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At least 19 recordsLinked to original sources
PubMed2025

Establishment of reverse genetics systems for Colorado tick fever virus.

The Colorado tick fever virus (CTFV), which has 12-segmented double-stranded RNA genomes, is a pathogenic arbovirus that causes severe diseases in humans. However, little progress has been made in the analysis of replication mechanisms and pathogenicity. This virological constraint is due to the absence of a reverse genetics system for CTFV; therefore, we aimed to establish the system. Initially, the efficacy of CTFV replication was investigated in various cell lines. CTFV was found to grow in many cell types derived from different hosts and organs. Subsequently, BHK-T7 cells stably expressing T7 RNA polymerase were transfected with plasmids encoding each of the 12 CTFV gene segments, expression plasmids encoding all CTFV proteins, and a vaccinia virus RNA-capping enzyme. Following transfection, the cells were co-cultured with Vero or HeLa cells. Using this system, we rescued monoreassortants and recombinant viruses harboring peptide-tagged viral proteins. Furthermore, an improved system using Expi293F cells expressing T7 RNA polymerase was established, which enabled the generation of recombinant reporter CTFVs. In conclusion, these reverse genetics systems for CTFV will greatly contribute to the understanding of viral replication mechanisms, pathogenesis, and transmission, ultimately facilitating the development of rational treatments and candidate vaccines.

Animals↗
PubMed1989

Co-circulation of multiple Colorado tick fever virus genotypes.

Colorado tick fever (CTF) virus, family Reoviridae, genus Orbivirus, contains 12 genes distinguishable by polyacrylamide gel electrophoresis (PAGE). Multiple genotypes of CTF virus were isolated at 3 field sites in Colorado in 1985. Five genotypes were found at Campos Cabin, 2 at Drake, and 6 at Rocky Mountain National Park. Virus isolations were made in 1985 from 6 patients with CTF. These isolates were distinct from each other and the field isolates. Although the CTF isolates were different by PAGE profile, the majority of the 12 genes were highly conserved among the 1985 isolates and a Florio isolate (FMA). Only genes 4 and 6 were variant among the 1985 CTF isolates and FMA, and no unique genes were identified. In 1986, a follow-up field survey was done at the Campos Cabin site. Of the 3 CTF PAGE genotypes obtained, 2 exhibited PAGE profiles which were different from the 1985 isolates. One isolate may have resulted from the reassortment of genes from 2 of the isolates circulating at Campos Cabin in 1985.

Animals↗
PubMed1987

Antigenic variants of Colorado tick fever virus.

Twenty strains of Colorado tick fever (CTF) virus, isolated from ticks, mammals and humans, and two antigenic relatives of CTF virus were compared in cross-neutralization tests. Viruses were tested using single-inoculation sera prepared in hamsters. Antigenic variation, as measured by differences detected in the neutralization test, was noted among the virus isolates identified as strains of CTF virus. The virus strains isolated from humans appeared to vary the most in serological reactions. The two antigenic relatives of CTF virus are clearly distinct from strains of CTF and are different from each other. Antigenic relationships between these two viruses were established using two sets of single-inoculation antisera and both complement fixation and neutralization tests. Six distinct antigenic variants of CTF virus isolated from humans and the virus strain from ticks (75V1906) that showed the least antigenic variation, were tested against 49 coded serum pairs from clinically diagnosed cases of CTF. Significant differences were noted in the number of convalescent-phase sera that reacted with each virus strain and in the number of seroconversions observed with each test virus strain. Convalescent phase sera that reacted with multiple virus strains often varied significantly in antibody titre from one virus strain to another. This indicates that, in some instances, antibody was probably produced in response to infection by different antigenic variants of CTF virus.

Animals↗
PubMed1976

Eyach--an arthropod-borne virus related to Colorado tick fever virus in the Federal Republic of Germany.

From Ixodes ricinus ticks collected in a tick-borne encephalitis focus in Baden-Württemberg, an agent pathogenic exclusively for suckling mice was isolated. Obviously a virus, it passed membrane filters with a pore width of 200 nm. It was resistant to ether and sodium deoxycholate, but not to chloroform. In the complement fixation test it showed a close, and in the neutralisation test a more one-sided relationship to Colorado tick fever virus.

Animals↗
PubMed1981

Experimental Colorado tick fever virus infection in Colorado mammals.

One hundred and twenty-four small mammals of six species were inoculated with three strains of Colorado tick fever (CTF) virus to define viremia and neutralizing (N) antibody responses. Adult Eutamias minimus and Eutamias embrinus, and juvenile Peromyscus maniculatus and Spermophilus lateralis, were highly susceptible to development of viremic infection. Adult S. lateralis and P. maniculatus were moderately susceptible (greater than or equal to 50% viremic). Five Sylvilagus nuttalli did not become viremic following experimental inoculation. Spermophilus richardsoni was also relatively resistant (less than or equal to 50% viremic). The longest duration of viremia (mean 15.8 days) and highest peak viremia levels (mean peak titer 10(3.9 plaque-forming units per ml) occurred in E. minimus. Adult E. umbrinus, juvenile S. lateralis, and juvenile P. maniculatus had moderate viremias. Adult S. lateralis and S. richardsoni often had short viremias during which virus was only intermittently detectable. N antibody production was most rapid in E. minimus in comparison with other species. In addition, N antibody persisted for 1 year in this species. In other species, many animals lost detectable antibody 5-11 months after infection. No significant differences were found in patterns of infection between three CTF virus strains. We conclude that of the six species inoculated, E. minimus is the best experimental host for CTF virus.

Animals↗
PubMed2004

Termination and read-through proteins encoded by genome segment 9 of Colorado tick fever virus.

Genome segment 9 (Seg-9) of Colorado tick fever virus (CTFV) is 1884 bp long and contains a large open reading frame (ORF; 1845 nt in length overall), although a single in-frame stop codon (at nt 1052-1054) reduces the ORF coding capacity by approximately 40 %. However, analyses of highly conserved RNA sequences in the vicinity of the stop codon indicate that it belongs to a class of 'leaky terminators'. The third nucleotide positions in codons situated both before and after the stop codon, shows the highest variability, suggesting that both regions are translated during virus replication. This also suggests that the stop signal is functionally leaky, allowing read-through translation to occur. Indeed, both the truncated 'termination' protein and the full-length 'read-through' protein (VP9 and VP9', respectively) were detected in CTFV-infected cells, in cells transfected with a plasmid expressing only Seg-9 protein products, and in the in vitro translation products from undenatured Seg-9 ssRNA. The ratios of full-length and truncated proteins generated suggest that read-through may be down-regulated by other viral proteins. Western blot analysis of infected cells and purified CTFV showed that VP9 is a structural component of the virion, while VP9' is a non-structural protein.

Animals↗
PubMed2003

Recombinant VP7-based enzyme-linked immunosorbent assay for detection of immunoglobulin G antibodies to Colorado tick fever virus.

VP6, VP7, VP9, VP10, VP11, and VP12 of Colorado tick fever virus (CTF virus), a virus member of the genus Coltivirus, family Reoviridae, were expressed in bacteria with the pGEX-4T-2 vector. A partial sequence of VP7 (designated pVP7) was chosen to elaborate an enzyme-linked immunosorbent assay (ELISA) for detecting anti-CTF virus immunoglobulin G (IgG) antibodies in humans. This was based on two observations: (i) among all expressed proteins, pVP7 showed the highest immunoreactivity to an anti-CTF virus hyperimmune ascitic fluid; (ii) to provide the highest selectivity of antibody detection, the expressed sequence was chosen within a region which is highly divergent (49% amino acid identity) from the homologous sequence of another coltivirus, the Eyach virus. The pVP7 ELISA was evaluated with 368 serum samples from French blood donors and found to provide 98.1% specificity. Assays with the Calisher set of human serum samples, positive for anti-CTF virus antibodies (C. H. Calisher, J. D. Poland, S. B. Calisher, and L. A Warmoth, J. Clin. Microbiol. 22:84-88, 1985), showed that the pVP7 ELISA provided 100% sensitivity for the tested population. After elaboration of recombinant-protein-based ELISAs for diagnosis of infections with members of the viral genera Orbivirus, Orthoreovirus, and Rotavirus, it was shown that a recombinant protein could be used to detect antibodies to the human pathogen Colorado tick fever virus.

Antibodies, Viral↗
PubMed2000

Sequence determination and analysis of the full-length genome of colorado tick fever virus, the type species of genus Coltivirus (Family Reoviridae).

The Colorado tick fever virus (CTFV) is the type species of genus Coltivirus, family Reoviridae. Its genome consisting of 12 segments of dsRNA was completely sequenced. It was found to be 29,174 nucleotides long (the longest of all Reoviridae genomes characterized to date). Conserved sequences at the 5' end (SACUUUUGY) and at the 3' end (WUGCAGUS) of the 12 segments were identified. The analysis of the putative proteins deduced from the nucleotide sequences permitted to identify functional motifs. In particular, the VP1 was identified unambiguously as the viral RNA dependent RNA pylmerase (RDRP) (VP1pol), with a GDD located at a similar position to Reoviridae RDRPs. In other genes, RGD cell-binding, NTPAse, single strand binding protein and kinase motifs were identified. Comparison with Reoviridae proteins showed significant similarities to RDRPs (CTFV-VP1) and sigma C protein of orthoreovirus (CTFV-VP6). Similarities to nonviral enzymatic proteins, such as methyltransferases, NTPAses, RNA replication factors, were also identified.

Animals↗
PubMed1964

PLAQUE ASSAY PROCEDURE FOR COLORADO TICK FEVER VIRUS.

Deig, E. Frank (University of California, Berkeley), and H. M. S. Watkins. Plaque assay procedure for Colorado tick fever virus. J. Bacteriol. 88:42-47. 1964.-A reproducible plaque assay procedure is described for the quantitation of Colorado tick fever virus in a cell line established from embryonic hamster tissue. Under the best conditions, plaques approximately 4 mm in diameter were formed after incubation at 37 C of 4 to 6 days. Several environmental variables in the procedure were studied. Efficiency was increased markedly by combining the virus during adsorption with serum proteins, and by carrying out this step at 25 C rather than at 37 C. The overlay medium used contained metabolites which promoted cell viability for periods greater than 1 week, and allowed plaques to develop. Plaque formation was relatively insensitive to a variation in pH between 7.1 and 8.1 (with optimal concentrations of bicarbonate). However, plaque development was inhibited with medium containing greater than 0.22% bicarbonate (at optimal pH), or when the initial pH was less than 7.0.

Adsorption↗
PubMed1985

Diagnosis of Colorado tick fever virus infection by enzyme immunoassays for immunoglobulin M and G antibodies.

An immunoglobulin M (IgM) capture enzyme immunoassay technique was adapted for the detection of antibody to Colorado tick fever virus in sera from 84 individuals for whom diagnosis had been confirmed by virus isolation or neutralization test. Titers were compared with those for IgG and neutralizing antibodies in these Colorado tick fever cases. IgM antibody titers were higher than neutralizing antibody titers, but neither appeared until 1 to 2 weeks after the onset of illness. Neutralizing antibodies were detected earlier than IgM antibodies, and both were detected with greater frequency than IgG antibodies. Late-convalescent-phase sera contained both neutralizing and IgG antibodies, but IgM was all but undetectable by 2 months after onset. Although the neutralization test may remain the serological test of choice, the enzyme immunoassay for IgM antibody offers a simple and more rapid method of serodiagnosis; the enzyme immunoassay is, however, less sensitive than the neutralization test. Furthermore, because there was a sharp decline in IgM antibody after 45 days, the presence of IgM antibody in a single serum sample provides a basis for the presumptive serodiagnosis of recent Colorado tick fever virus infection.

Adolescent↗
PubMed1975

Teratogenic effects of Colorado tick fever virus in mice.

The teratogenic effects in mice of Colorado tick fever (CTF) virus, a human pathogen, were demonstrated. Various routes of inoculation, gestational periods, and dosages of virus were studied. The incidence of stillbirth and neonatal death increased significantly after inoculation of pregnant mice with CTF virus. CTF virus crossed the placenta and replicated to high titers in the placental and fetal tissues. Mice that survived the first six weeks of life had neutralizing antibody to CTF virus. Neutralization tests, in which pregnant mice were inoculated with both CTF virus and specific antiserum to CTF virus, validated the teratogenic effects of CTF virus in mice.

Animals↗
PubMed1978

The development of Colorado tick fever virus within cells of the haemopoietic system.

Electron microscopic examination of haemopoietic liver tissue from mice infected in utero or when newborn showed inclusions of Colorado tick fever virus within erythroblasts, reticulocytes and erythrocytes. Inclusions were also seen within erythroblastoid cells undergoing mitosis. Other evidence of virus replication within erythropoietic cells was the presence of intracytoplasmic and intranuclear fibres, which have been shown to be associated with Colorado tick fever virus replication. The findings reported here support the hypothesis that virus replication within infected erythropoietic cells occurs concurrently with differentiation of the infected cell, resulting in the presence of virions within erythrocytes.

Animals↗
PubMed1993

Replication of Colorado tick fever virus within human hematopoietic progenitor cells.

Significant neutropenia, as well as thrombocytopenia and a mild anemia, occurs in patients infected with Colorado tick fever virus. In this study, human bone marrow CD34+ cells and KG-1a cells, a human hematopoietic progenitor cell line, were infected in vitro with Colorado tick fever virus. The time course and morphological appearance of viral replication in human progenitor cells were similar to those seen in erythroblasts and in HEL cells and suggest one possible mechanism for the clinical hematologic findings.

Animals↗