罗氏沼虾体内两种病毒颗粒的分离、纯化与核酸特性

从患肌肉白浊症状的罗氏沼虾幼苗体内分离纯化得到两种大小不同的病毒颗粒.这两种病毒颗粒均为对称的20面体结构,表面光滑,无囊膜,对氯仿不敏感.一种是直径为26nm～27nm的颗粒,在氯化铯中的密度为132g/cm3,病毒基因组含两段单链的RNA,分别为30kb和12kb,具有诺达病毒科成员的特征.一种是直径为14nm～16nm的颗粒,在氯化铯中的密度为133g/cm3,含有一段大小为09kb的单链RNA,拟为卫星病毒样颗粒或辅助病毒.     

罗氏沼虾诺达病毒的核酸检测及其部分序列分析

根据安替列群岛分离的罗氏沼虾诺达病毒株基因组序列(MrNV-ant),制备特异性核酸探针,设计特异性引物,用点杂交和RT-PCR的方法检测在中国境内分离的罗氏沼虾诺达病毒(MrNV-chin).点杂交的方法可以检测出少于26ng的患肌肉白浊症的组织样品中的病毒,或少于25ng的病毒RNA样品;RT-PCR可以检测出少于25pg的RNA样品.扩增的MrNV-chin RNA1序列长858个核苷酸,与MrNV-ant的核苷酸一致率为957%,两者翻译后的氨基酸序列的一致率为99.7%.扩增的MrNV-chin RNA 2序列长1121个核苷酸,与MrNV-ant的核苷酸一致率为92%,两者翻译后的氨基酸序列的一致率为93.2%.因此,MrNV-ant和MrNV-chin应属于同一种病毒的不同分离株.用两株罗氏沼虾诺达病毒的RNA聚合酶序列与其它6株诺达病毒RNA聚合酶序列比较后构建的进化树中,罗氏沼虾诺达病毒与Alphanodavirus的亲缘关系近于与Betanodavirus的亲缘,组成了一个新的分支.     

不同途径感染白斑综合征病毒 (WSSV) 对日本沼虾的致病性

日本沼虾肉质鲜美、经济价值高,但其对白斑综合征病毒(White spot syndrome virus,WSSV)的易感性尚不明确。采用流行病学调查、感染试验和荧光定量PCR(qPCR)等方法,研究了日本沼虾Macrobrachium nipponense对WSSV的易感性、感染发病率以及WSSV在其体内的增殖情况。结果表明:日本沼虾是WSSV的自然宿主,上海市附近省份地区120个样本的自然携带率为8.33%。日本沼虾可以通过注射、摄食、浸泡途径感染WSSV,10 d内累计感染率均为100%,累计死亡率分别为100%、75%和0%。其中注射途径感染日本沼虾,感染后5 d肌肉病毒含量达到感染后1 d的1 000倍,8 d死亡率达100%。用注射WSSV感染日本沼虾死亡率来测量病毒致死日本沼虾的半数致死剂量(LD_(50)),2.71×10~5病毒粒子/μL能使日本沼虾死亡率达到50%以上。在养殖生产中,日本沼虾可以通过摄食感染WSSV的病虾或死虾而感染WSSV,也能通过浸泡在含WSSV的水体中而感染,并因此成为一种传播媒介,从而影响了WSSV的迅速传播与致病。     

中国对虾中一种球状病毒的分离提纯与检测

从人工养殖生长比较缓慢的对虾与病虾的肝胰腺和消化道中,分离出一种大小为８０ｎｍ的球状病毒。同时采用超薄切片技术可在对虾中肠细胞中发现大量的球状病毒,这些病毒聚集成片,在同一细胞内的细胞器,如线粒体等均发生病变。此外,将分离提取的球状病毒进行感染试验,感染死亡率为６０％。并且从感染致死的对虾消化腺及肠道中,同样可以观察到大小相同、形态一致的球状病毒颗粒。     

罗氏沼虾肌肉白浊病病原研究

从患白浊病的罗氏沼虾虾苗病灶分离到10多株菌株,其中一株经人工感染证实为病原菌,经ATB Expression自动细菌鉴定仪测试,为木糖葡萄球菌,其主要特性为拟球状（直径0.8-1.1μm）,革兰氏阳性,无鞭毛,无荚膜,不产生芽孢,发酵葡萄糖、果糖、麦芽糖、乳糖、海藻糖、阿拉伯糖、蔗糖及甘露醇产酸,β-半乳糖甘酶、过氧化氢酶及脲酶阳性,氧化酶,血浆凝固醇、V-P反应,精氨酸双水解酶、鸟氨酸脱羧酶均为阴性。最适生长温度、盐度和pH值范围分别为28-37℃、0%-2%、6-9,对红霉素、卡那毒素、头孢氨苄及庆大霉素等药物敏感。     

罗氏沼虾野田村病毒中国株RT-LAMP-LFD快速检测方法的研究北大核心CSCD

罗氏沼虾白尾病(White tail disease,WTD)是流行于罗氏沼虾苗种阶段的重要疾病,该病病原被确定为罗氏沼虾野田村病毒(Macrobrachium rosenbergii Nodavirus,MrNV),为了建立罗氏沼虾野田村病毒中国分离株(MrNV-chin)的便捷、灵敏、特异的分子诊断方法,本研究根据罗氏沼虾野田村病毒RNA2基因保守序列的6个区域设计了4条特异性引物和1条检测探针,将环介导等温扩增技术与侧向流层析试纸(Lateral flow dipstick,LFD)相结合,建立了MrNV的RT-LAMP-LFD检测方法。结果表明,该方法的灵敏度是常规琼脂糖凝胶电泳的10倍;从核酸提取到检测结果判定仅需45min;反应特异性强,对其他虾类病毒WSSV、IHHNV、IMNV、TSV和YHV扩增结果均为阴性;操作简单,不需要复杂的仪器,在61℃水浴反应即可;利用LFD试纸显示结果,肉眼可直接观察判定。作为一种快速灵敏实用的分子诊断技术,该方法有利于罗氏沼虾白尾病的诊断与预防。     

罗氏沼虾野田村病毒中国株RT-LAMP-LFD快速检测方法的研究

罗氏沼虾白尾病(White tail disease,WTD)是流行于罗氏沼虾苗种阶段的重要疾病,该病病原被确定为罗氏沼虾野田村病毒(Macrobrachium rosenbergii Nodavirus,MrNV),为了建立罗氏沼虾野田村病毒中国分离株(MrNV-chin)的便捷、灵敏、特异的分子诊断方法,本研究根据罗氏沼虾野田村病毒RNA2基因保守序列的6个区域设计了4条特异性引物和1条检测探针,将环介导等温扩增技术与侧向流层析试纸(Lateral flow dipstick,LFD)相结合,建立了MrNV的RT-LAMP-LFD检测方法。结果表明,该方法的灵敏度是常规琼脂糖凝胶电泳的10倍;从核酸提取到检测结果判定仅需45min;反应特异性强,对其他虾类病毒WSSV、IHHNV、IMNV、TSV和YHV扩增结果均为阴性;操作简单,不需要复杂的仪器,在61℃水浴反应即可;利用LFD试纸显示结果,肉眼可直接观察判定。作为一种快速灵敏实用的分子诊断技术,该方法有利于罗氏沼虾白尾病的诊断与预防。     

壬基酚和雌二醇干扰罗氏沼虾卵黄蛋白原VTG基因表达的效应

对刚孵化后的罗氏沼虾(Macrobrachium rosenbergii)亲虾用高低两个浓度的壬基酚(nonylphenol,NP,100μg/L和0·01μg/L)和雌二醇(estradiol,E2,1μg/L和0·01μg/L)进行浸泡处理,分别于3d和5d对罗氏沼虾肝胰腺和卵巢中卵黄蛋白原(vitellogenin,VTG)基因表达变化进行半定量分析。结果显示,NP和E2能够提高罗氏沼虾肝胰腺和卵巢中卵黄蛋白原VTG基因的表达。100μg/LNP对罗氏沼虾表现出雌激素效应,0·01μg/LNP作用效果不明显;而E2则在1μg/L和0·01μg/L两个浓度下均对罗氏沼虾有较强的雌激素效应。在100μg/LNP作用下,卵巢VTG表达量3d和5d均保持较高水平,无明显下降,其他实验组均是3dVTG基因表达量升高,5d后表达量较3d表达量有所降低。结果表明,与其他动物一样,NP对罗氏沼虾具有内分泌干扰作用。     

罗氏沼虾源维氏气单胞菌的分离鉴定、毒力基因检测及组织病理学观察

为鉴定江苏省高邮市某养殖场罗氏沼虾(Macrobrachium rosenbergii)发病死亡的病原体,研究从患病濒死的罗氏沼虾肝胰腺中分离出一株优势菌WSQ-1,对其进行纯化培养、革兰氏染色、生理生化特征鉴定、16S rRNA及gyrB基因序列分析,鉴定为维氏气单胞菌(Aeromonas veronii)。回归感染实验表明,分离株WSQ-1对罗氏沼虾有较强致病性,且出现与自然发病类似的症状:感染虾活力减弱,食欲下降,肌肉发白; 72h半致死浓度(LD50)为2.51×10⁶ CFU/mL。组织病理学观察可见其肝胰腺组织病理损伤,并随感染浓度的增加或感染持续时间的延长而逐渐加重:低浓度或感染前期结缔组织间隙增大并积聚少量炎性细胞;继而肝小管管腔变形,肝细胞空泡化,部分组织坏死;高浓度或感染后期肝小管及结缔组织大面积坏死导致不可逆损伤。毒力基因检测显示该菌携带Acsv、AexT、Act、Aer、GcaT、Acg和OmpAI共7个毒力基因,表明分离菌具有较强致病性。药敏试验显示,该分离菌对头孢哌酮等16种药物敏感,对克拉霉素等10种药物耐药,仅对卡那霉素表现为中度敏...     

罗氏沼虾Rab11基因cDNA克隆及其组织表达分析

为了发掘罗氏沼虾(Macrobrachium rosenbergii)免疫相关基因, 研究Rab蛋白(Ras-related proteins inbrain)在罗氏沼虾免疫应答中发挥的作用, 研究采用RACE-PCR技术克隆了罗氏沼虾Rab11基因全长cDNA序列, 记为MrRab11。全长1381 bp, 包括226 bp的5'UTR, 511 bp的3'UTR和645 bp的开放阅读框, 编码214个氨基酸, 含有一个Rab结构域。氨基酸序列比对显示, 罗氏沼虾与冈比亚按蚊(Anopheles gambiae)、蚤状溞(Daphniapulex)、叶蝉(Homalodisca vitripennis) Rab11一致性分别为82%、83%和82%。软件预测, MrRab11编码的蛋白分子量约为23.75 kD, 等电点约为5.34。实时荧光定量表达分析表明, MrRab11基因在罗氏沼虾各组织中都有表达, 肝胰腺中的表达量最高, 其次是肌肉和肠道。在阴沟肠杆菌(Enterobacter cloacae)感染12h后, 罗氏沼虾肝胰腺中MrRab11的表达量上升, 显著高于对照组(P0.05), 推测这是MrRab11对阴沟肠杆菌的应激表达,MrRab11在肝胰腺中参与了罗氏沼虾免疫应答过程。     

Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis

It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis.     

The structure, reproduction and taxonomy of Vidalia and Osmundaria (Rhodophyta, Rhodomelaceae)

Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera.     

New or rare chromosome counts in Artemisia L. (Asteraceae, Anthemideae) and related genera from Kazakhstan

Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted.     

肝癌中HBV和HCV基因和抗原的分布及意义

采用原位分子杂交方法检测HCV RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细     

Selection with two alleles and complete dominance

For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment.