affected area outside the province.Conclusions:Based on the gene sequencing results and epidemiological investigation, the COVID-19 outbreak in Anyang city, Henan Province was a local epidemic and the source of it was a college student who returned to Anyang city from other province on December 28, 2021. These infections were linked to the same transmission chain as the existing local infection in other provinces.

Objective:To analyze the epidemiological characteristics and spatiotemporal characteristics of human brucellosis in Henan Province.Methods:Data of human brucellosis in Henan Province from 2005 to 2021 were collected through the China Disease Prevention and Control Information System, and a descriptive epidemiological method was used to analyze the epidemic profile of brucellosis in Henan Province and the three distribution characteristics. Global and local spatial autocorrelation were used to analyze the spatial distribution and the hot spots of brucellosis in Henan Province, respectively, and spatiotemporal scanning was used to analyze the spatiotemporal clustering regions of brucellosis in Henan Province.Results:A total of 39 862 brucellosis cases were reported in Henan Province from 2005 to 2021, with an average annual incidence of 2.44/100 000, and the number of cases showed an overall increasing trend each year (χ 2trend = 11 127.85, P = 0.001). The onset months were mainly concentrated from March to July, accounting for 59.00% (23 517/39 862), with May as the peak (5 478 cases). Cases of brucellosis were reported in 157 counties (cities, districts) of the province. The ratio of male to female was 2.52∶1.00 (28 542/11 320). Farmers were the main occupation, with 32 985 cases (82.75%). The age of onset was mainly 45 to 65 years old, with 20 226 cases (50.74%). The global spatial autocorrelation analysis showed that the global Moran's I was > 0, Z > 1.96, and P < 0.05 in all years except 2006 - 2008, showing spatial clustering. Further local spatial autocorrelation analysis was performed, and high-high and low-low clustering areas existed in 2012 - 2021 ( P < 0.01). Spatiotemporal scanning analysis showed that there was one spatiotemporal cluster in the high incidence area and two spatiotemporal clusters in the low incidence area. The high incidence cluster was centered in Neixiang County, covering 48 counties (cities, districts) including Song County and Ruzhou City, and the aggregation time was from 2014 to 2021. The two low incidence clusters were centered in Yongcheng City and Boai County, covering 58 and 18 counties (cities, districts), respectively, and the aggregation time was 2016 - 2021 and 2005 - 2012, respectively. Conclusion:The overall incidence of brucellosis in Henan Province is on the rise from 2005 to 2021, with middle-aged and elderly male farmers as the main affected population, and there are spatiotemporal clusters of brucellosis in Henan Province.

Objective:To analyze the genome characteristics and variations in nucleotides and amino acids of SARS-CoV-2 causing an outbreak in Henan Province in November 2021 and perform the traceability analysis.Methods:In this study, throat swab specimens from cases in the acute phase were collected and tested for the nucleic acids of SARS-CoV-2 by real-time fluorescent RT-PCR. SARS-CoV-2 nucleic acid-positive samples were subjected to high-throughput genome sequencing and whole-genome alignment analysis.Results:The median Ct values of ORF1ab gene and N gene in 70 positive specimens was 26.41 (15.58 to 39.27) and 24.43 (12.04 to 39.74), respectively. Compared with the sequence of Wuhan-Hu(NC_045512) reference strain, 47 to 49 nucleotide mutations sharing 47 nucleotide mutation and 41 amino acid mutations were found in 63 strains of successfully sequenced SARS-CoV-2. Nine nucleotide mutations and 12 amino acid mutations were found in the spike protein. The index case shared 47 mutations with the Russian imported cases in Henan Province on October 14 and the local cases in Jiangxi Province in October. Moreover, their genomes were highly homologous and they all belonged to the Delta variant (AY.122 evolutionary branch).Conclusions:Continuous monitoring of imported COVID-19 cases and prolonging the period of quarantine were needed to reduce the risk of local outbreak and epidemic caused by imported COVID-19 cases. Analysis of the genomic characteristics of SARS-CoV-2 and the variations in nucleotides and amino acids was conducive to trace the origin of COVID-19 outbreak quickly and provide reference for precise control.

Objective:To investigate the clinical characteristics of two Han families with familial vitreous amyloidosis (FVA) and the gene mutation.Methods:A pedigree investigation was performed.Two Han Chinese families with FVA treated in Xiangya Hospital of Central South University from January 2015 to December 2018 were collected.General examination and ophthalmic examination were performed among 112 members of the two families.Peripheral blood samples were collected from 32 family members (15 patients in MZ001 pedigree, 7 patients in MZ002 pedigree, and 5 persons with normal clinical phenotype from each pedigree) for DNA extraction, polymerase chain reaction (PCR) amplification, transthyretin ( TTR) gene screening and sequencing.Vitreous biopsy following three-channel 23-gauge pars plana vitrectomy was performed on the two probands in the two families.Vitreous specimens were sent for pathological examination.This study adhered to the Declaration of Helsinki.The study protocol was approved by an Ethics Committee of Xiangya Hospital of Central South University (No.201412463), and written informed consent was obtained from all subjects before any medical examination. Results:In MZ001, there were 15 cases of the 63 members presented bilateral vitreous opacity at an average age of (43.6±5.8) years.No lesion was found in nervous system, cardiovascular system, kidney or liver in general inspection.The vitreous of the proband (Ⅲ13) was so sticky that could not be totally removed during vitrectomy.The vitreous specimen showed positive Congo red staining.Ⅲ13 had elevated intraocular pressure after vitrectomy and was diagnosed as open-angle glaucoma.Gene sequencing revealed Gly83Arg mutation in the exon 3 of TTR gene.In MZ002, 7 cases of 49 members had bilateral vitreous opacity at an average age of (50.4±5.5) years, among which, 3 cases appeared symptoms of limb numbness and decreased muscle strength.The vitreous body of the proband (Ⅱ11) in MZ002 pedigree was looser and easier to remove during vitrectomy than that of Ⅲ13 in MZ001 pedigree.Vitreous specimen of Ⅱ11 was positive with Congo red staining.Gene sequencing revealed an Ala36Pro variant in the exon 3 of TTR gene. Conclusions:Gly83Arg or Ala36Pro mutation of TTR gene can cause FVA.Different mutations can lead to different clinical phenotypes such as age of onset, clinical symptoms and complications of other systems.

【Objective】 To retrospectively analyze the key performance indicators of two hepatitis B surface antigen (HBsAg) ELISA reagents used in blood stations, so as to provide basis for reagent selection from the perspective of laboratory performance. 【Methods】 The test results, reactive rate of initial/repeat test, concordance rate of initial and repeat test, and the NAT-yield implicated in ELISA-reactive samples with one assay from 2016 to 2019 were analyzed to compare the detection performance of two ELISA reagents. 【Results】 The total ELISA reactive rate of HBsAg was 0.39%(1 863/480 741), with 0.31% and 0.29% in two reagents, respectively. The initial reactive rate were 0.44% and 0.39%, concordance rate of initial and repeat test was 70.21% and 74.55%, respectively. 43 NAT-yield samples were implicated in ELISA-reactive samples with one assay. 【Conclusion】 There are differences underlying in the detection performance of the two reagents. In order to avoid blood waste and ensure blood safety, the key performance indexes and test results should be taken into consideration when selecting reagents.

Objective:To investigate the clinicopathological features of central nervous system (CNS) mesenchymal chondrosarcoma (MCS).Methods:Nine cases of CNS MCS were collected at the First Affiliated Hospital of Fujian Medical University from September 2010 to September 2020. The clinical,imaging,histopathological and immunohistochemical features were reviewed. NCOA2 gene rearrangement was evaluated by fluorescence in situ hybridization (FISH).Results:There were three male and six female patients, with age range of 1 to 59 years (median 31 years). Six cases were intracranial and three cases were intraspinal, and the tumors showed dural attachment. They were often diagnosed as meningioma basing on preoperative imaging. Microscopically, the tumors showed a characteristic biphasic histologic pattern composed of undifferentiated mesenchymal small cells and well-differentiated hyaline cartilage islands. The small cells area were positive for SOX9 (9/9), CD99 (8/9), and without BRG1 and INI1 deletion. The cartilaginous component expressed SOX9 (9/9) and S-100 protein (8/9). NCOA2 gene break apart signal was identified in five cases (5/5). Eight patients were followed up for 4-124 months. Three patients (3/8) had recurrences within one year and two patients died of the tumor.Conclusions:CNS MCS is an extremely rare malignant neoplasm with a propensity to dural involvement. Preoperative imaging has low diagnostic accuracy. CNS MCS should be differentiated from other CNS small round cell tumors and chondrosarcoma. FISH detection of NCOA2 gene rearrangement will assist the diagnosis of MCS.

Miao is an important ethnic minority in China. Miao medicine (MM) is a pivotal of traditional Chinese medicine. It is an accumulation of the Miao people's experience fighting against diseases for thousands of years and has been an important part of the Miao culture. The origin of MM is closely related to Miao culture, customs, and habits. MM gradually formed a traditional medical theoretical system with obvious regional characteristics, distinctive cultural characteristics, and unique efficacy. In this paper, we intro-duce and review the unique theoretical system of MM, including the basic theory of MM, the theory of medicine, nature, taste, and channel tropism, and the theory of entering the shelf. In addition, we summarize the development of MM to provide an overview of the theoretical characteristics and industrialisation of MM that is accessible to everyone.