Nucleotide sequence characterization and phylogenetic analysis of hantaviruses isolated in Shandong Province,China

Background China is the most severe endemic area of hemorrhagic fever with renal syndrome （HFRS） in the world with 30 000-50 000 cases reported annually, which accounts for more than 90% of total number of cases worldwide. The incidence rate of the syndrome in Shandong Province is one of the highest in China, which has ever reached 50 per 100 000 persons per year. However, the molecular characteristics of hantaviruses （HV） epidemic in Shandong Province remain unclear. Therefore it is useful to clarify nucleotide sequence and phylogenetic characteristics of HV isolated in Shandong Province in order to provide better advices to control and prevent HFRS. Methods RNAs were extracted from sera of clinically diagnosed patients and positive rodent lungs that were detected by indirect immunofluorescent assay （IFA）. Partial M segments of HV were amplified from the RNAs with reverse transcription nested polymerase chain reactions （nested PCR） using hantavirus genotype specific primers. The nested PCR products were sequenced and compared with those from previously epidemic isolates in Shandong and with other representative HV sequences from GenBank. Phylogenetic tree analyses were performed based on the sequences of the M genes. Results Thirty-four HV isolates in Shandong showed 67.1%-100% nucleotide identities. The nucleotide homologies among 6 Hantaan viruses （HTNV） isolates in Shandong were 78.1%-98.7%, while the homologies among 28 Seoul virus （SEOV） isolates in Shandong were 93.7%-100%. There were at least 3 subtypes HTNV （H2, H5, H9） and 2 subtypes SEOV （S2, S3） in Shandong Province. Conclusions In Shandong Province, the homologies of HTNV were lower and there were no predominant subtypes, while the homologies of SEOV were higher and S3 was the predominant subtype. The homologies of SEOV from rodents were higher than those from patients. The distribution of subtypes in Shandong was similar to that of the adjoining provinces. Phylogenetic analyses of the sequences showed geographic clustering of HV in Shandong.

Role of hepatic arterial ischaemia in biliary fibrosis following liver transplantation

Background In clinical liver transplantation, whether the delay of hepatic arterial ischaemia increases biliary fibrosis or not is controversial. We designed a liver transplantation model to test this controversy and explore its mechanism. Methods Twelve dogs were divided into two groups randomly： hepatic arterial ischaemia （HAI） and control groups. In HAI group, hepatic artery was perfused 60 minutes after portal perfusion, but in control group, hepatic arterial perfusion was simultaneous with portal perfusion. The pathological changes of intrahepatic bile ducts were observed. Transforming growth factor beta 1 （TGF-β1）, expressed in epithelial cells of intrahepatic bile duct, was detected by immunohistochemical streptoadividin-biotin complex method. Expressions of Smad3, P-Smad3 and the transcriptional levels of alpha smooth muscle actin （α-SMA） mRNA in intrahepatic bile ducts were detected by Western blotting and RT-PCR respectively.Results Compared with the control group, more collagen deposition and leucocytic infiltration could be seen in biliary vessel walls. Significantly more buffy particles, which are the proteins of TGF-β1, could be seen in biliary epithelial cells. P-Smad3 and α-SMA mRNA （as ratio to corresponding β-actin） in intrahepatic bile ducts were 1.82±0.18 and 1.86±0.73 respectively in HAI group, significantly higher than those in control group （0.59±0.09 and 0.46±0.18, respectively）. Conclusions Hepatic arterial ischaemia could increase the deposition of collagen fibres, trigger the transdifferentiation of myofibroblasts in intrahepatic bile duct and might result in biliary fibrosis by activating the TGF-β1 signalling pathway.