OBJECTIVE To establish a rapid and accurate PCR method for detecting 24 strains of Burkholderia cepacia complex(Bcc) by comparing three detection methods of loop-mediated isothermal amplification(LAMP), SYTO 9 dye method based on polymerase chain reaction(PCR) and TaqMan probe real-time fluorescent quantitative PCR method( TaqMan probe method). METHODS According to the molecular biological information of 24 strains of Bcc in the NCBI database, multiple candidate sequence fragments unique to Bcc were screened out, and specific primer and probe that could simultaneously detect 24 strains of Bcc were designed. At the same time, the detection methods of LAMP, SYTO 9 dye method based on PCR and Taqman probe were explored, and the optimal annealing temperature was optimized and screened. The 39 experimental strains were used to verify the Bcc detection method. RESULTS LAMP method could not effectively detect Bcc, SYTO 9 dye method and TaqMan probe method could effectively detect more than 20 strains of Bcc, while TaqMan probe method had higher amplification effect, better detection sensitivity, repeatability and stability, which could meet the requirements of this study. CONCLUSION In this study, a TaqMan probe method for rapid detection of Bcc was established. Compared with LAMP method and SYTO 9 dye method, this method has the advantages of fast, simple and high sensitivity, and provides technical support for the rapid detectionof Bcc.

OBJECTIVE To analyze the types and control measures of major microorganisms in pharmaceutical water systems, so as to provide guidance for effective control of pharmaceutical water systems. METHODS The main microbial species, abundance and harmfulness of drinking water, purified water and water for injection were reviewed, and the control measures on microorganisms in pharmaceutical water were discussed. RESULTS There were differences in the main microbial types in pharmaceutical water. Burkholderia cepacia complex and Ralstonia pickettii were conditioned pathogens in pharmaceutical water, thus causing certain biological safety hazards. CONCLUSION Pharmaceutical companies can strengthen the control of microorganisms in the water system by establishing microbial databases and common microbial strain banks at all levels. Trend analysis should to be conducted based on alert limits and action limits, so as to strengthen the control of microorganisms in the water system.

Patients with severe trauma require an extremely timely treatment and transfusion plays an irreplaceable role in the emergency treatment of such patients. An increasing number of evidence-based medicinal evidences and clinical practices suggest that patients with severe traumatic bleeding benefit from early transfusion of low-titer group O whole blood or hemostatic resuscitation with red blood cells, plasma and platelet of a balanced ratio. However, the current domestic mode of blood supply cannot fully meet the requirements of timely and effective blood transfusion for emergency treatment of patients with severe trauma in clinical practice. In order to solve the key problems in blood supply and blood transfusion strategies for emergency treatment of severe trauma, Branch of Clinical Transfusion Medicine of Chinese Medical Association, Group for Trauma Emergency Care and Multiple Injuries of Trauma Branch of Chinese Medical Association, Young Scholar Group of Disaster Medicine Branch of Chinese Medical Association organized domestic experts of blood transfusion medicine and trauma treatment to jointly formulate Chinese expert consensus on blood support mode and blood transfusion strategies for emergency treatment of severe trauma patients ( version 2024). Based on the evidence-based medical evidence and Delphi method of expert consultation and voting, 10 recommendations were put forward from two aspects of blood support mode and transfusion strategies, aiming to provide a reference for transfusion resuscitation in the emergency treatment of severe trauma and further improve the success rate of treatment of patients with severe trauma.

Objective:To compare the association between body mass index(BMI)trajectories deter-mined by different methods and the risk of overweight in early childhood in a prospective cohort study,and to identify children with higher risk of obesity during critical growth windows of early childhood.Methods:A total of 1 330 children from Peking University Birth Cohort in Tongzhou(PKUBC-T)were included in this study.The children were followed up at birth,1,3,6,9,12,18,and 24 months and 3 years of age to obtain their height/length and weight data,and calculate BMI Z-score.Latent class growth mixture modeling(GMM)and longitudinal data-based k-means clustering algorithm(KML)were used to determine the grouping of early childhood BMI trajectories from birth to 24 mouths.Linear regres-sion was used to compare the association between early childhood BMI trajectories determined by different methods and BMI Z-score at 3 years of age.The predictive performance of early childhood BMI trajecto-ries determined by different methods in predicting the risk of overweight(BMI Z-score＞1)at 3 years was compared using the average area under the curve(AUC)of 5-fold cross-validation in Logistic regres-sion models.Results:In the study population included in this research,the three-category trajectories determined using GMM were classified as low,medium,and high,accounting for 39.7％,54.1％,and 6.2％of the participants,respectively.The two-category trajectories determined using the KML method were classified as low and high,representing 50.3％and 49.7％of the participants,respectively.The three-category trajectories determined using the KML method were classified as low,medium,and high,accounting for 31.1％,47.4％,and 21.5％of the participants,respectively.There were certain differ-ences in the growth patterns reflected by the early childhood BMI trajectories determined using different methods.Linear regression analysis found that after adjusting for maternal ethnicity,educational level,delivery mode,parity,maternal age at delivery,gestational week at delivery,children's gender,and breastfeeding at 1 month of age,the association between the high trajectory group in the three-category trajectories determined by the KML method(manifested by a slightly higher BMI at birth,followed by rapid growth during infancy and a stable-high BMI until 24 months)and BMI Z-scores at 3 years was the strongest.Logistic regression analysis revealed that the three-category trajectory grouping determined by the KML method had the best predictive performance for the risk of overweight at 3 years.The results were basically consistent after additional adjustment for the high bound score of the child's diet balanced index,average daily physical activity time,and screen time.Conclusion:This study used different methods to identify early childhood BMI trajectories with varying characteristics,and found that the high trajectory group determined by the KML method was better able to identify children with a higher risk of overweight in early childhood.This provides scientific evidence for selecting appropriate methods to de-fine early childhood BMI trajectories.

Objective:To investigate the effects of prophylactic flow restriction for brachiocephalic arteriovenous fistula on postoperative high-flow-related complications and patency rate in patients undergoing hemodialysis.Methods:Clinical data of patients with end-stage renal disease who underwent brachiocephalic arteriovenous fistula surgery for hemodialysis from February 2017 to May 2022 in Department of Nephrology,Sir Run Run Shaw Hospital,Zhejiang University School of Medicine were retrospectively analyzed.During surgery,a 4-5 mm vascular suture loop was placed around the vein near the anastomosis as a flow restriction device in 43 patients(flow restriction group),while 42 patients did not receive the prophylactic flow restriction ring(control group).All patients were followed up for 1 to 5 years.The incidence rates of complications related to the hemodialysis access pathway,including distal ischemia syndrome,the formation of arteriovenous fistula aneurysms,thrombus,high-flow congestive heart failure,anastomosis of the vein within 1 cm of the anastomosis and cephalic arch stenosis,were compared between the two groups.The natural blood flow rate of the arteriovenous fistula,anastomosis size,the internal diameter of the vein near the anastomosis,primary patency rate,assisted primary patency rate,and secondary patency rate of the arteriovenous fistula,were also evaluated and compared between the two groups.Logistic regression analysis was used to investigate the factors affecting arteriovenous fistula patency rates,as well as the impact of the flow-restricting ring on postoperative factors.Results:Ultrasound measurements showed that the internal diameter of the vein at the site of the flow restriction ring in the flow restriction group was(3.7±0.6)mm at three months postoperatively,which was significantly smaller than the internal diameter of the narrowest part of the vein near the anastomosis in the control group[(4.1±1.0)mm,t=-2.416,P＜0.01].The postoperative anastomotic diameter and natural blood flow rate of the arteriovenous fistula in the flow restriction group were both significantly lower than those in the control group(both P＜0.05).Furthermore,the incidence rates of various complications in the flow restriction group were significantly lower than those in the control group(all P＜0.05).At 6,12,and 24 months postoperatively,the primary patency rate and assisted primary patency rate in the flow restriction group were significantly higher than those in the control group(both P＜0.05),while there was no significant difference in secondary patency rates between the two groups(P＞0.05).Binary logistic regression analysis indicated that age,diabetes,and natural blood flow rate of the arteriovenous fistula at 3 months postoperatively were independent risk factors for primary patency rate,while the flow restriction for brachiocephalic arteriovenous fistula was an independent protective factor for primary patency rate(P＜0.01 or P＜0.05).The application of flow restriction was negatively correlated with anastomotic diameter at 6 and 12 months,natural arteriovenous fistula blood flow,and the incidence rates of cephalic arch stenosis and aneurysm formation(all P＜0.05).Conclusion:The prophylactic constriction during brachiocephalic arteriovenous fistula surgery in patients undergoing hemodialysis can limit the size of the anastomosis and postoperative arteriovenous fistula blood flow,reducing complications such as cephalic arch stenosis and high-flow heart failure,and increasing primary patency rates of arteriovenous fistula and delay the reintervention of the fistula.

Objective:To observe the clinical efficacy of exercise-based acupuncture in treating ischemic stroke and its effects in improving the patient's motor function and daily living ability. Methods:A total of 106 patients with motor dysfunction due to cerebral infarction were randomized into a trial group and a control group,each consisting of 53 cases.Both groups received conventional rehabilitation;in addition,the control group was given point-toward-point acupuncture at scalp points,and the trial group was offered exercise-based acupuncture,3 times weekly for 4 consecutive weeks.Before treatment,after 2 and 4 weeks of treatment,and at the 2-month follow-up,the Fugl-Meyer assessment(FMA)scale,Lovett scale for muscle strength,modified Ashworth scale(MAS),and activities of daily living(ADL)scale were used to assess the patient's motor function,muscle strength,muscle tension,and daily living ability. Results:After treatment,both groups gained improvements in motor function and daily living ability;after 4-week treatment and at the 2-month follow-up,the trial group had higher FMA and ADL scores than the control group(P<0.05).The MAS score decreased after treatment in both groups;after 2 and 4 weeks of treatment and at the 2-month follow-up,the MAS score was lower in the trial group than in the control group(P<0.05).After the intervention,the Lovett score increased in both groups;after 4 weeks of treatment,the Lovett score was higher in the trial group than in the control group(P<0.05). Conclusion:Based on routine rehabilitation,exercise-based acupuncture and scalp point-toward-point acupuncture both can improve the motor function and daily living ability in ischemic stroke patients;exercise-based acupuncture performs better than scalp point-toward-point acupuncture.

Objective To analyze the characteristics of imported malaria epidemic from overseas in Wuhan, to explore the management mechanism of on-site cases, and to accumulate experience for the treatment of imported malaria in large cities after malaria elimination. Methods The epidemiological data on imported malaria from abroad during the period of malaria elimination (2010-2019) in Wuhan were collected. The gender, age and severe illness-related factors of the cases were analyzed. Based on the characteristics of the epidemic and the current situation of prevention and control, the content and experience of the “Municipal-District 24-7” case mechanism were discussed. Results The medical resources in Wuhan were the best in the central region, resulting in a large number of imported malaria cases, with a total of 474 cases reported from 2010 to 2019 (40.79% of the total number of cases in Hubei Province), including 359 cases of falciparum malaria, 36 severe cases and one death (the death rate was 0.28%). The patients were mainly young and middle-aged men aged 20 to 49 years old (97.26%). There were many referral cases (40.30%), and there was no seasonal clustering of cases reported. The undiagnosed proportion at the first visit was 44.85%, and the time of attack-diagnosis was 4 days or more in 61.00% of cases. The occurrence of severe cases was related to unconfirmed diagnosis at the first visit (χ²=35.46, P<0.001) and attack-diagnosis time (Z=-6.49, P<0.001). Conclusion Imported malaria occurs frequently in Wuhan, mainly falciparum malaria. However, “Municipal-District 24-7” case mechanism has effectively curbed the occurrence of severe and death cases and provided valuable experience for case management in similar cities in China.

The regulation of spermatogonial proliferation and apoptosis is of great significance for maintaining spermatogenesis. The single-cell RNA sequencing (scRNA-seq) analysis of the testis was performed to identify genes upregulated in spermatogonia. Using scRNA-seq analysis, we identified the spermatogonia upregulated gene origin recognition complex subunit 6 (Orc6), which is involved in DNA replication and cell cycle regulation; its protein expression in the human and mouse testis was detected by western blot and immunofluorescence. To explore the potential function of Orc6 in spermatogonia, the C18-4 cell line was transfected with control or Orc6 siRNA. Subsequently, 5-ethynyl-2-deoxyuridine (EdU) and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assays, flow cytometry, and western blot were used to evaluate its effects on proliferation and apoptosis. It was revealed that ORC6 could promote proliferation and inhibit apoptosis of C18-4 cells. Bulk RNA sequencing and bioinformatics analysis indicated that Orc6 was involved in the activation of wingless/integrated (Wnt)/ β-catenin signaling. Western blot revealed that the expression of β-catenin protein and its phosphorylation (Ser675) were significantly decreased when silencing the expression of ORC6. Our findings indicated that Orc6 was upregulated in spermatogonia, whereby it regulated proliferation and apoptosis by activating Wnt/β-catenin signaling.

【Objective】 To explore the risk factors of transfusion-related acute lung injury (TRALI). 【Methods】 The clinical symptoms, signs, imaging examinations, and laboratory test results of two patients with TRALI after blood transfusion were retrospectively analyzed, and human leukocyte antigen (HLA) genotyping of the patient and HLA antibodies typing of the plasma donors were performed. 【Results】 The clinical manifestations and laboratory parameters of two patients were consistent with those of TRALI after blood transfusion. After timely clinical respiratory support treatment, all patients were improved. Blood donors produced high titers of HLA-Ⅱ antibodies after pregnancy, including antibodies that specifically recognize the patient′s HLA antigen. 【Conclusion】 Two patients developed TRALI after platelet transfusion from a female blood donor, which was caused by HLA-Ⅱ antibodies.

OBJECTIVES: To explore the effect mechanism of electroacupuncture (EA) on functional constipation (FC) at the combined lower he-sea and front-mu points of large intestine based on enteric neuronal autophagy. METHODS: A total of 40 SPF Kunming mice were randomly divided into 5 groups (n = 8), i.e. a control group, a model group, an acupuncture group, a 3-methyl adenine (3-MA) group, and a 3-MA + acupuncture group. Except the control group, the FC model was established by gavage with compound diphenoxylate suspension for 14 days in the other 4 groups. After successful modeling, the mice of the acupuncture group and the 3-MA + acupuncture group received EA at bilateral "Tianshu" (ST 25) and "Shangjuxu" (ST 37), stimulated for 30 min with disperse-dense wave, 2 Hz/15 Hz of frequency, 1 mA of intensity. EA was delivered once daily. One course of treatment was composed of 5 days and 2 courses were needed, with an interval of 2 days. An intraperitoneal injection of 3-MA (15 mg/kg) was administered 30 min before EA in the mice of the 3-MA group and the 3-MA + acupuncture group, once daily. Before and after intervention, the time of the first black stool defecation and defecation behaviors in 6 h were observed in each group. After intervention, in every group, the small intestine propulsion rate was calculated, the colon tissue morphology was observed using HE staining, the ultrastructure of enteric neuronal autophagy was observed under transmission electron microscope, and the expressions of microtubule-associated protein 1 light chain 3 (LC3), Beclin-1 and neuronal nuclear antigen protein (NeuN) in neurons of colonic muscularis were determined by immunohistochemistry. RESULTS: Before intervention, when compared with those in the control group, the time of the first black stool defecation was prolonged (P<0.01, P<0.05), and numbers (P<0.01), wet weight (P<0.01, P<0.05) and water content (P<0.05, P<0.01) of stool in 6 h were reduced in the model, acupuncture, 3-MA and 3-MA + acupuncture groups. After intervention, compared with those in the control group, the time of the first black stool defecation was longer (P<0.05), and numbers (P<0.01), wet weight (P<0.01) and water content (P<0.01) of stool in 6 h were decreased in the model group. The time of the first black stool defecation was shortened (P<0.01), and numbers (P<0.01), wet weight (P<0.01) and water content (P<0.01) of stool in 6 h were increased in the acupuncture group when compared with those in the model group. The time of the first black stool defecation was extended (P<0.01), and numbers (P<0.01), wet weight (P<0.01) and water content (P<0.01) of stool in 6 h were declined in the 3-MA + acupuncture group in comparison with those in the acupuncture group. All layers of colon tissue were normal and intact in each group. When compared with the control group, the small intestine propulsion rate and the average optical density (OD) values of LC3, Beclin-1 and NeuN in neurons of colonic muscularis were decreased (P<0.01), and autophagosomes were dropped in the model group. In the acupuncture group, the small intestine propulsion rate and the average OD values of NeuN, LC3 and Beclin-1 in neurons of colonic muscularis increased (P<0.01,P<0.05), and autophagosomes were elevated when compared with those in the model group. The small intestine propulsion rate and the average OD values of NeuN, LC3 and Beclin-1 in neurons of colonic muscularis were dropped (P<0.05,P<0.01) in the 3-MA + acupuncture group in comparison with those in the acupuncture group. CONCLUSIONS Electroacupuncture may promote enteric neuronal autophagy and increase the number of neurons so that the intestinal motility can be improved and constipation symptoms can be relieved in FC mice.
Mice ; Animals ; Electroacupuncture ; Beclin-1 ; Acupuncture Points ; Constipation/therapy* ; Intestine, Small ; Autophagy ; Water