【作者】 黄诚；

【导师】 程安春； 汪铭书；

【作者基本信息】 四川农业大学 ， 预防兽医学， 2003， 硕士

【摘要】 根据国际基因库（GeneBank）1995年发布的鹅细小病毒（GPV）B株的全基因组序列，针对GPV的保守基因片段VP3设计一对引物，建立了一种用于检测GPV的特异的PCR方法，该法能够特异地扩增出预期大小约441bp片段。特异性实验证明，该法对鸭瘟病毒、鹅致病性大肠杆菌O₈和O₁、雏鹅新型病毒性肠炎病毒（NGVEV）呈阴性反应，仅能从鹅细小病毒（GPV）和番鸭细小病毒（MDPV）中扩增出特异性片段。灵敏性实验证明该方法的检测GPV核酸的灵敏度达到0.47pg。 运用所建立的特异的PCR方法，对人工感染鹅细小病毒后病毒在雏鹅体内的分布规律进行了研究，结果表明：病毒在雏鹅体内的分布规律因病毒感染的途径的不同而存在差异。经口感染组在感染后4小时不能从组织中检出病毒DNA，感染后8小时能从心脏、肝脏中检出病毒DNA、感染后24小时能从胸腺、胰腺、十二指肠、小肠、法氏囊、心脏、肝脏、脾脏、肾脏、血液中检出病毒DNA，感染后48小时后能够从直肠、骨髓、肺脏中检出病毒DNA，感染后72小时能从小脑、延髓、盲肠、肌肉、粪便中检出病毒DNA，感染后312小时后仍能从十二指肠、空肠、直肠、心脏、肝脏、肾脏、肌肉、粪便中检出病毒DNA。经皮下注射感染组在感染后4小时不能从组织中检出病毒DNA，感染后8小时能够从心脏，肝脏中检测出病毒DNA，感染后24小时能够从延髓、胸腺、胰腺、十二指肠、空肠、盲肠、法氏囊、心脏、肝脏、脾脏、肺脏、肾脏、血液、小脑、直肠、肌肉、粪便中检出病毒DNA，感染后48小时后能从骨髓中检测出病毒DNA，在感染后312小时后仍能从十二指肠、空肠、盲肠、心脏、肝脏、肾脏，粪便中检出病毒DNA。经滴鼻感染组在感染后4小时不能从组织中检出病毒DNA，感染后8小时后能够从心脏、肝脏中检出病毒DNA，感染后24小时能够从胸腺、十二指肠、直肠、空肠、肺脏、心脏、肝脏中检测出病毒DNA，感染后48小时能够从小脑、延髓、胰、盲肠、法氏囊、脾脏、肌肉、骨髓、粪便、血液中检出病毒DNA，在感染后312小时后仍能从胸腺、十二指肠、空肠、盲肠、直肠、心脏、肝脏、肾脏、粪便中检出病毒DNA。三个实验组的鹅在感染后83天均不能从体内检出病毒DNA。经皮下感染后病毒在雏鹅体内的分布速度较经口服和滴鼻感染后病毒在雏鹅体内分布速度快。各种受检组织中心脏和肝脏的检出率最高。经不同途径感染后，病毒均在感染后72小时在机体内各组织的分布最为广泛。本研究的三个实验组（口服，滴鼻，皮下注射），均未能从大脑、舌头、腺胃、食道中检出病毒DNA。更多还原

【Abstract】 Primers were designed based on the complete nucleotide sequence of goose parvovirus B strain that zadori subjected to Genebank in 1995,the PCR method for goose parvovirus detedtion was developed.Using this method we can amplify a differential fragment about 441bp from GPV sample ,but we cann’t get the same fragment from duck plague virus.Ecoli O1 and O8,MDPV, NGVEV samples.By this method a minimal sample of 0.47pg goose parvovirus nucleotide can be detected.Using the PCR method developed above,we studied the distribution regularity of virulent goose parvovirs in artifical infected gosling body.The result indicate that there exist difference among the three group(subcutaneously,orally,intranasally).The result of orally infected group: 4 hours after infected ,the GPV nucleotide cann’t be detedted from all samples collected; 8 hours afete infected, the GPV nucleotide can be detected from heart and liver;24 hours after infected, the GPV nucleotide can be detedted from thymus,pancreas,duodenum,small intestine,bursa fabricius ,heart,liver,spleen,kidney and blood;48 hours after infected, the GPV nucleotide can be firstly detected from rectum,bone marrow and lung;72 hours after infected , the GPV nucleotide can be detected from cerebellum ,mudulla oblongata,appendix,skeletal muscle,feces.312 hours after infected the GPV nucleotide still can be detected from duodenum,jejunum,rectum,heart,liver,kidney,skeletal muscle,feces.The result of subcutaneously infected group:4 hours after infected, the GPV nucleotide cann’t be detedted from all sample collected;8 hours after infected, the GPV nucleotide can be detected from heart and liver;24 hours after infected, the GPV nucleotide can be detected from mudulla oblongata,thymus,pancreas,duodenum,small intestine,appendix,bursa fabricius, heart, liver spleen,lung,kidney,blood,cerebellum,rectum,skeletal muscle,feces;48 hours after infected ,the GPV nucleotide can be firstly detected from bone marrow;312 hours after infected,the GPV nucleotide can still be detected from duodenum, small intestine,appendix,heart,liver,kidney,feces.The result of drop nose:4 hours after infected,the GPV nucleotide cann’t be detected from all sample collected,8 hours after infected,the GPV nucleotide can be detected from heart and liver.24 hours after infected,the GPV nucleotide can be detected from thymus,duodenum,rectum,jejunum,lung,cardiac muscle,48 hours after infected GPV nucleotide can be detected from cerebellum, medulla oblongata, Pancreas, cecum, bursa of Fabricius, spleen, Skeletal muscles, bone marrow, feces,blood.312 hours after infected GPV nucleotide can be detected from thymus, duodenum, jejunum, cecum, rectum, cardiac muscle, liver,Kidney,feces.during the period of this studies, we haven’t detected the GPV nucleotide from cerebrum,tongue,glandular stomache and gullet.83 days after infection,the virus DNA can’t be detected from any tissues of goslings.liver,carciac muscle, jejunum are the most suitable tissues for virus isolation.72 hours or so after infection GPV distribute most widely in gosling.更多还原