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多重RT-PCR检测猪口蹄疫病毒、猪水泡病病毒和猪水疱性口炎病毒的研究 总被引：1，自引：0，他引：1
建立了一种同时检测猪口蹄疫病毒(FMDV)、猪水泡病病毒(SVDV)和猪水疱性口炎病毒(VSV)三种病原体的多重RT-PCR方法。参照文献报道的基因序列,设计合成了三对特异性引物;PCR扩增条件进行优化后,用这三对引物对同一样品中的FMDV、SVDV、VSVRNA模板进行扩增,结果同时得到了三条特异性条带,大小与试验设计相符:FMDV(208bp)、SVDV(862bp)、VSV(638bp),且对猪瘟病毒(CSFV)、猪繁殖与呼吸综合症病毒(PRRSV)和猪传染性胃肠炎病毒(TGEV)核酸扩增结果为阴性;三种病毒RNA模板检出的最小量均为10fg。试验证明,此方法经济、快速、敏感、特异,可用于FMDV、SVDV和VSV这三种猪水泡性疾病的鉴别诊断及流行病学调查。 相似文献
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为建立一种鉴别猪塞内卡病毒A(SVA)、猪水疱病毒(SVDV)、口蹄疫病毒(FMDV)的方法，针对3种病毒的基因保守区域设计引物和探针，建立了可同时检测上述3种病毒的TaqMan三重荧光定量PCR方法。该方法仅能特异性扩增SVA和FMDV病毒基因以及SVDV阳性质粒，与猪繁殖与呼吸综合征病毒、经典猪瘟病毒、猪流行性腹泻病毒、猪圆环病毒2型、伪狂犬病病毒的阳性样品cDNA或DNA无交叉反应，对3种病毒核酸的最低检出限均为10 copies/μL,组内和组间变异系数均小于4%。应用该方法对2019-2022年采集的吉林省部分地区267份病料和本实验室留存的30份猪血清样品进行检测，未检出SVA、FMDV以及SVDV。建立的三重荧光PCR方法有利于对3种病毒进行有效的流行病学调查，可给快速准确鉴别致猪水疱症状病毒性疾病提供诊断方法。 相似文献
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为建立一种同时检测非洲猪瘟病毒（ASFV）和猪伪狂犬病病毒（PRV）的方法,本研究根据GenBank中的ASFV 72基因和PR V UL27基因的保守序列分别设计2对特异性引物,通过对双重PCR反应条件的优化,建立快速检测ASFV和PR V的双重PCR方法,并对该方法的特异性和敏感性进行验证。特异性试验结果显示,该方法对ASFV与PRV核酸的扩增能获得246 bp和401 bp的特异性目的片段,而对PRRSV、PEDV和CSFV的核酸扩增均为阴性;敏感性试验结果显示,该方法对ASFV和PRV核酸最低检测量分别为0.01 pg和0.12 pg。结果表明：建立的双重PCR方法具有特异性强、敏感性较好、快速简便等特点,可用于这两种病毒的同时检测和鉴别诊断。 相似文献
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猪群中7种高发传染病多重PCR诊断方法的建立 总被引：4，自引：0，他引：4
[目的]建立猪瘟、蓝耳病(又称猪繁殖与呼吸综合征)、圆环病毒2型感染、猪链球菌病、副猪嗜血杆菌病、猪多杀性巴氏杆菌感染和猪支原体感染7种猪病的多重PCR诊断方法.[方法]根据GenBank发表的核苷酸序列设计了分别针对猪瘟、蓝耳病、圆环病毒2型感染、猪链球菌病、副猪嗜血杆菌病、猪多杀性巴氏杆菌感染和猪支原体感染的2套多重PCR特异性引物,优化多重PCR反应条件后进行敏感性与特异性评价.[结果]建立的2套多重PCR诊断方法对其他常见猪病痛原的基因组无特异性扩增,检测病原基因组最低浓度可达到pg级.[结论]该研究建立的2套多重PCR诊断方法可以用于猪群中上述7种猪病痛原的单一感染或混合感染的鉴别诊断. 相似文献
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《中国预防兽医学报》2017,(2)
为建立同时检测非洲猪瘟病毒(ASFV)、水疱性口炎病毒(VSV)、猪口蹄疫病毒(FMDV),猪瘟病毒(CSFV)以及猪伪狂犬病病毒(PRV)的多重RT-PCR检测方法,本研究根据Gen Bank中登录的参考病毒株序列,选择各病毒的保守序列设计5对特异性引物,通过优化反应条件,建立了一种可以同时快速检测以上5种病毒的多重RT-PCR检测方法。结果显示,该方法对不同的细菌或病毒模板扩增结果均为阴性,特异性强;敏感性试验表明该方法对PRV、CSFV、ASFV、VSV和FMDV的核酸最少检出量分别为8.82×10~3拷贝/μL、6.87×10~4拷贝/μL、5.71拷贝/μL、4.93×10~4拷贝/μL和4.32×10~2拷贝/μL。以上结果表明该方法快速、灵敏、特异性强,对以上5种猪病病毒能够进行快速鉴别检测,为其临床诊断与流行病学调查提供了有效的检测方法。 相似文献
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《中国兽医学报》2019,(12):2293-2297
根据对GenBank中2015-2017年登录的18株国内塞内卡病毒(SVV)基因组序列比对分析结果,设计合成1对针对SVV 5’UTR区域的特异性引物和Taqman探针,通过优化荧光定量PCR反应条件,建立一种定量检测SVV的Taqman实时荧光定量PCR检测方法。该方法定量检测范围在10～10~9 copies/μL,检测下限为10 copies/μL,比常规PCR检测灵敏度高约1 000倍,同时检测口蹄疫病毒(FMDV)、猪水疱病病毒(SVDV)、猪水疱疹病毒(VSV)、圆环病毒(PCV)、猪伪狂犬病病毒(PRV)、猪流行性腹泻病毒(PEDV)、猪瘟病毒(CSFV)、猪繁殖与呼吸综合征病毒(PRRSV)均为阴性,特异性良好;对50份疑似病料同时进行荧光定量PCR检测和常规PCR检测,结果显示荧光定量PCR阳性检测率(38%)高于常规PCR阳性检测率(25%),证实本试验建立的SVV Taqman实时荧光定量PCR检测方法可实现对SVV的临床诊断和定量分析。 相似文献
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为建立可同时鉴别检测非洲猪瘟病毒(ASFV)以及猪圆环病毒2型(PCV2)、3型(PCV3)的三重PCR方法,根据GenBank中登录的ASFV、PCV2、PCV3相关基因保守序列,分别设计合成特异性引物,通过对引物浓度、退火温度、循环数等反应条件进行优化,建立了针对3种病原的三重PCR检测方法,并开展了敏感性、特异性及重复性试验。结果显示:引物最佳终浓度为0.375 pmol/μL,最佳退火温度为61℃,最佳循环数为35;该方法对ASFV、PCV2、PCV3的扩增目的条带分别为873、530和217 bp,对猪细小病毒和伪狂犬病病毒的扩增结果均为阴性;对ASFV、PCV2、PCV3重组质粒标准品的检出下限均为1×104 copies/μL;相同条件下重复性试验获得均匀一致的结果。结果表明,本研究建立的三重PCR检测方法具有特异性强,敏感性及重复性好等优点,可用于ASFV、PCV2和PCV3感染的快速鉴别诊断及流行病学调查。 相似文献
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为了建立一种针对猪水泡病毒(SVDV)的反转录环介导等温扩增技术(RT-LAMP),用于减少猪水泡病与口蹄疫因临床症状相似而造成的误诊,试验针对SVDV VP1基因序列设计4个LAMP引物,其中外侧引物用于SVDV PCR检测,并利用RT-LAMP和RT-PCR方法进行敏感性试验、特异性试验以及临床样品检测。结果表明:针对SVDV的RT-LAMP检测方法的敏感性是RT-PCR的10倍; RT-LAMP方法的特异性良好,与口蹄疫病毒无交叉反应;两种方法对临床样品检测的符合率为100%。说明试验建立的RT-LAMP方法具有敏感性高、特异性强、快速等特点,适合实验室和临床对SVDV的快速诊断。 相似文献
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根据基因库中的口蹄疫病毒（FM—DV）,猪水疱病病毒（SVDV）和水疱性口炎病毒（VSV）各基因序列,设计了与FMDV,SVDV和VSV互补的3对特异性引物,对样品中的cDNA模板进行了多重PCR扩增及反应条件的优化,结果同时得到与设计相符合的3条特异性条带,分别为189bp,125bp和300bp。用这3对引物对病毒样品cDNA模板进行多次扩增,均能稳定得到与设计相符合的3条特异性条带。本试验能特异、敏感、快速地鉴定FMDV,SVDV和VSV。 相似文献
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根据GenBank中的猪伪狂犬病病毒(PRV)gE、猪圆环病毒2型(PCV-2)ORF2、猪细小病毒(PPV)VP2基因序列,设计了3对引物,成功建立了检测PRV野毒株、PCV-2和PPV的多重PCR诊断方法,扩增产物分别为288 bp、419 bp、681 bp。敏感性、特异性试验结果显示,该PCR对3种病毒的最低核酸检测量分别为PRV 48.2 pg/L、PCV-2 36.7 pg/L、PPV 0.25 ng/L,而PRV(gE基因缺失株)、猪瘟病毒(CSFV)、猪繁殖与呼吸综合征病毒(PRRSV)、大肠杆菌的扩增结果均为阴性。对87份自然感染病猪样品的检测结果表明,该多重PCR检测结果与单一PCR检测结果完全符合。结果表明,该多重PCR方法具有很好的特异性和敏感性,可用于临床PRV野毒株和gE基因缺失疫苗株、PCV-2和PPV的检测。 相似文献
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魏静 《四川畜牧兽医学院学报》2009,(4):28-32
在现代法律秩序中,商会自治规范是制定法的基础和必要的补充,甚至在某些方面替代了制定法;商会自治规范主要包括商会组织规范、行为规范、惩罚规范以及争端解决规范等;其效力仅及于其内部成员;商会自治规范和制定法之间存在冲突,但也存在整合的基础。 相似文献
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本文概述了猪的毛色类型、猪的毛色遗传模式,着重综述了猪毛色基因分子基础的研究进展,指出存在问题并就未来发展方向做了思考。 相似文献
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REASONS FOR PERFORMING STUDY: Centesis of the bicipital bursa using an 8.9 cm long spinal needle has been reported but the alternative of employing a 3.8 cm long hypodermic needle requires validation. OBJECTIVE: To compare the efficacy of 2 different methods of centesis of the bicipital bursa and to evaluate the usefulness of ultrasonographic imaging to determine the location of solution administered when centesis of the bursa is attempted. METHODS: For Trial 1, 6 clinicians, who had no previous experience of centesis of the bicipital bursa, attempted to inject a solution composed of an aqueous radiopaque contrast medium and physiological saline solution (PSS) into the bicipital bursae of 2/12 horses using the previously described distal approach to inject one bursa and a proximal approach to inject the contralateral bursa. The bicipital tendon and bursa were examined ultrasonographically before and after injection; and both shoulders were examined radiographically to identify the location of the medium. In Trial 2, another 6 clinicians, also with no previous experience of centesis, repeated Trial 1, using 6 horses, but the radiopaque contrast medium was mixed with air instead of PSS. RESULTS: Accuracy of centesis using the proximal approach was 39% and that of the distal approach 28%. Ultrasonographic examination of the shoulder allowed the location of solution and air to be accurately predicted in all 12 shoulders examined. CONCLUSIONS: Clinicians who have had no previous experience performing centesis of the bicipital bursa are unlikely to be successful in centesis using either approach. Radiographic examination after injecting a radiopaque contrast medium may be necessary to assess the success of centesis especially if bursal fluid is not obtained during centesis. Injecting air along with the radiopaque contrast medium provides more accurate ultrasonographic confirmation of centesis and better radiographic definition than does injection without air. 相似文献
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Effects of size of ingestively masticated fragments of plant tissues on kinetics of digestion of NDF
Ingestively masticated fragments were collected and sized via sieving. Different sizes of esophageal masticate and ruminal digesta fragments, and ground fragments of larger masticated pieces were incubated in vitro, and undigested NDF remaining at intervals of up to 168 h of incubation was determined. The ruminal age-dependent time delay (tau) for onset of digestion of NDF was positively correlated (P < 0.004) with the mean sieve aperture estimated to retain 50% of the fragments between successive sieve apertures (MRA). Degradation rate of potentially degradable NDF (PDF) and level of indigestible NDF were not related (P > 0.10) to MRA of masticated and ground fragments. Estimates of tau were positively related to MRA, with slopes of bermudagrass < corn silage < ruminal fragments of corn silage. It was concluded that fragment size-, and consequently, ruminal age-dependent onset of PDF degradation of a mixture of different fragment sizes results in an age-dependent rate of degradation of the more rapidly degrading of two subentities of PDF. Models are proposed that assume a tau before onset of simultaneous degradation of PDF from two pools characterized as having gamma-modeled age-dependency and age-constant rates. The ruminal age-dependent pool seems to be associated with the faster-degrading pool, and its rate parameter increases with range in MRA in the population of fragments. Conceptually, the ruminal age-dependent rate parameter for PDF degradation seems to represent a composite of several effects: 1) effects of the size-dependent tau; 2) range in MRA of the population of ingestively masticated fragments; and 3) subentities of PDF that degrade via more rapid age-dependent rates compared with subentities of PDF that degrade via age-constant rates. The estimated fractional rates of ruminative comminution of ingestively masticated fragments (0.060 to 0.075/h) were of a magnitude similar to the mean fractional rates of PDF digestion (0.030 to 0.085/h), which implies that ruminative comminution may be first-limiting to fractional rate of PDF digestion. The in vivo roles of ingestive and ruminative mastication of fragments on PDF degradation must be considered in any kinetic system for estimating PDF digestion in the rumen. These results and others in the literature suggest that the rate of surface area exposure rather than intrinsic chemical attributes of PDF may be first-limiting to degradation rate of PDF in vivo. 相似文献
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乳酸杆菌作为益生菌广泛用于人和动物.本文综述了乳酸杆菌改善宿主健康的机制.乳酸杆菌可通过产生抗菌物质如乳酸、过氧化氢、细菌素,或者通过竞争营养或肠道黏附位点来抑制致病菌;通过诱导黏附素的分泌或阻止细胞凋亡而增强肠道的屏障功能,从而保护肠道.文章重点讨论了乳酸杆菌表面成分(表面蛋白、脂磷壁酸和肽聚糖)与肠道受体(C型凝集素受体、Toll样受体和Nod样受体),阐述了他们结合后启动免疫调节信号,调控肠道免疫功能以发挥改善健康作用的机制. 相似文献
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Billinghurst RC Brama PA van Weeren PR Knowlton MS McIlwraith CW 《American journal of veterinary research》2004,65(2):143-150
OBJECTIVE: To determine whether serum concentrations of biomarkers of skeletal metabolism can, in conjunction with radiographic evaluation, indicate severity of osteochondrosis in developing horses. ANIMALS: 43 Dutch Warmblood foals with varying severity of osteochondrosis. PROCEDURE: 24 foals were monitored for 5 months and 19 foals were monitored for 11 months. Monthly radiographs of femoropatellar-femorotibial and tibio-tarsal joints were graded for osteochondral abnormalities. Serial blood samples were assayed for 8 cartilage and bone biomarkers. At the end of the monitoring period, foals were examined for macroscopic osteochondrosis lesions. RESULTS: Temporal relationships were evident between certain serum biomarkers and osteochondrosis severity in foals during their first year. Biomarkers of collagen degradation (collagenase-generated neoepitopes of type-II collagen fragments, type-I and -II collagen fragments [COL2-3/4C(short)], and cross-linked telopeptide fragments of type-I collagen) and bone mineralization (osteocalcin) were positive indicators of osteochondrosis severity at 5 months of age. In foals with lesions at 11 months of age, osteochondrosis severity correlated negatively with COL2-3/4C(short) and osteocalcin and positively with C-propeptide of type-II procollagen (CPII), a collagen synthesis marker. Radiographic grading of osteochondrosis lesions significantly correlated with macroscopic osteochondrosis severity score at both ages and was strongest when combined with osteocalcin at 5 months and CPII at 11 months. CONCLUSIONS AND CLINICAL RELEVANCE: The ability of serum biomarkers to indicate osteochondrosis severity appears to depend on stage of disease and is strengthened with radiography. In older foals with more permanent lesions, osteochondrosis severity is significantly related to biomarker concentrations of decreased bone formation and increased cartilage synthesis. 相似文献