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目前国内对猪轮状病毒的研究较少,作者旨在对猪A群轮状病毒进行分离与鉴定,为后续猪轮状病毒致病机理和分子生物学特性研究奠定基础.用胰蛋白酶处理RT-PCR检测猪A群轮状病毒病原阳性的临床腹泻粪样,然后接种长成单层的MA-104细胞,进行病毒分离传代.再对分离毒株进行常规RT-PCR鉴定和电镜观察,并对分离毒株的VP6、VP7和VP8基因进行测序及序列分析.结果成功分离猪A群轮状病毒TM-a株,经序列同源性分析发现,与TM-a株VP6、VP7和VP8基因同源性最高的毒株分别为中国北京人源LL3354株、印度人源RMC321株和日本野猪源GUB-71株,其相似性为96.0％、95.1％和97.0％.该TM-a株轮状病毒不同基因表现出与人源轮状病毒和猪源轮状病毒的高度同源性,推测猪A群轮状病毒可能在人畜间传播,发生基因重组现象. 相似文献
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A群轮状病毒(GAR)是引起人类和多种动物腹泻的主要病原体,病原的分离与鉴定是轮状病毒(RV)流行病学、病原学等研究的基础.本研究采用接种MA 104细胞的方法,从内蒙古某猪场患病猪腹泻粪便中分离一株病毒,经3次蚀斑克隆纯化后,采用电镜观察、PCR鉴定和序列测定进行分析,结果表明该分离株为猪轮状病毒(PoRV).致病性试验表明该分离株能够引起仔猪急性腹泻,RT-PCR扩增其VP4、VP6和VP7的基因节段并进行序列测定,按照A群RV的最新分类方法,确定该分离株的VP6、VP7和VP4基因型分别为I5型、G9型和P[23]型.因此,将该分离株命名为Rotavirus A pig/China/NMTL/2009/Q9P[23]. 相似文献
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《畜牧与兽医》2016,(9):32-37
将1份经RT-PCR检测猪轮状病毒阳性的临床腹泻粪样感染Vero细胞,连续盲传8代,出现病变,成功分离到1株病毒,经RT-PCR检验和电镜观察,鉴定该病毒为轮状病毒,命名为GD-01-2015。扩增该病毒的VP4和VP7基因进行分型和系统进化分析,结果发现,其VP4基因为P[7]型,与来自韩国的猪源毒株K71同源性达到99.8%;其VP7基因为G5型,与来自韩国的猪源毒株06-6-1同源性达到99.7%。由此可以推测GD-01-2015株与韩国猪源毒株有相似的进化来源。根据轮状病毒分类委员会(RCWG)提出的A群轮状病毒的最新分类方法,GD-01-2015株基因型为G5P[7]。本研究为监测轮状病毒的流行状况及其疫苗的研制提供了理论依据。 相似文献
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轮状病毒是引起幼龄动物和儿童病毒性腹泻的主要病原,轮状病毒的分离鉴定为该病的流行病学调查和分子生物学特性研究奠定了基础。本研究采集北京某猪场腹泻发病仔猪肠内容物,将其接种MA104细胞,分离得到一株能产生明显细胞病变的病毒。对分离毒株进行胶体金、实时荧光定量RT-PCR和免疫荧光等方法鉴定,并对分离毒株的VP4、VP6和VP7基因进行测序及序列分析。结果表明,分离毒株为猪A群轮状病毒。按照A群轮状病毒的最新分类方法,确定该分离株VP4、VP6和VP7基因的基因型分别为P[13]型、I5型和G11型。因此,将该分离株命名为Rotavirus A pig/China/BJ/2015/G11P[13]。 相似文献
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轮状病毒是导致包括猪在内的多种新生动物腹泻的重要肠道病原。猪轮状病毒的致病性已经通过实验证实,在有腹泻的猪中经常可以检测出轮状病毒。轮状病毒属于呼肠孤病毒科轮状病毒属。早期的研究表明,所有的轮状病毒均具有抗原相关性,但近年来的研究发现不同的轮状病毒抗原性差异很大。在现有的A-G7群血清型不同的轮状病毒中有4群可感染猪(A、B、C、E),A群轮状病毒代表着公认的轮状病毒。1流行病学轮状病毒在猪群中广泛存在,美国学者Bridger等的血清学监测结果表明,77%～100%的成年猪为轮状病毒A、B、C群血清学阳性猪。我国学者对我国的… 相似文献
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为弄清2013年年初上海某猪场发生的仔猪腹泻疫情病因,本研究随机采集发病猪场的仔猪腹泻样品9份,利用RT-PCR方法对采集样品进行病原检测。结果显示,9份临床样品中有4份为猪轮状病毒阳性,而猪流行性腹泻病毒和猪传染性胃肠炎病毒均为阴性。根据GenBank公布的猪轮状病毒参考株序列设计两对引物,对其中阳性样品的VP6和VP7基因进行RT-PCR扩增、基因克隆和序列测定分析。结果显示,4份阳性样品的VP6基因大小为1356 bp,VP7基因大小为1100 bp;序列分析结果显示,VP6基因与A群代表株OSU的氨基酸同源性为64.1%~87.0%,与代表株Gottfriend的氨基酸同源性为26.3%~60.8%;VP7基因与不同G型代表株的氨基酸同源性为71.8%~97.3%。根据上述结果分析,我们推测引起此次仔猪腹泻疫情中的主要病原是由属于A群G9血清型的猪轮状病毒引起,本研究为此次仔猪腹泻疫情的诊断提供了依据。 相似文献
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《黑龙江畜牧兽医》2016,(23)
为了解猪轮状病毒(PoRV)贵州株NSP4基因的序列特征,试验根据Gen Bank上公布的猪轮状病毒NSP4基因序列设计1对特异性引物,扩增一株猪轮状病毒贵州流行株NSP4基因序列并进行序列测定及分析。结果表明:得到的碱基序列(562 bp)与预期结果相符,包含NSP4基因528 bp的开放阅读框,编码175 aa;与已知的14个国内外参考毒株NSP4核苷酸和推导氨基酸序列比较,同源性分别为89.1%~94.1%和95.4%~98.3%;氨基酸系统进化树结果显示,贵州流行株与国内流行株LL36755亲缘关系较近。说明试验成功克隆了PoRV贵州株NSP4基因,获得了其序列,并分析了其序列特征。 相似文献
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猪轮状病毒的分离与鉴定 总被引：1，自引：1，他引：0
将疑似感染猪轮状病毒的内蒙古某猪场的腹泻仔猪粪便样品在MA104细胞上分离培养,得到一株能产生明显细胞病变的毒株(命名为PRV L1株).经纯净性检测,该毒株无菌生长、无支原体污染及外源病毒污染.特异性检测结果显示该PRV L1株能被猪轮状病毒单特异性血清中和,并可被猪轮状病毒单克隆抗体识别,其VP7基因序列与G5型猪轮状病毒VP7基因序列同源性为99％.动物回归试验结果表明,该毒株口服攻击3日龄仔猪,可引起典型的猪轮状病毒病发病症状,并能在发病仔猪小肠内容物中检测到猪轮状病毒. 相似文献
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为分析鸽轮状病毒中国分离株的特性,采用高通量测序方法,测定从山东省青岛市某活禽交易市场采集的肉鸽粪便样品中分离的轮状病毒基因组,并对其进行特性分析与比较基因组研究。结果显示,G群轮状病毒鸽群分离株全基因包含11个节段,总长度为17 803 bp,具有与其他轮状病毒相似的结构和基因顺序。与24株轮状病毒代表性毒株的VP6氨基酸序列进化分析结果,以及其他5个结构蛋白氨基酸序列遗传进化分析结果均证实,该病毒属G群轮状病毒。该毒株与2011年从香港鸽群中分离的G群轮状病毒同源性最高,但衣壳糖蛋白VP7的氨基酸序列同源性仅为71.7%,抗原性存在较大差异。 相似文献
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The prevalence of porcine rotavirus infection in Venezuela was studied from November 1980 to November 1981. Rotaviruses were identified in 21.3 per cent of 286 diarrhoeic samples analysed by electron microscopy and in none of the 48 controls. Rotaviruses were readily detected throughout the year, which may correspond to the absence of seasonal climatic variations in a tropical country such as Venezuela. Rotavirus infection was frequently observed in pigs that were two to six weeks old and was rare in younger animals. The age distribution of rotavirus infection correlated well with the presence of circulating antibodies, as measured by the complement fixation test. Sucking pigs less than two weeks old appeared to be protected from the infection by passive immunity; no antibodies were detected in the two- to three-week-old group, coinciding with an increased susceptibility to rotavirus infection. The antigenic specificity of the rotavirus isolates was confirmed by enzyme-linked immunosorbent assay test, using antibodies directed against the group-specific antigen of human rotavirus. Polyacrylamide gel electrophoresis of viral double-stranded RNA revealed an heterogeneity on the electropherotype of selected samples obtained at different times during the study period. 相似文献
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A total of 113 diarrheic samples comprising of 68 buffalo calves and 45 cow calves were screened by RNA-PAGE for the detection
of presence of rotavirus. RNA-PAGE analysis of these samples revealed 11 (9.73%) was found positive for rotavirus. Out of
68 faecal samples of buffalo calves tested for viral gastroenteritis, 8 (11.76%) were found positive for rotavirus. Similarly,
out of 45 faecal samples of cattle calves tested for viral gastroenteritis, 3 (6.66%) was found positive for rotavirus. Rotavirus-positive
samples represented long electropherotype. All RNA-PAGE-positive faecal samples for rotavirus subjected to RT-PCR for VP7
gene, ten samples yielded a specific product of 1,013 bp of VP7 gene. All the PCR-positive samples of the present study were
subjected to genotyping with primers for G6, G8 and G10 genotype. All positive samples showed G10 genotype. This indicates
that G10 may be predominant genotype among bovine calves in Mumbai region in India. 相似文献
14.
Incidence of rotavirus in beef herds in Argentina 总被引：1，自引：0，他引：1
R C Bellinzoni N Mattion J L La Torre E A Scodeller 《Research in veterinary science》1987,42(2):257-259
Faecal samples were collected from 177 diarrhoeic and 40 healthy calves from 19 farms in Buenos Aires State, Argentina during 1984 and 1985. Samples were examined for rotavirus by enzyme-linked immunosorbent assay (ELISA) and polyacrylamide gel electrophoresis (PAGE) of their genomic RNA segments. Rotavirus was found in 95 samples of diarrhoeic calves (53 per cent) and in three healthy calves (7 per cent). All positive samples were tentatively classified as group A on the basis of electropherotype and ELISA test reactivity and exhibited 18 different genomic electropherotypic patterns. 相似文献
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Relative prevalence of typical and atypical strains among rotaviruses from diarrheic pigs in conventional swine herds 总被引：3，自引：0，他引：3
B H Janke J K Nelson D A Benfield E A Nelson 《Journal of veterinary diagnostic investigation》1990,2(4):308-311
Polyacrylamide gel electrophoresis was conducted on genomic RNA extracted from rotaviruses detected in diarrheic pigs from conventional swine herds. Ninety samples contained sufficient virus for RNA band visualization and genome classification. Genome profiles were characteristic of typical group A rotaviruses in 67.8% of the 90 samples, of group B rotaviruses in 10.0%, and of group C rotaviruses in 11.1%. In 11.1% of the samples, the presence of more than 11 bands suggested concurrent infection with more than 1 strain of rotavirus. In infections among nursing pigs, 76.4% were group A rotaviruses, 7.4% were group B, 7.4% were group C, and 8.8% were coinfections. In infections among weaned pigs, 40.9% were group A, 18.2% were group B, 22.7% were group C, and 18.2% were coinfections. Coelectrophoresis with prototype OSU and Gottfried strains revealed a great diversity in electropherotype among field strains of rotavirus. 相似文献
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R Bellinzoni N Mattion L Vallejos J L La Torre E A Scodeller 《Research in veterinary science》1987,43(1):130-131
In Argentina the presence of rotavirus was investigated in a chicken flock experiencing periodic episodes of diarrhoea during the winter of 1986. All the samples analysed were negative by the enzyme-linked immunosorbent assay (ELISA). However, when samples were observed by electron microscopy, particles which were indistinguishable from standard rotaviruses were detected in some samples. Ten of the 36 samples were positive after polyacrylamide gel electrophoresis (PAGE) analysis, all of them showing the same electropherotype. Based on these results these viruses were classified as rotavirus-like or atypical rotaviruses. 相似文献
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V. Parreo V. Costantini S. Cheetham J. Blanco Viera L. J. Saif F. Fernndez L. Leoni A. Schudel 《Zoonoses and public health》2001,48(9):713-720
Group A rotavirus (RV) and coronavirus (CV) are common viral pathogens associated with neonatal diarrhoea in numerous animal species. The purpose of this work was to investigate the presence of these viral agents in two farm populations of captured guanacos (Lama guanicoe) in the Argentinean Patagonia region, that developed severe diarrhoea outbreaks. Stool and serum samples were analysed for RV and bovine CV antigen and antibody by enzyme‐linked immunosorbent assay. Rotavirus was detected in faeces from two new‐born guanacos with acute diarrhoea, one in each farm. After electrophoretic analysis, each isolated strain, showed a distinctive long dsRNA electropherotype characteristic of group A rotaviruses (4:2:3:2). In addition, 95% (38 of 40) of the sampled animals were positive for RV antibodies, suggesting a high prevalence of RV infection in the populations tested. No evidence of CV circulation by antigen or antibody analysis was observed. To our knowledge, this is the first report of the detection and isolation of RV associated with neonatal diarrhoea in Lama guanicoe. 相似文献
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犬轮状病毒荧光定量RT—PCR检测方法的建立 总被引：1，自引：0，他引：1
目的:建立轮状病毒快速准确的检测方法,为研发诊断试剂盒和疫苗奠定基础。方法:用Primer5软件设计VP7引物,从采集的腹泻犬粪便样品抽提病毒RNA,用反转录聚合酶链式反应(RT-PCR)技术,将RNA反转录为cDNA,并进行PCR扩增,对扩增产物进行非变性聚丙烯酰胺凝胶电泳和测序,与代表株的VP7进行同源性比较。结果:从腹泻病犬粪便样品中抽提到了轮状病毒RNA,反转录后扩增产物为51bp的基因片段,经核苷酸序列分析,其PCR序列与犬轮状病毒VP7基因编码源序列的同源性为86.3%。结论:正确克隆了轮状病毒主要保护性抗VP7基因中抗原表位区,建立了犬轮状病毒实时定量诊断方法,为研制实时定量诊断试剂盒奠定了基础。 相似文献
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Martella V Ciarlet M Lavazza A Camarda A Lorusso E Terio V Ricci D Cariola F Gentile M Cavalli A Camero M Decaro N Buonavoglia C 《Veterinary microbiology》2005,111(1-2):117-124
An epidemiological survey was carried out to investigate the distribution of the VP7 and VP4 specificities of lapine rotaviruses (LRVs) in rabbitries from different geographical regions of Italy. Almost all the strains were characterized as P[22],G3, confirming the presence of the newly-recognized rotavirus P[22] VP4 allele in Italian rabbits. Only one P[14],G3 LRV strain was identified and two samples contained a mixed (P[14] + [22],G3) rotavirus infection. All the LRV strains analyzed exhibited a genogroup I VP6 specificity and a long dsRNA electropherotype. However, one of the P[14],G3 strains possessed a super-short pattern. Altogether, these data highlight the epidemiological relevance of the P[22] LRVs in Italian rabbitries. 相似文献
20.
E W Collisson T L Barber C M Shannon M C Kemp 《Journal of veterinary diagnostic investigation》1989,1(3):242-246
Two predominant electropherotypes of bluetongue virus (BTV) serotype 11 isolates from cattle during a 1981-1984 field study in eastern Colorado were characterized. The genomes of strains isolated from the first 2 years of the study had 1 predominant electropherotype (CO81), with the exception of 1 isolate that differed only in the migration of segment 3. A second electropherotype (CO83), with differences in the migration of 4 segments, coexisted in the same region during 1983 and 1984 with strains having the CO81 RNA profile. The genomes of CO81 and CO83 were also distinguishable from those of the US prototype of BTV 11. Analysis of the polypeptides of representative strains of each electropherotype by sodium dodecyl sulfate-polyacrylamide gel electrophoresis indicated that the proteins were very similar. The occurrence of the CO81 electropherotype was apparently the result of multiple viral infections since the positions of 7 segments had faint second bands and single-banded variants were isolated after serial plaque purifications. In addition, protein 7 of 1 of the CO81 isolates and protein 7 of the single-banded variant differed as shown by reverse phase-high performance liquid chromatography of 35S-methionine-labeled tryptic peptides. 相似文献