The complete genome sequence of J virus reveals a unique genome structure in the family Paramyxoviridae

J virus (J-V) was isolated from feral mice (Mus musculus) trapped in Queensland, Australia, during the early 1970s. Although studies undertaken at the time revealed that J-V was a new paramyxovirus, it remained unclassified beyond the family level. The complete genome sequence of J-V has now been determined, revealing a genome structure unique within the family Paramyxoviridae. At 18,954 nucleotides (nt), the J-V genome is the largest paramyxovirus genome sequenced to date, containing eight genes in the order 3'-N-P/V/C-M-F-SH-TM-G-L-5'. The two genes located between the fusion (F) and attachment (G) protein genes, which have been named the small hydrophobic (SH) protein gene and the transmembrane (TM) protein gene, encode putative proteins of 69 and 258 amino acids, respectively. The 4,401-nt J-V G gene, much larger than other paramyxovirus attachment protein genes sequenced to date, encodes a putative attachment protein of 709 amino acids and distally contains a second open reading frame (ORF) of 2,115 nt, referred to as ORF-X. Taken together, these novel features represent the most significant divergence to date from the common six-gene genome structure of Paramyxovirinae. Although genome analysis has confirmed that J-V can be classified as a member of the subfamily Paramyxovirinae, it cannot be assigned to any of the five existing genera within this subfamily. Interestingly, a recently isolated paramyxovirus appears to be closely related to J-V, and preliminary phylogenetic analyses based on putative matrix protein sequences indicate that these two viruses will likely represent a new genus within the subfamily Paramyxovirinae.     

A new model for Hendra virus encephalitis in the mouse

Hendra virus (HeV) infection in humans is characterized by an influenza like illness, which may progress to pneumonia or encephalitis and lead to death. The pathogenesis of HeV infection is poorly understood, and the lack of a mouse model has limited the opportunities for pathogenetic research. In this project we reassessed the role of mice as an animal model for HeV infection and found that mice are susceptible to HeV infection after intranasal exposure, with aged mice reliably developing encephalitic disease. We propose an anterograde route of neuroinvasion to the brain, possibly along olfactory nerves. This is supported by evidence for the development of encephalitis in the absence of viremia and the sequential distribution of viral antigen along pathways of olfaction in the brain of intranasally challenged animals. In our studies mice developed transient lower respiratory tract infection without progressing to viremia and systemic vasculitis that is common to other animal models. These studies report a new animal model of HeV encephalitis that will allow more detailed studies of the neuropathogenesis of HeV infection, particularly the mode of viral spread and possible sequestration within the central nervous system; investigation of mechanisms that moderate the development of viremia and systemic disease; and inform the development of improved treatment options for human patients.     

Analysis of Arabidopsis genome sequence reveals a large new gene family in plants

A detailed analysis of the currently available Arabidopsis thaliana genomic sequence has revealed the presence of a large number of open reading frames with homology to the stigmatic self-incompatibility (S) genes of Papaver rhoeas. The products of these potential genes are all predicted to be relatively small, basic, secreted proteins with similar predicted secondary structures. We have named these potential genes SPH (S-protein homologues). Their presence appears to have been largely missed by the prediction methods currently used on the genomic sequence. Equivalent homologues could not be detected in the human, microbial, Drosophila or C. elegans genomic databases, suggesting a function specific to plants. Preliminary RT-PCR analysis indicates that at least two members of the family (SPH1, SPH8) are expressed, with expression being greatest in floral tissues. The gene family may total more than 100 members, and its discovery not only illustrates the importance of the genome sequencing efforts, but also indicates the extent of information which remains hidden after the initial trawl for potential genes.     

Phylogenetic relationships within the harpacticoid copepod family Peltidiidae Sars, including the description of a new genus

With the discovery of Alteuthoides kootare gen. et sp. no v. , found living inside an hexactinellid sponge from offshore waters of New Zealand, and the synonymization herein of Paralteutha T. Scott with Eupelte Claus, the number of genera in the harpacticoid copepod family Peltidiidae Sars now stands at eight. In the morphology of the maxillipeds and the armature of the first leg exopods, Alteuthoides is most similar to Alteuthella (A. Scott) and Alteuthellopsis Lang. These shared apomorphies are regarded as convergent ecological specializations to a lifestyle as close associates of other marine invertebrates. Results of a cladistic analysis further assist in the interpretation of relationships within the family. Lang's speculation that Peltidium and Parapeltidium form an independent lineage from the ancestral peltidiid gains further support. Alteutha remains as the basal group to the second lineage in comparison with which all remaining genera are derived in some respect. Alteuthellopsis would appear to be the most advanced genus in the family. Because of recent additions to Neopeltopsis Hicks and Alteuthellopsis , revised diagnoses, together with a revised key to genera, are also presented.     

The structure of a rigorously conserved RNA element within the SARS virus genome

We have solved the three-dimensional crystal structure of the stem-loop II motif (s2m) RNA element of the SARS virus genome to 2.7-Å resolution. SARS and related coronaviruses and astroviruses all possess a motif at the 3′ end of their RNA genomes, called the s2m, whose pathogenic importance is inferred from its rigorous sequence conservation in an otherwise rapidly mutable RNA genome. We find that this extreme conservation is clearly explained by the requirement to form a highly structured RNA whose unique tertiary structure includes a sharp 90° kink of the helix axis and several novel longer-range tertiary interactions. The tertiary base interactions create a tunnel that runs perpendicular to the main helical axis whose interior is negatively charged and binds two magnesium ions. These unusual features likely form interaction surfaces with conserved host cell components or other reactive sites required for virus function. Based on its conservation in viral pathogen genomes and its absence in the human genome, we suggest that these unusual structural features in the s2m RNA element are attractive targets for the design of anti-viral therapeutic agents. Structural genomics has sought to deduce protein function based on three-dimensional homology. Here we have extended this approach to RNA by proposing potential functions for a rigorously conserved set of RNA tertiary structural interactions that occur within the SARS RNA genome itself. Based on tertiary structural comparisons, we propose the s2m RNA binds one or more proteins possessing an oligomer-binding-like fold, and we suggest a possible mechanism for SARS viral RNA hijacking of host protein synthesis, both based upon observed s2m RNA macromolecular mimicry of a relevant ribosomal RNA fold.     

Eggplant latent viroid,the candidate type species for a new genus within the family Avsunviroidae (hammerhead viroids)

Viroids, small circular RNAs that replicate independently and in most cases incite diseases in plants, are classified into the families Pospiviroidae, composed of species with a central conserved region (CCR) and without hammerhead ribozymes, and Avsunviroidae, composed of three members lacking CCR but able to self-cleave in both polarity strands through hammerhead ribozymes. Here we report the biological and molecular properties of Eggplant latent viroid (ELVd). Purified circular ELVd induces symptomless infections when inoculated into eggplant seedlings. ELVd can be transmitted horizontally and through seed. Sequencing 10 complete cDNA clones showed that ELVd is a circular RNA of 332 to 335 nucleotides with high variability. This RNA can adopt a quasi-rod-like secondary structure of minimal free energy and alternative foldings that permit formation of stable hammerhead structures in plus and minus strands. The ribozymes are active in vitro and, most likely, in vivo. Considering the ELVd properties to be intermediate between those of the two genera of family Avsunviroidae, we propose ELVd as the type species of a third genus with the name ELAVIROID:     

Emaravirus属新病毒:中国枣树花叶伴随病毒全基因组测序

[目的]2016年以来,新疆阿克苏等地区出现了一种新的枣树病害,严重威胁当地及周边红枣产业。本研究旨在鉴定引起此次病害的病原,探究病原体的传播方式,为生物防治策略的开发提供研究基础。[方法]对发病植株进行小RNA测序以鉴定病原体;对新鉴定的病毒,通过RNAseq和反转录PCR获取病毒全序列;体外表达重组的病毒结构蛋白并制备特异性抗体,通过Western斑点杂交法在发病植株中确证病毒蛋白;收集发病区域的媒介昆虫,通过反转录PCR在昆虫体内检测病毒的基因组,鉴定可能的传毒介体。[结果]本研究鉴定一种新的欧洲山梣环斑病毒属病毒为新疆新发枣树病害可能的病原体,命名为中国枣树花叶伴随病毒(Chinese date mosaic-associated virus,CDMaV)。CDMaV是一种多分段单链RNA病毒,基因组由5条负义RNA组成;RNA1-RNA5大小分别为7160、2224、1230、1493、971 nt,每条基因组RNA的互补链包含一个开放阅读框,共编码5个蛋白,依次为依赖RNA的RNA聚合酶、包膜糖蛋白、核衣壳蛋白和两个未知功能蛋白。在枣树寄生虫枣瘿螨体内扩增到病毒序列,表明该病毒可能以枣瘿螨为介体在枣树间进行传播。[结论]本研究为新疆新发枣树病害鉴定了相关病原体CDMaV,完成CDMaV全基因组测序,并鉴定枣瘿螨为可能的传毒介体。鉴定病原体和传播介体是建立病害防治方法的必要基础。     

Full genome sequencing and genetic characterization of Eubenangee viruses identify Pata virus as a distinct species within the genus Orbivirus

Eubenangee virus has previously been identified as the cause of Tammar sudden death syndrome (TSDS). Eubenangee virus (EUBV), Tilligery virus (TILV), Pata virus (PATAV) and Ngoupe virus (NGOV) are currently all classified within the Eubenangee virus species of the genus Orbivirus, family Reoviridae. Full genome sequencing confirmed that EUBV and TILV (both of which are from Australia) show high levels of aa sequence identity (>92%) in the conserved polymerase VP1(Pol), sub-core VP3(T2) and outer core VP7(T13) proteins, and are therefore appropriately classified within the same virus species. However, they show much lower amino acid (aa) identity levels in their larger outer-capsid protein VP2 (<53%), consistent with membership of two different serotypes - EUBV-1 and EUBV-2 (respectively). In contrast PATAV showed significantly lower levels of aa sequence identity with either EUBV or TILV (with <71% in VP1(Pol) and VP3(T2), and <57% aa identity in VP7(T13)) consistent with membership of a distinct virus species. A proposal has therefore been sent to the Reoviridae Study Group of ICTV to recognise 'Pata virus' as a new Orbivirus species, with the PATAV isolate as serotype 1 (PATAV-1). Amongst the other orbiviruses, PATAV shows closest relationships to Epizootic Haemorrhagic Disease virus (EHDV), with 80.7%, 72.4% and 66.9% aa identity in VP3(T2), VP1(Pol), and VP7(T13) respectively. Although Ngoupe virus was not available for these studies, like PATAV it was isolated in Central Africa, and therefore seems likely to also belong to the new species, possibly as a distinct 'type'. The data presented will facilitate diagnostic assay design and the identification of additional isolates of these viruses.     

Organization of a family of highly repetitive sequences within the human genome

Recombinant clones containing the highly repetitive human DNA sequence approximately 340 base-pairs in length obtained after EcoRI digestion (αRI-DNA) were cloned in plasmid pAT153. Two clones contained a single copy of the αRI-DNA sequence, and the third had an insert with two copies of the sequence in tandem. When radioactive recombinant DNA was hybridized to total human DNA partially digested with EcoRI, a series of multiple bands was obtained up to 22 repeats in length, demonstrating that the αRI-DNA sequences occur in tandem arrays in the genomic DNA. A reassociation analysis using isolated insert DNA from one of the recombinant clones showed that the family of sequences is repeated 22,000 times in the human genome. Clones containing the αRI-DNA sequence were also isolated from a library of human genomic DNA in bacteriophage λ. Using these clones it was shown that, in at least some cases, the repetitive element is bounded by DNA less abundant than the αRI sequence.     

Paraulopidae and Paraulopus, a new family and genus of aulopiform fishes with revised relationships within the order

A species group hitherto concealed within Chlorophthalmus (Chlorophthalmidae) is described as a new family and genus of Aulopiformes, Paraulopidae and Paraulopus, respectively. Paraulopus clearly belongs in Aulopiformes owing to an enlarged uncinate process on the second epibranchial, the absence of a swimbladder, and fusion of the medial processes of the pelvic girdle, but characterized by having the following combination of characters: the fourth basibranchial with a long tail but no gap separating fourth basibranchial and fifth ceratobranchial, epipleural bones distributed from posterior portion of abdominal vertebra to anterior portion of caudal vertebra, and flesh specimens of most species with paired olive spots dorsally on body. The phylogenetic position of Paraulopus is defined by a cladistic analysis of 101 morphological characters, in 21 genera of Aulopiformes. In a single most parsimonious tree, Paraulopus and Chlorophthalmus are in two different major clades, there being four major clades in all, roughly corresponding to the four suborders of Aulopiformes. Paraulopidae is included in the suborder Synodontoidei. In addition, Bathysauroides is transferred to Chlorophthalmoidei from Giganturoidei, and Bathysauroides and Bathysauropsis are elevated to familial status. Received: January 13, 2001 / Revised: September 27, 2001 / Accepted: October 15, 2001     

The relationship of the genus Schizopera Sars within the family Diosaccidae (Copepoda: Harpacticoida)

A review of Schizopera Sars indicates that four species are so much more primitive in the antenna and leg setation that they should be removed to a new genus, Eoschizopera, which can be considered as a direct and immediate ancestor of Schizopera. A further, new, species is described in this new genus. The relationships within this branch of the family Diosaccidae are discussed and the scheme of family evolution proposed by Lang is modified to include Eoschizopera and other genera not considered by him (Goffinella, Protopsammotopa, Psammotopa, Actopsyllus, Balucopsylla, Schizoperoides). Actopsyllus hartmannorum Kunz is removed to a further new genus- Helmutkunzia.     

A new genus and two new species of Pennellidae (Copepoda: Siphonostomatoida) and an analysis of evolution within the family

Summary A new pennellid genus,Exopenna, is established on the basis of two specimens of a new species,E. crimmeni, recovered from a deep-sea cod,Antimora, caught in the North East Atlantic. It is ectoparasitic and is diagnosed by the combination of a straight body, tightly coiled egg sacs and flattened, plate-like antennary processes. Another new species,Peniculus elongatus, is described from Australian waters and new records ofP. fistula andLernaeenicus ramosus further increase the known pennellid fauna of that region. A preliminary analysis of the phylogenetic relationships of the pennellid genera is undertaken and a partially resolved cladogram produced which identifies the main generic groupings. ac]19850522     

The crystal structure of cricket paralysis virus: the first view of a new virus family.

Numerous small, RNA-containing insect viruses are currently classified as picornaviruses, or as 'picorna-like', since they superficially resemble the true picornaviruses. Considerable evidence now suggests that several of these viruses are members of a distinct family. We have determined the gene sequence of the capsid proteins and the 2.4 A resolution crystal structure of the cricket paralysis virus. While the genome sequence indicates that the insect picorna-like viruses represent a distinct lineage compared to true picornaviruses, the capsid structure demonstrates that the two groups are related. These viral genomes are, thus, best viewed as composed of exchangeable modules that have recombined.     

The concept of genus within the family Phytoseiidae (Acari: Parasitiformes): historical review and phylogenetic analyses of the genus Neoseiulus Hughes

Systematic studies on the family Phytoseiidae were first conducted at the beginning of the 20^th century but increased greatly after the Second World War. Various classifications have been proposed based on different characters such as: dorsal, ventral, and leg chaetotaxy; the shape of ventrianal and sternal shields; the shape of the insemination apparatus (spermatheca) and spermatodactylus; the number of teeth on the movable digit of chelicera; and dorsal and ventral adenotaxy. The genus concepts developed over the last five decades can be divided into two main categories or hypotheses. The first, supported mainly by Chant and McMurtry, focuses on dorsal and ventral chaetotaxy, and the genera so defined usually include a great number of species. The second category, proposed by Athias‐Henriot, considers the shape of the insemination apparatus as the key character, and the genera so defined usually include a limited number of species. From a diagnostic point of view, both classifications have a valid structure, but the question investigated herein was: which of the two classifications or hypotheses fits phylogenetic evolution? To answer this, we conducted molecular phylogenetic analyses (using the genes ITS and 12S rRNA) on the genus Neoseiulus, which has been subjected to classification based on the two main genus concepts. The results showed that the first hypothesis (Chant and McMurtry) leads to polyphyly of the genus Neoseiulus, while the second (Athias‐Henriot) leads to paraphyly of the genus. The results show that acarologists who first decided that the insemination apparatus was of evolutive importance could be correct as the shape of the insemination apparatus seems to better fit evolutive clades than dorsal and ventral chaetotaxy. The morphology of this organ, however, must be more accurately studied to better define homologies. The present paper investigates the two main hypotheses proposed until now for classification of Phytoseiidae and thereby opens the way for improved classification. © 2012 The Linnean Society of London, Zoological Journal of the Linnean Society, 2012, 165 , 253–273.     

Testing genome skimming for species discrimination in the large and taxonomically difficult genus Rhododendron

Standard plant DNA barcodes based on 2–3 plastid regions, and nrDNA ITS show variable levels of resolution, and fail to discriminate among species in many plant groups. Genome skimming to recover complete plastid genome sequences and nrDNA arrays has been proposed as a solution to address these resolution limitations. However, few studies have empirically tested what gains are achieved in practice. Of particular interest is whether adding substantially more plastid and nrDNA characters will lead to an increase in discriminatory power, or whether the resolution limitations of standard plant barcodes are fundamentally due to plastid genomes and nrDNA not tracking species boundaries. To address this, we used genome skimming to recover near-complete plastid genomes and nuclear ribosomal DNA from Rhododendron species and compared discrimination success with standard plant barcodes. We sampled 218 individuals representing 145 species of this species-rich and taxonomically difficult genus, focusing on the global biodiversity hotspots of the Himalaya-Hengduan Mountains. Only 33% of species were distinguished using ITS+matK+rbcL+trnH-psbA. In contrast, 55% of species were distinguished using plastid genome and nrDNA sequences. The vast majority of this increase is due to the additional plastid characters. Thus, despite previous studies showing an asymptote in discrimination success beyond 3–4 plastid regions, these results show that a demonstrable increase in discriminatory power is possible with extensive plastid genome data. However, despite these gains, many species remain unresolved, and these results also reinforce the need to access multiple unlinked nuclear loci to obtain transformative gains in species discrimination in plants.     

Genomic and proteomic characterization of the broad-host-range Salmonella phage PVP-SE1: creation of a new phage genus

(Bacterio)phage PVP-SE1, isolated from a German wastewater plant, presents a high potential value as a biocontrol agent and as a diagnostic tool, even compared to the well-studied typing phage Felix 01, due to its broad lytic spectrum against different Salmonella strains. Sequence analysis of its genome (145,964 bp) shows it to be terminally redundant and circularly permuted. Its G+C content, 45.6 mol%, is lower than that of its hosts (50 to 54 mol%). We found a total of 244 open reading frames (ORFs), representing 91.6% of the coding capacity of the genome. Approximately 46% of encoded proteins are unique to this phage, and 22.1% of the proteins could be functionally assigned. This myovirus encodes a large number of tRNAs (n=24), reflecting its lytic capacity and evolution through different hosts. Tandem mass spectrometric analysis using electron spray ionization revealed 25 structural proteins as part of the mature phage particle. The genome sequence was found to share homology with 140 proteins of the Escherichia coli bacteriophage rV5. Both phages are unrelated to any other known virus, which suggests that an "rV5-like virus" genus should be created within the Myoviridae to contain these two phages.     

The mitochondrial genome is large and variable in a family of plants (cucurbitaceae)

The genome sizes of mitochondrial DNA from darkgrown (etiolated) shoots of several higher plants were determined by reassociation kinetics and restriction analysis. Kinetic complexities obtained from reassociation kinetics measured spectrophotometrically indicate a mitochondrial genome size of 1600 Md for muskmelon, 1000 Md for cucumber, 560 Md for zucchini squash and 220 Md for watermelon (four species in the cucurbit family), as well as 240 Md for pea and 320 Md for corn. The kinetic curves also reveal the presence (except in corn) of sequences of a few magadaltons of complexity, reiterated about 10-50 times and representing 5%- 10% of the DNA in each mitochondrial genome. Molecular weight summation of fragments resulting from digestion with restriction endonucleases Sal I and Kpn I give genome size estimates similar to those obtained from reassociation kinetics, except for muskmelon and cucumber, for which the large number of fragments of similar size limits our estimate to at least 500 Md. The number of mitochondrial genomes per diploid cell is estimated to be about 110 to 140 for muskmelon, zucchini and watermelon. We consider the possible evolutionary mechanisms by which the mitochondrial genome has grown within the cucurbit family and the possible reasons for the existance of a seven to eight-fold range in mitochondrial genome size among such closely related species.     

The characterization of complete mitochondrial genome and phylogenetic relationship within Rattus genus (Rodentia: Muridae)

The genus Rattus is one of the main pest genus of rodent. Most species of the genus carry all kinds of pathogenic bacteria to human being. They are traditionally considered to be a least understood group. The complete mitochondrial genome of the White-Footed Indochinese Rat, Rattus nitidus was determined in this study. The characterization of mitochondrial genomes of Rattus genus was also analyzed based on comprehensive comparison. The result of evolutionary patterns of protein-coding genes (PCGs) suggested purifying selection was the predominant evolutionary forces in the mitochondrial genomes of Rattus genus. The NADH dehydrogenase 4 gene (ND4) showed a highly elevated Ka/Ks ratio compared to the other protein-coding genes, which indicated ND4 was most likely under relaxed selection pressure. Phylogenetic analysis provided a well-supported outline of Rattus genus, and revealed two groups in the genus. R. nitidus had a sister relationship with R. norvegicus.     

Sequence variation within a nonstructural region of the hepatitis G virus genome.

Nine sets of nested PCR primers from a 2.6-kb region of the hepatitis G virus (HGV) genome at nucleotide positions 5829 to 8421 were designed and used to analyze serum specimens obtained from patients with community-acquired non-A, non-B hepatitis who were HGV RNA positive. One set of primers was found to be most efficient in detecting HGV and was subsequently used to test 162 HCV-positive and 11 HCV-negative plasma units obtained from individual paid donors. HGV RNA was detected in 30 (17.3%) plasma units, 2 of which were found among the 11 HCV-negative specimens. A complete set of nine PCR fragments was obtained from two patients with community-acquired acute non-A, non-B hepatitis and from four paid donors. All PCR fragments were sequenced and were shown to have a nucleotide similarity of 85.9 to 92.3% and a derived amino acid similarity of 96.0 to 99.0%. The majority of nucleotide changes occurred in the third position of codons. The HGV nucleotide and protein sequences obtained in this study were compared with HCV sequences. Based on this analysis the 2.6-kb fragment was predicted to encode the C-terminal part of the putative NS4b, the entire NS5a, and almost the complete NS5b proteins. Putative protease cleavage sites separating these proteins were also predicted. In serial specimens obtained from the two HGV-infected patients, no significant variations were found in the HGV nucleotide and derived amino acid sequences over time. The HGV sequences obtained from one patient showed no changes over 6 months, whereas more than 99.0% homology was observed for sequences from the second patient over 2.5 years. Heterogeneity analysis performed on 10 sequences obtained in this study and corresponding regions from 6 known full-size sequences of the HGV genomes demonstrated notable discrete heterogeneity consistent with the existence of HGV genetic groups or types.