Multiple virus resistance in transgenic plants conferred by the human dsRNA-dependent protein kinase

We have developed a new strategy for engineering resistance to multipleviruses in plants. The strategy exploits the human double stranded (ds)RNA-dependent protein kinase (PKR). PKR is one of theinterferon-induced enzymes. It confers viral resistance in mammals byinhibitingviral replication through the inactivation of the translational initiationfactor, eIF-2, upon activation by dsRNA. The humanPKR gene was fused to the promoter of theArabidopsis blue copper binding protein gene(BCB) that is induced rapidly in response to wounding. Thechimeric gene cassette was introduced into tobacco plants. Expression of thePKR gene in transgenic tobacco plants was demonstrated byRNA gel blot analysis and autophosphorylation assay of anM _r 68,000 protein. The transgenic plantsexpressing the PKR gene showed significantly reduced viralsymptoms or no viral symptoms at all, when challenged by different plant RNAviruses, such as Cucumber mosaic virus, Tobaccoetch virus, or Potato virus Y. Thus, expressionof a single component in the human interferon pathway, thePKR gene, can effectively confer resistance to multipleviruses in transgenic plants.     

Effect of deleterious mutation-accumulation on the fitness of RNA bacteriophage MS2

Abstract.— RNA viruses show the highest mutation rate in nautre. It has been extensively demonstrated that, in the absence of purifying selection, RNA viruses accumulate deleterious mutations at a high rate. However, the parameters describing this accumulation are, in general, poorly understood. The present study reports evidences for fitness declines by the accumulation of deleterious mutations in the bacteriophage MS2. We estimated the rate of fitness decline to be as high as 16% per bottleneck transfer. In addition, our results agree with an additive model of fitness effects.     

Intron Conservation in a UV-Specific DNA Repair Gene Encoded by Chlorella Viruses

Large dsDNA-containing chlorella viruses encode a pyrimidine dimer-specific glycosylase (PDG) that initiates repair of UV-induced pyrimidine dimers. The PDG enzyme is a homologue of the bacteriophage T4-encoded endonuclease V. The pdg gene was cloned and sequenced from 42 chlorella viruses isolated over a 12-year period from diverse geographic regions. Surprisingly, the pdg gene from 15 of these 42 viruses contain a 98-nucleotide intron that is 100% conserved among the viruses and another 4 viruses contain an 81-nucleotide intron, in the same position, that is nearly 100% identical (one virus differed by one base). In contrast, the nucleotides in the pdg coding regions (exons) from the intron-containing viruses are 84 to 100% identical. The introns in the pdg gene have 5′-AG/GTATGT and 3′-TTGCAG/AA splice site sequences which are characteristic of nuclear-located, spliceosomal processed pre-mRNA introns. The 100% identity of the 98-nucleotide intron sequence in the 15 viruses and the near-perfect identity of an 81-nucleotide intron sequence in another 4 viruses imply strong selective pressure to maintain the DNA sequence of the intron when it is in the pdg gene. However, the ability of intron-plus and intron-minus viruses to repair UV-damaged DNA in the dark was nearly identical. These findings contradict the widely accepted dogma that intron sequences are more variable than exon sequences. Received: 13 May 1999 / Accepted: 20 August 1999     

Infectibility and sensitivity of UK raspberry, blackberry and hybrid berry cultivars to Rubus viruses

Six blackberry or hybrid berry cultivars and 19 raspberry cultivars were assessed for their infectibility with, and sensitivity to, graft inoculation with 10 distinct viruses found infecting Rubus in the UK. Cultivars were grafted with each of, two isolates of the pollen borne raspberry bushy dwarf virus (RBDV), five aphid borne viruses: black raspberry necrosis, raspberry leaf mottle (RLMV), raspberry leaf spot (RLSV), rubus yellow net and raspberry vein chlorosis (RVCV); and isolates of the nematode transmitted nepoviruses, arabis mosaic, raspberry ringspot, strawberry latent ringspot and tomato black ring. All tested cultivars were infectible with a resistance breaking isolate of RBDV but only about half of that number with the Scottish type isolate of the virus. The raspberry cvs Autumn Bliss, and occasionally Glen Garry and Glen Prosen, developed leaf yellowing symptoms following infection with RBDV, but none of the other infected cultivars showed obvious leaf symptoms when kept in a heated glasshouse during the growing season. All tested cultivars were infectible with each of the four viruses transmitted in nature by the aphid, Amphorophora idaei. Most were infected symptomlessly, but seven cultivars developed severe leaf spotting symptoms due to infection with RLMV or RLSV. All but one of the raspberry cultivars were infectible with RVCV, which is transmitted in nature by the aphid Aphis idaei, and almost all infected plants developed leaf symptoms; only one of the hybrid berry or blackberry cultivars tested was infected with RVCV. In tests with the four nepoviruses, all tested cultivars, except Tummelberry, were infectible with at least one or more of these viruses. However, cultivars responded differently to challenge inoculation with different isolates of individual nepoviruses. Several cultivars developed chlorotic leaf mottling following infection with some nepovirus isolates. The implications of these results for virus control are discussed in the light of the changing pattern of virus and virus vector incidence in the UK.     

Molecular mimicry and the autoimmune response to the peripheral nerve myelin PO glycoprotein

In the Lewis rat immunisation with the myelin PO glycoprotein can induce an inflammatory demyelinating disease of the peripheral nervous system, experimental allergic neuritis (EAN), which has many clinical and histopathological parallels with the human disease the Guillain-Barre syndrome. In view of the reported association of GBS with a number of infectious agents we have investigated whether molecular mimicry may occur between microbial antigens and the PO protein that could possibly trigger a similar pathogenic autoimmune response in man. A computer search of the available protein sequence data bases identified several absolute sequence homologies between PO and viral proteins that involve five or more consecutive amino acid residues. Four of these sequence homologies involved viral pathogens previously associated with the Guillain-Barre syndrome, namely Epstein-Barr virus (EBV), cytomegalovirus (CMV), Varicella zoster virus (VZV) and human immunodeficiency virus I (HIV I). Although, sequence homologies were also found between viral peptides and the neuritogenic determinants of PO, residues 56–71 and 180–199, these homologies proved incapable of eliciting EAN in the Lewis rat. These observations are discussed with reference to the role that molecular mimicry between T cell epitopes on pathogen derived antigens and the PO protein may play in the pathogenesis of the Guillain-Barre syndrome.Abbreviations EAN Experimental allergic neuritis - EAE experimental allergic encephalomyelitis - PNS peripheral nervous system - CNS central nervous system - MBP myelin basic protein - GBS Guillain Barre syndrome - CFA complete Freund's Adjuvant - LPC lysophosphatidyl choline - VZV Varicella zoster virus - CMV cytomegalovirus - EBV Epstein Barr virus - HIV I human immunodeficiency virus I Special issue dedicated to Dr. Alan N. Davison     

饮用水水源与自来水中病毒和指示细菌存在水平的比较研究

自来水中人肠道病毒的存在已引起人们极大的关切和忧虑。本文报道了武汉东湖水和以东湖为水源的自来水中病毒和指示细菌的存在水平。水源水经过预加氯消毒、絮凝沉淀、砂滤和最后加氯消毒处理而成的自来水中细菌总数检测范围是41—500/升,总大肠菌是2—13/升、粪大肠菌为0—4/升、大肠菌噬菌体是0—13.6PFU/升,平均2.48PFU/升,肠道病毒为0—78PFU/升,平均为6.7PFU/升。水源水经制水工艺处理去除指示细菌、大肠菌噬菌体和肠道病毒的效率分别为97.95—99.99％,90.63％和53.18％。这种结果说明自来水制水工艺能有效地降低指示细菌,大肠菌噬菌体,而去除肠道病毒效率较低,揭示肠道病毒对环境压专的耐受性明显地比指示细菌强。     

Time scale of Poxvirus evolution

Unlike in vertebrates and RNA viruses, the molecular clock has not been estimated so far for DNA viruses. The extended conserved central region (102 kb) of the orthopoxvirus genome and the DNA polymerase gene (3 kb) were analyzed in viruses representing several genera of the family Poxviridae. Analysis was based on the known dating of the variola virus (VARV) transfer from Western Africa to South America and previous data on the phylogenetic relatedness of modern West African and South American isolates of VARV. The mutation accumulation rate was for the first time estimated for these DNA viruses at (0.9–1.2) × 10^?6 substitutions per site per year. It was assumed that poxviruses diverged from an ancestor approximately 500,000 years ago to form the recent species and that the ancestor of the genus Orthopoxvirus emerged approximately 300,000 years ago and gave origin to the modern species approximately 14,000 years ago.     

不同容量胸段经椎间孔硬膜外注射阻滞范围的CT影像研究及诊断性阻滞的可行性分析

摘要 目的：探讨不同容量对胸段经椎间孔硬膜外注射（TFEI）药液扩散范围和镇痛效果的影响以及胸段TFEI用于诊断性阻滞的可行性。方法：选择2021年1月至2022年12月南京大学医学院附属鼓楼医院收治的胸段带状疱疹相关疼痛患者140例,随机分为4组,实施单次TFEI并分别注入不同容量含造影剂局麻药（A组：0.2 mL;B组：0.5 mL;C组：1.0 mL;D组：2 mL）,CT扫描并观察造影剂在硬膜外向头侧、尾侧及总扩散节段,造影剂在椎间孔、同侧椎旁间隙、同侧及对侧硬膜外间隙扩散情况,判断是否为选择性神经根阻滞。评估注射前、注射后30分钟及24小时视觉模拟评分（VAS）。结果：头侧和尾侧扩散节段以及总扩散节段数,D组最多,A组最少（P<0.05）;C组、D组造影剂扩散≥3个节段发生率明显高于B组（P<0.05）;C组、D组病例造影剂扩散至同侧椎旁间隙和对侧硬膜外间隙的发生率明显高于A组、B组（P<0.05）,仅A组37.1%的病例实现选择性神经根阻滞,其余各组均无选择性阻滞病例。注射后30分钟,C、D组VAS评分显著低于A、B组（P<0.05）;注射后24小时,D组VAS评分显著低于A、B组（P<0.05）。结论：胸段带状疱疹相关疼痛患者TFEI药液扩散范围随注射容量的增加而扩大,且在硬膜外倾向于头侧扩散,2 mL容量单次TFEI可阻滞3个以上的神经节段,获得良好的镇痛效果。胸段TFEI行诊断性阻滞的可行性较差。     

Darwin’s Galapagos gourd: providing new insights 175 years after his visit

The year 2010 marks the 175th anniversary of Charles Darwin’s visit to the Galapagos Islands. A recent paper by J. C. Briggs, ‘Darwin’s biogeography’ (Journal of Biogeography, 2009, 36 , 1011–1017), summarizes Darwin’s contributions to the field of biogeography, stressing the importance of his natural history specimens. Here, we illustrate how a plant collected by Darwin during his visit to Floreana and not collected since can provide insights into dispersal to oceanic islands as well as extinction of island plants, based on ancient DNA from Darwin’s herbarium specimen.