Search

Citrus yellow mottle-associated virus (CiYMaV) belonging to the subgenus Mandarivirus within the genus Potexvirus, was first identified in 2018 from Pakistan (CiYMaV-PK), where it is endemic in several regions. Here, three full-length cDNA clones (pCiYMaV-FL-1, pCiYMaV-FL-18, and pCiYMaV-FL-22) corresponding to the genomic RNA of CiYMaV were constructed and then agroinfiltrated on Chandler pummelo (Citrus grandis) seedlings using the vacuum infiltration method. All the inoculated plants developed severe vein yellowing, leaf mottling, and dwarfing symptoms by 40 days post-infiltration (dpi). The results of a direct tissue blot immunoassay and reverse transcription polymerase chain reaction detection showed 94.7–100% infection rates of pCiYMaV-FL at 60 dpi. Despite there being no observed difference among the three clones in the severity of symptom, pCiYMaV-FL-22 showed the highest accumulation levels of viral RNA and coat proteins. Moreover, pCiYMaV-FL-22 successfully infected seven other citrus varieties and induced symptoms in five of them. Transmission electron microscopy identified the presence of filamentous virus particles in extracts from systemic leaves of the plants infected with pCiYMaV-FL-22 at 6-months post-infiltration. Taken together, the results indicate that Koch's postulates were fulfilled for CiYMaV in citrus plants. This is the first report of full-length infectious cDNA clones of CiYMaV, and thus, the data provide a basis for further study of the molecular mechanisms of virus infection and the development of a viral vector to express foreign genes in citrus plants.

SARS-CoV-2 is an enveloped virus responsible for the Coronavirus Disease 2019 (COVID-19) pandemic. Here, single viruses were analyzed by atomic force microscopy (AFM) operating directly in a level 3 biosafety (BSL3) facility, which appeared as a fast and powerful method to assess at the nanoscale level and in 3D infectious virus morphology in its native conformation, or upon inactivation treatments. AFM imaging reveals structurally intact infectious and inactivated SARS-CoV-2 upon low concentration of formaldehyde treatment. This protocol combining AFM and plaque assays allows the preparation of intact inactivated SARS-CoV-2 particles for safe use of samples out of level 3 laboratory to accelerate researches against the COVID-19 pandemic. Overall, we illustrate how adapted BSL3-AFM is a remarkable toolbox for rapid and direct virus analysis based on nanoscale morphology. [ABSTRACT FROM AUTHOR]

An all-atom theory of viral structural transitions (STs) is developed based on a multiscale analysis of the N-atom Liouville equation. The approach yields an understanding of viral STs from first principles and a calibrated interatomic force field. To carry out the multiscale analysis, we introduce slow variables characterizing the whole-virus dynamics. Use of the “nanocanonical ensemble” technique and the fundamental hypothesis of statistical mechanics (i.e., the equivalence of long-time and ensemble averages) is shown to imply a Fokker-Planck equation yielding the coarse-grained evolution of the slow variables. As viral STs occur on long time scales, transition state theory is used to estimate the energy barrier of transition between free energy wells implied by observed hysteresis in viral STs. Its application to Nudaurelia capensis ω virus provides an upper bound on the free energy barrier when a single dilatational order parameter is used. The long time scale of viral STs is shown to follow from the aggregate effect of inertia, energy barrier, and entropic effects. Our formulation can be generalized for multiple order parameter models to account for lower free energy barrier pathways for transition. The theory with its all-atom description can be applied to nonviral nanoparticles as well. [ABSTRACT FROM AUTHOR]

In this paper, the mechanisms of overcharging of a colloidal macroion in the presence of multivalent counterions are investigated by means of Monte Carlo simulations. This computational technique appears as a powerful tool for probing the validity of semianalytical models developed for this issue. In particular, the simulations performed are compared with the predictions of two different models based on the one component plasma (OCP) theory. Therein, the multivalent ionic atmosphere confined at the macroion surface is approximated by a two-dimensional Wigner crystal. These kinds of models are largely used in the literature since (in some cases) they present quite simple equations to describe the electric double layer (EDL) of macroions with different geometries in the presence of much smaller (but still multivalent) ions. In this sense, charge inversion phenomena of membranes, polyelectrolytes, DNA molecules, etc., are straightforwardly predicted in terms of these expressions. Unfortunately, comparisons between these predictions and experimental results are scarce, mostly due to the difficulty to reproduce the experimental conditions in the laboratory. Accordingly, the goal of the present paper is to simulate EDLs under real conditions (in which overcharging phenomena are expected to happen) and use the results obtained in this way for comparing with those obtained from OCP models. [ABSTRACT FROM AUTHOR]

Recent discoveries of new large DNA viruses reveal high diversity in their morphologies, genetic repertoires, and replication strategies. Here, we report the novel features of Medusavirus, a large DNA virus newly isolated from hot spring water in Japan. Medusavirus with a diameter of 260 nm shows a T=277 icosahedral capsid with unique spherical-headed spikes on its surface. It has a 381 kb genome encoding 461 putative proteins, 86 of which have their closest homologs in Acanthamoeba castellanii, whereas 279 (61%) are ORFans. The virus lacking the genes of DNA topoisomerase II and RNA polymerase showed that the DNA replication takes place in the host nucleus while the progeny virions are assembled in the cytoplasm. Furthermore, Medusavirus encoded all of five types of histones (H1, H2A, H2B, H3, and H4) and one DNA polymerase, which are phylogenetically placed at the root of the eukaryotic clades. By contrast, the host amoeba encoded many Medusavirus homologs including the major capsid protein. These facts strongly suggested that amoeba is indeed the most promising natural host of Medusavirus, and lateral gene transfers have taken place repeatedly and bidirectionally between the virus and its host since the early stage of their co-evolution. Medusavirus reflects the traces of direct evolutionary interactions between the virus and eukaryotic hosts, which may be caused by sharing the DNA replication compartment and evolutionarily long lasting viral-host relationships. Based on its unique morphological characteristics and phylogenomic relationships with other known large DNA viruses, we propose that Medusavirus forms a new family Medusaviridae. [ABSTRACT FROM AUTHOR]

This paper examines the vital role played by electron microscopy toward the modern definition of viruses, as formulated in the late 1950s. Before the 1930s viruses could neither be visualized by available technologies nor grown in artificial media. As such they were usually identified by their ability to cause diseases in their hosts and defined in such negative terms as "ultramicroscopic" or invisible infectious agents that could not be cultivated outside living cells. The invention of the electron microscope, with magnification and resolution powers several orders of magnitude better than that of optical instruments, opened up possibilities for biological applications. The hitherto invisible viruses lent themselves especially well to investigation with this new instrument. We first offer a historical consideration of the development of the instrument and, more significantly, advances in techniques for preparing and observing specimens that turned the electron microscope into a routine biological tool. We then describe the ways in which the electron microscopic images, or micrographs, functioned as forms of new knowledge about viruses and resulted in a paradigm shift in the very definition of these entities. Micrographs were not mere illustrations since they did the work for the electron microscopists. Drawing extensively on primary publications, we adduce the role of the new instrument in understanding the so-called eclipse phase in virus multiplication and the unexpected spinoffs of data from electron microscopy in naming and classifying viruses. Thus, we show that electron microscopy functioned not only to provide evidence, but also arguments in facilitating a reordering of the world that it brought into the visual realm. [ABSTRACT FROM AUTHOR]

Abstract Orthohantaviruses (family Hantaviridae order Bunyavirales) are emerging pathogens with a significant impact on human health. They are transmitted via aerosolized excreta of rodents which also act as reservoir hosts, constituting a unique route for dispersion. Dobrava-Belgrade and Puumala orthohantaviruses have been previously reported from Anatolia, in rodents, case reports and occasional outbreaks. We have collected rodents at several locations during a surveillance study in eastern Anatolia. The specimens were morphologically-identified and various tissues were screened via a generic orthohantavirus reverse transcription polymerase chain reaction assay. DNA barcoding via mitochondrial cytochrome b sequencing was performed in rodents with detectable orthohantavirus sequences. High throughput sequencing was performed for viral genome characterization. Fifty rodents were collected and identified morphologically as Microtus spp. (96%) and Apodemus spp. (4%). Orthohantavirus sequences were detected in lung and spleen or liver tissues of 4 voles (8%), barcoded as Microtus obscurus. The virus sequences were identified as Tula orthohantavirus (TULV) and near-complete genomic segments of the prototype viral genome, tentatively named as the Tula orthohantavirus-Turkey (TULV-T), could be characterized. Putative open reading frames for viral nucleocapsid and a nonstructural protein on the S segment, glycoproteins G1 and G2 on the M segment and viral replicase on the L segment were identified on the TULV-T. Several minor sequence variants were further characterized. No evidence of recombination could be detected and pairwise comparisons displayed over 95% amino acid sequence identities to various Eurasian TULV strains. Phylogenetic analyses revealed distinct clustering of all genome segments from previously-characterized TULV strains via various approaches and models. Here, TULV-T constituted a novel lineage, forming an intermediate among Asian and European TULV lineages. This report describes the initial documentation of TULV circulation and its potential reservoir in Anatolia. The extent of virus dispersion, alternate hosts or outcomes of human exposure require elucidation. Highlights • We describe the initial documentation of Tula orthohantavirus in Anatolia • Near-complete genome of the prototype strain is characterized • Potential reservoir voles are identified • Viral genome constitute a novel lineage, an intermediate among Asian and European strains [ABSTRACT FROM AUTHOR]

Abstract Pinworms are important parasitic nematodes in animals and humans, and many species are of medical and veterinary importance. The genus Syphabulea is a poorly known group of pinworms. The systematic position of Syphabulea in Oxyuridae remains unclear. Moreover, there is still a paucity of detailed information on some morphological aspects of the type species S. tjanschani (Ablasov, 1962). In the present study, the detailed morphology of S. tjanschani was studied using light microscopy and, for the first time, scanning electron microscopy. The results revealed three traits useful for the characterization of the species including: cuticular depression around excretory pore, non-prominent labial teeth and number of adhesive ridges in three ventral mammelons. The ribosomal [small ribosomal DNA (18S), internal transcribed spacer (ITS) and large ribosomal DNA (28S)] and mitochondrial [cytochrome c oxidase subunit 1 (cox 1)] target regions of S. tjanschani were also firstly amplified and sequenced for future use in the molecular identification of this poorly known species. In addition, in order to investigate the systematic position of Syphabulea in Oxyuridae, the phylogenetic analyses of representatives of the Oxyuridae were performed using maximum likelihood (ML) and maximum parsimony (MP) methods based on 18S, 28S and 18S + 28S + cox 1 sequence data, respectively. The phylogenetic results based on different sequence data all supported the genus Syphabulea to be a member of the subfamily Syphaciinae. The phylogenetic analysis based on 28S sequence data also showed Syphabulea as sister to the genus Syphatineria. Our present study represents the first attempt to resolve the systematic position of the genus Syphabulea using phylogenetic analyses based on molecular data. Highlights • S. tjanschani was redescribed based on newly material. • SEM observations revealed some previously unreported morphological features. • Genetic characterization of S. tjanschani was firstly reported. • Phylogenetic results supported Syphabulea to be a member of Syphariinae. [ABSTRACT FROM AUTHOR]

The spindle-shaped virion morphology is common among archaeal viruses, where it is a defining characteristic of many viral families. However, structural heterogeneity intrinsic to spindle-shaped viruses has seriously hindered efforts to elucidate the molecular architecture of these lemon-shaped capsids. We have utilized a combination of cryo-electron microscopy and X-ray crystallography to study Acidianus tailed spindle virus (ATSV). These studies reveal the architectural principles that underlie assembly of a spindleshaped virus. Cryo-electron tomography shows a smooth transition from the spindle-shaped capsid into the tubular-shaped tail and allows low-resolution structural modeling of individual virions. Remarkably, higher-dose 2D micrographs reveal a helical surface lattice in the spindle-shaped capsid. Consistent with this, crystallographic studies of themajor capsid protein reveal a decorated four-helix bundle that packs within the crystal to form a four-start helical assembly with structural similarity to the tube-shaped tail structure of ATSV and other tailed, spindle-shaped viruses. Combined, this suggests that the spindle-shaped morphology of the ATSV capsid is formed by a multistart helical assembly with a smoothly varying radius and allows construction of a pseudoatomic model for the lemon-shaped capsid that extends into a tubular tail. The potential advantages that this novel architecture conveys to the life cycle of spindle-shaped viruses, including a role in DNA ejection, are discussed. [ABSTRACT FROM AUTHOR]

Summary: Background: Cutaneous warts have a cure rate after therapy of no more than approximately 50%. Recently, we developed and validated a standard assessment tool for warts (Cutaneous WARTS diagnostic tool, CWARTS) based on phenotypical characteristics. Objectives: To assess whether patient and morphological wart characteristics predict the human papillomavirus (HPV) type in a specific wart and whether these characteristics as well as the HPV type predict a favourable treatment response. Methods: Photographs were used to score nine morphological wart characteristics using the newly developed CWARTS tool. Genotyping of 23 wart‐associated HPV types was performed using the hyperkeratotic skin lesion–polymerase chain reaction/multiplex genotyping assay. The results were correlated with a favourable response to treatment with monochloroacetic acid, cryotherapy or a combination of cryotherapy and salicylic acid. Odds ratios were calculated using logistic regression in a generalized estimating equations model. Results: Black dots (capillary thrombosis) strongly predicted the presence of any HPV type in a wart. From all characteristics tested, the HPV type most strongly predicted the treatment response when the warts were treated with monochloroacetic acid or a combination of cryotherapy and salicylic acid with a significantly decreased treatment response if the warts contained HPVs of the alpha genus (HPV2, HPV27 or HPV57). When cryotherapy alone was used for common warts, HPV type did not play a role, but cryotherapy was less effective in the presence of callus and when the wart was located deeper in the skin. Conclusions: Morphological characteristics of the warts and the HPV genotype influence treatment outcome and thus potentially influence future treatment decisions for common and plantar warts. [ABSTRACT FROM AUTHOR]

We propose that viruses with geometric defects are not necessarily flawed viruses. A geometric defect may be a reactive site. Defects may facilitate assembly, dissociation, or accessibility of cellular proteins to virion components. In single molecule studies of hepadnavirus assembly, defects and overgrowth are common features. Icosahedral alphaviruses and flaviviruses, among others, have capsids with geometric defects. Similarly, immature retroviruses, which are non-icosahedral, have numerous "errors". In many viruses, asymmetric exposure of interior features allows for regulated genome release or supports intracellular trafficking. In these viruses, the defects likely serve a biological function. Commonly used approaches for spherical virus structure determination use symmetry averaging, which obscures defects. We suggest that there are three classes of asymmetry: regular asymmetry as might be found in a tailed phage, irregular asymmetry as found, for example, in defects randomly trapped during assembly, and dynamic asymmetry due to Brownian dynamics of virus capsids. Awareness of their presence and recent advances in electron microscopy will allow unprecedented investigation of capsid irregularities to investigate their biological relevance. [ABSTRACT FROM AUTHOR]

The human poliovirus receptor (PVR) is a cell surface protein with a multitude of functions in human biology. PVR was initially identified as the receptor for the human poliovirus and recent discoveries have given a greater insight into both its morphology and its function. Alternative splicing of the PVR gene results in a total of 4 alternatively spliced isoforms. Two of these isoforms lack a complete transmembrane domain and are considered soluble and block viral infection; the remaining two transmembrane isoforms differ only at their extreme C-terminal domains resulting in differential localization in epithelia and polarity of viral infection. In addition to its role as a receptor for the human poliovirus, several native biological functions have also been uncovered. PVR is an important cell adhesion protein and is involved in the transendothelial migration of leukocytes. Through its interactions with CD226 and TIGIT, transmembrane proteins found on leukocytes, PVR is a key regulator of the cell-mediated immune response. As PVR is differentially regulated in a broad spectrum of cancers, it has a strong potential for clinical use as a biomarker. PVR is also a possible target for novel cancer therapies. Utilizing its natural tropism for PVR, a genetically modified form of the live attenuated poliovirus vaccine is currently being tested for its ability to locate and destroy certain tumors. These recent studies emphasize the importance of PVR in human biology and demonstrate its utility beyond being a viral receptor protein. [ABSTRACT FROM AUTHOR]

In this study, we present the characterization and genomic data of three Achromobacter phages belonging to the family Siphoviridae. Phages 83-24, JWX and JWF were isolated from sewage samples in Paris and Braunschweig, respectively, and infect Achromobacter xylosoxidans, an emerging nosocomial pathogen in cystic fibrosis patients. Analysis of morphology and growth parameters revealed that phages 83-24 and JWX have similar properties, both have nearly the same head and tail measurements, and both have a burst size between 85 and 100 pfu/cell. In regard to morphological properties, JWF had a much longer and more flexible tail compared to other phages. The linear double-stranded DNAs of all three phages are terminally redundant and not circularly permutated. The complete nucleotide sequences consist of 81,541 bp for JWF, 49,714 bp for JWX and 48,216 bp for 83-24. Analysis of the genome sequences showed again that phages JWX and 83-24 are quite similar. Comparison to the GenBank database via BLASTN revealed partial similarities to Roseobacter phage RDJL phi1 and Burkholderia phage BcepGomr. In contrast, BLASTN analysis of the genome sequence of phage JWF revealed only few similarities to non-annotated prophage regions in different strains of Burkholderia and Mesorhizobium. [ABSTRACT FROM AUTHOR]

The capsid domain (CA) of the retroviral Gag protein is a primary determinant of Gag oligomerization, which is a critical step for immature Gag lattice formation and virus particle budding. Although the human immunodeficiency virus type 1 (HIV-1) CA carboxy-terminal domain (CTD) is essential for CA-CA interactions, the CA CTD has been suggested to be largely dispensable for human T-cell leukemia virus type 1 (HTLV-1) particle biogenesis. To more clearly define the roles of the HTLV-1 CA amino-terminal domain (NTD) and CA CTD in particle biogenesis, we generated and analyzed a panel of Gag proteins with chimeric HIV-1/HTLV-1 CA domains. Subcellular distribution and protein expression levels indicated that Gag proteins with a chimeric HIV-1 CA NTD/HTLV-1 CA CTD did not result in Gag oligomerization regardless of the parent Gag background. Furthermore, chimeric Gag proteins with the HTLV-1 CA NTD produced particles phenotypically similar to HTLV-1 immature particles, highlighting the importance of the HTLV-1 CA NTD in HTLV-1 immature particle morphology. Taken together, these observations support the conclusion that the HTLV-1 CA NTD can functionally replace the HIV-1 CA CTD, but the HIV-1 CA NTD cannot replace the HTLV-1 CA CTD, indicating that the HTLV-1 CA subdomains provide distinct contributions to Gag-Gag oligomerization, particle morphology, and biogenesis. Furthermore, we have shown for the first time that HIV-1 and HTLV-1 Gag domains outside the CA (e.g., matrix and nucleocapsid) impact Gag oligomerization as well as immature particle size and morphology. [ABSTRACT FROM AUTHOR]

The critical roles of viruses in nutrient cycling are well established for marine ecosystems, but less is known for freshwater viruses about diversity, seasonality, and relationships with productivity and ecosystem trophic status. This study aimed to examine virus morphotypes and abundance in relation to prokaryotic cell abundance and chlorophyll a concentrations in three seasonal temperate freshwater sites representing eutrophic, mesotrophic, and oligotrophic status in Lake Michigan watershed. We found strong correlations between virus-like particle abundance (VLPA) and prokaryote abundance (PA) and chlorophyll concentration across the trophic gradient ( r = 0.744, P < 0.001, n = 123), but without consistent seasonal patterns. TEM showed Myoviridae, Podoviridae, Siphoviridae, and non-tailed virus morphotypes. The eutrophic site was distinct with one- to two-fold higher PA and VLPA than mesotrophic and oligotrophic sites. But the eutrophic and mesotrophic sites had more similar VLPA:PA ratios (eutrophic 26, mesotrophic 25.5, oligotrophic 4.1). The oligotrophic site also had a lower proportion of Podoviridae and more Siphoviridae than the other sites. The similarity between eutrophic and mesotrophic sites, but a distinctive oligotrophic site, suggests that the effects of terrestrial runoff as sources of viruses and hosts may be important, along with P limitation and trophic status, in determining virus-host relationships in freshwaters. [ABSTRACT FROM AUTHOR]

The standard virus classification scheme for arenaviruses and bunyaviruses shifted dramatically when several groups reported the detection and isolation of divergent groups of viruses in a variety of insect collections. Although these viral families can differ in terms of morphology, structure and genetics, recent findings indicate these viruses may have a shared evolutionary origin. To determine the phylogenetic relations among these families, we inferred phylogenetic trees using three methods. The Maximum Likelihood and Bayesian trees were rooted as suggested by the (molecular clock-rooted) BEAST tree. Our results highlight a noteworthy relation among these viral supergroups of different genome organizations. Our study suggests that the best scenario is the existence of at least three monophyletic supergroups, all of them well supported. The recent data indicate that these viruses are evolutionarily and genetically interconnected. While these supergroups appear to be closely related in our phylogenetic analysis, other viruses should be investigated in future research. In sum, our results also provide insights into the classification scheme, thereby providing a new perspective about the fundamental questions of family origins, diversity and genome evolution. [ABSTRACT FROM AUTHOR]

Light microscopy is a straightforward and highly portable imaging approach that is used for the detection of parasites, fungi, and bacteria. The detection of individual virus particles has historically not been possible through this approach. Thus, characterization of virus particles is typically performed using high-energy approaches such as electron microscopy. These approaches require purification of virions away from its normal milieu, significant levels of expertise, and only count a small number of particles at a time. To correct these deficiencies we created a platform that allows label-free, point-of-need virus imaging and counting. We adapted a multiplex-capable, interferometric imaging technique to a closed-system that allows real-time particle detection in complex mixtures. To maximize virus particle binding we constructed a disposable device with a constant flow rate of ∼3 μl min−1. Biosafety was achieved by having a sealable sample addition port. Using this platform we were able to readily identify virus binding in a 20 minute experiment. Sensitivity was comparable to laboratory-based assays such as ELISA and plaque assay, and showed equal or better sensitivity against paper-based assays designed for point-of-need use. Our results demonstrate a platform that can be used for rapid multiplexed detection and visualization of whole virus particles. We envision this technology as a sample-to-answer platform for detection and visualization of viruses without the need for prior labeling. This would enable both research investigation of virus particle behavior and morphology and have the potential to be used in a diagnostic context, where direct imaging from samples such as blood and urine would be valuable. [ABSTRACT FROM AUTHOR]

Mud Crab Reovirus (MCRV) is a major pathogen of cultured mud crab Scylla paramamosain. Early studies showed that the mortality rate caused by MCRV was high. However, current studies show that MCRV has been widely present in cultured mud crab ponds, and the mortality rate has dramatically decreased, suggesting that it has become more infectious but less virulent. In this study, an epidemic MCRV strain named MCRV-NH was isolated from a representative farming areas of mud crabs to reveal the intrinsic molecular mechanism of the virulence variation of MCRV. Electron microscopy results showed that MCRV-NH was similar to the early reported MCRV or Scylla serrata reovirus (Ss RV) in terms of morphology and structure. Viral infection experiments showed that no mud crab was dead during the infection period of 11 days even though MCRV-NH could rapidly proliferate in mud crabs at 21–23 °C, demonstrating that the mortality rate caused by MCRV-NH significantly decreased. Sequencing results of the MCRV-NH genome showed that eleven nucleotides were deleted in the sequencing region, and its nucleotide sequence identities with those of MCRV and Ss RV were 99.43% and 99.33%, and the numbers of corresponding nucleotide mutations were 150 and 126, respectively. Evolutionary analysis showed that MCRV-NH belonged to the same species as MCRV and Ss RV, and MCRV-NH should be a variant. In comparison with the open reading frames (ORFs) of MCRV and Ss RV, the ORFs of MCRV-NH had no nucleotide deletions, but the numbers of nucleotide mutations were 100 and 123, and the numbers of amino acid mutations were 32 and 35, respectively. Eight mutated amino acids compared with the two reoviruses were located at the predicted active sites of the viral proteins. Among them, three mutations resulted in the generation of new phosphorylation sites, and two mutations led to the loss of the original phosphorylation sites. Meanwhile, a heptapeptide repeat sequence in VP9 disappeared, and a streptococcal histidine triad protein in VP6 and a Big-1 (bacterial Ig-like domain 1) domain in VP11 were generated due to key amino acid mutations in these viral proteins, which are involved in the infection and replication of MCRV. These changes in the active sites of the viral proteins may be responsible for the MCRV virulence variation. This study provided essential evidence to support the conclusion that MCRV-NH was a variant of MCRV with weaker pathogenicity and revealed the underlying possible molecular mechanism of the virulence variation of MCRV-NH. • MCRV-NH was isolated from a representative farming areas of mud crabs. • MCRV-NH genome showed >99% identities with those of MCRV and Ss RV. • MCRV-NH was a variant of MCRV with weak pathogenicity. • Key amino acid mutations may be responsible for less virulence of MCRV-NH. [ABSTRACT FROM AUTHOR]

Electron cryo-tomography (cryoET) is currently the only technique that allows the direct observation of proteins in their native cellular environment. Sub-volume averaging of electron tomograms offers a route to increase the signal-to-noise of repetitive biological structures, such improving the information content and interpretability of tomograms. We discuss the potential for sub-volume averaging in highlighting and investigating specific processes in situ , focusing on microtubule structure and viral infection. We show that (i) in situ sub-volume averaging from single tomograms can guide and complement segmentation of biological features, (ii) the in situ determination of the structure of individual viruses is possible as they infect a cell, and (iii) novel, transient processes can be imaged with high levels of detail. [ABSTRACT FROM AUTHOR]

A hemagglutinating virus isolate designated 11OG0352, was obtained from a duck fecal sample. Genetic and virological analyses indicated that it might represent a novel serotype of avian paramyxovirus (APMV). Electron micrographs showed that the morphology of the virus particle was similar to that of APMV. The complete genome of this virus comprised 15,444 nucleotides complying with the paramyxovirus “rule of six” and contains six open reading frames (3′-N-P-M-F-HN-L-5′). The phylogenetic analysis of the whole genome revealed that the virus was a member of the genus Avulavirus , but that it was distinct from APMV-1 to APMV-13. Although the F-protein cleavage site was TREG K ↓L, which resembles a lentogenic strain of APMV-1, the K residue at position -1 of the cleavage site was first discovered in APMV members. The phosphoprotein gene of isolate 11OG0352 contains a putative RNA editing site, 3′-AUUUUCCC-5′ (negative sense) which sequence differs from that of other APMVs. The intracerebral pathogenicity index test did not detect virulence in infected chicks. In hemagglutination inhibition (HI) tests, an antiserum against this virus did not detectably react with other APMVs (serotypes 1–4, 6–9) except for low reciprocal cross-reactivity with APMV-6. We designated this isolate, as APMV-14/duck/Japan/11OG0352/2011 and propose that it is a novel APMV serotype. The HI test may not be widely applicable for the classification of a new serotype because of the limited availability of reference antisera against all serotypes and cross-reactivity data. The nucleotide sequence identities of the whole genome of 11OG0352 and other APMVs ranged from 46.3% to 56.1%. Such comparison may provide a useful tool for classifying new APMV isolates. However, the nucleotide sequence identity between APMV-12 and APMV-13 was higher (64%), which was nearly identical to the lowest nucleotide identity (67%) reported in subgroups within the serotype. Therefore, consensus criteria for using whole genome analysis should be established. [ABSTRACT FROM AUTHOR]

Metavisitor is a software package that allows biologists and clinicians without specialized bioinformatics expertise to detect and assemble viral genomes from deep sequence datasets. The package is composed of a set of modular bioinformatic tools and workflows that are implemented in the Galaxy framework. Using the graphical Galaxy workflow editor, users with minimal computational skills can use existing Metavisitor workflows or adapt them to suit specific needs by adding or modifying analysis modules. Metavisitor works with DNA, RNA or small RNA sequencing data over a range of read lengths and can use a combination of de novo and guided approaches to assemble genomes from sequencing reads. We show that the software has the potential for quick diagnosis as well as discovery of viruses from a vast array of organisms. Importantly, we provide here executable Metavisitor use cases, which increase the accessibility and transparency of the software, ultimately enabling biologists or clinicians to focus on biological or medical questions. [ABSTRACT FROM AUTHOR]

The capacity to transition between distinct morphological forms is a key virulence trait for diverse fungal pathogens. A poignant example of a leading opportunistic fungal pathogen of humans for which an environmentally responsive developmental program underpins virulence is Candida albicans. C. albicans mutants that are defective in the transition between yeast and filamentous forms typically have reduced virulence. Although many positive regulators of C. albicans filamentation have been defined, there are fewer negative regulators that have been implicated in repression of filamentation in the absence of inducing cues. To discover novel negative regulators of filamentation, we screened a collection of 1,248 C. albicans homozygous transposon insertion mutants to identify those that were filamentous in the absence of inducing cues. We identified the Rho1 GAP Lrg1, which represses filamentous growth by stimulating Rho1 GTPase activity and converting Rho1 to its inactive, GDP-bound form. Deletion of LRG1 or introduction of a RHO1 mutation that locks Rho1 in constitutively active, GTP-bound state, leads to filamentation in the absence of inducing cues. Deletion of the Rho1 downstream effector PKC1 results in defective filamentation in response to diverse host-relevant inducing cues, including serum. We further established that Pkc1 is not required to sense filament-inducing cues, but its kinase activity is critical for the initiation of filamentous growth. Our genetic analyses revealed that Pkc1 regulates filamentation independent of the canonical MAP kinase cascade. Further, although Ras1 activation is not impaired in a pkc1Δ/pkc1Δ mutant, adenylyl cyclase activity is reduced, consistent with a model in which Pkc1 functions in parallel with Ras1 in regulating Cyr1 activation. Thus, our findings delineate a signaling pathway comprised of Lrg1, Rho1 and Pkc1 with a core role in C. albicans morphogenesis, and illuminate functional relationships that govern activation of a central transducer of signals that control environmental response and virulence programs. [ABSTRACT FROM AUTHOR]

Unlike human immunodeficiency virus type 1 (HIV-1)-infected humans, African-origin, natural simian immunodeficiency virus (SIV) hosts, such as African green monkeys (AGMs), sustain nonpathogenic SIV infections and rarely vertically transmit SIV to their infants. Interestingly, chronically SIV-infected AGMs have anatomically compartmentalized SIV variants in plasma and milk, whereas humans and SIV-infected rhesus monkeys (RMs), Asian-origin nonnatural SIV hosts, do not exhibit this compartmentalization. Thus, it is possible that AGM SIV populations in milk have unique phenotypic features that contribute to the low postnatal transmission rates observed in this natural host species. In this study, we explored this possibility by characterizing the infectivity, tropism, and neutralization susceptibility of plasma and milk SIVsab env variants isolated from chronically SIVsab92018ivTF-infected AGMs. AGM plasma and milk SIVsab env pseudovirus variants exhibited similar infectivities, neutralization susceptibilities to autologous and heterologous plasma, and chemokine coreceptor usages for cell entry, suggesting similar abilities to initiate infection in a new host. We also assessed the cytokine milieu in SIV-infected AGM milk and compared it to that of SIV-infected RMs. MIP-1β, granulocyte colony-stimulating factor (G-CSF), interleukin-12/23 (IL-12/23), and IL-13 trended significantly higher in SIV-infected AGM milk than in that of RMs, while IL-18 and IL-6 trended significantly higher in SIV-infected RM milk than in that of AGMs. Taken together, our findings imply that nonviral maternal factors, such as the cytokine milieu, rather than unique characteristics of SIV populations in the milk contribute to the low postnatal transmission rates observed in AGMs. [ABSTRACT FROM AUTHOR]

Inhalation of infected brain homogenate results in transepithelial transport of prions across the nasal mucosa of hamsters, some of which occurs rapidly in relatively large amounts between cells (A. E. Kincaid, K. F. Hudson, M. W. Richey, and J. C. Bartz, J. Virol 86:12731-12740, 2012, doi:http://dx.doi.org/10.1128/JVI.01930-12). Bulk transepithelial transport in the nasal cavity has not been studied to date. In the present study, we characterized the frequency, size, and specificity of the intercellular spaces that mediate the bulk transport of inhaled prions between cells of mice or hamsters following extranasal inoculation with mock-infected brain homogenate, different strains of prion-infected brain homogenate, or brain homogenate mixed with India ink. Infected or mock-infected inoculum was identified within lymphatic vessels of the lamina propria and in spaces of>5 μm between a small number of cells of the nasal mucosa in>90% of animals from 5 to 60 min after inhalation. The width of the spaces between cells, the amount of the inoculum within the lumen of lymphatic vessels, and the timing of the transport indicate that this type of transport was taking place through preexisting spaces in the nasal cavity that were orders of magnitude wider than what is normally reported for paracellular transport. The indiscriminate rapid bulk transport of brain homogenate in the nasal cavity results in immediate entry into nasal cavity lymphatics following inhalation. This novel mechanism may underlie the recent report of the early detection of prions in blood following inhalation and has implications for horizontal prion transmission. [ABSTRACT FROM AUTHOR]

The Gag protein is the main retroviral structural protein, and its expression alone is usually sufficient for production of viruslike particles (VLPs). In this study, we sought to investigate--in parallel comparative analyses--Gag cellular distribution, VLP size, and basic morphological features using Gag expression constructs (Gag or Gag-YFP, where YFP is yellow fluorescent protein) created from all representative retroviral genera: Alpharetrovirus, Betaretrovirus, Deltaretrovirus, Epsilonretrovirus, Gammaretrovirus, Lentivirus, and Spumavirus. We analyzed Gag cellular distribution by confocal microscopy, VLP budding by thinsection transmission electron microscopy (TEM), and general morphological features of the VLPs by cryogenic transmission electron microscopy (cryo-TEM). Punctate Gag was observed near the plasma membrane for all Gag constructs tested except for the representative Beta- and Epsilonretrovirus Gag proteins. This is the first report of Epsilonretrovirus Gag localizing to the nucleus of HeLa cells. While VLPs were not produced by the representative Beta- and Epsilonretrovirus Gag proteins, the other Gag proteins produced VLPs as confirmed by TEM, and morphological differences were observed by cryo-TEM. In particular, we observed Deltaretrovirus-like particles with flat regions of electron density that did not follow viral membrane curvature, Lentivirus- like particles with a narrow range and consistent electron density, suggesting a tightly packed Gag lattice, and Spumaviruslike particles with large envelope protein spikes and no visible electron density associated with a Gag lattice. Taken together, these parallel comparative analyses demonstrate for the first time the distinct morphological features that exist among retrovirus- like particles. Investigation of these differences will provide greater insights into the retroviral assembly pathway. [ABSTRACT FROM AUTHOR]

Okra (Abelmoschus esculentus) is an economically important plant in China which is widely used in food, medicine and manufacturing. In September 2014, diseased samples of okra were collected in Yanqing District, Beijing City, China. Symptoms were brown to brownish black, sunken cavities on the leaves and fruits. The disease occurred on okra plants with an incidence of 35–55% in different fields. According to morphological and cultural characteristics, and sequence analysis of the internal transcribed spacer (ITS) and actin (ACT) regions, the pathogen was identified asBoeremia exigua(Desm.) Aveskamp, Gruyter & Verkley. Pathogenicity of the isolated fungus was conducted on okra fruits and on potted plants. Symptoms similar to those occurring in the field were reproduced. This is the first report of Boeremia spot of okra caused byBoeremia exiguain China. [ABSTRACT FROM PUBLISHER]

A new foliar disease was observed on baby lima bean (Phaseolus lunatus) in fields across western New York State, USA. The disease occurred in 10 fields with variable incidence and severity. Symptoms were initially necrotic, tan spots on leaves with red to reddish brown irregular margins that coalesced to encompass the entire leaf and cause abscission. Pycnidia were observed within the lesions. Isolations from diseased leaves yielded several pycnidial fungi, including aDidymellaspecies. These isolates were characterized by morphology and sequencing of multiple reference genes (internal transcribed spacer (ITS), partial actin, β-tubulin (tub2), translation elongation factor 1-α (TEF), 28S rDNA large subunit (LSU),rpb2and calmodulin). A four gene phylogeny (ITS,tub2, LSU andrpb2) showed that the isolates from baby lima bean belonged to a well-supported clade that contained the type culture ofDidymella americana. Pathogenicity of the isolates on three commonly grown cultivars of baby lima bean was confirmed. Symptoms that developed on inoculated plants were similar to those observed on diseased plants in the field. This is the first report ofD. americanaon baby lima bean. [ABSTRACT FROM PUBLISHER]

Jumbo phages infecting Ralstonia solanacearum were isolated in Thailand (ϕRSL2) and Japan (ϕRSF1). They were similar regarding virion morphology, genomic arrangement, and host range. Phylogenetic and proteomic tree analyses demonstrate that the ϕRSL2 and ϕRSF1 belong to a group of evolutionary related phages, including Pseudomonas phages ϕKZ, 201ϕ2-1 and all previously described ϕKZ-related phages. Despite conserved genomic co-linearity between the ϕRSL2 and ϕRSF1, they differ in protein separation patterns. A major difference was seen in the detection of virion-associated-RNA polymerase subunits. All β- and β′-subunits were detected in ϕRSF1, but one β′-subunit was undetected in ϕRSL2. Furthermore, ϕRSF1 infected host cells faster (latent period: 60 and 150 min for ϕRSF1 and ϕRSL2, respectively) and more efficiently than ϕRSL2. Therefore, the difference in virion-associated-RNA polymerase may affect infection efficiency. Finally, we show that ϕRSF1 is able to inhibit bacterial wilt progression in tomato plants. [ABSTRACT FROM AUTHOR]

An emergent viral pathogen termed severe fever with thrombocytopenia syndrome virus (SFTSV) is responsible for thousands of clinical cases and associated fatalities in China, Japan, and South Korea. Akin to other phleboviruses, SFTSV relies on a viral glycoprotein, Gc, to catalyze the merger of endosomal host and viral membranes during cell entry. Here, we describe the postfusion structure of SFTSV Gc, revealing that the molecular transformations the phleboviral Gc undergoes upon host cell entry are conserved with otherwise unrelated alpha- and flaviviruses. By comparison of SFTSV Gc with that of the prefusion structure of the related Rift Valley fever virus, we show that these changes involve refolding of the protein into a trimeric state. Reverse genetics and rescue of site-directed histidine mutants enabled localization of histidines likely to be important for triggering this pH-dependent process. These data provide structural and functional evidence that the mechanism of phlebovirus– host cell fusion is conserved among genetically and pathophysiologically distinct viral pathogens. [ABSTRACT FROM AUTHOR]

Dengue virus (DENV) infects ∼400 million people annually, and there is no available vaccine or therapeutics. It is not clear why candidate vaccines provide only modest protection. In addition to the presence of four different dengue serotypes, there is also structural heterogeneity in DENV infectious particles, even within a strain. This severely complicates the development of vaccines and therapeutics. The currently known different morphologies of DENV are: immature, partially mature, compact mature, and expanded mature forms of the virus. In this review I describe these forms of the virus, their infectivity, and how antibodies could recognize these morphologies. I also discuss possible vaccine and antibody therapeutic formulations to protect against all morphologies. [ABSTRACT FROM AUTHOR]

The procedure for phase extension (PX) involves gradually extending the initial phases from low resolution (e.g. ∼8 Å) to the high-resolution limit of a diffraction data set. Structural redundancy present in the viral capsids that display icosahedral symmetry results in a high degree of non-crystallographic symmetry (NCS), which in turn translates into higher phasing power and is critical for improving and extending phases to higher resolution. Greater completeness of the diffraction data and determination of a molecular replacement solution, which entails accurately identifying the virus particle orientation(s) and position(s), are important for the smooth progression of the PX procedure. In addition, proper definition of a molecular mask (envelope) around the NCS-asymmetric unit has been found to be important for the success of density modification procedures, such as density averaging and solvent flattening. Regardless of the degree of NCS, the PX method appears to work well in all space groups, provided an accurate molecular mask is used along with reasonable initial phases. However, in the cases with space group P1, in addition to requiring a molecular mask, starting the PX at a higher resolution (e.g. 6 Å) overcame the previously reported problems due to Babinet phases and phase flipping errors. [ABSTRACT FROM PUBLISHER]

Atomic models of small icosahedral RNA viruses were first derived from X-ray crystal structures between 1978 and 1990. In that same period, electron cryo-microscopy with image reconstructions (cryoEM) was being developed, and viruses were among the early successful subjects of study. By 1990 it was clear that many important virus-related structures (e.g. antibodies and receptor molecules bound to viruses) would be difficult or impossible to crystalize but could be imaged by cryoEM at low to moderate resolution. Combining crystal structures of Fab fragments, receptor molecules and virus particles with cryoEM structures of the complexes allowed accurate ‘pseudo-atomic' models to be derived and resulted in a fundamental understanding of the chemistry for complex formation. Later, cryoEM produced structures of enveloped viruses that could not be crystallized, but the ecto domains of subunits forming the capsids were crystallized and fitted into the cryoEM density as rigid models. This created highly informative ‘pseudo-atomic' models of the virions. In this review we track the early development of these methods with specific examples described in more detail and also provide a more comprehensive table of references that illustrate the breadth and depth of combining X-ray crystallography and cryoEM. [ABSTRACT FROM PUBLISHER]

In many plant and bacterial viruses, a filamentous nucleocapsid makes up the entire virus particle. These viruses usually form fibres naturally, but they do not crystallize. Fibre diffraction has therefore been the method of choice for structural studies, aided in recent years by cryo-electron microscopy (EM) and solid state nuclear magnetic resonance (ssNMR). Filamentous viruses, particularly tobacco mosaic virus (TMV), have been important in developing fibre diffraction methods, and fibre diffraction allowed TMV to be among the first virus structures determined. Structures of several viruses related to TMV and several filamentous bacterial viruses have been determined at resolutions of 3 Å or better, and lower resolution structures have been determined by fibre diffraction, sometimes in conjunction with other methods, for many other, unrelated, filamentous viruses. [ABSTRACT FROM PUBLISHER]

The article offers information on Virology research. Topics discussed include significance of virology in the study of ecology, evolution of viruses, interaction among viruses and other microorganisms; morphology and physiology of viruses; and information on Metabolism in viruses. It also mentions human virus interaction.

Thecurrent work investigates the thermoresponsive in situ chiralto nonchiral ordering transformation of a rodlike virus in the naturallyassembled statethe chiral nematic liquid crystal (CLC) phase.We take this as an elegant example of reconfigurable self-assembly,through which it is possible to realize in situ transformation fromone assembled state to another without disrupting the preformed assemblyin general or going through a secondary assembling procedure of thedisassembled building blocks. The detailed investigation presentedhere reveals many unique characteristics of the thermoresponsive 3Dchiral ordering of rodlike viruses induced by heat stress. The chiralto nonchiral ordering transformation is highly reversible in the temperaturerange of up to 60 °C and can be repeated many times. There existsa critical temperature around 40 °C which is independent of theionic strength and virus concentration. Such reconfigurable orderingin the CLC phase stems from the intrinsic structure change of constituentcoat proteins without disrupting the structural integrity of the virus,as revealed by three analytical techniques targeting levels rangingfrom the molecular, secondary conformation of the constituent proteinsto the whole single virus, respectively. Such structural flexibility,also termed polymorphism, is relative to the survival strategies ofa biological organism such as the virus and can be transformed intovery precious material properties. The potential of the virus-basedCLC phase as the chiral matrix to regulate chiro-optical propertiesof gold nanorods is also presented. [ABSTRACT FROM AUTHOR]