Objective:This study aimed to explore the molecular mechanisms of the maladaptation of H3N2 influenza virus in chicken embryos, provide a theoretical basis for the restoration of H3N2 influenza vaccine production in chicken embryos.Methods:Samples of respiratory secretions from patients with influenza-like symptoms (Influenza-like Illness, ILI) caused by H3N2 influenza virus were inoculated into chicken embryos and Madin-Darby Canine Kidney cells (MDCK), respectively. After isolating the virus, hemagglutination experiments were conducted to detect hemagglutination titers and hemagglutination inhibition experiments were used to compare antigenic differences; further, whole-genome sequencing of H3N2 influenza virus was performed using second-generation high-throughput gene sequencing (Next Generation High-Throughput Gene Sequencing, NGS), and key amino acid sites of mutations were identified through sequence alignment; combined with sialic acid receptor binding experiments, the differences in the binding of wild-type and mutant receptor binding sites (RBS) to sialic acid receptors were compared; finally, molecular docking and molecular dynamics simulation method were used to explore the specific molecular mechanisms of how mutation sites affect the differences in the affinity of the RBS pocket for sialic acid receptors.Results:The hemagglutination assay result indicated that both chicken embryos and MDCK cells could isolate the influenza virus, and the hemagglutination inhibition test showed that no antigenic differences were produced in the isolated strains. NGS analysis revealed that the H3N2 virus underwent an F195Y mutation in the (RBS) region of the hemagglutinin (HA) protein after adaptation through chicken embryo passages. Receptor-binding experiments demonstrated that the F195Y mutation enhanced the virus′s binding ability to α2, 3-linked sialic acid glycan (Neu5Acα2-3Galβ1-4GlcNAcβ-PAA, 3′SLN), while the mutation did not affect the affinity of the RBS pocket for α2, 6-linked sialic acid glycan (Neu5Acα2-6Galβ1-4GlcNAcβ-PAA, 6′SLN). Molecular docking and molecular dynamics simulation result indicate that the F195Y mutation, by replacing a hydrophobic amino acid with a hydrophilic one, leads to a significant decrease in the structure of the RBS pocket, enhancing the binding stability of the H3N2 influenza virus with α2, 3-sln. This is specifically manifested by an increase in binding time and an increase in the number of hydrogen bonds at the RBS site with the receptor. Furthermore, the F195Y mutation does not alter the binding of the virus to other receptors.Conclusions:The F195Y mutation in the RBS pocket of H3N2 influenza virus is a key site affecting the viral chicken embryo inadaptability.

Objective To explore the safety and feasibility of transumbilical single-site laparoscopic partial splenectomy for pediatric splenic cysts.Methods A retrospective analysis was conducted on 5 cases of splenic cysts from January 2021 to April 2024,with cyst diameters ranging from 6 cm to 11 cm.A vertical incision approximately 2 cm in length was made at the umbilicus,and three 5 mm trocars were inserted in a"triangle"shape,one for observation and two for operation.Splenectomy of the upper pole(4 cases)or lower pole(1 case)was performed.Results The operations were successfully completed in all the 5 cases without conversion to open surgery.One case required intraoperative transfusion of suspended red blood cells.The operation time was 100-250 min(mean,162 min),the intraoperative blood loss was 30-200 ml(mean,72 ml),the postoperative drainage time was 2-3 d,and the postoperative hospital stay was 4-8 d(mean,6.4 d).No complications were observed.One case had mild thrombocytosis(434×109/L)at one week postoperatively,which returned to normal upon re-examination two weeks later.Postoperative pathological examination confirmed splenic cysts.Follow-ups ranged for6-44 months,with an average of 28.4 months.No complications such as thrombocytosis,cyst recurrence,residual splenic necrosis,venous thrombosis,or incisional hernia were reported.Conclusion Transumbilical single-site laparoscopic partial splenectomy is safe and feasible for treating pediatric splenic cysts.

Objective: To analyze respiratory syncytial virus(RSV) outbreaks surveillance results and the epidemiological characteristics in kindergarten and school in Shenzhen during 2017-2023 , so as to provide a scientific reference for control and prevention of RSV. Methods: Epidemiological data and surveillance results of RSV outbreaks in kindergarten and school from 2017 to 2023 were collected for descriptive analyses. Results: A total of 31 RSV outbreaks were identified in kindergarten and school in 2017-2023 in Shenzhen, 346 cases were reported, the average incidence rate was 22.02%. The most annual RSV outbreaks were reported in 2020 with 14 outbreaks, followed by 8 outbreaks in 2023. A total of 64.52% of RSV outbreaks were identified in kindergarten with rest occurring in primary school or middle school. The greatest monthly count of outbreak was 18 (58.06%) in September, followed by 3 outbreaks (9.68%) in March and October. A total of 244 swab samples were collected, 169 samples were positive for respiratory viruses, the positive rate was 69.26%, 121 samples were positive for RSV,from 31 respiratory syncytical virus outbreaks 57 and samples were positive for other respiratory viruses(9 samples were positive for two respiratory viruses). A toral of 14(45.16%) outbreaks are caused by RSV alone, 17 outbreaks (54.84%) were caused by RSV and other respiratory viruses. Conclusions Most RSV outbreaks in kindergarten and school are reported after 2020 in Shenzhen, most RSV outbreaks occur in kindergarten, peak seasons of RSV outbreaks are autumn and spring.

Objective To explore the safety and feasibility of transumbilical single-site laparoscopic partial splenectomy for pediatric splenic cysts.Methods A retrospective analysis was conducted on 5 cases of splenic cysts from January 2021 to April 2024,with cyst diameters ranging from 6 cm to 11 cm.A vertical incision approximately 2 cm in length was made at the umbilicus,and three 5 mm trocars were inserted in a"triangle"shape,one for observation and two for operation.Splenectomy of the upper pole(4 cases)or lower pole(1 case)was performed.Results The operations were successfully completed in all the 5 cases without conversion to open surgery.One case required intraoperative transfusion of suspended red blood cells.The operation time was 100-250 min(mean,162 min),the intraoperative blood loss was 30-200 ml(mean,72 ml),the postoperative drainage time was 2-3 d,and the postoperative hospital stay was 4-8 d(mean,6.4 d).No complications were observed.One case had mild thrombocytosis(434×109/L)at one week postoperatively,which returned to normal upon re-examination two weeks later.Postoperative pathological examination confirmed splenic cysts.Follow-ups ranged for6-44 months,with an average of 28.4 months.No complications such as thrombocytosis,cyst recurrence,residual splenic necrosis,venous thrombosis,or incisional hernia were reported.Conclusion Transumbilical single-site laparoscopic partial splenectomy is safe and feasible for treating pediatric splenic cysts.

Objective:This study aimed to explore the molecular mechanisms of the maladaptation of H3N2 influenza virus in chicken embryos, provide a theoretical basis for the restoration of H3N2 influenza vaccine production in chicken embryos.Methods:Samples of respiratory secretions from patients with influenza-like symptoms (Influenza-like Illness, ILI) caused by H3N2 influenza virus were inoculated into chicken embryos and Madin-Darby Canine Kidney cells (MDCK), respectively. After isolating the virus, hemagglutination experiments were conducted to detect hemagglutination titers and hemagglutination inhibition experiments were used to compare antigenic differences; further, whole-genome sequencing of H3N2 influenza virus was performed using second-generation high-throughput gene sequencing (Next Generation High-Throughput Gene Sequencing, NGS), and key amino acid sites of mutations were identified through sequence alignment; combined with sialic acid receptor binding experiments, the differences in the binding of wild-type and mutant receptor binding sites (RBS) to sialic acid receptors were compared; finally, molecular docking and molecular dynamics simulation method were used to explore the specific molecular mechanisms of how mutation sites affect the differences in the affinity of the RBS pocket for sialic acid receptors.Results:The hemagglutination assay result indicated that both chicken embryos and MDCK cells could isolate the influenza virus, and the hemagglutination inhibition test showed that no antigenic differences were produced in the isolated strains. NGS analysis revealed that the H3N2 virus underwent an F195Y mutation in the (RBS) region of the hemagglutinin (HA) protein after adaptation through chicken embryo passages. Receptor-binding experiments demonstrated that the F195Y mutation enhanced the virus′s binding ability to α2, 3-linked sialic acid glycan (Neu5Acα2-3Galβ1-4GlcNAcβ-PAA, 3′SLN), while the mutation did not affect the affinity of the RBS pocket for α2, 6-linked sialic acid glycan (Neu5Acα2-6Galβ1-4GlcNAcβ-PAA, 6′SLN). Molecular docking and molecular dynamics simulation result indicate that the F195Y mutation, by replacing a hydrophobic amino acid with a hydrophilic one, leads to a significant decrease in the structure of the RBS pocket, enhancing the binding stability of the H3N2 influenza virus with α2, 3-sln. This is specifically manifested by an increase in binding time and an increase in the number of hydrogen bonds at the RBS site with the receptor. Furthermore, the F195Y mutation does not alter the binding of the virus to other receptors.Conclusions:The F195Y mutation in the RBS pocket of H3N2 influenza virus is a key site affecting the viral chicken embryo inadaptability.

Objective: To analyze the monitoring results and characteristics of human metapneumovirus (HMPV) outbreaks in Shenzhen in kindergartens and schools during 2017-2023, so as to provide a evidence for prevention and control of respiratory diseases. Methods: Based on the epidemiological data and laboratory test results of HMPV outbreaks in kindergartens and schools of Shenzhen during 2017 to 2023, collected from the influenza monitoring network platform, Mann Kendall test was applied for trend analysis pf HMPV. Results: A total of 11 HMPV outbreaks were identified in kindergartens and schools from 2017 to 2023 for 128 cases with an average incidence rate of 26.61% in 2018. The most yearly HMPV outbreaks were reported in 2020 being 4 outbreaks, followed by 2 outbreaks in 2018. A total of 72.73% (8 outbreaks) of HMPV outbreaks were identified in kindergartens with 27.27% (3 outbreaks) occurring in primary schools. The monthly greatest count of outbreak was 3(27.27%) in March and December, followed by 2 outbreaks (18.18%) in April and June. A total of 98 swab samples were collected, and 80 samples were positive for respiratory viruses with a positive rate of 81.63%; among them, 67 samples were positive for HMPV alone, 10 samples were positive for other respiratory viruses alone, and 3 samples were positive both for HMPV and other respiratory virus. Among the 11 human metalung virus cases, 6 HMPV outbreaks were caused by human metapneumovirus alone, and 5 outbreaks were caused by HMPV and other respiratory viruses. Except for 2020, the percentage of HMPV outbreaks in influenza like illness outbreaks increased with year ( Z=2.25, P =0.02). Conclusion HMPV outbreaks have been reported every year from 2017 to 2023 in kindergartens and schools in Shenzhen, most of HMPV in kindergartens, and the peak seasons of HMPV outbreaks are spring and winter.

Objective:To investigate the related risk factors of surgical appendectomy after endoscopic retrograde appendicitis therapy (ERAT) in children.Methods:From September 2019 to November 2023 at the Children's Hospital Affiliated to Shandong University,the data from all related children with appendectomy after ERAT were analyzed. The general situation and main clinical manifestations of the children were collected. According to the effect of ERAT,the patients were divided into transfer to surgical group and appendicitis recurrence group by the reasons and time of transfer to surgical treatment. The indexes such as ERAT and reoperation process and pathological types of appendix after operation were collected,and the clinical experience was summarized.Results:Among the 242 children who underwent ERAT,32 cases underwent appendectomy again,including 19 males and 13 females,with an average age of（9.16±2.77）years,and the reoperation rate was 13.2%. The clinical manifestation was abdominal pain (32 cases,100.0%),mainly right lower abdominal pain (28 cases,87.5%),and the main duration was less than 3 months (30 cases,93.8%). Abdominal ultrasound or CT before ERAT mainly indicated appendicitis (11 cases,34.4%) and appendicitis complicated with appendiceal calculus (11 cases,34.4%). There were 9 children in transferred to surgical group,of which 5 cases were transferred to surgery because of ERAT intubation failure,and 4 cases were transferred to surgery because of appendix perforation or abscess. Postoperative pathology showed acute suppurative appendicitis in 5 cases and acute gangrenous appendicitis in 4 cases. There were 23 children in appendicitis recurrence group,whose abdominal pain was relieved after ERAT,but their symptoms were repeated after discharge. After clinical evaluation,they were diagnosed as recurrent appendicitis and underwent surgical appendectomy. Most of them were reoperated within 6 months after ERAT (21 cases,91.3%).Postoperative pathology was mainly chronic appendicitis (10 cases,43.5%).Conclusion:Acute suppurative appendicitis and acute gangrenous appendicitis are prone to catheterization or endoscopic failure during ERAT,and there is a high risk of surgical transfer. ERAT may relieve the symptoms of chronic appendicitis in children. If the symptoms are repeated,surgery is still needed to remove the appendix.

Objective:To understand the epidemiological characteristics of human respiratory syncytial virus (HRSV) among cases presenting with influenza-like illness (ILI) in Shenzhen City from 2019 to 2023.Methods:Respiratory specimens were collected from two national sentinel hospitals in Shenzhen from March 2019 to December 2023, specifically targeting cases of ILI. The real-time PCR method was used for the detection and genotyping of HRSV. Basic demographic information was collected and used for the epidemiological analysis.Results:A total of 9 278 respiratory specimens of influenza-like cases were collected and detected, with a total positive rate of 4.77% (443/9 278) for HRSV. In 2021 (8.48%, 167/1 970), the positive rate of HRSV was significantly higher than in 2019 (3.35%, 52/1 552), 2022 (1.80%, 39/2 169), and 2023 (4.49%, 133/2 960), and the difference was statistically significant ( χ 2=102.395, P<0.001). The prevalence of HRSV was mainly in summer and early autumn (September), and there was an abnormal increase in the positive rate of HRSV in winter 2022. The highest positive rate of HRSV was in children under five years old (9.84%, 330/335). The typing results showed that in 2022, the prevalence of HRSV-A was predominant (71.79%, 28/39), and in 2023, HRSV-A and HRSV-B subtypes coexisted. Conclusions:The prevalence of HRSV in Shenzhen from 2019 to 2023 has obvious seasonality, mainly in summer and early autumn. Children under five years old are the main population of HRSV infections.

Objective:To understand the epidemiological characteristics of human respiratory syncytial virus (HRSV) among cases presenting with influenza-like illness (ILI) in Shenzhen City from 2019 to 2023.Methods:Respiratory specimens were collected from two national sentinel hospitals in Shenzhen from March 2019 to December 2023, specifically targeting cases of ILI. The real-time PCR method was used for the detection and genotyping of HRSV. Basic demographic information was collected and used for the epidemiological analysis.Results:A total of 9 278 respiratory specimens of influenza-like cases were collected and detected, with a total positive rate of 4.77% (443/9 278) for HRSV. In 2021 (8.48%, 167/1 970), the positive rate of HRSV was significantly higher than in 2019 (3.35%, 52/1 552), 2022 (1.80%, 39/2 169), and 2023 (4.49%, 133/2 960), and the difference was statistically significant ( χ 2=102.395, P<0.001). The prevalence of HRSV was mainly in summer and early autumn (September), and there was an abnormal increase in the positive rate of HRSV in winter 2022. The highest positive rate of HRSV was in children under five years old (9.84%, 330/335). The typing results showed that in 2022, the prevalence of HRSV-A was predominant (71.79%, 28/39), and in 2023, HRSV-A and HRSV-B subtypes coexisted. Conclusions:The prevalence of HRSV in Shenzhen from 2019 to 2023 has obvious seasonality, mainly in summer and early autumn. Children under five years old are the main population of HRSV infections.

Objective To investigate the safety and feasibility of transumbilical single-site laparoscopic treatment of Meckel's diverticulum in children.Methods A total of 122 patients with Meckel's diverticulum were treated with transumbilical single-site laparoscopic treatment in our department from January 2019 to June 2023.A longitudinal incision was made through the umbilical cord,and 5 mm trocars were inserted into the left and right sides of the incision.The laparoscope and non-invasive intestinal forceps were inserted via the observation and operation holes to probe the abdominal cavity and locate the diverticulum.After the umbilical cord incision was expanded,the diverticulum was pulled out from the abdominal cavity for external resection and anastomosis.Results All the 122 children were diagnosed by laparoscopy.There were 4 cases who were diagnosed as having acute gangrenous appendicitis preoperatively,but were confirmed as having Meckel's diverticulum with perforation under laparoscope intraoperatively.There were 38 cases of lower digestive tract bleeding,48 cases of inflammatory reaction,12 cases of intussusception,8 cases of intestinal obstruction caused by cord and band compression,and 12 cases of peritonitis caused by perforation.All the operations were successfully completed,and the operation time was 58-125 min,with an average of 78 min.Pathological examination after surgery showed Meckel's diverticulum in all the cases.Two cases of incision infection were cured after dressing change treatment.The patients were given a liquid diet 2-4 d after surgery and discharged 5-9 d later.A total of 119 patients were followed up for 3-18 months after surgery,and the surgical results were satisfactory.There were no serious complications such as rectal bleeding,anastomotic stenosis,adhesive intestinal obstruction,or incisional hernia after surgery.Conclusions Transumbilical single-site laparoscopic treatment of Meckel's diverticulum in children is safe and feasible.Meckel's diverticulum combined with perforation or intestinal obstruction is not a contraindication of transumbilical single-site laparoscopic surgery.