Objective To analyze the virus subtypes, molecular evolutional and molecular transmission network features of the first confirmed mpox case in Huai’an City, Jiangsu Province, so as to provide insights into understanding of the transmission and evolution dynamics of mpox virus and formulation of the mpox control strategy in the city. Methods Genomic DNA was extracted from swabs of the first confirmed mpox case’s skin lesions in Huai’an City, and the amplicon sequencing library was constructed using the hypersensitive mpox virus whole-genome capture kit. High-throughput sequencing was performed using the GridION X5 nanopore sequencer on the Nanopore sequencing platform, and single nucleotide polymorphism (SNP) analysis of mpox virus genome sequences was performed following sequence assembly. In addition, phylogenetic analysis, genetic genealogy and molecular traceability analysis were performed. Results The virus whole genome sequence of the first confirmed mpox case was successfully obtained by high-throughput sequencing, with a full length of 197 182 bp, and was named hMpxV/China/JS-HA01/2023, which belonged to the clade IIb (West African clade) lineage B.1.3. Compared with the mpox virus reference sequence MPXV-M5312_HM12_Rivers-001 (GenBank accession number: NC_063383), the genome sequence of the Huai’an virus isolate carried 86 SNPs, including 40 SNPs in the coding region as non-synonymous mutations and 73 SNPs as nucleotide mutations caused by APOBEC3 (APOBEC3). Of the 97 mpox virus gene sequences, 79 sequences were included in the molecular network (81.44%), and the threshold of the genetic distance accessed to the network was 0.35/10⁵. There were two large molecular transmission clusters and one scattered cluster in the molecular transmission network of the mpox virus, andthehMpxV/China/JS-HA01/2023 sequence was located in the large cluster. The 97 gene sequences formed 92 haplotypes, including three shared haplotypes Hap_4, Hap_6 and Hap_38, and an exclusive haplotype Hap_1 of hMpxV/China/JS-HA01/2023 generated from mutation of the exclusive haplotype Hap_43, while the exclusive haplotype Hap_43 was generated from mutation of the shared haplotype Hap_38. Conclusions The whole genome sequence of the mpox virus isolated from the first confirmed mpox case in Huai’an City has been successfully obtained, and the molecular evolutionary and molecular transmission network characteristics of the virus have been preliminarily understood.

Objective To prepare and characterize the mouse polyclonal antibody against the dense granule protein 24 (GRA24) of Toxoplasma gondii, and explore its preliminary applications. Methods The GRA24 coding sequences of different T. gondii strains were aligned using the MEGA-X software, and the dominant peptide of the GRA24 protein was analyzed with the Protean software. The base sequence encoding this peptide was amplified using PCR assay and ligated into the pET-28a vector, and the generated GRA24 truncated protein was transformed into Escherichia coli BL21. After induction by isopropyl-beta-D-thiogalactopyranoside (IPTG), the expression and purification of the recombinant GRA24 protein was analyzed using sodium dodecyl sulfate - polyacrylamide gel electrophoresis (SDS-PAGE). BALB/c mice were immunized by subcutaneous injection with the purified recombinant GRA24 truncated protein to generate the polyclonal antibody, and the titer of the polyclonal antibody was measured using enzyme linked immunosorbent assay (ELISA). The specificity of the polyclonal antibody was tested using Western blotting, and the intracellular localization of the polyclonal antibody was investigated using immunofluorescence assay (IFA). Results SDS-PAGE showed successful construction of the recombinant expression plasmid, and Coomassie brilliant blue staining showed the generation of the high-purity recombinant GRA24 truncated protein. ELISA measured that the titer of the polyclonal antibody against the GRA24 truncated protein was higher than 1:208 400, and Western blotting showed that the polyclonal antibody was effective to recognize the endogenous GRA24 proteins of different T. gondii strains and specifically recognize the recombinant GRA24 truncated protein. Indirect IFA showed that the GRA24 protein secreted 16 hour following T. gondii invasion in host cells. Conclusions The polyclonal antibody against the T. gondii GRA24 protein has been successfully prepared, which has a widespread applicability, high titers and a high specificity. This polyclonal antibody is available for Western blotting and IFA, which provides the basis for investigating the function of the GRA24 protein.

Objective To explore the automated identification and diameter measurement methods for inferior vena cava (IVC) based on clinical ultrasound images of IVC. Methods An automated identification and localization method based on topology and automatic tracking algorithm was proposed. Tracking algorithm was used for identifying and continuously locating to improve the efficiency and accuracy of measurement. Tests were conducted on 18 sets of ultrasound data collected from 18 patients in intensive care unit (ICU),with clinicians' measurements as the gold standard. Results The recognition accuracy of the automated method was 94.44％ (17/18),and the measurement error of IVC diameter was within the range of±1.96s (s was the standard deviation). The automated method could replace the manual method. Conclusion The proposed IVC automated identification and localization algorithm based on topology and automatic tracking algorithm has high recognition success rate and IVC diameter measurement accuracy. It can assist clinicians in identifying and locating IVC,so as to improve the accuracy of IVC measurement.

The chronic low-pressure hypoxic environment in high-altitude areas is closely associated with the development of epilepsy， which is associated with various factors such as neuronal death， inflammatory response， and gene expression. In order to quickly diagnose epilepsy in high-altitude areas， the research on epilepsy-related biomarkers has attracted more and more attention. This article reviews the research on biomarkers for epilepsy in high-altitude areas， in order to provide new ideas for the early diagnosis of epilepsy in high-altitude areas and opportunities for exploring potential intervention measures.
Biomarkers

Humans ; Aged ; Acute Kidney Injury ; Risk Factors ; Creatinine

ObjectiveThe contamination of foodborne pathogens in ready-to-eat fruits and vegetables in Shanghai was analyzed to provide a scientific basis for food safety， risk assessment and related supervision. MethodsFrom June to September 2021， a total of 143 batches of12 kinds of ready-to-eat fruits and vegetables such as lettuce， chicory， and cherry tomatoes were collected from farmers’ markets， supermarkets， and e-commerce platforms. The total number of bacterial colonies， Salmonella， Listeria monocytogenes， Staphylococcus aureus， Cronobacter spp. and Shiga toxin-producing Escherichia coli in the samples were tested according to National Food Contamination and Harmfulness Risk Monitoring Manual. ResultsAmong the 143 batches， foodborne pathogens were detected in 68 batches， with a total detection rate of 47.55% （68/143）. A total of 79 strains of foodborne pathogens were detected. The detection rate of Staphylococcus aureus was the highest （32.87%， 47/143）， followed by Cronobacter spp. （20.98%， 30/143）， Salmonella （0.70%， 1/143）， Listeria monocytogenes （0.70%， 1/143）， Shiga toxin-producing Escherichia coli （0.00%）. Furthermore， the detection rate was higher in different ready-to-eat fruits and vegetables： chicory （17.33%）， cucumber （17.14%）， cherry tomatoes （16.00%）， and honeydew melon （15.38%）， respectively. Meanwhile， the contamination rate of pathogens in ready-to-eat fruits and vegetables from farmers’ markets， supermarkets， and e-commerce platforms was relatively high. ConclusionReady-to-eat fruits and vegetables in Shanghai are contaminated by foodborne pathogens. The prevention and control of the contamination of post-harvest fruits and vegetables should be strengthened to reduce the risk of foodborne disease outbreaks.

Objective:To investigate the effect of sodium octanoate on renal-intestinal ischemia- reperfusion injury (IRI) after resuscitation from traumatic cardiac arrest in pigs.Methods:Twenty-two miniature piglets with a body weight of (37.6±2.5)kg were divided into three groups according to the random-number table method: normal group ( n=7), IRI group ( n=7) and IRI-treated group ( n=8). A renal-intestinal IRI model of the pig was established by allowing femoral artery to bleed through blood pump at a rate of 2 ml·kg -1·min -1 until cardiac arrest, followed by whole blood transfusion through the femoral vein at a rate of 5 ml·kg -1·min -1 after observation for 6 minutes, and 50% of total blood loss was reinfused before resuscitation. Both the IRI group and IRI-treated group were with IRI model, while normal group was just monitored without induction of IRI. Besides, IRI-treated group was injected intravenously with sodium octanoate (30 mg/kg) for 1 hour at 5 minutes after restoration of spontaneous circulation (ROSC). (1) The rate of resuscitation success, survival rate at 4, 24 hours after resuscitation, blood loss when reaching cardiac arrest criteria and resuscitation time when reaching the ROSC criteria were compared in the three groups. (2) Levels of serum creatinine (SCr), urea nitrogen (BUN), intestinal fatty acid binding protein (iFABP) and diamine oxidase (DAO) were measured before resuscitation and at 1, 2, 4, 24 hours after resuscitation. (3) The animals were sacrificed at 24 hours post-resuscitation to harvest renal and intestinal tissues rapidly. TUNEL test was applied for the cellular apoptosis index. Prussian blue was used to detect the rate of iron deposition. Western blot analysis was used to measure levels of glutathione peroxidase 4 (GPX4) and acyl-CoA synthetase long-chain family member4 (ACSL4). Results:In three groups, all pigs survived. There was no significant difference in blood loss or resuscitation time between IRI group and IRI-treated group (all P>0.05). There was no significant difference in levels of SCr, BUN, iFABP or DAO before resuscitation and at 1, 2, 4, 24 hours after resuscitation in normal group (all P>0.05). But their levels were gradually increased at 1, 2, 4, 24 hours after resuscitation from that before resuscitation in IRI group and IRI-treated group (all P<0.01). Among three groups, levels of SCr, BUN, iFABP and DAO had no significant difference before resuscitation (all P>0.05), but showed obvious increase in IRI group and the IRI-treated group at 1, 2, 4, 24 hours after resuscitation compared with normal group, especially in IRI group (all P<0.01). In normal group, IRI group and IRI-treated group after 24 hours for resuscitation, the cellular apoptosis index of renal tissues was (2.3±0.8)%, (44.0±5.4)% and (13.8±4.3)%; the cellular apoptosis index of intestinal tissues was (2.6±0.9)%, (61.3±10.4)% and (20.8±3.7)%; the rate of iron deposition of renal tissues was (0.6±0.1)%, (3.9±1.0)% and (1.7±0.3)%; the rate of iron deposition of intestinal tissues was (0.8±0.1)%, (4.9±0.9)% and (2.1±0.5)% (all P<0.01). The cellular apoptosis index and rate of iron deposition of both renal and intestinal tissues were the highest in IRI group. The renal-intestinal expression of GPX4 in IRI group and IRI-treated group was lower than that in normal group at 24 hours after resuscitation (all P<0.05), with the lowest in IRI group. The renal-intestinal expression of ACSL4 in IRI group and IRI-treated group was higher than that in normal group at 24 hours after resuscitation (all P< 0.01), with the highest in IRI group. Conclusion:Sodium octanoate can reduce renal-intestinal IRI after resuscitation from traumatic cardiac arrest in pigs, the mechanism for which is probably due to that sodium octanoate can inhibit cellular apoptosis and reduce ferroptosis by regulating the expression levels of GPX4 and ACSL4.

Objective:To evaluate the effect of obesity on the dose-effect relationship of remimazolam when combined with alfentanil in painless gastroscopy.Methods:American Society of Anesthesiologists physical status Ⅰor Ⅱ patients of both sexes, scheduled for elective painless gastroscopy, aged 18-64 yr, were divided into 2 groups according to the body mass index (BMI): normal (BMI 19-24 kg/m 2) group and obese (BMI≥28 kg/m 2) group.Alfentanil 5 μg/kg combined with remimazolam was given intravenously in all the patients, and the dose of remimazolam was determined by the modified Dixon′s up-and-down method.The initial dose of remimazolam was 0.25 mg/kg, and each time the dose was increased or decreased by 0.05 mg/kg based on the sedative effect.The response was defined as positive when the responses that affected the operation of examination developed during insertion of the gastroscope and within the first 2 min of examination such as swallowing, bucking or body movement.This process was repeated until the seventh intersection occurred.The 50% effective dose (ED 50), 95% effective dose (ED 95), and 95% confidence interval ( CI) of remimazolam were calculated by probit method. Results:There were 26 patients in normal group and 18 patients in obese group.The ED 50 (95% CI) of remimazolam was 0.196 (0.087-0.274) mg/kg, and the ED 95 (95% CI) was 0.322 (0.256-1.397) mg/kg in normal group.The ED 50 (95% CI) of remimazolam was 0.125 (0.102-0.148) mg/kg, and the ED 95 (95% CI) was 0.161 (0.141-0.242) mg/kg in obese group.The ED 50 and ED 95 were significantly lower in obese group than in normal group ( P<0.001). Conclusions:Obesity increases the potency of remimazolam when combined with alfentanil 5 μg/kg in the patients undergoing painless gastroscopy.

Objective:To explore the association of DNA methylation with immune response to hepatitis B (HepB) vaccine in Han nationality children from Guangxi province.Methods:A total of 263 children aged 8-9 months who had completed HepB immunization program were recruited from three hospitals in Guangxi province by using unmatched case-control method. Children with the HepB surface antibody concentration(Anti -HBs)<100 mIU/ml was set as the case group and ≥100 mIU/ml as the control group. Multiplex PCR and heavy sulfite sequencing were used to treat the samples. Illumina platform was used for high-throughput DNA methylation sequencing of IFNG gene target regions and CpG sites. Unconditional logistic regression was used to analyze the association between cytosine-phospho-guanosine DNA methylation at 18 loci of IFNG gene and HepB immune response level. Results:There were 104 children in the case group and 159 in the control group. The median ( Q1, Q3) level of anti -HBs in two groups were 62.34 (30.06, 98.88) mIU/ml and 1 089.10 (710.35, 1 233.45) mIU/ml. The methylation levels of IFNG_1 gene 44 and 93 locus in the case group were higher than those in the control group ( P<0.05). The unconditional logistic regression model showed that the DNA methylation level of IFNG_1 gene at 44 ( OR=1.18, 95% CI: 1.03-1.35) and 93 ( OR=1.21, 95% CI: 1.07-1.38) locus was associated with the HepB response level. Conclusion:The changes of DNA methylation at locus 44 and 93 of IFNG_1 gene may be relevant factors affecting the response level of HepB in Han nationality children from Guangxi province.

The laccase (PpLAC) gene family members in peach fruit were identified and the relationship between their expression pattern and chilling induced browning were investigated. The study was performed using two varieties of peaches with different chilling tolerance, treated with or without exogenous γ-aminobutyric acid (GABA) during cold storage. Twenty-six genes were screened from the peach fruit genome. These genes were distributed on 6 chromosomes and each contained 5-7 exons. The PpLAC gene family members shared relatively similar gene structure and conserved motifs, and they were classified into 7 subgroups based on the cluster analysis. Transcriptome sequencing revealed that the expression levels of PpLAC7 and PpLAC9 exhibited an increasing pattern under low temperature storage, and displayed a similar trend with the browning index of peach fruit. Notably, GABA treatment reduced the degree of browning and inhibited the expression of PpLAC7 and PpLAC9. These results suggested that PpLAC7 and PpLAC9 might be involved in the browning of peach fruit during cold storage.
Food Storage ; Fruit/genetics* ; Laccase/genetics* ; Prunus persica/genetics*