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Applied Microbiology and Biotechnology Apr 2012Virus-like particles (VLPs) are shell-like viruses that lack virus-specific genetic materials. Many viral-structured proteins can assemble into VLPs, which mimic the... (Review)
Review
Virus-like particles (VLPs) are shell-like viruses that lack virus-specific genetic materials. Many viral-structured proteins can assemble into VLPs, which mimic the overall structure of virus particles and can elicit strong immune responses in a host. Dengue viruses (DENVs), from the genus Flavivirus, are transmitted to humans through the bites of an infected Aedes mosquito. DENVs cause several diseases that prevailed mainly in tropical and subtropical areas. However, effective treatment measures and preventive strategies for dengue diseases are still lacking. The present minireview summarized the assembly and maturation of DENVs, the strategies and effective factors for dengue VLP construction, and the application of DENV VLPs.
Topics: Animals; Defective Viruses; Dengue; Dengue Virus; Humans
PubMed: 22382168
DOI: 10.1007/s00253-012-3958-7 -
PloS One 2019Most viruses are known to spontaneously generate defective viral genomes (DVG) due to errors during replication. These DVGs are subgenomic and contain deletions that...
Most viruses are known to spontaneously generate defective viral genomes (DVG) due to errors during replication. These DVGs are subgenomic and contain deletions that render them unable to complete a full replication cycle in the absence of a co-infecting, non-defective helper virus. DVGs, especially of the copyback type, frequently observed with paramyxoviruses, have been recognized to be important triggers of the antiviral innate immune response. DVGs have therefore gained interest for their potential to alter the attenuation and immunogenicity of vaccines. To investigate this potential, accurate identification and quantification of DVGs is essential. Conventional methods, such as RT-PCR, are labor intensive and will only detect primer sequence-specific species. High throughput sequencing (HTS) is much better suited for this undertaking. Here, we present an HTS-based algorithm called DVG-profiler to identify and quantify all DVG sequences in an HTS data set generated from a virus preparation. DVG-profiler identifies DVG breakpoints relative to a reference genome and reports the directionality of each segment from within the same read. The specificity and sensitivity of the algorithm was assessed using both in silico data sets as well as HTS data obtained from parainfluenza virus 5, Sendai virus and mumps virus preparations. HTS data from the latter were also compared with conventional RT-PCR data and with data obtained using an alternative algorithm. The data presented here demonstrate the high specificity, sensitivity, and robustness of DVG-profiler. This algorithm was implemented within an open source cloud-based computing environment for analyzing HTS data. DVG-profiler might prove valuable not only in basic virus research but also in monitoring live attenuated vaccines for DVG content and to assure vaccine lot to lot consistency.
Topics: Algorithms; Animals; Chromosome Mapping; DNA Primers; Datasets as Topic; Defective Viruses; Genome, Viral; High-Throughput Nucleotide Sequencing; Humans; Molecular Typing; Mumps virus; Parainfluenza Virus 5; Real-Time Polymerase Chain Reaction; Sendai virus; Sensitivity and Specificity
PubMed: 31100083
DOI: 10.1371/journal.pone.0216944 -
Virus Research Apr 2014Long-term surviving sugar beet plants were investigated after beet curly top virus infection to characterize defective (D) viral DNAs as potential symptom attenuators....
Long-term surviving sugar beet plants were investigated after beet curly top virus infection to characterize defective (D) viral DNAs as potential symptom attenuators. Twenty or 14 months after inoculation, 20 D-DNAs were cloned and sequenced. In contrast to known D-DNAs, they exhibited a large range of sizes. Deletions were present in most open reading frames except ORF C4, which encodes a pathogenicity factor. Direct repeats and inverted sequences were observed. Interestingly, the bidirectional terminator of transcription was retained in all D-DNAs. A model is presented to explain the deletion sites and sizes with reference to the viral minichromosome structure, and symptom attenuation by D-DNAs is discussed in relation to RNA interference.
Topics: Beta vulgaris; DNA, Viral; Defective Viruses; Geminiviridae; Genes, Viral; Sequence Deletion
PubMed: 24530983
DOI: 10.1016/j.virusres.2014.01.028 -
Virology Dec 1990Defective interfering virus particles (DIP) frequently play an important part in viral persistence in vitro, and may in some instances modify a virus infection in vivo,...
Defective interfering virus particles (DIP) frequently play an important part in viral persistence in vitro, and may in some instances modify a virus infection in vivo, causing attenuation or persistence of the infection. To explain certain aspects of the growth of these mutants in vitro, other factors have been invoked such as interferon, mutations in the wild-type virus or the infected cells, or other substances released by infected cells that attenuate the infection. We present here a simple model of the growth of DIP in vitro which shows that (a) the observed population dynamics of DIP can readily be explained without invoking such extrinsic factors; (b) the initial multiplicity of infection of DIP is the principal determinant of the outcome of infection in both single- and repeated-passage cultures; and (c) in a long-term culture in vitro, the criterion used to decide the time of virus passage directly determines how long the standard virus, DIP, and cells survive. This model may be used with minor modifications to predict the behavior in vitro of other mutant viruses with a dominantly interfering phenotype.
Topics: Biological Evolution; Defective Viruses; Viral Interference; Virus Physiological Phenomena; Virus Replication; Viruses
PubMed: 2238471
DOI: 10.1016/0042-6822(90)90150-p -
Journal of Virology Oct 1978A temperature-sensitive group II mutant of influenza virus, ts-52, with a presumed defect in viral RNA synthesis, readily produced von Magnus-type defective interfering...
A temperature-sensitive group II mutant of influenza virus, ts-52, with a presumed defect in viral RNA synthesis, readily produced von Magnus-type defective interfering virus (DI virus) when passed serially (four times) at high multiplicity in MDBK cells. The defective virus (ts-52 DI virus) had a high hemagglutinin and a low infectivity titer, and strongly interfered with the replication of standard infectious viruses (both ts-52 and wild-type ts+) in co-infected cells. Progeny virus particles produced by co-infection of DI virus and infectious virus were also defective and also had low infectivity, high hemagglutinating activity, and a strong interfering property. Infectious viruses ts+ and ts-52 were indistinguishable from ts-52 DI viruses by sucrose velocity or density gradient analysis. Additionally, these viruses all possessed similar morphology. However, when the RNA of DI viruses was analyzed by use of polyacrylamide gels containing 6 M urea, there was a reduction in the amount of large RNA species (V1 to V4), and a number of new smaller RNA species (D1 to D6) with molecular weights ranging from 2.9 X 10(5) to 1.05 X 10(5) appeared. Since these smaller RNA species (D1 to D6) were absent in some clones of infectious viruses, but were consistently associated with DI viruses and increased during undiluted passages and during co-infection of ts-52 with DI virus, they appeared to be a characteristic of DI viruses. Additionally, the UV target size of interfering activity and infectivity of DI virus indicated that interfering activity was 40 times more resistant to UV irradiation than was infectivity, further implicating small RNA molecules in interference. Our data suggest that the loss of infectivity observed among DI viruses may be due to nonspecific loss of a viral RNA segment(s), and the interfering property of DI viruses may be due to interfering RNA segments (DIRNA, D1 to D6). ts-52 DI virus interfered with the replication of standard virus (ts+) at both permissive (34 degrees C) and nonpermissive temperatures. The infectivity of the progeny virus was reduced to 0.2% for ts+ and 0.05% for ts-52 virus without a reduction in hemagglutinin titer. Interference was dependent on the concentration of DI virus. A particle ratio of 1 between DI virus (0.001 PFU/cell) and infectious virus (1.0 PFU/cell) produced a maximal amount of interference. Infectious virus yield was reduced 99.9% without any reduction of the yield of DI viruses Interference was also dependent on the time of addition of DI virus. Interference was most effective within the first 3 h of infection by infectious virus, indicating interference with an early function during viral replication.
Topics: Cell Line; Defective Viruses; Influenza A virus; Mutation; RNA, Viral; Temperature; Viral Interference; Viral Proteins; Virus Replication
PubMed: 702654
DOI: 10.1128/JVI.28.1.375-386.1978 -
Blood Oct 1996Adult T-cell leukemia (ATL), an aggressive neoplasm of mature helper T cells, is etiologically linked with human T lymphotropic virus type I (HTLV-1). After infection,... (Comparative Study)
Comparative Study
Adult T-cell leukemia (ATL), an aggressive neoplasm of mature helper T cells, is etiologically linked with human T lymphotropic virus type I (HTLV-1). After infection, HTLV-I randomly integrates its provirus into chromosomal DNA. Since ATL is the clonal proliferation of HTLV-I-infected T lymphocytes, molecular methods facilitate the detection of clonal integration of HTLV-I provirus in ATL cells. Using Southern blot analyses and long polymerase chain reaction (PCR) we examined HTLV-I provirus in 72 cases of ATL, of various clinical subtypes. Southern blot analyses revealed that ATL cells in 18 cases had only one long terminal repeat (LTR). Long PCR with LTR primers showed bands shorter than for the complete virus (7.7 kb) or no bands in ATL cells with defective virus. Thus, defective virus was evident in 40 of 72 cases (56%). Two types of defective virus were identified: the first type (type 1) defective virus retained both LTRs and lacked internal sequences, which were mainly the 5' region of provirus, such as gag and pol. Type 1 defective virus was found in 43% of all defective viruses. The second form (type 2) of defective virus had only one LTR, and 5'-LTR was preferentially deleted. This type of defective virus was more frequently detected in cases of acute and lymphoma-type ATL (21/54 cases) than in the chronic type (1/18 cases). The high frequency of this defective virus in the aggressive form of ATL suggests that it may be caused by the genetic instability of HTLV-I provirus, and cells with this defective virus are selected because they escape from immune surveillance systems.
Topics: Adult; Blotting, Southern; Cell Transformation, Viral; DNA, Neoplasm; DNA, Viral; Defective Viruses; Genome, Viral; Human T-lymphotropic virus 1; Humans; Immunologic Surveillance; Leukemia-Lymphoma, Adult T-Cell; Polymerase Chain Reaction; Preleukemia; Proviruses; Repetitive Sequences, Nucleic Acid
PubMed: 8874205
DOI: No ID Found -
Defective RNAs inhibit the assembly of influenza virus genome segments in a segment-specific manner.Virology Feb 1996Four avian influenza viruses have been generated, each containing a single extra defective RNA segment in addition to the eight standard segments. Three of the extra...
Four avian influenza viruses have been generated, each containing a single extra defective RNA segment in addition to the eight standard segments. Three of the extra RNAs were derived from segment 1 and the fourth from segment 2. Chick embryo fibroblast cells were infected with each virus, and a wild-type virus. Virus RNA was quantified in extracts of virus-infected cells and in virus released by 10 hr postinfection using reverse transcription and by Northern blot analysis. In the case of two of the viruses the presence of the defective RNA did not markedly affect the accumulation of virus RNA within the infected cell, but significantly and selectively reduced the amount of the "parent" segment in released virus. This effect was reduced in a third virus. In a fourth virus, defective RNA was found to be present at a low-input multiplicity and results were varied. Mixed infections of one of the viruses with a closely related wild-type virus resulted in reduction of the corresponding vRNA segment of the nondefective virus. We conclude that assembly of influenza virus segments is not a purely random process.
Topics: Animals; Base Sequence; Blotting, Northern; Cell Line; Chick Embryo; Defective Viruses; Genome, Viral; Influenza A virus; Molecular Sequence Data; RNA, Viral; Transcription, Genetic; Virus Assembly; Virus Replication
PubMed: 8607262
DOI: 10.1006/viro.1996.0068 -
Journal of Virology Jun 1993Defective interfering (DI) RNA genomes of poliovirus which contain in-frame deletions in the P1 capsid protein-encoding region have been described. DI genomes are...
Defective interfering (DI) RNA genomes of poliovirus which contain in-frame deletions in the P1 capsid protein-encoding region have been described. DI genomes are capable of replication and can be encapsidated by capsid proteins provided in trans from wild-type poliovirus. In this report, we demonstrate that a previously described poliovirus DI genome (K. Hagino-Yamagishi and A. Nomoto, J. Virol. 63:5386-5392, 1989) can be complemented by a recombinant vaccinia virus, VVP1 (D. C. Ansardi, D. C. Porter, and C. D. Morrow, J. Virol. 65:2088-2092, 1991), which expresses the poliovirus capsid precursor polyprotein, P1. Stocks of defective polioviruses were generated by transfecting in vitro-transcribed defective genome RNA derived from plasmid pSM1(T7)1 into HeLa cells infected with VVP1 and were maintained by serial passage in the presence of VVP1. Encapsidation of the defective poliovirus genome was demonstrated by characterizing poliovirus-specific protein expression in cells infected with preparations of defective poliovirus and by Northern (RNA) blot analysis of poliovirus-specific RNA incorporated into defective poliovirus particles. Cells infected with preparations of defective poliovirus expressed poliovirus protein 3CD but did not express capsid proteins derived from a full-length P1 precursor. Poliovirus-specific RNA encapsidated in viral particles generated in cells coinfected with VVP1 and defective poliovirus migrated slightly faster on formaldehyde-agarose gels than wild-type poliovirus RNA, demonstrating maintenance of the genomic deletion. By metabolic radiolabeling with [35S]methionine-cysteine, the defective poliovirus particles were shown to contain appropriate mature-virion proteins. This is the first report of the generation of a pure population of defective polioviruses free of contaminating wild-type poliovirus. We demonstrate the use of this recombinant vaccinia virus-defective poliovirus genome complementation system for studying the effects of a defined mutation in the P1 capsid precursor on virus assembly. Following removal of residual VVP1 from defective poliovirus preparations, processing and assembly of poliovirus capsid proteins derived from a nonmyristylated P1 precursor expressed by a recombinant vaccinia virus, VVP1 myr- (D. C. Ansardi, D. C. Porter, and C. D. Morrow, J. Virol. 66:4556-4563, 1992), in cells coinfected with defective poliovirus were analyzed. Capsid proteins generated from nonmyristylated P1 did not assemble detectable levels of mature virions but did assemble, at low levels, into empty capsids.(ABSTRACT TRUNCATED AT 400 WORDS)
Topics: Capsid; Defective Viruses; Genetic Complementation Test; Genome, Viral; HeLa Cells; Humans; Mutagenesis; Poliovirus; Protein Precursors; RNA, Viral; Recombinant Proteins; Vaccinia virus; Virion
PubMed: 8388519
DOI: 10.1128/JVI.67.6.3684-3690.1993 -
Journal of Virology Jul 1975We have characterized the virus progeny and its DNA from plaque-purified and undiluted passages of herpes simplex virus 1 in HEp-2 cells. Secifically, (i) infectious... (Comparative Study)
Comparative Study
We have characterized the virus progeny and its DNA from plaque-purified and undiluted passages of herpes simplex virus 1 in HEp-2 cells. Secifically, (i) infectious virus yields declined progressively in passages 1 through 10 and gradually increased at passages 11 through 14. The yields correlated with PFU/particle ratios. (ii) In cells infected with virus from passages 6 through 10, there was an overproduction of an early viral polypeptide (no. 4) and a delay in the synthesis of late viral proteins. In addition, the virus in these passages interfered with the replication of a nondefective marker virus. Cells infected with passage 14 virus produced normal amounts of polypeptide 4 and, moreover, this virus showed minimal interfering capacity. (iii) In addition to DNA of density 1.726 g/cm-3, which was the sole component present in viral progeny of passage 0, passages 6 through 14 contained one additional species (p 1.732) and in some instances (passages 6 and 10) also DNA of an intermediate buoyant density. The ratio of p 1.732 to p 1.726 DNA increased to a maximum of 4 in passages 6 through 9 and gradually decreased to 1 in passages 10 through 14. (iv) p 1.732 DNA cannot be differentiated from p 1.726 DNA with respect to size; however, it has no Hin III restriction enzyme cleavage sites and yields only predominantly two kinds of fragments with molecular weights of 5.1 x 10-6 and 5.4 x 10-6 upon digestion with EcoRI enzyme. (v) Partial denaturation profiles of purified p 1.732 DNA from passage 14 revealed the presence of two types of tandemly repeated units corresponding roughly in size to the EcoRI fragments and situated in different molecules. (vi) In addition to the two kinds of p 1.732 molecules consisting of tandem repaeat units of different sizes, other evidence for the diversity of defective DNA molecules emerged from comparisons of specific infectivity and interfering capacity of the progeny from various passages. The data suggest that some of the particles with DNA of normal buoyant density (1.726) must also be defective since the capacity to interfere and to produce an excess of polypeptide 4 did not appear to be proportional to the amount of high-buoyant-density defective DNA. The data suggest that defective interfering particles are replaced by defective particles with diminished capacity to interfere and that more than one species of defective DNA molecules evolves on serial preparation of HSV.
Topics: Carcinoma, Squamous Cell; Cell Line; Centrifugation, Density Gradient; DNA Restriction Enzymes; DNA, Viral; Defective Viruses; Electrophoresis, Polyacrylamide Gel; Formaldehyde; Humans; Laryngeal Neoplasms; Microscopy, Electron; Nucleic Acid Denaturation; Peptide Biosynthesis; Simplexvirus; Viral Interference; Viral Proteins; Virus Replication
PubMed: 166202
DOI: 10.1128/JVI.16.1.153-167.1975 -
Journal of Virology Nov 1974BHK cells infected with defective-interfering passages of Sindbis virus accumulate a species of RNA (20S) that is about half the molecular weight of the major viral mRNA...
BHK cells infected with defective-interfering passages of Sindbis virus accumulate a species of RNA (20S) that is about half the molecular weight of the major viral mRNA (26S). We have performed competitive hybridization experiments with these species of RNA and have established that 20S RNA contains approximately 50% of the nucleotide sequences present in 26S RNA. Our further studies, however, demonstrate that 20S RNA is unable to carry out the messenger function of 26S RNA. We found very little of the defective RNA associated with polysomes in vivo. In addition, it was unable to stimulate protein synthesis in vitro under conditions in which 26S RNA was translated. We have also examined viral RNA synthesis in BHK cells infected with standard or defective-interfering passages of Sindbis virus. This comparison suggests that defective partioles do not synthesize a functional replicase.
Topics: Animals; Base Sequence; Cell Line; Cell-Free System; Cricetinae; Defective Viruses; Electrophoresis, Polyacrylamide Gel; Kidney; Nucleic Acid Hybridization; Phosphorus Radioisotopes; Polyribosomes; Protein Biosynthesis; RNA, Messenger; RNA, Viral; Sindbis Virus; Sulfur Radioisotopes; Tritium; Uridine; Viral Interference; Viral Proteins; Virus Replication
PubMed: 4473568
DOI: 10.1128/JVI.14.5.1189-1198.1974