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Objective:To investigate the whole genome characteristics of coxsackievirus A4 (CVA4) circulating in Qingdao city.Methods:Four CVA4 isolates circulating in Qingdao city during 2013 to 2015 were selected. Whole genome sequences of these strains were amplified by one-step reverse transcription-polymerase chain reaction (RT-PCR). Sequence alignment and phylogenetic analysis were performed using MEGA7.0 software package. Genetic recombination analysis was performed using similarity plots 3.5.1 software package.Results:Phylogenetic analysis showed that based on the sequences of the whole genome and P1, P2 and P3 regions, HS312/QD/CHN/2013 and HS605/QD/CHN/2014 strains together with the early domestic isolates belonged to the same clade, while FY218/QD/CHN/2015 strain and CV-A4/P1033/2013/China strain collected in Wenzhou in 2013 formed another clade in each phylogenetic tree. HS144/QD/CHN/2014 strain belonged to the same clade as HS312/QD/CHN/2014, HS605/QD/CHN/2014 and the early domestic CVA4 isolates in the phylogenetic tree based on the P1 region, but formed a separate clade in the phylogenetic trees based on the whole genome, P2 region and P3 region. Genetic recombination analysis revealed that there was genetic recombination between HS144/QD/CHN/2014 strain and the CVA2 strain of CV-A2/P373/2013/China isolated in mainland China in 2013 in the region of 2C-3D (5 081-7 301); FY218/QD/CHN/2015 and CV-A4/P1033/2013/China strains were highly homologous and recombination signal sequences were detected in the region of 2A-2B (3 821-4 161) between the two strains and the CVA2 strain of CV-A2/P373/2013/China.Conclusions:The CVA4 isolates circulating in Qingdao city presented obvious genetic diversity at the genome-wide level.
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Objective@#To investigate the etiology spectrum of hand foot and mouth disease (HFMD) and to analyze the molecular characteristics of Coxsackievirus A10 and A6 in Qingdao in 2014.@*Methods@#Throat swabs of HFMD cases were tested for total enteroviruses (EVs), EV-A71, CV-A16, CV-A10 and CV-A6 by multiplex real time RT-PCR. Other EV serotypes were identified through the sequences of partial VP1 gene. The full-length of VP1 gene of CV-A10 and CV-A6 were amplified and sequenced. The sequences were phylogenetically analyzed using MEGA 5.0 software package.@*Results@#A total of 1727 HFMD patients were detected in 2014 and 11serotypes of enteroviruses were identified. EV-A71(38.0%, 410/1078), CV-A16(28.8%, 311/1078), CV-A10(14.1%, 152/1078)and CV-A6(3.2%, 34/1078)were the most dominant pathogen in 2014 in Qingdao. Proportions of CV-A10 in enterovirus infected children varied dramatically in different ages(χ2=15.19, P=0.001), showing a downtrend with age. Proportion of CV-A10 in inpatients was much higher than that in outpatients(χ2=49.1, P <0.001), while 60% of those inpatients showed nervous system damage symptoms, with pathological reflex or disappearance of physiological reflex. Phylogenetic analysis of complete VP1 sequences showed that all CV-A10 strains identied in this study belonged to genotype C, 71.7% of which located in branch C5; all CV-A6 strains belonged to genotype D while 83.3% of which located in branch D5.@*Conclusions@#EV-A71, CV-A16, CV-A10 and CV-A6 were the prevalent pathogens of HFMD in Qingdao in 2014. CV-A10 was more frequently detected in lower age group with HFMD, which can cause damage to nervous system. Strains of branch C5 in genotype C were the dominant strains of CV-A10 circulated in Qingdao in 2014 while strains of branch D5 in genotype D were the major strains of CV-A6.
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Objective@#To analyze the molecular epidemiology of norovirus (NoV) genotype GⅡ.15 in Qingdao City.@*Methods@#One thousand four hundred and twelve stool samples were collected from suspected NoV infected patients and detected by real-time polymerase chain reaction (PCR). Open reading frame (ORF)1-ORF2 and VP1 gene were amplified by reverse transcription (RT)-PCR and sequenced for genotyping, evolutionary analysis and homology modeling.@*Results@#Seven cases of GⅡ.15 type were detected including four sporadic cases and one outbreak.The VP1 gene was highly homologous and had little variation compared with early strain J23/US/1999. The differences of amino acids between strains in Qingdao City were mainly asparagine/asparticacid(N/D)300 and proline/serine(P/S)302.Homology modeling suggested that VP1 of GⅡ.15 strain was composed of S domain and P domain (P1 subdomain included 224-276 and 431-555, P2 subdomain included 277-430). S domain contained eight anti-parallel β-sandwiches and two α-helixes, and P1 subdomain contained one α-helix and seven β-strands, and the P2 subdomain folded into a compact barrel-like structure consisting of six β-strands.Argnine (R)-glycine (G)-valine (V)-motif (289-291) and three specific loci including glutarnine (Q)313, asparagine (N)349 and Q389 were located in the P2 subdomain, with NGR-motif (265-267) located at 22nd upstream of RGV-motif.Site I (SNR-alanine(A)- histidine(H)357-361), Site Ⅱ (D388) and Site Ⅲ (G454, G455) were the main characteristic sites of histo-blood group antigens (HBGA) binding interface, which may be similar to the binding pattern of GⅡ.4 type VA387 and HBGA.@*Conclusion@#Although GⅡ.15 type NoV evolves very slowly, it may still have the risk to become an epidemic strain, which needs to be monitored and further studied.
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Objective To analyze the molecular epidemiology of norovirus (NoV) genotype G Ⅱ.15 in Qingdao City.Methods One thousand four hundred and twelve stool samples were collected from suspected NoV infected patients and detected by real-time polymerase chain reaction (PCR).Open reading frame (ORF) I-ORF2 and VP1 gene were amplified by reverse transcription (RT)-PCR and sequenced for genotyping,evolutionary analysis and homology modeling.Results Seven cases of GⅡ.15 type were detected including four sporadic cases and one outbreak.The VP1 gene was highly homologous and had little variation compared with early strain J23/US/1999.The differences of amino acids between strains in Qingdao City were mainly asparagine/asparticacid(N/D) 300 and proline/serine (P/S) 302.Homology modeling suggested that VP1 of GⅡ.15 strain was composed of S domain and P domain (P1 subdomain included 224-276 and 431-555,P2 subdomain included 277-430).S domain contained eight anti-parallel β3-sandwiches and two α-helixes,and P1 subdomain contained one α-helix and seven β3-strands,and the P2 subdomain folded into a compact barrel-like structure consisting of six β-strands.Argnine (R)-glycine (G)-valine (V)-motif (289-291) and three specific loci including glutarnine (Q)313,asparagine (N)349 and Q389 were located in the P2 subdomain,with N GR-motif (265-267) located at 22nd upstream of RGV-motif.Site Ⅰ (SNR-alanine (A)-histidine (H)357-361),Site Ⅱ (D388) and Site IⅢ (G454,G455) were the main characteristic sites of histo-blood group antigens (HBGA) binding interface,which may be similar to the binding pattern of G Ⅱ.4 type VA387 and HBGA.Conclusion Although G Ⅱ.15 type NoV evolves very slowly,it may still have the risk to become an epidemic strain,which needs to be monitored and further studied.
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Carcinoma of pancreatic body and tail is a high invasive disease with a low resectability rate.It was once believed that celiac axis infiltration usually contraindicated resection.Distal pancreatectomy with en bloc celiac axis resection (DP-CAR) is described as a new treatment method of this disease.In recent years,more and more literatures have reported this operation,but they were case reports or small sample retrospective study,the results of which differed according to the different treatments and perioperative managements in different centers.The advantages and disadvantages of DP-CAR are still controversial.Research progress of DP-CAR is reviewed in this article.
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Objective To investigate the molecular epidemiological characteristics of hemaggluti-nin (HA) and neuraminidase (NA) of influenza B viruses (IBV) isolated in Qingdao from 2011 to 2018. Methods A total of 12236 samples of influenza-like cases in Qingdao from 2011 to 2018 were collected to extract viral RNAs. All samples were screened for influenza A viruses ( IAV) and IBV by one-step multiplex real-time RT-PCR. Lineages of IBV were identified. One hundred and eighty-two strains of IBV were select-ed to amplify HA and NA genes by RT-PCR and then analyzed by sequencing. Phylogenetic analysis and variation analysis of genes and amino acids were carried out. Results IBV was detected almost every year in Qingdao from 2011 to 2018. The positive rate was only slightly lower than that of IAV ( 4. 99％ vs 6. 21％). B/Victoria linkage had two prominent epidemic years (2011-2012, 2015-2016), while B/Yama-gata linkage had three (2013-2014, 2014-2015, 2017-2018). Most of the infected people were children un-der 10 years old, and the people infected with the two lineages had similar age characteristics. Phylogenetic analysis of HA genes showed clusters in Victoria clades of 1A and 1B and Yamagata clades of 2 and 3. IBV of Yamagata lineage had more amino acid mutation sites than those of Victoria lineage in HA genes with grea-ter genetic diversity. The B/Yamagata strains had 12 amino acid mutations and the B/Victoria strains had seven in four major epitopes. In the receptor binding sites, two amino acid mutations were detected in the B/Yamagata strains and three in the B/Victoria strains. In Qingdao, 26 strains of IBV were intra-lineage reas-sortments, mostly of the B/Victoria lineage, and 23 strains were inter-lineage reassortments, mostly between HA-B/Yamagata and NA-B/Victoria strains. A possible resistant strain to NA inhibitor was found. Conclu-sions The significance of IBV in seasonal influenza should not be neglected. Amino acid substitution, in-sertion/deletion and gene reassortment were the main strategies for the natural evolution of IBV. Influenza surveillance was of great importance and influenza vaccine strains needed to be updated in time.
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Objective@#To illuminate the gene characteristics and clinical characterization of Coxsackievirus B5 (CV-B5) strains isolated from patients with sevre hand, foot and mouth disease (HFMD) in Qingdao city.@*Methods@#A total of 1 844 patients of HFMD were consecutively admitted to Qingdao Women and Children's Hospital from 2013 to 2014. Information of the study population described above was collected retrospectively. The samples were collected from at least 1 site (throat swab, cerebrospinal fluid), which viral nucleic acid extracted and the entire VP1 gene sequences of CV-B5 isolates were amplified and sequenced, then the homology and phylogeny analysis were conducted by MEGA7.0. The prototype Faulkner strain and other VP1 amino acid sequences were derived from the GenBank database.@*Results@#A total of 8 CV-B5 positive cases were obtained, including 4 males and 4 females; 6 severe hospitalized cases and 2 outpatients. The age of 6 hospitalized patients ranged from 3 to 48 months, with a median of 26 months. For the six inpatients, fever, convulsions vomiting, diarrhea and rash were the main clinical manifestation, and all combined with viral encephalitis. Compared with the prototype strain Faulkner, in the VP1 region,the nucleotide and the amino acid homologies was 77.3%-78.8% and 95.5%-97.0% respectively. Five out of the six severe cases with substitution of serine (S) to asparagine (N) at amino acid site 95 in the VP1 region. The sequences of 8 CV-B5 strains were classified into genogroup D.@*Conclusion@#Hand, foot and mouth disease associated with CV-B5 virus infection can result in nervous system involvement and the main complication was viral encephalitis. The CV-B5 strains associated with severe hand, foot and mouth disease had high nucleotide homology and present a certain regional aggregation.
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Objective To investigate the molecular characteristics of coxsackievirus A12 ( CV-A12) and to understand the clinical manifestations of severe hand, foot and mouth disease (HFMD) caused by CV-A12 in Qingdao. Methods Throat swabs of HFMD, herpangina and influenza-like cases from 2011 to 2016 were detected for enteroviruses ( EVs) in Qingdao. Human rhabdomyosarcoma ( RD) and human la-ryngeal carcinoma (Hep-2) cells were used for virus proliferation and CV-A12 strains were identified through a semi-nest RT-PCR. The full-length of VP1 gene of CV-A12 strains was sequenced and phylogenetically an-alyzed using MEGA7. 0 software package. Clinical data of severe HFMD cases positive for CV-A12 were col-lected and analyzed. Results CV-A12-positive HFMD, herpangina and influenza-like cases accounted for 0. 3％(18/6798), 1. 2％(2/169) and 0. 1％(1/676) in Qingdao, respectively. Most of the HFMD caused by CV-A12 in children were mild before 2013 (84. 6％, 11/13), while hospitalized severe cases with neurological symptoms (100％, 5/5) became more common after 2013. Phylogenetic analysis of the VP1 region revealed that CV-A12 strains worldwide could be divided into two genotypes, A and B. All of the CV-A12 strains successfully sequenced in Qingdao from 2011 to 2016 belonged to genotype B, and 88. 9％(16/18) of them belonged to subgenotype B2. All hospitalized severe cases of CV-A12-caused HFMD after 2013 were associated with strains in branch B2b of subgenotype B2. Conclusion CV-A12 was one of the pathogens causing HFMD, herpangina and influenza-like illness in children in Qingdao. Strains of genotype B2 were the predominant CV-A12 strains circulating in Qingdao in recent years. CV-A12-caused HFMD might complicated by nervous system damage.
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Gastric resection can cause a multifactorial clinical manifestations of dyspepsia,such as flatulence,diarrhea,weight loss,and fat diarrhea.Exocrine pancreatic insufficiency (EPI) is one of the possible mechanisms of fat maldigestion following gastric surgery,the main causes may be related to rapid gastric emptying;asynchrony between gastric emptying and bilio-pancreatic secretion due to new tracts of various reconstructions;bacterial overgrowth after gastrectomy and so on.Oral pancreatic enzyme replacement therapy (PERT) is the mainstay of treatment for EPI,due to lack of available evidence so far,the efficacy and safety of pancreatic enzyme substitution in patients following gastric resection remains unclear and cannot be generally recommended.This review will sum up the revelant studies addressing EPI and PERT after gastric resection in recent years,and summarizes the mechanisms,clinical diagnostic methods and PERT treatment perscription of EPI after gastrectomy to improve the cognition of clinicans.
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Objective To analyze the molecular epidemiological characteristics of norovirus (NoV) outbreaks in Qingdao between 2014 and 2016.Methods Stool samples were collected from NoV outbreaks between January 2014 and December 2016 and detected by real-time RT-PCR.NoV open reading frame 1 (ORF1) and ORF2 were partially amplified by RT-PCR.The amplified products were further analyzed by gene sequencing and genotyping.Phylogenetic analysis was conducted by using MEGA 6.0 software package.Results A total of 23 NoV outbreaks, involving 260 cases, were reported during 2014 to 2016.Of all collected stool samples, 128 were positive for NoV including 6 of genogroupⅠ (GⅠ) and 122 of genogroupⅡ (GⅡ).All positive samples were genotyped into 6 genotypes, which were GⅡ.P17-GⅡ.17, GⅡ.P12-GⅡ.3, GⅡ.P7-GⅡ.6、GⅡ.P2-GⅡ.2, GⅠ.Pb-GⅠ.6 and GⅡ.Pg-GⅡ.12.The 23 outbreaks included both single infections and mixed genotype infections, which were 11 of GⅡ.17 single infection, 4 of GⅡ.3 single infection, 3 of GⅡ.17 and GⅡ.3 mixed infection, 2 of GⅡ.17 and GⅡ.6 mixed infection, 1 of GⅠ.6 single infection, 1 of GⅡ.17 and GⅡ.2 mixed infection and 1 of GⅡ.17 and GⅡ.12 mixed infection.Conclusion NoV was an important pathogen responsible for viral diarrhea outbreaks in Qingdao.Several different genotypes were detected.The newly variant GⅡ.P17-GⅡ.17 was the predominant epidemic strain causing norovirus outbreaks in Qingdao during 2014 to 2016.
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Objective@#To analyze the molecular epidemiology of Norovirus(NoV) infection among sporadic hospitalized adults with diarrhea in Qingdao, 2015.@*Methods@#Four hundred and nine stool samples were collected from hospitalized adults with diarrhea and detected by Real-time RT-PCR. For genotyping, ORF1 and ORF2 were partially amplified by RT-PCR and sequenced.@*Results@#18.1%(74/409) of stool samples were positive for NoV genogroup I(GI) (10/74) and genogroup II(G II) (64/74). Fifty-three positive samples of GII were sequenced and divided into 4 genotypes, including GII.Pe-GII.4(26/53), GII.P17-GII.17(19/53), GII.P12-GII.3(7/53) and GII.P16-GII.13(1/53). All GII.4 strains were the variants of GII.4-Sydney-2012. From May to Aug in 2015, GII.4 only accounted for 5.3%(1/19) and GII.17 were the major epidemic strains(68.4%, 13/19). But from Sept to Dec in 2015, GII.17 decreased substantially to 10.9%(6/55) and GII.4 became the most predominated strains (45.5%, 25/55).@*Conclusions@#NoV is an important pathogen responsible for viral diarrhea among adults in Qingdao. GII.4-Sydney-2012 and the newly variant GII.P17-GII.17 were the predominant epidemic strains in Qingdao, 2015.
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Objective To investigate the etiology of hand,foot and mouth disease (HFMD) and to analyze the genetic characteristics of three prevalent enteroviruses in Qingdao in 2015. Methods City-wide surveillance data in 2015 were used to analyze the epidemiological characteristics of HFMD in Qingdao. RNA samples extracted from throat swab of HFMD cases were examined for general enteroviruses (EVs) such as enterovirus 71(EV71), coxsackievirus A16 (CA16) and CA6 by real-time RT-PCR. Full-length VP1 genes of EV-positive specimens were amplified and sequenced. Sequencing results were phylogenetically analyzed using MEGA 7.0 software package. Results A total of 804 patients with HFMD were identified from 1 176 patients in 2015. CA6 (41.4%),EV71 (31.6%) and CA16 (15.3%) were the predominant EVs causing HFMD. Children 5 years of age and under accounted for 80.3% of the 804 HFMD cases. CA6 was responsible for 48.9% of HFMD cases in children under 3 years old. The epidemic subtypes of EV71 and CA16 in Qingdao were C4a and B1b,respectively. Twenty-eight randomly selected CA6 strains were all classified into type A genome. Conclusion CA6, EV71 and CA16 were the prevalent pathogens causing HFMD in Qingdao in 2015. CA6 became the most predominant pathogen,mainly targeting children under 3 years old. C4a remained the prevailing subtype of EV71,while different from the co-prevalence of B1a and B1b subtypes in the past,B1b became the predominant subtype of CA16. CA6 strains circulating in Qingdao in 2015 mainly belonged to type A genome and evolved into multiple smaller branches. However, CA6 strains isolated in Qingdao in 2015 and 2013 located in different branches.
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Hepatolenticular degeneration was one of the rare several genetic metabolic diseases in clinic that could be cured by liver transplantation method, developing slowly and being irreversible. Metabolic disorders of copper lead to abnormal copper accumulation in various of tissues and organs. So that, the disease′s clinical manifestations were lacking in specificity and many patients missed the best opportunity of drug treatment. With the maturity of technologies and innovation of theory of liver transplantation, there were more and more methods that will be applied to personalized treatment. In this paper, a review of the research progress in the treatment of hepatolenticular degeneration with liver transplantation was made with reference to the relevant literature at home and abroad.
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Objective To explore the application value and clinical efficacy of one half layer pancreaticojejunostomy with the posterior wall of pancreas reinforced in pancreaticoduodenectomy.Methods The retrospective cross-sectional study was conducted.The clinical data of 17 patients with pancreatic neoplasms and ampullar neoplasms who underwent pancreaticoduodenectomy at the Second Affiliated Hospital of Harbin Medical University from May to September 2015 were collected.One half layer pancreaticojejunostomy with the posterior wall of pancreas reinforced method was applied to the digestive tract reconstruction after pancreaticoduodenectomy in the 17 patients.Observation indicators included:(1)surgical situations:surgical procedures,operation time,time of pancreaticojejunostomy,volume of intraoperative blood loss,tumor sizes,(2) postoperative situations:recovery time of gastrointestinal function,postoperative complications,duration of postoperative hospital stay,(3) postoperative pathological examinations,(4) follow-up.Patients were followed up by outpatient examinations including color Doppler ultrasound or abdominal computed tomography (CT) and telephone interview detecting abdominal pain or distention and general situations (diet,sleep) up to October 2015.Measurement data were represented as average (range).Results (1) Surgical situations:all the 17 patients underwent successful operations without perioperative death,including 16 undergoing radical pancreaticoduodenectomy and 1 undergoing pancreaticoduodenectomy and left liver resection.The average operation time,average pancreaticojejunostomy time,average volume of intraoperative blood loss and average tumor size were 276 minutes (range,230-440 minutes),12 minutes (range,9-16 minutes),310 mL (range,200-950 mL) and 3.25 cm2(range,1.92-5.60 cm2),respectively.(2) Postoperative situations:the average recovery time of gastrointestinal function was 3 days (range,1-7 days).Three patients had postoperative complications,including 1 patient with pancreatic fistula (Grade A) and 2 patients with delayed gastric emptying,and all of them had been healed after symptomatic and supportive treatments.The results of T-tube cholangiography or CT before hospital discharge showed that there was no leakage around the anastomoses.The average duration of postoperative hospital stay was 10 days(range,6-20 days).(3) The postoperative pathological examinations showed 5 patients of pancreatic ductal adenocarcinomas,4 of common bile duct ampulla area adenocarcinomas,3 of duodenal papillary adenocarcinomas,3 of pancreatic intraductal papillary mucinous neoplasms and 2 of duodenal ampullary adenocarcinomas.(4) Followup:all the 17 patients were followed up for 1-4 months and the abdominal color Doppler ultrasound or CT showed that there was no evidence of tumor recurrence or leakage around anastomoses.Conclusion One half layer pancreaticojejunostomy with the posterior wall of pancreas reinforced is safe and feasible,and it can reduce the rate of pancreatic fistula successfully.
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Neuroendocrine tumors (NETs) is a rare and heterogeneous group of tumors with widely varying morphologies and behaviors. Due to their rarity and heterogeneity, progress in improving its treatment has been slow. Pancreatic neuroendocrine tumors (pNETs) is a subset of NETs, previously known as islet cell tumors, occupies 3% of the primary pancreatic tumors with the annual incidence rate of (1-2)/100 000. In recent years, it is very necessary to improve the diagnosis and treatment of pNETs.
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Pancreatic ductal carcinoma accounts for 85％-90％ in pancreatic cancer, followed by the pancreatic cellendocrine tumors and pancreatic acinar cell carcinoma.In addition, mesenchymal cell carcinoma of pancreas is rare, and a mixed pancreatic cancer from 3 cells carcinoma is extremely rare.And pancreatic cancer always assume hypovascular tumor,spontaneous rupture of pancreatic cancer is rarely reported.A patient with a mixed duct-acinar-islet cell tumor of pancreas was admitted to the Second Affiliated Hospital of Harbin Medical University in June 2014, and underwent emergent operation of intraabdominal bleeding due to spontaneous rupture of tumor.The follow-up was done up to January 20, 2015.The patient died of intraabdominal widespread implantation metastasis of pancreatic cancer.A mixed duct-acinar-islet cell tumor of pancreas is extremely rare and easy to metastasis and diffusion of tumor with a rapid disease progression and a poor prognosis of patient,while operation is the key to terminate deterioration of the condition, and is the last line of defense to save lives.
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Hepatocellular carcinoma (HCC) combined with inferior vena cava (IVC) tumor thrombosis is regarded unresectable.Most of the patients received non-surgical treatment or gave up treatment,and the prognosis of these patients is poor.As the development of surgical treatment,the success rate of surgical treatment for HCC combined with IVC tumor thrombosis is increasing yearly.In May of 2012,one patient with HCC combined with IVC tumor thrombosis received hepatic Ⅴ,Ⅶ and Ⅷ segmentectomy + tumor thrombosis removal from the IVC at the Second Affiliated Hospital of Harbin Medical University.Preoperative computed tomography showed space-occupying lesions in the segments Ⅴ,Ⅶ and Ⅷ,and the IVC was filled with tumor thrombi.The volume of the left liver was 489 cm3,which was under the limit for survival.In order to preserve the remnant liver,right hepatectomy with reservation of hepatic segment Ⅵ,and the tumor thrombi in the IVC were removed with total hepatic vascular exclusion.The patient was recovered with no tumor recurrence or metastasis at postoperative month 18.
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For patients with malignant pancreatic cancer combined with vascular invasion,radical pancreaticoduodenectomy with vascular resection and anastomosis is the treatment of choice.Because this procedure is difficult to manage and with high risks,it is a great challenge to surgeons.A 50-year old patient with pancreatic head cancer whose portal vein and superior mesenteric vein were involved received radical pancreaticoduodenectomy in the Second Affiliated Hospital of Harbin Medical University.In the surgery,the tumor and its surrounding tissues were dissected,and then the portal vein and splenic vein were reconstructed.The patient was discharged at the 10th day after the surgery with favorable prognosis.
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Objective To investigate the effective route and proper method in transfecting gene into liver graft mediated by adeno-associated virus vector.Methods Three routes including hepatic artery,portal vein and hepatic artery+portal vein,and 3 methods,i.e.routine,circulation and clamping were employed for infusion.The best infusion route and method of gene transfection into liver graft were determined by observing the color change of liver and detecting liver function and transfoetion rate of liver cells.The safety of these methods was evaluated.Results In all the infusion procedures,the color of the liver grafts turned from red to white,no apparent color differenee of the livers and no enlargement nor mottling were observed under surgical microscope.The liver color was back to normal immediately after blood flow was restored.No significantly statistical differences of the ALT values were observed among all the groups(F=0.343,1.265,0.055,P＞0.05).Adeno-associated virus vectors coding for the enhanced green fluorescence protein(AAV2-EGFP)were successfully transfected into liver cells by the 3 infusion routes 1 week later,and the difierences of transfection rates via the 3 routes had no statistical significance(F=0.080,0.091,0.045,P＞0.05).The transfoction rate of AAV2-EGFP was the highest at any time points when using the clamping method,and then followed by circulation method and routine method,with statistical differenee(F=3.880,2.976,5.129,P＜0.05).The transfection rates of AAV2-EGFP were increased progressively and peaked at the 6th week,and then they were decreased gradually.Conclusions Infusion via hepatic artery is the effective route for gene transfection and clamping the vessels can elevate the transfection rate of AAV2-EGFP.All procedures were performed without detectable liver injury.The transfection of gene into liver graft mediated by adeno-associated virus vector is a slow and persistent process.