Isometric Particles Research Articles

Identification, characterization and full-length sequence analysis of a novel dsRNA virus isolated from the arboreal ant Camponotus yamaokai.

A novel dsRNA virus was identified from the arboreal ant Camponotus yamaokai. The complete nucleotide sequence analysis of the virus revealed that the virus consisted of 5704 bp with two ORFs. ORF1 (3084 nt) encoded a putative capsid protein. ORF2 (1977 nt) encoded a viral RNA-dependent RNA polymerase (RdRp). ORF2 could be translated as a fusion with the ORF1 product by a - 1 frameshift in the overlapping ORF1. Phylogenetic analyses based on the RdRp revealed that the virus from C. yamaokai was most likely a novel totivirus, but it was not closely related to the previously known totiviruses in arthropods. Transmission electron microscopy revealed isometric virus particles of ~30 nm diameter in the cytoplasm, which was consistent with the characteristics of the family Totiviridae. The virus was detected by reverse transcription-PCR in all caste members and developmental stages of ants, including eggs, larvae, pupae, adult workers, alates (male and female) and queens. To our knowledge, this is the first report of a member of the family Totiviridae in a hymenopteran; the virus was designated Camponotus yamaokai virus.

Fully resolved simulations of the flow through a packed bed of cylinders: Effect of size distribution

Fully resolved simulations of the flow through a fixed bed of pellets are performed to better understand the effect of the local microstructure on the energy loss, i.e., pressure drop through the bed. Both mono-disperse and poly-disperse systems as well as spherical and cylindrical pellets (solid particles) are investigated. Using a DEM-DLM/FD simulation method inspired by “Wachs, A. (2009). A DEM-DLM/FD method for direct numerical simulation of particulate flows: sedimentation of polygonal isometric particles in a Newtonian fluid with collisions. Comput. Fluids 38(8), 1608–1628” and implemented here in a Finite Volume context with second order reconstruction of the particle boundary as in “Rahmani, M., Wachs, A. (2014). Free falling and rising of spherical and angular particles. Phys. Fluids 26, 083301”, we evidence that the computed solution converges nicely with mesh refinement and provide guidelines on the grid size to guarantee a satisfactory level of accuracy. Based on these trustworthy simulation results, we investigate the impact of the particle shape as well as the degree of poly-dispersity in the system on the pressure drop over the fixed bed in the viscous regime. Unprecedented simulation results on the flow through a bed of poly-disperse cylinders indicate that the correlation for poly-disperse spheres suggested in “Van der Hoef, M.A., Beetstra, R., Kuipers, J.A.M. (2005). Lattice-Boltzmann simulations of low-Reynolds-number flow past mono- and bidisperse arrays of spheres: results for the permeability and drag force. J. Fluid Mech. 528, 233–254” may still be valid for cylinders of moderate aspect ratio.

Activity-induced phase separation and self-assembly in mixtures of active and passive particles.

We investigate the phase behavior and kinetics of a monodisperse mixture of active (i.e., self-propelled) and passive isometric Brownian particles through Brownian dynamics simulations and theory. As in a purely active system, motility of the active component triggers phase separation into a dense and a dilute phase; in the dense phase, we further find active-passive segregation, with "rafts" of passive particles in a "sea" of active particles. We find that phase separation from an initially disordered mixture can occur with as little as 15% of the particles being active. Finally, we show that a system prepared in a suitable fully segregated initial state reproducibly self-assembles an active "corona," which triggers crystallization of the passive core by initiating a compression wave. Our findings are relevant to the experimental pursuit of directed self-assembly using active particles.

Derivation of occupational exposure levels (OELs) of low-toxicity isometric biopersistent particles: How can the kinetic lung overload paradigm be used for improved inhalation toxicity study design and OEL-derivation?

BackgroundConvincing evidence suggests that poorly soluble low-toxicity particles (PSP) exert two unifying major modes of action (MoA), in which one appears to be deposition-related acute, whilst the other is retention-related and occurs with particle accumulation in the lung and associated persistent inflammation. Either MoA has its study- and cumulative dose-specific adverse outcome and metric. Modeling procedures were applied to better understand as to which extent protocol variables may predetermine any specific outcome of study. The results from modeled and empirical studies served as basis to derive OELs from modeled and empirically confirmed directions.ResultsThis analysis demonstrates that the accumulated retained particle displacement volume was the most prominent unifying denominator linking the pulmonary retained volumetric particle dose to inflammogenicity and toxicity. However, conventional study design may not always be appropriate to unequivocally discriminate the surface thermodynamics-related acute adversity from the cumulative retention volume-related chronic adversity. Thus, in the absence of kinetically designed studies, it may become increasingly challenging to differentiate substance-specific deposition-related acute effects from the more chronic retained cumulative dose-related effects.ConclusionIt is concluded that the degree of dissolution of particles in the pulmonary environment seems to be generally underestimated with the possibility to attribute to toxicity due to decreased particle size and associated changes in thermodynamics and kinetics of dissolution. Accordingly, acute deposition-related outcomes become an important secondary variable within the pulmonary microenvironment. In turn, lung-overload related chronic adversities seem to be better described by the particle volume metric. This analysis supports the concept that ‘self-validating’, hypothesis-based computational study design delivers the highest level of unifying information required for the risk characterization of PSP. In demonstrating that the PSP under consideration is truly following the generic PSP-paradigm, this higher level of mechanistic information reduces the potential uncertainty involved with OEL derivation.

Detection of a new ilarvirus infecting wild radish in New Zealand

A new ilarvirus was detected in wild radish in New Zealand. Numerous isometric particles (O 35 nm) were seen by electron microscopy. Partial sequences for RNA1, RNA2 and RNA3 were obtained. Based on sequence data, the virus belongs to the Ilarviruses subgroup 1, which members normally cause diseases of economic importance in their hosts, however it is unknown if the virus contributed to the severe symptoms observed in the plant also infected with Turnip mosaic virus.

Optimal online soft sensor for product quality monitoring in propylene polymerization process

Abstract In the real-time propylene polymerization manufacturing process, melt index (MI), as the key product quality variable, is hard to be measured on-line, which brings difficulties to the control and optimization of this process. However, a large amount of data of other relative process variables in this process can be routinely recorded online by the distributed control system (DCS). An optimal soft-sensor of least squares support vector machine (LS-SVM) is therefore proposed to implement the on-line estimation of MI with the above real-time DCS records, where LS-SVM is employed for developing a data-driven model of the above industry process. In view of that the input variable selection and parameter setting are crucial for the learning results and generalization ability of LS-SVM, the nonlinear isometric feature mapping technique and particle swarm optimization algorithm are then structurally integrated into the model to search the optimal values of those parameters. Considering the process time-varying nature, an online correction strategy is further switched on to update the modeling data and revise the model configuration parameters via adaptive behavior. Finally, the explored soft sensor model is illustrated with a real plant of propylene polymerization, and the results show the predictive accuracy and validity of the proposed systematic approach.

First Report of Cucumber mosaic virus Subgroup IA Isolate Infecting Yucca aloifolia in Italy.

Yucca aloifolia L. (Spanish bayonet), family Asparagaceae, is the type species of the genus Yucca. It is native to Mexico and the West Indies and is appreciated worldwide as an ornamental plant. In 2013, during a survey for viruses in ornamental plants in the Campania region of southern Italy, symptoms consisting of bright chlorotic spots and ring spots 1 to 3 mm in diameter with some necrotic streaks were observed on leaves of two plants of Y. aloifolia growing in a nursery located in the Pignataro Maggiore municipality, Caserta Province. Cucumber mosaic virus (CMV) infection was suspected because the symptoms resembled those caused by CMV in Yucca flaccida (1). A range of herbal plant indicators was inoculated with sap extracts of symptomatic Y. aloifolia plants and developed symptoms indicative of CMV. Furthermore, 30 nm isometric virus particles were observed in the same Y. aloifolia sap extracts by transmission electron microscopy. The identity of the virus was confirmed by positive reaction in ELISA tests with CMV polyclonal antisera (Bioreba) conducted on sap extracts of symptomatic Y. aloifolia plants and systemically infected symptomatic hosts (i.e., Nicotiana tabacum, N. glutinosa, Cucumber sativus cv. Marketer, Solanum lycopersicum cv. San Marzano). The presence of CMV in the two naturally infected Y. aloifolia and other mechanically inoculated plants was further verified by reverse transcription (RT)-PCR. Total RNAs were extracted with the E.Z.N.A. Plant RNA Kit (Omega Bio-Tek), according to the manufacturer's instructions. RT-PCR was carried out with the ImProm-II Reverse Transcription System first-strand synthesis reaction (Promega) using the primer pair CMV1 and CMV2 (2). These primers amplify part of the CP gene and part of the 3'-noncoding region of CMV RNA3 and were designed to produce amplicons of different sizes to distinguish CMV isolates belonging to subgroups I or II (3). RT-PCR products were obtained from both naturally infected Y. aloifolia and mechanically inoculated plants as well as from PAE1 isolate of CMV (2), used as positive control, but not from healthy plants. Based on the length of the amplicons obtained (487 bp), the CMV isolate from Y. aloifolia (named YAL) belonged to subgroup I (3). The amplified RT-PCR products were purified with QIAquick PCR Purification Kit (Qiagen), cloned in the pGEMT vector (Promega), and three independent clones were sequenced at MWG (Ebersberg, Germany). Sequences obtained from the two CMV-infected Y. aloifolia plants were identical. This sequence was deposited at GenBank (Accession No. HG965199). Multiple alignments of the YAL sequence with sequences of other CMV isolates using MEGA5 software revealed highest percentage of identity (98.9%) with the isolates Z (AB369269) and SO (AF103992) from Korea and Japan, respectively. Moreover, the YAL isolate was identified as belonging to subgroup IA, based on the presence of only one HpaII restriction site in the 487-bp sequence, as previously proposed (2). Although CMV seems to not be a major threat currently for the production of Y. aloifolia, because the farming of this plant is performed using vegetative propagation, particular attention should be given to the presence of the virus in donor mother plants in order to avoid the dispersion of infected plants that could serve as sources for aphid transmission to other susceptible plant species. To our knowledge, this is the first report of CMV infection of Y. aloifolia in the world.

Impact of needle-like crystals on wet and solid-lipid extrusion processes

The aim of the current work was to investigate systematically the impact of needle-like drug crystals on both wet and solid-lipid extrusion processes. Co-rotating twin-screw extrusion, through 0.5, 0.7 and 1.0mm multi-holed capillary dies, was therefore used to assess the processability of formulations containing up to 80wt.% of three needle-like, poorly soluble active ingredients, i.e. mesalamine, nimesulide and praziquantel. Their impact on both wet and solid-lipid extrusions was evaluated prior to and after an appropriate micronisation step aimed at reducing their particle size, and obtaining more isometric particles. Occlusion of die holes and a steady increase in extrusion pressure, whose extent was found to be dependent on both the drug loading and the morphology of the drug's crystal, was observed with all the active ingredients tested. A change in particle morphology, combined to a reduction in particle size, allowed the occlusion of die holes to be considerably reduced and/or prevented, and smooth and robust extrusion processes to be achieved.

Microstructure Evolution in Lithium-Ion Battery Electrode Processing

The electrode microstructure and composition play an important role in determining the performance of the lithium ion battery (LIB). To this end, the processing of the multi-phase slurry consisting of active nanoparticles, conductive additives, binder and solvent determines the electrochemical properties and performance of the electrode. Mixing components in the slurry homogenously is one essential requirement to process the high quality electrode. Additionally, it is necessary to make these components cooperate very well with each other.1-4 Fundamentally, slurry properties and drying methods mainly determine the microstructure and the performance of the cathode composite. Viscosity of the slurry is an important property, which is determined by the concentration and the solid loading of each component, the interaction force among nanoparticles in the slurry, and the properties of active material nanoparticles that include the size of the nanoparticle, size distribution and the surface area.5 One common mechanism of the drying process is solvent evaporation from the surface of the substrate. The evaporation induced assembly is a popular scheme to make nanoparticles form desirable microstructures, and this assembly is controlled by interactions among nanoparticles and solvents, evaporation rates, mobility of nanoparticles and the morphology of nanoparticles.In our present work, a morphology-detailed mesoscale model has been developed to gain fundamental understating of the influence of active particle morphology, size, volume fraction, solvent evaporation rated, and multi-phase (active particle, conductive additive, binder and solvent) interaction. Our results demonstrate that smaller isometric active particles tend to form favorable aggregation with conductive additive particles. Two regimes, namely spontaneous aggregation and evaporation induced

Presentation of epitope sequences from foreign viruses on the surface of apple latent spherical virus particles

Apple latent spherical virus (ALSV) has small isometric particles that are comprised of two single-stranded RNA species (RNA1 and RNA2) and three capsid proteins (Vp25, Vp20, and Vp24). We constructed ALSV vectors for presenting foreign peptides on the surface of virus particles. In these vectors, peptides can be fused to either of two C-terminal regions of Vp20 (amino acid positions between G171 and P172 or between P172 and L173) or the C-terminus (T192) of Vp24. An ALSV vector presenting the epitope sequences of the coat protein (CP) of zucchini yellow mosaic virus (ZYMV) could systemically infect host plants and was specifically recognized by antiserum against ZYMV by ELISA, immunoelectron microscopy, and immunoblotting. RT-PCR showed that the epitope sequences up to 20 amino acids were stably maintained in the chimeric ALSV for more than 10 serial passages and at least six months. Purified chimeric ALSV particles induced an immune response and the production of antibodies against ZYMV-CP in rabbits. The ALSV vector was also used for expression of an epitope from VP1 of foot-and-mouth disease virus.

Western Flower Thrips (Frankliniella occidentalis) Transmits Maize Chlorotic Mottle Virus

AbstractMaize chlorotic mottle virus (MCMV) is one of the co‐infection pathogens that cause corn (maize) lethal necrosis, but the transmission mechanism of MCMV is not yet clear. In order to determine the ability of western flower thrips (Frankliniella occidentalis; WFT) to transmit MCMV, imported maize seeds from Thailand were germinated in an insect‐free greenhouse and the seedlings were tested for the transmission by WFT of chlorotic mottle virus disease. The thrips (WFT), starved for 48 h then allowed to feed for 30 min on maize plants infected with MCMV or asymptomatic maize plants, were transferred to healthy seedlings. After 35 days, the seedlings with WFT from diseased maize plants showed chlorotic mottle symptoms, whereas seedlings with WFT from asymptomatic maize plants remained healthy. A single band of 711 bp was amplified by RT‐PCR using primers MCMV‐F/MCMV‐R from the MCMV‐infected plants and WFT collected from the diseased plants. Sequencing of the amplified product and further sequence comparison indicated that the two viruses from both sources showed 99% similarity of nucleotides and they should be regarded as identical. In addition, isometric particles c. 30 nm in diameter, characteristic of MCMV, were found in the WFT samples from diseased maize plants. Thus, it is concluded that WFT transmits MCMV. Our findings suggest that corn lethal necrosis disease can be controlled or minimized by the eradication of WFT from the field or greenhouses.

Microstructure Evolution in Lithium-Ion Battery Electrode Processing

The processing induced active particle assembly determines the internal microstructure and resultant performance of the electrode in a lithium-ion battery. A morphology-detailed mesoscale model has been developed to gain fundamental understanding of the influence of active particle morphology, size, volume fraction, solvent evaporation, and multi-phase (active particle, conductive additive, binder and solvent) interaction. Our results demonstrate that smaller isometric active particles tend to form favorable aggregation with conductive additive particles. Two regimes, namely spontaneous aggregation and evaporation induced aggregation, have been identified. Low solvent evaporation rate promotes spontaneous aggregation resulting in an enhanced interfacial area than that in evaporation-induced aggregation. The influence of active material morphology and volume fraction on conducting pathway formation has been conjectured.

Molecular characterization of a novel victorivirus from the entomopathogenic fungus Beauveria bassiana

New Zealand isolates of the entomopathogenic fungus Beauveria were examined for the presence of dsRNAs and virus-like particles. Seven out of nine isolates contained one or more high-molecular-weight dsRNAs and all seven contained isometric virus particles ranging in size from 30 to 50 nm. B. bassiana isolate ICMP#6887 contained a single dsRNA band of ~6 kb and isometric virus-like particles of ~50 nm in diameter. Sequencing revealed that the virus from ICMP#6887 had a genome of 5,327 nt with two overlapping ORFs coding for a putative coat protein (CP) and an RNA-dependent RNA-polymerase (RdRp). The sequence showed a highest CP identity of 58.3 % to Tolypocladium cylindrosporum virus 1 (TcV1) and a highest RdRp identity of 48.8 % to Sphaeropsis sapinea RNA virus 1 (SsRV1). Since both TcV1 and SsRV1 belong to the genus Victorivirus, the new virus from B. bassiana ICMP#6887 was tentatively assigned the name Beauveria bassiana victorivirus 1 (BbVV1-6887).

Mycoviruses ofBotrytis cinereaisolates from different hosts

Botrytis cinerea (teleomorph Botryotinia fuckeliana) is a necrotrophic plant pathogenic fungus that causes grey mould and enormous economic losses worldwide in different crops. Control of B. cinerea is difficult due to the appearance of fungicide-resistant isolates, and the diversity in virulence due to genetic variability and, perhaps, the infection with mycoviruses or fungal viruses. The discovery of mycoviruses and their possible application as biocontrol agents, as well as their use as tools to study the plant–pathogen interaction, has encouraged their study in B. cinerea. Herein, we have analysed the occurrence of mycoviruses in Spanish B. cinerea isolates to approach a better understanding of the interactions among viruses, fungi and plants in this pathosystem. Fifty-five percent of the B. cinerea isolates analysed contained double-stranded RNA (dsRNA) elements, and the number of dsRNA elements, their relative concentration and size were variable among isolates. Some of these dsRNAs were related to the presence of virus like rod or isometric particles, and to cellular degeneration and malformed mitochondria. We have also demonstrated that a 3 kb dsRNA present in 55% of the isolates having dsRNA elements was a mycovirus genome. Partial sequence of that mycovirus presented high identity in nucleotide and amino acid sequence with Botrytis cinerea mitovirus 1 (BcMV1). Analysis of the genetic distance within Spanish BcMV1 sequences showed the existence of different isolates of this mitovirus inside the Spanish B. cinerea population analysed. This is the first report of the variability of dsRNA elements and the partial genome sequence of a mitovirus associated with Spanish B. cinerea isolates and the genetic diversity within Spanish isolates of BcMV1.

A fundamental study of the extent of meaningful application of Maxwell's and Wiener's equations to the permeability of binary composite materials. Part I: A numerical computation approach

In studies of the permeability of composite materials, consisting of solid isometric particles A dispersed in a polymeric matrix B, Maxwell's classical equation, rigorously valid only for very low fractional volumes (vA) of spherical particles, is often used for data analysis up to much higher vA values. Theoretical justification for this practice can be provided, only up to vA≈0.5, by current analytical models based on cubic lattices of congruent spheres. Replacing spheres by cubes yields a model covering the full composition range vA=0−1, but lacking convenient general analytical tractability. Accordingly, to explore unrestrictedly the practical validity of the Maxwell equation, a simple cubic lattice-of-cubes model was combined with an appropriate numerical computation tool. The results establish satisfactory applicability of the Maxwell equation for isometric particles A, at all compositions and over a wide range of component permeability ratios (α=PA/PB=0−100).The practical applicability of the corresponding classical equation of Wiener (which extends Maxwell's treatment to anisometric particles via the value of a single geometrical parameter AW) was similarly explored, using appropriate model s.c. lattices of (a) unidimensionally anisometric particles (transverse square rods of varying length) or (b) bidimensionally anisometric particles (transverse square platelets of varying area). The computations covered the same vA and α ranges as above, as well as a range of aspect ratios in each of the cases (a) and (b). The results obtained enable determination of values of AW, independent of vA and α, linked directly to the aspect ratio of the embedded particles, and demonstrate very good practical applicability of the Wiener equation (with the proper AW value) under all conditions studied.

Collective stringlike motion of semiflexible filamentous particles in columnar liquid crystalline phases

We study, by means of Brownian dynamics simulations, heterogeneous dynamics in a dense columnar phase of monodisperse hard filamentous particles, and find that in a background of barely moving particles, some particles occasionally engage in a fast coherent string-type motion similar to what is observed in glassy states of isometric particles. This fast motion is triggered by the exchange of particles between two or more columns at different positions in the columns, which leads to sudden displacement of particles between these positions. The distribution of particle displacements shows a pronounced peak at one particle length. We perform our simulations with particles of different persistence lengths and find that for more flexible particles, the number of jump events increases. As the number of particles in the columns increases with system size for a given linear fraction of particles in the columns, the peak in the distribution becomes wider and, for sufficiently large systems, the peak disappears completely. This is associated with the increase in the magnitude of fluctuations in the motion of particles as the system size increases. Our simulation results explain recent experimental observations on single-particle motion in dense columnar phases in aqueous dispersions of filamentous virus particles.

Molecular detection of an isolate of Cucumber mosaic virus infecting Oxalis corymbosa in Aligarh, India

Plants of Oxalis corymbosa displaying typical viral symptoms were observed during spring 2010 in plant nurseries in central India. The virus was mechanically transmissible to indicator plants and induced symptoms similar to those incited by Cucumber mosaic virus (CMV). Electron microscopic examinations of symptomatic leaf samples revealed the presence of isometric virus-like particles about 28–30 nm in diameter. Molecular detection methods allowed the identification of the pathogen as a CMV. The results were confirmed by reverse transcription polymerase chain reaction with specific CMV primers, and digestion with a restriction enzyme. The coat protein gene was 95 and 99% identical at the nucleotide and amino acid levels, respectively, to the published sequences of CMV, high amino acid sequence homology in multiple alignment analysis and closest phylogenetic relationship with Lucknow isolate (JX570732) of CMV. This represents the first record for CMV-infecting O. corymbosa in Aligarh India.

First Report of Broad bean wilt virus-2 in Pepper in the Czech Republic.

In August 2012, field-grown pepper (Capsicum annuum) plants were inspected for the presence of viral diseases in Moravia, Czech Republic. Leaves taken from plants showing virus-like symptoms were examined for the presence of pepper viruses by double antibody sandwich (DAS)-ELISA. Of the 80 collected samples, 22 were infected with Cucumber mosaic virus, 17 with Potato virus Y, 13 with Broad bean wilt virus-1, and 11 with Broad bean wilt virus-2 (BBWV-2). The BBWV-2 positive samples were collected near the villages of Hrádek and Tasovice in Znojmo District and Loděnice in Brno-Country District. Six samples positive for BBWV-2 were also infected with one or two other viruses. BBWV-2 was mechanically transmitted from the field-collected samples where it was present alone onto Chenopodium quinoa plants. After systemic mottling emerged on indicator plants, symptomatic leaves were collected and the presence of BBWV-2 was confirmed by electron microscopy and reverse transcriptase (RT)-PCR. Isometric particles of about 25 nm corresponding in size and shape to the BBWV-2 virions (2) were observed. Total RNA was extracted from infected C. quinoa leaves, and RT-PCRs with primer pairs specific to four BBWV-2 genome regions (1) were carried out using oligo-dT and random hexamer mixtures for the cDNA synthesis. Amplicons of expected sizes, ~400 bp for Fab5'R1, ~460 bp for Pol2, ~500 bp for MP4, and ~250 bp for SCP were obtained for all samples. To the best of our knowledge, this is the first report of a natural occurrence of BBWV-2 in the Czech Republic.

Molecular characterization of a trisegmented chrysovirus isolated from the radish Raphanus sativus

Radish (Raphanus sativus L.) is cultivated worldwide and is of agronomic importance. dsRNAs associated with partitiviruses were previously found in many R. sativus varieties. In this study, three large dsRNAs from radish were cloned using a modified single primer amplification technique. These three dsRNAs—of lengths 3638, 3517 and 3299 bp—shared conserved untranslated terminal regions, and each contained a major open reading frame putatively encoding the chrysoviral replicase, capsid protein and protease respectively. Isometric virus-like particles (VLP), approximately 45nm in diameter, were isolated from the infected radish plants. Northern blotting indicated that these dsRNAs were encapsidated in the VLP. The virus containing these dsRNA genome segments was named Raphanus sativus chrysovirus 1 (RasCV1). Phylogenetic analysis revealed that RasCV1 is a new species of the Chrysoviridae family and forms a plant taxon with another putative plant chrysovirus, Anthurium mosaic-associated virus (AmaCV). Furthermore, no fungal mycelia were observed in radish leaf tissues stained with trypan blue. These results indicated that RasCV1 is most likely a plant chrysovirus rather than a chrysovirus in symbiotic fungi. An exhaustive BLAST analysis of RasCV1 and AmaCV revealed that chrysovirus-like viruses might widely exist in eudicot and monocot plants and that endogenization of chrysovirus segments into plant genome might have ever happened.

A wild-type Botrytis cinerea strain co-infected by double-stranded RNA mycoviruses presents hypovirulence-associated traits

BackgroundBotrytis cinerea CCg378 is a wild-type strain infected with two types of double-stranded RNA (dsRNA) mycoviruses and which presents hypovirulence-associated traits. The objectives of the present study were to characterize the mycoviruses and investigate their relationship with the low virulence degree of the fungal host.ResultsB. cinerea CCg378 contains five dsRNA molecules that are associated with two different types of isometric viral particles of 32 and 23 nm in diameter, formed by structural polypeptides of 70-kDa and 48-kDa, respectively.The transfection of spheroplasts of a virus-free strain, B. cinerea CKg54, with viral particles purified from the CCg378 strain revealed that the 2.2-kbp dsRNAs have no dependency on the smaller molecules for its stable maintenance in the fungal cytoplasm, because a fungal clone that only contains the 2.2-kbp dsRNAs associated with the 32-nm particles was obtained, which we named B. cinerea CKg54vi378. One of the 2.2 kbpdsRNA segments (2219 bp) was sequenced and corresponds to the gene encoding the capsid protein of B. cinerea CCg378 virus 1 (Bc378V1), a putative new member of the Partitiviridae family.Furthermore, physiological parameters related to the degree of virulence of the fungus, such as the sporulation rate and laccase activity, were lower in B. cinerea CCg378 and B. cinerea CKg54vi378 than in B. cinerea CKg54. Additionally, bioassays performed on grapevine leaves showed that the CCg378 and CKg54vi378 strains presented a lower degree of invasiveness on the plant tissue than the CKg54 strain.ConclusionsThe results show that B. cinerea CCg378 is coinfected by two mycoviruses and that the 2.2-kbp dsRNAs correspond to the 32-nm mycovirus genome, which would be a new member of the Partitiviridae family as it has the typical pattern of partitiviruses. On the other hand, the results suggest that the hypovirulence of B. cinerea CCg378 could be conferred by both mycoviruses, since the fungal clone B. cinerea CKg54vi378 presents an intermediate virulence between the CKg54 and CCg378 strains. Therefore, the putative partitivirus would be partially contributing to the hypovirulence phenotype of the CCg378 strain.