Objective:To accurately ascertain the whole genome sequencing and the composition of open reading frames (ORFs) of non-replicating Tiantan strain of vaccinia virus (NTV) using next-generation long-read sequencing technology.Methods:NTV, obtained from our laboratory stock, was amplified and purified on chicken embryo fibroblast cells(CEFs), and the full-length genomic nucleic acid of NTV was extracted. The PacBio HiFi sequencing platform was utilized for de novo assembly to obtain the complete genomic sequence of NTV. Using a homology annotation strategy, we identified its ORF composition and compared it with known non-replicating vaccinia virus strains. Results:The total length of NTV′s genome was 171 729 bp, with a GC content of 33%. Its unique inverted terminal repeat (ITR) region comprised hairpin structures, two tandem repeat regions, and three non-repeat regions. NTV contained 166 ORFs, with major differences observed in the ITR and its surrounding regions when compared to MVA-BN and NYVAC. These three strains shared a common set of 138 ORFs. NTV encoded six unique ORFs related to virus evasion of host antiviral response.Conclusions:This study accurately determines the whole genome sequencing and ORFs composition of NTV, and reveals its similarities and differences with other replication-deficient vaccinia virus strains, which pave a way for the development and application of the next generation of monkeypox vaccines and novel viral vectors.

Objective:To construct recombinant influenza viruses expressing Gaussia luciferase (Gluc) with different influenza virus backbones and analyze their growth characteristics, genetic stability, ability to express Gluc and in vitro anti-influenza drug activity. Methods:The C-terminal of PR8NA was modified by inserting the porcine teschovirus-2A autocleavage peptide (P2A) and the Gluc-coding gene. Recombinant viruses, PR8NAGluc/PR8 and PR8NAGluc/WSN, were rescued using the eight-plasmid system of influenza virus reverse genetics, with seven plasmids derived from A/Puerto Rico/8/34(PR8) (H1N1) and A/WSN/1933 (WSN) H1N1. The genetic stability of the recombinant viruses was verified by RT-PCR. The fluorescence activity and the growth kinetics of the two recombinant viruses were compared. The correlation between the fluorescence activity of PR8NAGluc/WSN and median tissue culture infective dose (TCID 50), and the anti-drug activity of PR8NAGluc/WSN against oseltamivir, favipiravir, and Lianhua Qingwen in vitro were also analyzed. Results:The Gluc-expressing recombinant viruses constructed using PR8 and WSN backbones were successfully rescued by reverse genetics. Compared with the PR8 backbone, the WSN backbone significantly improved the fluorescence activity of Gluc. Moreover, the PR8NAGluc/WSN virus expressed stably in embryonated egg, and its replication kinetics was slightly lower than that of wild type. The fluorescence activity of PR8NAGluc/WSN virus had a good correlation with its TCID 50. The PR8NAGluc/WSN virus was sensitive to oseltamivir, favipiravir and Lianhua Qingwen. Conclusions:The recombinant virus with a WSN backbone exhibited higher fluorescence expression intensity as compared with the recombinant virus with a PR8 backbone. This study provided reference for high-throughput screening of anti-influenza drugs and the development of influenza virus vector vaccines.

Objective    To summarize the experience and lessons of right ventricular decompression in children with pulmonary atresia and intact ventricular septum (PA/IVS) and to reflect on the strategies of right ventricular decompression. Methods    The clinical data of 12 children with PA/IVS who underwent right ventricular decompression in our hospital from March 2015 to December 2019 were reviewed retrospectively. There were 10 males and 2 females with a median age at the time of surgery was 5 d (range, 1-627 d). Correlation analysis between the pulmonary valve transvalvular pressure gradient and changes in Z score of tricuspid valves after decompression was performed. Results    One patient died of refractory hypoxemia due to circulatory shunt postoperatively and family members gave up treatment. There were 2 (16.67%) patients received postoperative intervention. The pulmonary transvalvular gradient after decompression was 31.95±21.75 mm Hg. Mild pulmonary regurgitation was found in 7 patients, moderate in 2 patients, and massive in 1 patient. The median time of mechanical ventilation was 30.50 h (range, 6.00-270.50 h), and the average duration of ICU stay was 164.06±87.74 h. The average postoperative follow-up time was 354.82±331.37 d. At the last follow-up, the average Z score of tricuspid valves was 1.32±0.71, the median pressure gradient between right ventricle and main pulmonary artery was 41.75 mm Hg (range, 21-146 mm Hg) and the average percutaneous oxygen saturation was 92.78%±3.73%. Two children underwent percutaneous balloon pulmonary valvoplasty at 6 and 10 months after surgery, respectively, with the rate of reintervention-free of 81.8%. There was no significant correlation between pulmonary transvalvular gradients after decompression and changes in Z score of tricuspid valves (r=–0.506, P=0.201). Conclusion    For children with PA/IVS, the simple pursuit of adequate decompression during right ventricular decompression may lead to  severe pulmonary dysfunction, increase the risk of ineffective circular shunt, and induce refractory hypoxemia. The staged decompression can ensure the safety and effectiveness for initial surgery and reduce the risk of postoperative death.

Objective:To review and analyze the clinical experience of children with aortic valve stenosis and/or insufficiency treated with autologous pulmonary valve for aortic valve replacement procedure(Ross operation) with ePTFE artificial valve for right ventricular outflow tract reconstruction.Methods:From 2015 to 2020, 8 cases of aortic stenosis and/or aortic insufficiency treated by Ross operation in our center were collected, with an age of 0.5-13.2 years old. 4 cases of aortic stenosis were diagnosed preoperatively, 3 cases of aortic stenosis with aortic insufficiency, and 1 case of infective endocarditis involving the aortic valve. The operation was carried out in three steps: Harvest autologous pulmonary valve; the diseased aortic valve was resected and autologous pulmonary valve was transplanted to the aorta by aortic root transplantation; the right ventricular outflow tract was reconstructed by a handmade ePTFE artificial flap blood vessel.Results:In 6 cases, the right ventricular outflow tract was reconstructed by hand-sewn ePTFE trileaflets, and artificial univalve in 2 cases, no death occurred during operation; all patients were cured and discharged. The patients were followed up for 1 to 36 months, with mean of(12.63±12.19) months. There was no long-term death or valvular complications. During follow-up echocardiography indicated 1 case of moderate aortic regurgitation, 1 case of mild-moderate regurgitation, and moderate regurgitation was found in 2 patients with artificial single pulmonary valve. For the remaining patients, they were mild aortic regurgitation, and a trivial or mild pulmonary artery regurgitation with hand-sewn three-leaflets ePTFE artificial vessel; All patients were followed up at the last time with a peak pressure of(6.63±3.46) mmHg(1 mmHg=0.133 kPa) across the aortic valve. The left ventricular outflow tract and aortic annulus shrank slightly after surgery(the diameter of one patient with Ross-Konno operation increased), but the annulus diameter increased with age. There was no need for further intervention.Conclusion:The Ross operation is safe for the treatment of aortic valve disease, it has good hemodynamic effect, and the autologous pulmonary artery has growth potential, especially suitable for children and young patients. Hand-sewn ePTFE with trileaflet vessels for reconstruction of right ventricular outflow tract performs well in anti-regurgitation function in the short term or may be used as a replacement material for the homograft/heterograft vessel, but longer follow-up and more cases are needed.

Objective:To discuss the effect of modified ultrafiltration combined with sequential infusion of blood products, such as platelets and cryoprecipitation, on perioperative coagulation function in neonates undergoing cardiovascular surgery under extracorporeal circulation.Methods:A retrospective analysis was performed on 83 neonates who underwent cardiovascular surgery from January 2017 to December 2019. They were divided into the control group (conventional ultrafiltration, 51 cases) and the treatment group (modified ultrafiltration + sequential infusion of blood products, 32 cases).Results:The age of treatment group was significantly younger than that of the control group ( P<0.05). The extracorporeal circulation time of the treatment group was significantly longer than that of the control group. Compared with the preoperative data, post-APTT, post-PT and post-INR were increased significantly in the control group, platelet count and FIB were decreased significantly ( P<0.01), while there was no significant difference in the treatment group. Compared with the treatment groups, post-APTT, post-PT and post-INR were significantly increased in the control group, and postoperative platelet count and FIB were significantly decreased ( P<0.05). The drainage of control group was significantly higher than that of the treatment group at 12 h and 24 h after surgery ( P<0.01). Conclusion:The application of modified ultrafiltration combined with sequential infusion of blood products can significantly improve perioperative coagulation and reduce mediastinal bleeding in neonatal after cardiovascular surgery.

Objective To review the clinical experience with the Norwood stage Ⅰprocedure.Methods Between June 2016 to October 2018, totally 5 neonates underwent Norwood stage Ⅰ procedure.There were 3 boys and 2 girls, weighing(2. 98 ±0.60)kg with median of 2.95 kg.Age at surgeries ranged from 1 to 8 days.All 5 cases underwent the Norwood stage Ⅰprocedure under deep hypothermic circulatory arrest, including 3 cases of modified Blalock-Taussig shunts (MBTS) and 2 ca-ses of RV-PA shunt.Results The third case was successfully closed the chest on postoperative day 2 and extubated, but died from DIC due to severe infection on the postoperative day 6.The fifth case died from pericardiol tamponade at 10 hours after the operation.The first,second and fourth cases were followed up 1 month after discharge with NYHA Ⅰ, but the first and second cases dropped out of follow-up due to some personal resaons.The fourth case underwent the bidirectional Glenn procedure 9 monthes after the stage Ⅰ procedure and recovered smoothly.Conclusion The application of these modified methods and te-chiques based on the classical Norwood procedure help to accumulate experience in sugical treatment of HLHS in China .

Objective Norwood Stage Ⅰ is the standard procedure to cope with hypoplastic left heart syndrome (HLHS),which continues to be the most challenging congenital heart disease.The aim of this study is to retrospectively compare the classical perioperative management of Norwood Stage] with the modified strategy.Methods Between June 2010 and November 2017,totally 10 patients with HLHS underwent the standard Norwood Stage Ⅰ procedure.They are stratified to two tiers:Group A,from June 2010 to August 2014,there were 5 boys.Age at surgeries ranged from 29 to 75 days,and weight 2.57-3.50 kg with median of 3.13 kg.Group B,from August 2014 to November 2017,there were 4 boys and 1 girl.Age at surgeries ranged from 6 to 22 days,and weight from 2.0-3.1 kg.In Group A,all 5 cases underwent the standard Norwood Stage.procedure under deep hypothermic circulatory arrest,including 4 cases of modified Blalock-Taussig shunt (MBTS) and 1 case of RV-PA shunt.In Group B,all 5 cases adopted side graft technique and RV-PA shunt,aortic arch and ascending aorta were reconstructed with treated bovine pericardial patch.Group B used incubators to adjust systemic vascular resistance instead of vasodilators.Results Group A's early mortality is 40％;Group B's early mortality is 20％,1 case died of tamponade.Conclusion The standard Norwood Stage Ⅰ procedure is a complex procedure,which demands multidisplinary cooperation,to palliatively correct HLHS.To adjust and find a suitable perioperative managements can improve the results.Sharing experiences on perioperative managements of Norwood Stage Ⅰ between heart centers in China will be helpful to decrease the mortality and morbidity in relatively short period.

Objective To analysis the the strategy of repairing coarctation of aorta with hypoplastic aortic arch in neo-nate.Methods Collected consecutive 24 neonates suffered coarctation of aorta with hypoplastic aortic arch form January 2015 to March 2017,12 patients were underwent aortic arch reconstruction with extremely extended end-to-end anastomosis under cardiopulmonary bypass(CPB) through the media sternotomy approach(CPB group), while another 12 cases were under coarc-tation repair using end-to-end anastomosis or end-to-side anastomosis through left posterolateral thoracotomy approach ( control group).The postoperative blood pressure,echocardiography and cardiac CT scan were used to evaluate the result of the aortic arch rconstruction.The pressuregradientof the upper limb and lower limb,flow velocity of the anastomoses, aortic arch morphol-ogy,rate of the residual abstruction were compared between the two groups.Results Mechanical ventilation time[ CPB group (17 ±27)h vs.control group(44 ±52)h, P＜0.05], ength of stay in ICU[CPB group(3.75 ±1.36)days vs.control group (6.54 ±5.08)days, P＜0.05], all patients were followed up for 1－12 months,the aortic residual pressure[CPB group(14 ± 10)mmHg (1 mmHg＝0.133 kPa) vs.control group(26 ±17)mmHg,P ＜0.05) ], flow velocity of the anastomoses[CPB group(2.32 ±0.78)m/s, control group(1.55 ±0.99)m/s, P ＜0.05 ], pressuregradientof the upper limb and lower limb [CPB group (6.67 ±3.49)mmHg, control group(7.41 ±4.29)mmHg, P＞0.05].There was no died in two gruops.Con-clusion Anastomosis with end to end techinique under cardiopulmonary bypass may achieve better effect in neonates with co-arctation of aorta with hypoplastic aortic arch,it has better duplcity, operability and early curative effect,but the long-term effect need the multicenter and large sample-volume study to explore.

Objective:To observe the influence of sevoflurane pretreatment in the lung function of the infants during heart operation by cardiopulmonary bypass (CPB), and to explore its lung protection and possible mechanism.Methods:Sixty infants with ventricular septal defect were enrolled at age less than 1 year old and randomly assigned to pretreatment group and control group (n=30).After the induction of general anesthesia and tracheal intubation,the patients in pretreatment group received continuous inhalation of 1.0 MAC sevoflurane until the beginning of CPB.Inhale sevoflurane was absent in control group.The duration of ventilator support of the infants in two groups was recorded.The Pplate,CL,OI,A-aDO2 ,RI,the number of leukocytes and neutrophils segregated in lung of the patients were compared between two groups at the four time points T0 (before aorta clamping),T1 (30 min after aorta declamping),T2,and T3 (2 h and 6 h after CPB).Results:Compared with control group,the duration of ventilator support of the infants in pretreatment group was obviously shortened (P <0.05).In each group,the CL and OI were significantly decreased (P < 0.05 or P < 0.01),while the Pplate, A-aDO2 ,RI,the number of leukocyte and neutrophils segregated in lung were significantly increased (P <0.05 or P <0.01)at T1,T2,T3 time points compared with T0 time point.The CL and OI in pretreatment group were significantly increased (P <0.05 or P <0.01);the Pplate,A-aDO2,RI,the number of leukocytes and neutrophils segregated in lung in pretreatment group were significantly decreased at T1,T2,and T3 (P <0.05 or P <0.01) compared with control group.Conclusion:Sevoflurane pretreatment might play a role in decreasing the leukocyte adhesion and protecting the lung function in the infants during opening heart operation by CPB.

Objective: To investigate the effect of cold self-blood cardioplegia with ulinastatin on immature myocardial cell apoptosis and protein expressions of Bcl-2, Bax in ventricular septal defect (VSD) infants.
Methods: A total of 60 infants received VSD repairing operation with cardiopulmonary bypass (CPB) in our hospital were summarized. The patients were randomly divided into 2 groups:Test group, the infants received cold self-blood cardioplegia with ulinastatin when aortic cross-clamp was closed. Control group, the infants received cold self-blood cardioplegia when aortic cross-clamp was closed. n=30 in each group. The right atrium tissue was collected before CPB and 10 min after releasing aortic cross-clamp. The index of myocardial cell apoptosis was observed by TUNEL method, and the protein expressions of Bcl-2, Bax were examined by immunohistological method.
Results: Both groups showed the higher index of myocardial cell apoptosis at 10 min after releasing aortic cross-clamp than 5 min before CPB, and the apoptosis index in Test group was lower than that in Control group, all P<0.05. The protein expressions of Bcl-2 and Bax were obviously increased at 10 min after releasing aortic cross-clamp than 5 min before CPB in both groups. Compared with Control group, Test group presented the higher Bcl-2 protein expression and lower Bax protein expression, all P<0.05.
Conclusion: Cold self-blood cardioplegia with ulinastatin could protect immature myocardum from ischemia-reperfusion injury in VSD infants during CPB operation in clinical practice.