Strangles,caused by Streptococcus equi subsp.equi,remains one of the most prevalent and high-incidence infectious diseases in intensive donkey farms,posing a significant threat to the healthy development of the donkey industry.Vaccination serves as an effective measure for the pre-vention and control of the disease,however,there is currently no attenuated vaccine against this disease in China.To provide a candidate strain for the development of a live attenuated strangles vaccine,this study focused on a wild-type S.equi subsp.equi strain isolated from donkeys.Using homologous recombination gene knockout technology,the aroA gene(encoding 5-enolpyru-vylshikimate-3-phosphate synthase)and the sag A gene(encoding the precursor of streptolysin S toxin)were sequentially deleted to construct a double-gene-deletion strain(ΔsagA/aroA).The virulence and key biological characteristics of the mutant strain were systematically evaluated.TheΔsagA/aroA strain was successfully generated,exhibiting complete loss of hemolytic activity and maintaining stable genetic inheritance over 60 consecutive passages.Electron microscopy revealed that the mutant retained morphological characteristics compared to the wild-type strain,and its growth rate was significantly slower(P＜0.000 1).Virulence assessment using a challenge dose of 1× 105 CFU/0.2 mL(the minimum fully lethal dose of the wild-type strain)demonstrated markedly attenuated virulence in the mutant.Immunization trials with 1 ×104 CFU/0.2 mL of theΔsagA/aroA strain revealed a increase in ELISA antibody titers by day 7 post-vaccination,and higher levels at days 14 and 21.Notably,antibody levels in the experimental group were significant-ly higher than those in the control group(P＜0.000 1).These findings confirm that the double-gene-deletion strain S.equi subsp.equi ΔsagA/aroA exhibits reduced virulence while retaining im-munogenicity,which suggested it can be used as a promising candidate strain for further develop-ment of a live attenuated strangles vaccine.

Objective:To understand the evolution and variation characteristics of the H1N1 influenza virus in Wuxi City from 2018 to 2023.Methods:Real time PCR was used to perform nucleic acid testing on throat swab samples of influenza like cases sent to sentinel hospitals for testing. The influenza A (H1N1) pdm09 positive samples were subjected to cell culture, and nucleic acid was extracted from strains with a red blood cell agglutination test (HA) ≥1∶8. The whole genome sequence was amplified, and a library was constructed. The MiSeq sequencer was used for sequencing on the machine. Using NC_026431.1 as a reference strain, we analyzed the offline data using CLC Genomics Workbench (Version 23) software. MEGA 7.0 software was used to construct a phylogenetic tree, and NetNGlyc 1.0 Server software was used to predict N-glycosylation sites.Results:The nucleotide and amino acid homology between 38 strains of A (H1N1) pdm09 influenza virus from 2018 to 2023 were 96.06%-100% and 96.12%-100%, respectively. From February to May 2023, all 12 strains of A (H1N1) pdm09 had two amino acid mutation sites occurring in the HA antigenic determinant cluster, namely the Ca region (A203T) and the Cb region (K71Q). No mutations were found in the HA receptor binding site and NA resistance site. The strains from January to June 2018 belong to the 6B. 1A evolutionary branch, the strains from December 2018 to January 2020 belong to three evolutionary branches: 6B. 1A. 1, 6B. 1A. 5a, and 6B. 1A. 7, and the strains from February to May 2023 belong to the 6B. 1A. 5a. 2a evolutionary branch. 38 strains of A (H1N1) pdm09 HA gene all have 7 potential N-glycosylation sites, while NA gene has 7-8 potential N-glycosylation sites.Conclusions:There are characteristic amino acid mutation sites of H1N1 influenza A in Wuxi City from 2018 to 2023. The emergence of these mutation sites may affect the virus′s transmission and antigenic changes.

Strangles,caused by Streptococcus equi subsp.equi,remains one of the most prevalent and high-incidence infectious diseases in intensive donkey farms,posing a significant threat to the healthy development of the donkey industry.Vaccination serves as an effective measure for the pre-vention and control of the disease,however,there is currently no attenuated vaccine against this disease in China.To provide a candidate strain for the development of a live attenuated strangles vaccine,this study focused on a wild-type S.equi subsp.equi strain isolated from donkeys.Using homologous recombination gene knockout technology,the aroA gene(encoding 5-enolpyru-vylshikimate-3-phosphate synthase)and the sag A gene(encoding the precursor of streptolysin S toxin)were sequentially deleted to construct a double-gene-deletion strain(ΔsagA/aroA).The virulence and key biological characteristics of the mutant strain were systematically evaluated.TheΔsagA/aroA strain was successfully generated,exhibiting complete loss of hemolytic activity and maintaining stable genetic inheritance over 60 consecutive passages.Electron microscopy revealed that the mutant retained morphological characteristics compared to the wild-type strain,and its growth rate was significantly slower(P＜0.000 1).Virulence assessment using a challenge dose of 1× 105 CFU/0.2 mL(the minimum fully lethal dose of the wild-type strain)demonstrated markedly attenuated virulence in the mutant.Immunization trials with 1 ×104 CFU/0.2 mL of theΔsagA/aroA strain revealed a increase in ELISA antibody titers by day 7 post-vaccination,and higher levels at days 14 and 21.Notably,antibody levels in the experimental group were significant-ly higher than those in the control group(P＜0.000 1).These findings confirm that the double-gene-deletion strain S.equi subsp.equi ΔsagA/aroA exhibits reduced virulence while retaining im-munogenicity,which suggested it can be used as a promising candidate strain for further develop-ment of a live attenuated strangles vaccine.

Objective:To understand the evolution and variation characteristics of the H1N1 influenza virus in Wuxi City from 2018 to 2023.Methods:Real time PCR was used to perform nucleic acid testing on throat swab samples of influenza like cases sent to sentinel hospitals for testing. The influenza A (H1N1) pdm09 positive samples were subjected to cell culture, and nucleic acid was extracted from strains with a red blood cell agglutination test (HA) ≥1∶8. The whole genome sequence was amplified, and a library was constructed. The MiSeq sequencer was used for sequencing on the machine. Using NC_026431.1 as a reference strain, we analyzed the offline data using CLC Genomics Workbench (Version 23) software. MEGA 7.0 software was used to construct a phylogenetic tree, and NetNGlyc 1.0 Server software was used to predict N-glycosylation sites.Results:The nucleotide and amino acid homology between 38 strains of A (H1N1) pdm09 influenza virus from 2018 to 2023 were 96.06%-100% and 96.12%-100%, respectively. From February to May 2023, all 12 strains of A (H1N1) pdm09 had two amino acid mutation sites occurring in the HA antigenic determinant cluster, namely the Ca region (A203T) and the Cb region (K71Q). No mutations were found in the HA receptor binding site and NA resistance site. The strains from January to June 2018 belong to the 6B. 1A evolutionary branch, the strains from December 2018 to January 2020 belong to three evolutionary branches: 6B. 1A. 1, 6B. 1A. 5a, and 6B. 1A. 7, and the strains from February to May 2023 belong to the 6B. 1A. 5a. 2a evolutionary branch. 38 strains of A (H1N1) pdm09 HA gene all have 7 potential N-glycosylation sites, while NA gene has 7-8 potential N-glycosylation sites.Conclusions:There are characteristic amino acid mutation sites of H1N1 influenza A in Wuxi City from 2018 to 2023. The emergence of these mutation sites may affect the virus′s transmission and antigenic changes.

Objective:To study the influence of G protein-coupled estrogen receptor 1 (GPER1) specific agonist G1 and antagonist G2 in epilepsy susceptibility of temporal lobe epileptic rats.Methods:Sixty rats were randomly divided into control group, G1 treatment group and G15 treatment group ( n=20). Rats in the latter two groups were intraperitoneally injected with GPER1 agonist G1 (10 μg) or antagonist G15 (40 μg) for a consecutive 12 d. Lithium chloride pilocarpine epilepsy models were prepared in the 3 groups. The behavior manifestations of these rats were observed within 1 h of intraperitoneal injection of pilocarpine; Racine grading was used to evaluate the severity of epileptic seizures every 5 min; the latency of epileptic seizures (Racine grading IV) and epileptic seizure grading at different time points in the 3 groups were compared. The EEG monitoring was performed to these rats, and EEG data were recorded from 10 min before pilocarpine injection to 2 h after pilocarpine injection; EEG time-frequency was analyzed by Fast-Fourier transform (FFT); distribution of brain electrical energy and changes of θ and α wave energy during 20 min of epileptic status were compared among the 3 groups. Results:(1) As compared with that in the control group and G1 treatment group, the latency of epileptic seizures in the G15 treatment group was significantly shortened ( P<0.05); 15 and 20 min after pilocarpine injection, the epileptic seizure grading of rats in G1 treatment group was statistically lower than that in control group ( P<0.05); 15-35 min after pilocarpine injection, the epileptic seizure grading of rats in G15 treatment group was significantly higher than that in control group ( P<0.05). (2) As compared with those in the control group, rats in the G1 treatment group had smaller brain wave amplitude, while the rats in the G15 treatment group had earlier seizure time, larger brain wave amplitude and higher frequency. There were no obvious changes in the amount of brain electrical energy between the G1 treatment group and control group; while the amount of brain electrical energy in the G15 treatment group 2 h after pilocarpine injection was higher than that in the control group. As compared with those in the control group and G1 treatment group, the θ and α wave energy values of rats in the G15 treatment group were significantly increased within 20 min of epileptic status ( P<0.05). Conclusion:Activation level of GPER1 might be associated with susceptibility to epileptic seizures, and specific inhibition of GPER1 activation can enhance the susceptibility to epilepsy and increase the energy values of specific frequency bands during epilepsy.

Objective To study the antipyretic effect of Paracetamol Tablets,Compound Paracetamol and Amantadine Hydrochloride Tablets,Compound Dextromethorphan Hydrobromide Tablets,and Chaiqin Qingning Capsules on the fever model induced by LPS and dry yeast in rats.Methods Fever was induced by ip injecting LPS (100 μg/kg) or sc injecting dry yeast (20％) in rats.We observed the changes of temperature of the rats after administration of Paracetamol Tablets,Compound Paracetamol and Amantadine Hydrochloride Tablets,Compound Dextromethorphan Hydrobromide Tablets (the acetaminophen contents were 205.67,102.83,and 51.42 mg/kg)and Chaiqin Qingning Capsules (1110.60,555.30,and 277.65 mg/kg).Maximum temperature rise height (△T) and temperature response index (TRI) were calculated,and the curve of average rise in temperature was drawn.Results Each dose group of Paracetamol Tablets,Compound Paracetamol and Amantadine Hydrochloride Tablets,Compound Dextromethorphan Hydrobromide Tablets,and Chaiqin Qingning Capsules had obvious antipyretic effect on the fever model induced by LPS and dry yeast in rats,and there was a certain dose-effect relationship.Conclusion Paracetamol Tablets,Compound Paracetamol and Amantadine Hydrochloride Tablets,Compound Dextromethorphan Hydrobromide Tablets,and Chaiqin Qingning Capsules has certain antipyretic effect on LPS and dry yeast fever model in rats,and on the whole,the Western medicine acts rapid but continue for a short time,while the traditional Chinese medicine acts slow but continues for a long time.

Objective:Esophageal cancer has high morbidity in China, and surgery is the main treatment for this disease. Postop-erative nutrition is also important for the patients. In this article, we discuss the possibility of retrosternal route gastrostomy feeding (RGF). Methods: The data of 127 esophageal cancer patients between 2011 and 2013 were retrospectively analyzed. RGF was per-formed in all the patients, and post-operation complications were studied. Results: Bowel obstruction, catheter displacement, and wound infection did not occur. Conclusion:RGF is a safe and effective nutrition method for patients who underwent retrosternal recon-struction.

Objective: In order to improve the technology of Chinese medicine extraction, experiments were made with large pore size ultrafiltration (LPS UF) membranes in the process of Chinese medicine production, and the feasibility of replacing traditional alcohol sedimentation with LPS UF method in the production of compound Chinese medicine was also explored. Methods: The water extraction liquid of Shenbao Mixture with alcohol percolation extract was ultrafiltered. Icariin as index component was determined. Results: Component of Icariin was reserved as 90%, this technology was similar to alcohol extraction. Conclusion: The experiments on the extraction of “Shengbao” mixture show that the LPS UF membranes with MWCO above 100,000 were more effective in retaining the effective ingredients and removing the precipitates.