Molecular epidemiology of molluscum contagiosum.
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Adolescent↗
Biomedical subjects
G Darai
Publications and source records attributed to G Darai.
Analysis of the primary structure of the long terminal repeat and the gag and pol genes of the human spumaretrovirus.
The nucleotide sequence of the human spumaretrovirus (HSRV) genome was determined. The 5' long terminal repeat region was analyzed by strong stop cDNA synthesis and S1 nuclease mapping. The length of the RU5 region was determined and found to be 346 nucleotides long. The 5' long terminal repeat is 1,123 base pairs long and is bound by an 18-base-pair primer-binding site complementary to the 3' end of mammalian lysine-1,2-specific tRNA. Open reading frames for gag and pol genes were identified. Surprisingly, the HSRV gag protein does not contain the cysteine motif of the nucleic acid-binding proteins found in and typical of all other retroviral gag proteins; instead the HSRV gag gene encodes a strongly basic protein reminiscent of those of hepatitis B virus and retrotransposons. The carboxy-terminal part of the HSRV gag gene products encodes a protease domain. The pol gene overlaps the gag gene and is postulated to be synthesized as a gag/pol precursor via translational frameshifting analogous to that of Rous sarcoma virus, with 7 nucleotides immediately upstream of the termination codons of gag conserved between the two viral genomes. The HSRV pol gene is 2,730 nucleotides long, and its deduced protein sequence is readily subdivided into three well-conserved domains, the reverse transcriptase, the RNase H, and the integrase. Although the degree of homology of the HSRV reverse transcriptase domain is highest to that of murine leukemia virus, the HSRV genomic organization is more similar to that of human and simian immunodeficiency viruses. The data justify classifying the spumaretroviruses as a third subfamily of Retroviridae.
Amino Acid Sequence↗
Identification and characterization of the repetitive DNA element in the genome of insect iridescent virus type 6.
The genome of the Chilo iridescent virus (CIV) was analyzed for existence of repetitive DNA sequences by DNA-DNA hybridization using a defined and complete gene library of the viral genome (209 kbp) and by heteroduplex mapping. These experiments revealed the presence of repetitive DNA elements in the CIV genome, which are located in the EcoRI fragment H and in the EcoRI DNA fragment C at the coordinates 0.535 to 0.548 (EcoRI/Pstl DNA fragment, 2.7 kbp) and 0.920 to 0.944 (PvuII CIV DNA fragment L, 5.1 kbp), respectively. The DNA nucleotide sequence (2708 bp) of the EcoRI/Pstl subfragment was determined. The comparative analysis of the DNA sequences of this particular region of the viral genome with the DNA sequences of the PvuII DNA fragment L (5064 bp) revealed the presence of several DNA sequences within the EcoRI/Pstl subfragment of the EcoRI CIV DNA fragment H which show homology to DNA sequences of the PvuII DNA fragment L. For example, a DNA element (box A, 91 bp) is located at nucleotide positions 1981 to 2072 of the EcoRI CIV DNA fragment H which are complementary (greater than 90%) to the nine regions of the PvuII DNA fragment L (L-boxes 1 to 9). Furthermore heteroduplex mapping revealed the existence of a stem-loop structure (stem, 65 +/- 10 bp and loop, 652 +/- 80 bp) at the genome coordinates 0.571 to 0.582 (2.5 kbp, HindIII/EcoRI subfragment of the EcoRI CIV DNA fragment H). This indicates that an inverted repeat sequence is located at this region of the viral genome. The DNA nucleotide sequence of this subfragment was determined (2555 bp) which confirmed the data obtained from electron microscopy. An inverted repeat DNA sequence located at nucleotide positions 304 and 1011 is able to form this type of stem-loop structure.
Base Sequence↗
DNA nucleotide sequence analysis of the PvuII DNA fragment L of the genome of insect iridescent virus type 6 reveals a complex cluster of multiple tandem, overlapping, and interdigitated repetitive DNA elements.
The DNA nucleotide sequence of the PvuII DNA fragment L (0.920 to 0.944 map units (m.u.] of the genome (209 kbp) of insect iridescent virus type 6 was determined. The size of this DNA fragment was 5064 bp with a base composition of 39.79% G + C and 60.21% A + T. The DNA sequence contained many perfect direct repeats of sizes up to 145 bp. In addition to these repetitions, a cluster of four imperfect repetitive DNA elements (R1 to R4) with a complex structural arrangement was detected. R1, R2, and R3 existed in duplicate (two boxes (B] between nucleotide positions 271 and 3466) and their size were as follows: R1-B1/B2 (567/568 bp), R2-B1/B2 (917/931 bp), and R3-B1/B2 (92/88 bp). The R4 repetitive element was found in 12 boxes (between bases 1301 and 4417), which were interrupted at nucleotide positions 1883 to 2236 and 3341 to 3587. These interruptions define three segments (S) harboring boxes B1 to B3 (S1), B4 to B8 (S2), and B9 to B12 (S3). The size of the individual boxes was found to be 239, 233, 107, 244, 222, 242, 242, 148, 240, 242, 242, and 102 bp for R4-B1 to B12, respectively. Five open reading frames (ORFs of 118 to 333 amino acid (AA) residues) were detected. The analysis of the amino acid sequences of the largest ORF revealed that the deduced amino acid sequence of the putative gene product contained two repetitions TR1 (three domains of 50 AA) and TR2 (two domains of 74 AA). Sequences of 43 amino acid residues of ORF 5 (160 to 202 AA) were homologous within the majority of ORFs. A consensus sequence-MANL(X)6 IGSSST(X)6 L(X)1 LGS(X)1 LQISG(X)2 L(X)1 VN- was found in all five ORFs. Although classical canonical and noncanonical transcriptional start signals were detectable, polyadenylation signals were not observed.
Amino Acid Sequence↗
Nucleotide sequence analysis of the env gene and its flanking regions of the human spumaretrovirus reveals two novel genes.
Recombinant clones that represent the 3' part of the genome of the human spumaretrovirus (foamy virus) were established from viral DNA and from DNA complementary to viral RNA. The recombinant clones were characterized by blot hybridizations and nucleotide sequence analysis. The deduced protein sequence of the clones at their 5' ends was found to be homologous to the 3' domain of retroviral reverse transcriptases. Downstream of a small intergenic pol-env region a long open reading frame of 985 amino acid residues was identified that according to its genomic location, size, glycosylation signals, and hydrophobicity profile closely resembles the lentiviral env genes. The spumaretroviral env gene is followed by two open reading frames, termed bel-1 and bel-2 which are located between env and the long terminal repeat region. The long terminal repeat of 1259 nucleotides is preceded by a polypurine tract and contains the canonical signal sequences characteristic for transcriptional regulation of retroviruses. The provisional classification of the spumaretrovirus subfamily is discussed.
Amino Acid Sequence↗
Genomic characterization of a poxvirus isolated from a child.
A poxvirus was isolated from a six-year-old girl. The comparative analyses of the genome of this isolate (H-CP-LSax) which were carried out using the restriction endonucleases BamHI, HindIII, KpnI, MluI, NcoI, SacI, and SmaI revealed that this isolate is a member of the genus orthopoxvirus. Since the girl had never been vaccinated against smallpox, and had close contact to domestic animals, including cats, rabbits and guinea pigs, the genome of H-CP-LSax virus was genetically analysed in comparison with other known orthopoxviruses. The analysis demonstrates clearly that the HindIII cleavage pattern of H-CP-LSax DNA is different from the HindIII DNA cleavage patterns of vaccinia virus, cowpox virus, rabbit poxvirus, cat poxvirus, ectromelia virus, and okapi poxvirus. Surprisingly, it was found that the HindIII and SmaI cleavage patterns of the DNA of one out of six elephant poxviruses which were analysed under the same conditions were virtually identical to the HindIII and SmaI cleavage patterns of H-CP-LSax DNA. Although SmaI and HindIII digestion of both virus genomes gave the same fragment patterns, the viral DNAs can be distinguished from each other by the restriction endonucleases SacI, BamHI, and KpnI, which also show high similarities in the fragmentation patterns of both viruses. The results obtained in this study indicate three possibilities concerning the origin of H-CP-LSax virus. Firstly that the H-CP-LSax virus originated from an unknown animal species. Secondly, that this virus is a variant of elephant poxvirus in which the HindIII and SmaI sites are extremely conserved, and finally that H-CP-LSax can be a recombinant virus of unknown origin.
Child↗
HSV-1 virulence for mice by the intracerebral route is encoded by the BamHI-L DNA fragment containing the cell fusion gene.
The phenotype of pathogenicity by direct intracerebral inoculation of herpes simplex virus type 1 (HSV-1) was mapped in the viral genome. This phenotype could be rescued by cotransfection of unit length HSV-1 DNA of an avirulent strain with the BamHI fragment L (0.70-0.738 map units) cloned from a virulent strain. The virulence function was localized in the 2.0 Kb NruI-BamHI fragment in the right-hand side of BamHI-L, the same region that encodes a virus cell-fusion gene (3). Transduction of virulence was linked with the phenotype of a larger plaque size. It is concluded that a neurovirulence function resides in the BamHI-L fragment of the HSV-1 genome, closely linked to the viral gene for cell fusion.
Animals↗
Molecular cloning and physical mapping of the genome of insect iridescent virus type 6: further evidence for circular permutation of the viral genome.
A defined and complete gene library of the Chilo iridescent virus (CIV) genome was established. The CIV DNA was cleaved with restriction endonucleases EcoRI, NcoI, SphI, and BamHI or double digested with BamHI/SalI and the resulting DNA fragments were inserted into the corresponding sites of the bacterial vectors pACYC184, pKm2, pL-ES-C3, and pAT153 using T4 DNA ligase. All cloned fragments were identified by digestion of the recombinant plasmids with different restriction enzymes and checked by hybridization of recombinant plasmid to viral DNA. This analysis revealed that sequences representing 100% of the viral genome were cloned into the EcoRI site of pACYC184. Although the CIV genome is linear, all 32 EcoRI fragments have been cloned directly. This suggests that the CIV genome is circularly permuted. In addition, NcoI(72%), SphI(40.7%), BamHI (11.6%), and BamHI/SalI(39.7%) DNA fragments of the viral genome were inserted into the corresponding sites of pKm2, pL-ES-C3, and pAT153, respectively. The physical map of the viral genome was constructed using the established gene library for restriction enzymes ApaI, BamHI, EcoRI, NcoI, SalI, and SmaI. Although the CIV genome is linear, this analysis revealed that the restriction maps of the viral genome are circular. This finding supports the hypothesis that the CIV genome is circularly permuted.
Cloning, Molecular↗
Identification and nucleotide sequence analysis of the repetitive DNA element in the genome of fish lymphocystis disease virus.
The genome of the fish lymphocystis disease virus (FLDV) was screened for the existence of repetitive DNA sequences using a defined and complete gene library of the viral genome (98 kbp) by DNA-DNA hybridization, heteroduplex analysis, and restriction fine mapping. A repetitive DNA sequence was detected at the coordinates 0.034 to 0.057 and 0.718 to 0.736 map units (m.u.) of the FLDV genome. The first region (0.034 to 0.057 m.u.) corresponds to the 5' terminus of the EcoRI FLDV DNA fragment B (0.034 to 0.165 m.u.) and the second region (0.718 to 0.736 m.u.) is identical to the EcoRI DNA fragment M of the viral genome. The DNA nucleotide sequence of the EcoRI FLDV DNA fragment M was determined. This analysis revealed the presence of many short direct and inverted repetitions, e.g., a 18-mer direct repetition (TTTAAAATTTAATTAA) that started at nucleotide positions 812 and 942 and a 14-mer inverted repeat (TTAAATTTAAATTT) at nucleotide positions 820 and 959. Only short open reading frames were detected within this region. The DNA repetitions are discussed as sequences that play a possible regulatory role for virus replication. Furthermore, hybridization experiments revealed that the repetitive DNA sequences are conserved in the genome of different strains of fish lymphocystis disease virus isolated from two species of Pleuronectidae (flounder and dab).
Animals↗
Determination of the nucleotide sequence flanking the deletion (0.762 to 0.789 map units) in the genome of an intraperitoneally avirulent HSV-1 strain HFEM.
Herpes simplex virus type 1 (HSV-1) strain HFEM which harbours a deletion of 4.1 kbp in its genome (0.762 to 0.789 map units, HpaI DNA fragment P of HSV-1), is apathogenic for mice and tree shrews by the intraperitoneal application route. The exact position of this deletion was determined by DNA sequence analysis. This analysis was performed using the recombinant plasmid pU18HSHF-XmI-B which harbours the flanking genome regions (0.752 to 0.762 and 0.789 to 0.7895 map units) of the deletion in the genome of HSV-1 HFEM, and the recombinant plasmids pU18HSF-XmI-B, pU18HSF-AS, and pHSF-BB-BsH-D, harbouring particular regions of the genome of the virulent HSV-1 strain F at the coordinates 0.752 to 0.761, 0.786 to 0.790, and 0.762 to 0.771, respectively. The comparison of the DNA sequence of this region with the DNA sequences of the corresponding genome regions of the pathogenic HSV-1 strain F and HSV-1 strain 17 showed that the 5' end of the deletion in the genome of HSV-1 HFEM starts at the nucleotide position 3774 of the BamHI DNA fragment B from HSV-1/17. This position is 71 bp upstream of the UL/RL junction of the HSV-1 genome. The 3' terminus of the deletion ends at the nucleotide position 7226 of the BamHI DNA fragment B from HSV-1/17. The position is within the incomplete ninth repetitive box (ACTCC-CACGCACCCCC) and is located 36 bp upstream of the 3' end of the IE 110 mRNA.
Animals↗
Molecular cloning of the genome of human spumaretrovirus.
DNA of human spumaretrovirus (HSRV) was cloned from both cDNA and from viral DNA into phage lambda and bacterial plasmid vectors. The recombinant plasmids harboring viral DNA were characterized by Southern blot hybridization and restriction mapping. Physical maps were constructed from cDNA and found to be colinear with the restriction maps obtained from viral DNA. The recombinant clones isolated contained viral DNA inserts which range in size from 2.2 kb to 15.4 kb. The recombinant clones allowed to construct a physical map of the complete HSRV provirus of 12.2 kb.
Cloning, Molecular↗
Influencing of ionizing radiation on herpes simplex virus and its genome.
The effects of cobalt-60 gamma-rays, 10 MeV electrons and 52 MeV deutrons on the survival of plaque-forming ability has been studied in various strains of herpes simplex virus (HSV). The results show that the D0 for the loss of plaque-forming ability in different HSV strains lies in the range 1-3 kGy. Irradiation of isolated HSV-1 DNA with cobalt-60 gamma-rays resulted in damage, as indicated by electrophoresis of purified viral DNA and by restriction endonuclease analysis, at doses of 1 kGy, with complete loss of structure at doses above 4 kGy. The infectivity of the irradiated naked DNA was lost at doses above 4 kGy, but after irradiation of the intact virus some plaque-forming ability was retained after doses of 10 or even 40 kGy. Thus the organization within the viral capsid may play a protective role by modifying the severity of the radiation damage, and preserving at least some degree of infectivity.
Cobalt Radioisotopes↗
Nucleotide sequence analysis of a cloned DNA fragment from human cells reveals homology to retrotransposons.
During molecular cloning of proviral DNA of human spumaretrovirus, various recombinant clones were established and analyzed. Blot hybridization revealed that one of the recombinant plasmids had the characteristic features of a member of the long interspersed repetitive sequences family. The DNA element was analyzed by restriction mapping and nucleotide sequencing. It showed a high degree of amino acid sequence homology of 54.3% when compared with the 5'-terminal part of the pol gene product of the murine retrotransposon LIMd. The 3' region of the cloned DNA element encodes proteins with an even higher degree of homology of 67.4% in comparison to the corresponding parts of a member of the primate KpnI sequence family.
Amino Acid Sequence↗
Analysis of the genome of molluscum contagiosum virus by restriction endonuclease analysis and molecular cloning.
Virions of molluscum contagiosum virus (MCV), a member of the poxviridae, were isolated directly from lesions of individual patients and characterized by restriction enzyme analysis. The comparative analysis of the cleavage patterns and Southern blot hybridization of 14 independently isolated virus samples revealed that MCV isolates can be classified into two different types. The majority of MCV isolated from clinically typical skin lesions (13 of 14) showed similar DNA cleavage patterns and were termed MCV type 1, whereas one isolate derived from a vaginal lesion showed a completely different DNA cleavage pattern and therefore was termed MCV type 2. For detailed investigation of the viral genome, a defined gene library of MCV DNA sequences was established. The Bam HI DNA fragments of the viral genome of MCV type 1 prototype isolate 1/80 was inserted into the bacterial plasmid vector pAT153. With the exception of terminal fragments (fragments A and B) of the viral genome, all other DNA fragments were cloned. All cloned Bam HI DNA fragments were individually identified by digestion of the recombinant plasmid DNA with different restriction enzymes and screened by hybridization of plasmid DNA to viral DNA.
Adult↗
Comparative analysis of the genomes of orthopoxviruses isolated from elephant, rhinoceros, and okapi by restriction enzymes. Brief report.
Orthopoxviruses from different zoo-kept mammalian species including Elephas maximus (8 isolates), Ceratotherium simum (1 isolate), and Okapia johnstoni (2 isolates) were characterized by restriction enzyme analysis of the viral genome. The four enzymes BamHI, MluI, NcoI, and SalI were found to be optimal for strain differentiation.
Animals↗
Insect iridescent virus type 6 induced toxic degenerative hepatitis in mice.
The toxic effect of insect iridescent virus type 6 - chilo iridescent virus - (CIV) was investigated using Balb/c mice (strain ByJ Ico and Kisslegg). The animals were inoculated with CIV intraperitoneally (1 X 10(9) to 9.2 X 10(11) TCID50/animal). The animals which were administered with 1 X 10(11) to 9 X 10(11) TCID50 of CIV per animal, developed acute clinical illness and died during 18 to 80 h post infection. Histopathological and electronmicroscopic examinations of the liver tissues of those animals which died and/or were sacrificed when moribund showed acute degenerative hepatitis leading to death. No evidence for viral replication was found in the liver cells affected. A mortality rate between 21.1% and 100% was recorded for CIV, depending on the strain and number of mice used and the dose of virus administered. The toxic effect of CIV was eliminated or reduced extensively using heat denaturation or treatment of CIV with sodium dodecylsulphate or proteinase K. This indicates that the nature of the factor causing toxic degenerative cell damage is a protein.
Animals↗
Concatemeric forms of intracellular Tupaia herpesvirus DNA.
Kinetics of Tupaia herpesvirus strain 2 (THV-2) DNA synthesis was measured by labeling with radioactive thymidine and determining the amount of newly synthesized DNA. Viral DNA synthesis reached a maximum between 24 to 36 hr postinfection, preceded by a transient stimulation of host cell DNA synthesis. Blot hybridizations of labeled recombinant plasmids harboring inserts of both genomic termini of THV-2 and of individual viral terminal DNA fragments to DNA of virus-infected cells and combined restriction enzyme analysis of labeled intracellular viral DNA revealed the presence of linear concatemeric and/or circular viral DNA molecules.
Animals↗
A sequence in HpaI-P fragment of herpes simplex virus-1 DNA determines intraperitoneal virulence in mice.
The virulence of herpes simplex virus-1 (HSV-1) strains by the intraperitoneal (ip) route of injection in mice depends on the presence of an intact sequence in the HpaI DNA fragment P within coordinates 0.762 to 0.787. Deletion of the HpaI-P region (e.g., strain HFEM) abrogates the ability of the virus to infect mice by the ip route without affecting pathogenicity by the intracerebral (ic) route. A recombinant virus (M1C1) derived from DNA of the HSV-1 HFEM strain and the MLUIDNA fragment (coordinates 0.761 to 0.796) spanning the HpaI-P sequence of the pathogenic strain F regained pathogenicity for mice by the ip route.
Animals↗