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PubMed1990

Oxophenarsine, an antisyphilis drug inhibits HIV-1-specific protein synthesis in acutely and persistently infected lymphocytes.

The development of drugs that can inhibit both acute and persistent anti-human immunodeficiency virus type 1 (HIV-1) infection is a major goal in the treatment of patients with acquired immunodeficiency syndrome (AIDS). Most of the anti-HIV-1 drugs reported thus far, such as azidothymidine (AZT), inhibit acute HIV-1 infection but have no antiviral effect against persistent infection. We report here that oxophenarsine (3-amino-4 hydroxyphenylarsineoxide hydrochloride), an antisyphilus drug inhibits HIV-1 production in acutely infected peripheral blood lymphocytes (PBL) and persistently infected T cells. In acutely infected PBL and H9 T cells, the drug is effective at concentrations as low as 0.07-0.15 micrograms/ml with no significant cytotoxicity at concentrations of 6.0 micrograms/ml or below. It does not inhibit HIV-1 reverse transcriptase at doses up to 60 micrograms/ml. The drug has no effect on HIV-1-specific DNA and RNA synthesis. However, it inhibits HIV-1 protein synthesis in both acutely and persistently infected cells.

Anti-Infective Agents↗
PubMed2001

Sensitivity and specificity of the complement fixation test for detection of cattle persistently infected with Anaplasma marginale.

The complement fixation (CF) test commonly is used to identify cattle infected with Anaplasma marginale prior to interstate or international movement. Estimates of the accuracy of the CF test in detecting animals persistently infected with A. marginale vary widely. In this study, the sensitivity and specificity of the CF test for detection of carrier animals was determined using serum from 232 cattle previously defined as A. marginale positive or negative by nested polymerase chain reaction methods and hybridization. Considering results from 2 independent laboratories and interpreting a 1:5 suspect reaction as positive, the best estimate of CF test sensitivity was 20%, with a specificity of 98%. Using a 1:10 cutoff, sensitivity decreased to 14% and specificity increased to 99%. Results of this study indicate that the CF test is ineffective for identifying cattle persistently infected with A. marginale and thus is inadequate for anaplasmosis regulatory and surveillance programs.

Anaplasma↗
PubMed1993

Persistent infection of human erythroblastoid cells by poliovirus.

Human erythroblastoid K562 cells have been recently described as a relatively nonpermissive host for poliovirus replication. During investigations of virus-induced cytopathic effects in this cell line, we discovered that poliovirus easily established persistently infected cultures in K562-Mu cells. In these cultures, most cells remained viable, with overall viability continuously maintained between 67 and 92% over 3 months. Infected K562 cells continued to grow, usually without any major periods of crisis in the culture or large diminutions in cell growth rate. K562-Mu cells produced a slower onset of virus production than observed in HeLa cells, and virus titers in culture supernates rapidly stabilized at levels between 10(5) and 10(6) PFU/ml. In infectious center or limiting dilution assays, only about 10% of K562 cells produced infectious virus after 2 days. However, when assays were extended to 3 to 5 days, most K562 cells in the culture scored as infectious centers, suggesting productive infection of all cells in the culture with delayed kinetics of virus production. Cultures of infected K562 cells could not be cured of virus by prolonged incubation with high-titer neutralizing antibody. Pulse-label SDS-PAGE analysis of infected cultures detected moderate levels of virus protein synthesis which peaked at 9-12 hr postinfection; however, little or no shutoff of host protein synthesis was observed at any time point during infection. Immunoblot analysis with antisera to the p220 subunit of eIF-4F demonstrated extensive but incomplete cleavage of p220 in infected K562 cells at times which correlated with peak viral protein synthesis. Taken together, the results demonstrate a persistent infection in which host cell shutoff does not occur despite significant viral protein synthesis and extensive early degradation of p220.

Adsorption↗
PubMed2000

Single amino acid differences are sufficient for CD4(+) T-cell recognition of a heterologous virus by cattle persistently infected with bovine viral diarrhea virus.

Cattle that are persistently infected (PI) with one strain of bovine viral diarrhea virus (BVDV) can resolve infection with a second, antigenically heterologous strain but not the homologous strain. Since CD4(+) T cells are thought to be critical for the resolution of acute BVDV infection (Howard et al., 1992, Vet. Immunol. Immunopathol. 32, 303-314), we have examined the recognition of a heterologous virus (NADL) by CD4(+) T cells from Pe515-PI animals. The immune response of non-PI control cattle challenged with NADL or Pe515ncp was strain cross-reactive, whereas Pe515-PI animals responded to NADL only. The immune repertoire of both groups included NS3, which differs by approximately 1% (9/683) amino acids between these two viruses. Lymphoproliferative responses to proteins and synthetic peptides corresponding to three nonconservative differences in NS3 demonstrated that CD4(+) T cells from non-PI control animals responded well to proteins but poorly to the peptides from both viruses. In contrast, PI animals were responsive to heterologous proteins and peptides but nonresponsive to the homologous equivalents. A single amino acid difference between the two sequences was sufficient to allow responsiveness.

Amino Acid Sequence↗
PubMed1997

Dexamethasone inhibits virus production and the secretory IgA response in oesophageal-pharyngeal fluid in cattle persistently infected with foot-and-mouth disease virus.

Cattle persistently infected with foot-and-mouth disease virus were treated with dexamethasone to suppress the immune system in an attempt to influence the level of virus recovery from oesophageal pharyngeal (probang) samples. Twelve carrier cattle were assigned to one of three groups: control; 0.1 mg/kg dexamethasone; and 0.5 mg/kg dexamethasone. Groups 2 and 3 were injected intramuscularly three times weekly for 3 weeks with dexamethasone between days 33 and 56 post-infection with foot-and-mouth disease virus (FMDV). Cattle in both groups developed a leucocytosis, neutrophilia and lymphopenia. The secretory IgA response to FMDV infection was inhibited following, but not during, dexamethasone treatment between days 70 and 98 post-infection (P < 0.05). FMDV recovery from probang samples was reduced between days 40 and 64 post-infection (P < 0.05) during treatment with either 0.1 or 0.5 mg/kg dexamethasone. Following cessation of dosing with dexamethasone virus recovery returned to control levels. These observations suggest dexamethasone inhibits shedding of FMDV in a reversible manner which may be related to its immunosuppressive, anti-inflammatory or physiological actions.

Animals↗
PubMed2005

Neuroblastoma cell-adapted yellow fever virus: mutagenesis of the E protein locus involved in persistent infection and its effects on virus penetration and spread.

Persistent infection of mouse neuroblastoma NB41A3 cells with yellow fever 17D virus generates viral variants which exhibit defective cell penetration, poor cell-to-cell spread, small plaque size and reduced growth efficiency, caused by substitution of glycine for aspartic acid or glutamic acid at positions 360 and 362 in the envelope protein. These positions occur within a charge cluster, Asp360-Asp361-Glu362, located in domain III, near its interface with domain I. To characterize further the molecular basis for the variant phenotype, a series of mutant viruses containing substitutions at position 360, 361 and 362, were studied for effects on the cell culture properties typical of the neuroblastoma-adapted variant. Most substitutions at position 360 gave rise to viruses that were very defective in cell penetration, growth efficiency and cell-to-cell spread, whereas substitution with glutamic acid yielded a virus indistinguishable from parental yellow fever 17D. Substitution with lysine was not tolerated and substitution with asparagine resulted in frequent wild-type revertants. A glycine residue was not tolerated at position 361, but substitution at 362 yielded a small plaque virus, similar to the effect of substitution at position 360. These data indicate that the yellow fever virus E protein contains a locus within domain III where a negative-charge cluster is important for optimal function of this domain in virus-cell interactions beyond the stage of virus attachment. Modelling predictions suggest that the mutations alter the local properties of the loop within domain III, and may compromise interactions of this domain with an adjacent region of domain I during conformational changes that occur in the E protein in association with virus entry.

Amino Acid Substitution↗
PubMed1993

Enhanced production of morbillivirus gene-specific RNAs following induction of the cellular stress response in stable persistent infection.

Previous in vitro work demonstrated the incorporation of the major inducible 70k heat shock protein (i.e., 72k HSP) into the biologically active light nucleocapsid (L-NC) variant of canine distemper virus (CDV). Here, in vitro induction of the cellular stress response, characterized by elevated cytoplasmic and intranuclear 72k HSP, enhanced L-NC expression in mink lung cells supporting stable persistent infection by raccoon-origin CDV. Increases in L-NC were correlated to increased viral RNA production in cell-free transcriptional assays. The enhanced production of viral transcripts within infected cells following stress response induction was confirmed by slot blot and Northern blot analysis of total cellular RNA and was reflected in increased total viral protein production. Post-shock increases in viral fusion (F) gene transcripts and F protein were associated with dramatic increases in viral cytopathic effect. Modest induction of cell-free infectious viral progeny was also documented. A similar effect of the cellular stress response upon viral protein expression, cytopathic effect, and cell-free infectious progeny release was demonstrated in murine neuroblastoma cells persistently infected with a canine CDV isolate. Alterations of the persistent viral phenotype were independent of the specific mechanism of stress-response induction (i.e., heat or sodium arsenite), supporting the role of the stress response and not a particular stressor in mediating these changes. These results document the ability of the cellular environment to alter persistent viral RNA metabolism, thereby altering the infection phenotype.

Animals↗
PubMed1991

Progeny of sheep persistently infected with border disease virus.

Most lambs affected with border disease die early in life but those which survive gradually loose their body tremors and their fleece abnormalities become less clear. Seven female lambs persistently infected with border disease virus were reared to maturity and bred from when they were 2 to 3 years old. Two failed to conceive but five gave birth to 6 live lambs with clinical signs of border disease characterized by hairy and pigmented fleece with or without body tremors. The epidemiological significance of persistently infected sheep is discussed.

Animals↗
PubMed1989

Persistent infections of a field strain of rabies virus in murine neuroblastoma (NA-C1300) cell cultures.

Rabies virus from the brain of a striped skunk (Mephitis mephitis) from Ontario was inoculated into murine neuroblastoma (NA-C1300) cell cultures. These cultures were incubated and the cells were subcultured every three to four days. The presence of viral antigen in the cell cultures was monitored by direct immunofluorescent staining and in the culture fluids by titration in either baby hamster kidney (BHK/C13) or NA cells or in experimental mice. The virus-infected NA cultures evolved from an initial high viral concentration in supernatant fluid through a period of decreasing titers of infectious virus in the supernatant fluids to a final phase where no infectious virus has been found following cell culture and animal inoculation methods attempted although the persistently infected cells remained 95-100% viral nucleocapsid antigen-positive. Possible mechanisms involved in the perpetuation of this infection are discussed. This is the first report of a persistent infection of cell cultures by a field strain of rabies virus.

Animals↗
PubMed1995

Effect of non-H-2-linked genes on anti-virus immune responses and long-term survival in mice persistently infected with E-55+ murine leukemia virus.

We have previously demonstrated that BALB/c-H-2k (BALB.K) mice are susceptible to the development of thymic lymphoma induced by E-55+ murine leukemia virus (MuLV). In the present studies, C57BL/10-H-2k (B10.BR) mice were found to be resistant to E-55+ MuLV-induced lymphoma despite the fact that these mice become persistently infected. This resistance to lymphomagensis is mediated by the anti-virus immune response since immunosuppressed mice progress to develop disease. The protective immune response in B10.BR mice is bimodal with respect to time after virus infection. The early immune response results in a dramatic decrease in the number of virus-infected cells within 4-8 weeks after infection. This decrease in virus-infected cells occurs in immunocompetent mice from strains that are either resistant (B10.BR) or susceptible (BALB.K) to E-55+ MuLV-induced disease. Subsequently, susceptible mice develop an increase in infected cells, whereas no increase in infected cells occurs in resistant mice despite the fact that they are persistently infected. This later phase of resistance in B10.BR appears to be mediated by T cells. Since B10.BR and BALB.K both express the H-2k haplotype, resistance appears to be mediated by a non-H-2-linked gene(s). (BALB.K x B10.BR)F1 mice are resistant to disease development, indicating resistance is a dominant trait.

Animals↗
PubMed1984

Genomic alterations associated with persistent infections by equine infectious anaemia virus, a retrovirus.

The unique periodic nature of equine infectious anaemia (EIA) is believed to result from the ability of the infecting virus. EIAV, to undergo relatively rapid antigenic variations which circumvent host immune responses resulting in distinct virus populations in sequential clinical episodes in the persistently infected horse. This model was examined by oligonucleotide mapping comparisons of the RNA genomes of selected isolates of EIAV. Variations in oligonucleotide maps could be reproducibly demonstrated (i) after adaptation of the laboratory strain of EIAV to replication in a pony, (ii) after serial passage of virus between two ponies, and (iii) after a prolonged persistent infection in a single pony. In the latter case, the two isolates examined were recovered from different clinical episodes and were shown to be antigenic variants. In contrast, no variations in RNA structure could be detected in oligonucleotide maps of virus isolated after prolonged passage in tissue culture. Thus, these results support our concept that EIAV is a highly mutable virus, which may given rise to antigenic variants in the presence of immune pressures. The degree of variation observed between oligonucleotide maps is similar to that observed previously between variants of visna virus. These similarities between EIAV an visna suggest that genomic point mutations producing antigenic variants may be a more important mechanism of retrovirus persistence than was previously recognized.

Animals↗
PubMed1997

Generation of cytopathogenic subgenomic RNA of classical swine fever virus in persistently infected porcine cell lines.

Two biological clones (A.1 and B.2) of the classical swine fever virus strain Alfort/187 and the recombinant virus vA187-1, derived from a cDNA clone of Alfort/187, were used to establish persistently infected cultures of the swine kidney cell lines SK-6 and PK-41. It was found that 100% of the cells in the passaged cultures were positive for viral antigen throughout the course of the experiment. Additionally, supernatants collected upon passaging of the cells continuously contained high titers of infectious virus. In six separate cultures persistently infected with either the biological clones or the recombinant virus, a cytopathic effect occurred spontaneously between passage 8 and 94. The cytopathogenic agent in the supernatants of these cultures could be passaged repeatedly, suggesting the generation of a mutant virus. Analysis of RNA from such cultures revealed the presence of a subgenomic viral RNA of approximately 8 kilobases (kb). In all six cases, this RNA had an identical internal deletion of 4764 nucleotides, including the region coding for all structural proteins. The subgenomic RNA replicated and was packaged in the presence of wild-type virus. Cells infected with cytopathogenic virus contained increased amounts of the viral protein NS3 thought to be involved in pestivirus cytopathogenicity.

Animals↗
PubMed1979

Regulation of persistent infection with herpes simplex virus in vitro by hydrocortisone.

About 1% of Raji cells showed sensitivity to herpes simplex virus type 2 (HSV-2) infection when tested by infectious center assays or immunofluorescence tests, and the percentage did not change during cell passage. The addition of hydrocortisone to Raji cells persistently infected with HSV-2 caused a marked increase in virus production and in the number of HSV-producing cells. In the case of HSV-1, it was shown that the addition of hydrocortisone was required to maintain persistent infection. These observations suggest that control of replication of HSV-1 and HSV-2 in these cells is regulated by different mechanisms.

Burkitt Lymphoma↗
PubMed1989

Antigenic diversity of bovine viral diarrhea-mucosal disease (BVD-MD) viruses recently isolated from persistently infected cattle and mucosal disease, and serologic survey on bovine sera using antigenically different BVD-MD viruses.

Twenty nine recent isolates of bovine viral diarrhea-mucosal disease (BVD-MD) virus, 17 from persistently infected cattle and 12 from mucosal disease, were compared antigenically with the reference strains by a serum neutralization test. The reference viruses were divided into 2 groups, tentatively designated as N and K, based on the antigenic relationships in the cross-neutralization test. Antigenic properties of recent isolates were considerably different with the sources of virus isolation. Seventeen isolates recovered from persistently infected cattle were divided into 3 groups in the neutralization test using antisera to the reference strains; 12 and 2 were considered as the possible members of groups K and N, respectively, and the others belonged to neither group. On the other hand, 10 of 12 isolates recovered from mucosal disease were considered as the possible members of group N, and the others were classified into neither group. Interestingly, none of BVD-MD viruses isolated from cases of mucosal disease belonged to group K. The results of serologic survey on sera collected from 713 cattle at the Hokkaido provinces in 1974 to 1988 indicated that infections of cattle with BVD-MD viruses other than group K were prominent before 1981. Cattle infected with group K BVD-MD virus were first detected in 1982, and increased in number thereafter. The results obtained in this study suggested that BVD-MD viruses with various antigenic properties spread widely among cattle herds, and also a possibility that clinical manifestations in cattle infected with BVD-MD viruses may differ with their antigenic properties.

Animals↗
PubMed1986

The persistent infection of a canine thymus cell line by equine infectious anaemia virus and preliminary data on the production of viral antigens.

Equine infectious anaemia virus (EIAV) was adapted to the Cf2Th cell line, a heterologous malignant line from canine thymus. A persistent infection was monitored for 100 serial passages by demonstrating the presence of virus and viral antigens at each 10th passage by electron-microscopy, immunodiffusion and immunofluorescence. Chromosome analysis of EIAV-infected cells indicated they had a karyotype resembling the control cells of similar passage history. Virus-infected cells, grown in roller cultures for 65 days without subculturing, continuously produced viral antigens into supernatant fluids which were harvested every 3-4 days. Antigen peaks were observed at approximately 12-day intervals. Immunoprecipitin lines of identity were demonstrated between ether-extracted antigens from virus-infected canine cell line and known positive EIAV antigen extracted from infected equine spleen and a commercial source. Replication of a non-oncogenic retrovirus (EIAV) resulted in the continuous release of viral antigens from a persistently infected and infinite cell line.

Animals↗
PubMed2000

Electron microscopy of buffalo green monkey kidney cells persistently infected with hepatitis A virus and immunolocalization of HAV antigens.

Studies were carried out to analyse the ultrastructural changes and the distribution of hepatitis A virus (HAV)/antigens at subcellular level in buffalo green monkey kidney (BGMK) cells persistently infected with HM-175 strain of HAV. HAV infected BGMK cells showed distinct abnormalities in the endoplasmic reticulum and cytoplasmic membrane as compared to uninfected cells. The abnormalities were characterized by wavy arrays, structures like myelin, annulate lamellae, cytoplasmic inclusion bodies and vesicles. The wavy arrays within the cytoplasm of the host cells appeared to represent degenerating membranes. A complex myelin like body was found in close association with a group of virus like particles. Annulate lamellae like structures involving single paired membrane were detected infrequently whereas the cytoplasmic vesicles were numerous in these cells. An indirect immunogold technique was utilized to localize the HAV antigenin infected cells. A high density immunogold label for HIV like particles was predominantly detected in cytoplasmic vesicles. These results suggest a strong association of membrane substructure in vesicle forms with the compartmentalized replication of HAV within persistently infected host cells.

Animals↗
PubMed1994

Genetic variation of foot-and-mouth disease virus during persistent infection in cattle.

Genetic variation of foot-and-mouth disease virus O1 Campos has been analyzed in consecutive isolates recovered over a one- or two-year period from four cattle with experimental persistent infection. Comparisons of RNase T1 two-dimensional maps and nucleotide sequences of the VP1-coding region revealed a continual, although irregular, increase in the fixation of mutations as the infection progressed. Most changes were not conserved in consecutive isolates. These results, together with the substantial rates of genomic variation observed between some pairs of strains recovered at close time periods, suggested the coexistence of heterogeneous populations in which variants evolve independently from each other, and predominate at irregular time intervals. Furthermore, non-related patterns of variation were observed in the four animals. Similarly, genetic diversity of representative strains from major serotype O outbreaks in endemic disease regions of southeastern Brazil and central eastern Argentina which occurred between 1958 and 1983, suggested that outbreak strains are also likely to represent fluctuations of heterogeneous populations which evolve independently from each other. The possible role of persistent infections in the introduction of variant populations in the field is discussed.

Amino Acid Sequence↗
PubMed2005

Bovine viral diarrhea virus in alpaca: abortion and persistent infection.

An alpaca herd in eastern Ontario experienced vague signs of illness, including anorexia and lethargy in 9 animals, 2.5 months after the addition of a chronically ill cria and his dam to the farm. Subsequently 2 alpaca had early pregnancy loss; one aborted at 5.5 months gestation and the other at 7 months gestation. Seventeen were found to have serum antibody to bovine viral diarrhea virus (BVDV), with highest titers to BVDV type 1. The fetus that was aborted at 5.5 months gestation, 3 months after the clinical outbreak, was found to be positive for BVDV on immunohistochemical staining, and noncytopathic BVDV type 1b was isolated. Of the 13 cria born alive that season, a single male underweight alpaca cria, born 9 months after the clinical illnesses, was infected with BVDV type 1b. The cria was positive for BVDV at birth, at 3 and 26 days of age and continued to be positive for noncytopathic BVDV using virus isolation, nested reverse transcription PCR, antigen detection ELISA, and immunohistochemical staining until euthanasia at 46 days of age. The cria remained serum antibody negative to both BVDV type 1 and type 2. A diagnosis of persistent infection was made. This is the first report describing persistent infection with BVDV in an alpaca cria.

Abortion, Veterinary↗