Characterization of a new cell line from caudal fin of koi, Cyprinus carpio koi, and first isolation of cyprinid herpesvirus 3 in China

A new continuous cell line (KCF-1) from caudal fin of koi, Cyprinus carpio koi, was developed and sub-cultured more than 100 passages since the present study was initiated in March 2006. KCF-1 predominantly consisted of short fibroblast-like cells and grew well in Dulbecco's modified Eagle medium (DMEM) supplemented with 10% fetal bovine serum (FBS). Chromosome analysis revealed that 56% of the KCF-1 cells maintained normal diploid chromosome number (2n = 100) at Passage 82. Using the KCF-1 cell line, a strain of cyprinid herpesvirus 3 (designated as CyHV-3-QY08) was isolated from the diseased koi. CyHV-3-QY08 continuously propagated in the KCF-1 cells, as confirmed by immunofluorescence assay (IFA) and transmission electron microscopy (TEM). KCF-1 cells infected with CyHV-3-QY08 produced typical cytopathic effects characterized by severe vacuolation, deformation of nuclei, and marginalization of the nuclear chromatin, which are consistent with those of previous reports. CyHV-3-QY08 was purified and subsequently analyzed by SDS-PAGE and TEM. The results showed that the purified virions contained two types of morphologies and were composed of more than 30 obvious viral polypeptides. An infectivity experiment revealed that CyHV-3-QY08 could cause 100% mortality in the infected koi. Based on the genome sequence of CyHV-3-I/U, the CyHV-3(I/U)-ORF136 homologue in CyHV-3-QY08 was cloned and sequenced. Multiple sequence alignments of CyHV-3-I/U-ORF136 homologues showed that CyHV-3-QY08 belonged to the typical Asian genotype. The CyHV-3(I/U)-ORF136 homologue seems to be a novel molecule marker, which can be used to distinguish Asia isolates from Europe-America strains.     

肠道病毒衣壳蛋白致病性研究进展

肠道病毒是人类感染率很高的一种病毒,其衣壳蛋白在病毒的致病性方面起着重要的作用.近年来,大量的研究工作致力于肠道病毒农壳蛋白的致病性研究.此文就衣壳蛋白的基因组结构、基因分型、与病毒受体的相互作用和疫苗研制等方面作了综述.     

经皮肤损伤感染马尔尼菲青霉菌致病力动物实验研究

目的研究马尔尼菲青霉菌经皮肤损伤途径感染小鼠致病力情况。方法将马尔尼菲青霉菌野生株和人感染株孢子悬液分别注入鼠尾皮内,观察接种后发病情况,于第15,50 d分批处死、解剖。结果马尔尼菲青霉菌导致小鼠皮肤感染发病率为100%,而85%小鼠皮损可自行消退痊愈,15%小鼠出现皮肤播散性感染;组织病理示:皮损处细胞性炎症反应在皮损中显著。从发病时间和早期病变严重程度比较,野生株致病力显著强于人感染株(P<0.05);但是从后期病变严重程度和自行痊愈率比较,野生株和人感染株致病力差异无显著性(P>0.05)。结论马尔尼菲青霉菌可以经皮肤损伤引起小鼠致病,其引起机体剧烈细胞免疫应答反应是致病力重要因素之一;野生株和人感染株感染早期致病力有差异,预后无差异。     

腹泻便中成团肠杆菌的分离与鉴定

本文报告了自腹泻便中检出的13株成团肠杆菌生化鉴定,分子生物学检测,药敏试验及致病性研究结果。生化鉴定结果与文献基本相符,分子生物学检测该组细菌DNA同源性为72％～100％,证明是同种细菌。致病性研究发现部分菌株可引起动物和生物模型病变,与人的腹泻关系密切,14种药物敏感试验说明该组细菌的抗药谱很广,临床上可用的敏感药物不多,发生感染后治疗将十分困难。     

Cloning and further sequence analysis of the spike gene of attenuated porcine epidemic diarrhea virus DR13

The spike (S) gene of the attenuated porcine epidemic diarrhea virus (PEDV) DR13 was cloned and sequenced to further explore the functions of wild type PEDV and attenuated PEDV. Sequencing revealed a single large ORF of 4,149 nucleotides encoding a protein of 1,382 amino acids with predicted M _r of 151 kDa. The coding region of the S gene of attenuated PEDV DR13 had 20 nucleotide changes that appeared to be significant determinants of function in that they produced changes in its predicted amino acid sequence. Notably, attenuated PEDV DR13 has previously been found to exhibit reduced pathogenicity in pigs. The regions containing these 20 nucleotide changes may therefore be crucial for PEDV pathogenicity. The attenuated PEDV DR13 S protein contains 28 Asn-Xaa-Ser/Thr sequons, 21 asparagines that are predicted to be N-glycosylated and a stretch of highly hydrophobic residues at positions 1,327–1,347, which is predicted to form an α-helix and to function as a membrane anchor. One (from N to K at 378) of the changes in the deduced amino acid sequence destroyed N-linked glycosylation sites, while another change (from N to S at 114) created a new one at a different location. These alterations in N-linked glycosylation sites reflected 3 nucleotide changes, which were related to the above-mentioned nucleotide changes and are suggested to influence the pathogenicity of attenuated PEDV DR13. Attenuated PEDV DR13 has 96.5, 96.4, 96.1, 93.9, 93.5 and 96.6% DNA sequence identities with CV777, Br1/87, JS-2004-2, Spk1, Chinju99 and parent DR13, respectively. Likewise, it shares 95.7, 95.4, 95.6, 92.0, 91.6 and 95.7% identity with those genes at the deduced amino acid sequence level. Phylogenetic analysis suggested that attenuated PEDV DR13 is closely related to CV777, Br1/87, JS-2004-2 and parent DR13, rather than to Spk1 and Chinju99 and is especially close to the Chinese PEDV strain JS-2004-2. Nucleotide sequence data reported is available in the GenBank database under the Accession Nos. DQ462404 and DQ862099.     

Molecular characterization of three novel murine noroviruses

Murine noroviruses (MNV) comprise a group of newly recognized pathogens infecting laboratory mice. The first reported murine norovirus, murine norovirus 1 (MNV-1), produces a transient infection with a short duration of fecal shedding after infection of immunocompetent laboratory mice. Our laboratory subsequently isolated three novel murine noroviruses, murine norovirus 2 (MNV-2), murine norovirus 3 (MNV-3), and murine norovirus 4 (MNV-4), that have markedly different pathogenicity from MNV-1 by producing persistent infections and prolonged fecal shedding in infected immunocompetent mice. In this study, the nucleotide sequences and the predicted amino acid sequences of the three novel murine noroviruses were determined and compared to each other, MNV-1, and other previously described human and animal noroviruses. The three novel murine norovirus strains were shown to be related to each other and MNV-1 by sequence and phylogenetic analysis even though MNV-2, MNV-3 and MNV-4 all display markedly different biologic behavior from that of MNV-1. The nucleotide sequence data reported in this paper have been submitted to the GenBank nucleotide sequence database and have been assigned the accession numbers DQ223041, DQ223042, and DQ223043.     

牙龈卟啉单胞菌rag位点基因分型与细菌毒力的关系研究

目的：探讨牙龈卟啉单胞菌rag位点基因分型与细菌毒性的关系。方法将不同rag基因位点的牙龈卟啉单胞菌W83(rag-1型)、A011/9(rag-2型)、QM220(rag-3型)和ATCC33277(rag-4型)菌株分别刺激中性多形核白细胞,未受刺激的中性多形核白细胞作为对照。采用RT-PCR检测FcγⅢb受体基因,流式细胞术检测中性多形核白细胞凋亡情况,试剂盒检测毒力因子。结果 W83组 FcγⅢb基因相对表达量显著高于其他组( P <0．05),其次为QM220组,显著高于A011/9和ATCC33277组(P<0.05)。流式细胞术检测发现,ATCC33277组细胞凋亡率显著高于其他各组(P<0.05),A011/9组凋亡率显著高于W83组(P<0.05)。各基因分型组LPS、LBP、mCD14、MMP-8 MMP-9、IL-8以及IL-1β均高于对照组(P<0.05)。W83组上述指标显著高于其他各基因型组(P<0.05),QM220组次之,与 A011/9、ATCC33277组比较差异有统计学意义(P <0.05)。 A011/9组在 LPS、LBP、mCD14方面高于ATCC33277组(P<0.05),而MMP-8、MMP-9两者差异无统计学意义(P>0.05)。结论牙龈卟啉单胞菌菌株中rag位点基因分型的细菌毒性rag-1型最强,rag-3型次之,较弱的为rag-2型和rag-4型。     

新型禽源人流感病毒（H7N9）的小鼠感染模型和传播模型建立

目的建立新型禽源人流感病毒（H7N9）小鼠模型和可能的H7N9致病性和传播力研究。方法H7N9病毒感染小鼠,并与同居小鼠合笼,研究同居小鼠的临床指征变化,病毒复制情况,病毒在组织中的分布,以及病理变化。通过观察同居小鼠的发病情况等方面研究H7N9病毒在同笼小鼠中的传播能力。结果研究表明H7N9病毒能有效地感染小鼠并造成致死,可以通过直接接触传播感染小鼠并引起病理等改变。结论建立了H7N9小鼠模型,并对小鼠通过接触传播感染进行了初步研究,为深入研究传播力奠定了基础。     

无症状高尿酸血症患者尿酸盐晶体致病性相关因素研究进展

无症状高尿酸血症（AH）是以血清尿酸水平升高为特征的一种代谢性疾病。当血清尿酸水平超过饱和值后,尿酸盐晶体就会析出并沉积在关节滑膜、软骨及肾脏等组织诱发急性炎性反应。尿酸盐晶体是用于诊断痛风的金标准,但并非所有尿酸盐晶体沉积的AH患者最终都会发展为痛风。尿酸盐晶体沉积最终是否会引起痛风发作取决于尿酸盐晶体相关致病性基因、机体的免疫状态、软骨损伤以及滑液环境等多重因素的共同作用。明确尿酸盐晶体在AH发展为痛风过程中的致病性相关因素及其触发机制,能为AH患者中痛风高危人群的早期筛查、分层管理及个体化治疗提供依据。     

致病性奈瑟菌属免疫球蛋白Al蛋白酶最新研究进展

一直以来,免疫球蛋白A1蛋白酶因其对人类IgA1的特异性切割导致黏膜免疫受损而倍受关注。随着生物学和免疫学研究的不断发展,近年来致病性奈瑟菌属IgA1蛋白酶的研究也在不断地深入,尤其是IgA1蛋白酶切割性能与IgA1铰链区变异之间的关系、自身输出的模式以及诱导T细胞免疫等新概念,使得研究者对IgA1蛋白酶有了更进一步的认知。