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differential requirements for alternative splicing and nuclear export functions of equine infectious anemia virus rev protein.the rev protein of equine infectious anemia virus (erev) exports unspliced and partially spliced viral rnas from the nucleus. like several cellular proteins, erev regulates its own mrna by mediating an alternative splicing event. to determine the requirements for these functions, we have identified erev mutants that affect rna export or both export and alternative splicing. mutants were further characterized for subcellular localization, nuclear-cytoplasmic shuttling, and multimerization. none o ...19989632773
hyperglobulinemia and lymphocyte subset changes in naturally infected, inapparent carriers of equine infectious anemia virus.to determine blood protein concentration, immunoglobulin concentration, and lymphocyte profiles in equine infectious anemia virus (eiav) seropositive, naturally infected horses without clinical signs of disease.19989706205
analysis of the polymerization kinetics of homodimeric eiav p51/51 reverse transcriptase implies the formation of a polymerase active site identical to heterodimeric eiav p66/51 reverse transcriptase.homodimeric eiav p51/51 and heterodimeric eiav p66/51 reverse transcriptase were purified in order to compare the different modes of dna synthesis supported by the enzymes. analysis of the dimerization behavior of the eiav enzymes indicates that the dimer stability of eiav reverse transcriptase enzymes is higher than that of their hiv-1 reverse transcriptase counterparts. eiav p51/51 polymerizes dna distributively whereas dna synthesis by eiav p66/51 is processive. steady-state and pre-steady-st ...19989724526
disease induction by virus derived from molecular clones of equine infectious anemia virus.equine infectious anemia virus (eiav), a macrophage-tropic lentivirus, causes persistent infections of horses. a number of biologic features, including the rapid development of acute disease, the episodic nature of chronic disease, the propensity for viral genetic variation, and the ability for many infected animals to eventually control virus replication, render eiav a potentially useful model system for the testing of antiretroviral therapies and vaccine strategies. the utility of the eiav sys ...19989420249
development and characterization of an in vivo pathogenic molecular clone of equine infectious anemia virus.an infectious nonpathogenic molecular clone (19-2-6a) of equine infectious anemia virus (eiav) was modified by substitution of a 3.3-kbp fragment amplified by pcr techniques from a pathogenic variant (eiav(pv)) of the cell culture-adapted strain of eiav (eiav(pr)). this substitution consisted of coding sequences for 77 amino acids at the carboxyl terminus of the integrase, the s1 (encoding the second exon of tat), s2, and s3 (encoding the second exon of rev) open reading frames, the complete env ...19989445039
application of equine infectious anemia virus core proteins produced in a baculovirus expression system to serological diagnosis.equine infectious anemia virus (eiav) core proteins were obtained from a baculovirus expression system. recombinant baculoviruses (rbvs) highly expressed the gag precursor and p26 antigens in an rbv-infected sf21 cell culture supernatant. enzyme-linked immunosorbent assay (elisa) and agar gel immunodiffusion (agid) were conducted using the expressed proteins to detect antibodies from experimentally infected horses. the expressed antigens showed low background levels, high specificity and sensiti ...19979492183
maturation of the cellular and humoral immune responses to persistent infection in horses by equine infectious anemia virus is a complex and lengthy process.equine infectious anemia virus (eiav) provides a natural model system by which immunological control of lentivirus infections may be studied. to date, no detailed study addressing in parallel both the humoral and cellular immune responses induced in horses upon infection by eiav has been conducted. therefore, we initiated the first comprehensive characterization of the cellular and humoral immune responses during clinical progression from chronic disease to inapparent stages of eiav infection. u ...19979094660
characterization and mutational studies of equine infectious anemia virus dutpase.the macrophage tropic lentivirus, equine infectious anemia virus (eiav), encodes a dutpase in the pol gene that is required for efficient replication in macrophages. two naturally occurring variants of the enzyme were expressed as recombinant proteins in escherichia coli; metal chelate affinity chromatography was used to purify histidine-tagged recombinant enzymes to greater than 80% homogeneity in a single chromatographic step. biochemical and enzymatic analyses of these preparations suggest th ...19979187238
localized sequence heterogeneity in the long terminal repeats of in vivo isolates of equine infectious anemia virus.the role of in vivo long terminal repeat (ltr) sequence variation of the lentivirus equine infectious anemia virus (eiav) has not been explored. in this study, we investigated the heterogeneity found in the ltr sequences from seven eiav-seropositive horses: three horses with clinical disease and four horses without any detectable signs of disease. ltr sequences were targeted in this study because the ltr u3 enhancer region of tissue culture-derived isolates has been identified as one of the few ...19979188555
mutations in hiv reverse transcriptase which alter rnase h activity and decrease strand transfer efficiency are suppressed by hiv nucleocapsid protein.structural studies of authentic hiv reverse transcriptase (rt) suggest a role for the p51 carboxyl terminus in forming an active rnase h conformation [rodgers, d. w., gamblin, s. j., harris, b. a., ray, s., culp, j. s., hellmig, b., woolf, d. j., debouck, c. & harrison, s. c. (1995) proc. natl. acad. sci. usa 92, 1222-1226]. we have purified mutant rt heterodimers containing deletion of 5, 9, or 13 amino acids from the p51 carboxyl terminus. these "selectively deleted" heterodimers have been ana ...19979192628
flow cytometric method for detecting thiazole orange-positive (reticulated) platelets in thrombocytopenic horses.to evaluate a method for detecting thiazole orange-positive (to+, reticulated) platelets in equine blood, using flow cytometry.19979328660
elevation of cytokines associated with the thrombocytopenia of equine infectious anaemia.thrombocytopenia is a common finding in infection with equine infectious anaemia virus (eiav), a lentivirus with some homology to human immunodeficiency virus (hiv). the thrombocytopenia of eia, like that in some hiv patients, appears to have a multifactorial pathogenesis. to investigate the decreased platelet production seen in experimental eia, the levels of three potential negative regulators of platelet production--tumour necrosis factor-alpha (tnf-alpha), transforming growth factor-beta (tg ...19979349475
frequency of memory cytotoxic t lymphocytes to equine infectious anemia virus proteins in blood from carrier horses.horses with equine infectious anemia virus (eiav) have episodes of viremia and disease; however, most eventually become inapparent carriers. a possible mechanism of control is cytotoxic t lymphocytes (ctl). to evaluate ctl in inapparent carriers with low viral loads, peripheral blood mononuclear cells (pbmc) were stimulated in vitro with autologous eiav-infected pbmc and human il-2 to detect memory ctl (ctlm). in initial studies, three carriers had ctlm and one of these had low-level effector ct ...19979375012
isolation of an 11-kda protein associated with the topoisomerase i activity from equine infectious anemia virus.we have previously demonstrated the presence of topoisomerase i (topo i) activity in purified retroviral particles (i.e., human immunodeficiency virus type 1, equine infectious anemia virus-eiav and moloney murine leukemia virus). in our present work, an attempt was made to determine the nature and origin of the protein that is associated with this activity. for that purpose we have isolated the topo i activity from equine infectious anemia virus cores and showed that a major protein band of an ...19968607786
inhibitory activity of the equine infectious anemia virus major 5' splice site in the absence of rev.the major 5' splice site of equine infectious anemia virus (eiav) conforms to the consensus 5' splice site in eight consecutive positions and is located immediately upstream of the gag aug. our results show that the presence of this 5' splice site on the eiav gag mrna decreases gag production 30- to 60-fold. this is caused by inefficient nuclear mrna export and inefficient mrna utilization. inhibition could be overcome by providing human immunodeficiency virus type 1 rev/rev-responsive element, ...19968648699
detection of equine infectious anemia viral rna in plasma samples from recently infected and long-term inapparent carrier animals by pcr.control of equine infectious anemia (eia) is currently based on detection of anti-eia virus (eiav) antibodies. however, serologic diagnostic methods may give false-negative results in infected horses that fail to respond adequately or are in the early stages of infection. we developed a reverse transcriptase nested pcr (rt-npcr) assay for the detection of viral gag gene sequences in plasma from eiav-infected horses. the ability of rt-npcr to detect field strains of eiav was investigated by assay ...19968735102
structural studies of the equine infectious anemia virus trans-activator protein.trans-activator (tat) proteins are necessary components for the completion of the t replication cycle of lentiviruses. the three-dimensional structure of the equine infectious anemia virus (eiav) tat protein (e-tat) was studied with cd spectroscopy, nmr spectroscopy, and restrained molecular-dynamics calculations. no stable elements of regular secondary structure were detected, but the sequence regions responsible for nucleic acid binding showed helix-forming tendency, e-tat exhibits a flexible ...19968797834
initiation of (-) strand dna synthesis from trna(3lys) on lentiviral rnas: implications of specific hiv-1 rna-trna(3lys) interactions inhibiting primer utilization by retroviral reverse transcriptases.initiation of minus (-) strand dna synthesis was examined on templates containing r, u5, and primer-binding site regions of the human immunodeficiency virus type 1 (hiv-1), feline immunodeficiency virus (fiv), and equine infectious anemia virus (eiav) genomic rna. dna synthesis was initiated from (i) an oligoribonucleotide complementary to the primer-binding sites, (ii) synthetic trna(3lys), and (iii) natural trna(3lys), by the reverse transcriptases of hiv-1, fiv, eiav, simian immunodeficiency ...19968816751
structure of equine infectious anemia virus proteinase complexed with an inhibitor.equine infectious anemia virus (eiav), the causative agent of infectious anemia in horses, is a member of the lentiviral family. the virus-encoded proteinase (pr) processes viral polyproteins into functional molecules during replication and it also cleaves viral nucleocapsid protein during infection. the x-ray structure of a complex of the 154g mutant of eiav pr with the inhibitor hby-793 was solved at 1.8 a resolution and refined to a crystallographic r-factor of 0.136. the molecule is a dimer ...19968844837
a primary production deficit in the thrombocytopenia of equine infectious anemia.the purpose of this study was to identify the mechanisms responsible for the thrombocytopenia that develops following infection of horses by the lentivirus equine infectious anemia virus (eiav). immunocompetent arabian foals and arabian foals with severe combined immunodeficiency (scid), which lack functional b and t lymphocytes, were experimentally infected with eiav. levels of viremia and a number of clinical and hematologic parameters were examined prior to and following infection. thrombocyt ...19968892906
analysis of the long terminal repeat from a cytopathic strain of equine infectious anemia virus.sequential passage of the tissue culture-adapted prototype strain of eiav in fetal donkey dermal (fdd) cell cultures generated a virus stock which exhibits cytopathic effects in fdd cell cultures. in this study, the effects of the long terminal repeat (ltr) region on virus replication and cytopathogenicity were examined. the fdd-adapted virus ltr was found to contain a number of base pair mutations and a large insertion within the u3 region in comparison with the previously characterized ltr, la ...19968918926
retroviruses and systemic lupus erythematosus.in some animal models of autoimmune diseases the roles of exogenous and endogenous retroviruses are clearly defined. in ungulates caprine arthritis encephalitis virus, equine infectious anemia virus or maedi-visna virus infections cause a well-defined autoimmune disease and the appearance of seropositivity of the animals is of diagnostic value. likewise, in mrl lpr/lpr mice insertion of a retrotransposon into the fas gene could clearly be shown to cause survival of autoreactive lymphocytes. desp ...19968930671
nmr analysis of the trans-activation response (tar) rna element of equine infectious anemia virus.transcription of lentiviral dna in the host cell is regulated by an interaction between the viral tar rna stem-loop and the viral tat protein. here we present a model of the three-dimensional structure of the tar rna stem-loop of the equine infectious anemia virus (eiav), derived from two- and three-dimensional nmr data. this 25 nucleotide rna consists of an a-form helical stem capped by two u-g base pairs and a four-nucleotide loop. two loop cytidines are stacked into the loop interior and like ...19957479065
production of monoclonal antibodies in horses. 19957550692
structural studies of hiv-1 tat protein.tat (trans-activator) proteins are early rna binding proteins regulating lentiviral transcription. these proteins are necessary components in the life cycle of all known lentiviruses, such as the human immunodeficiency viruses (hiv) or the equine infectious anemia virus (eiav). tat proteins are thus ideal targets for drugs intervening with lentiviral growth. the consensus rna binding motif (tar, trans-activation responsive element) of hiv-1 is well characterized. structural features of the 86 am ...19957723010
structural analysis of the principal immunodominant domain of the feline immunodeficiency virus transmembrane glycoprotein.in the transmembrane envelope glycoprotein (tm) of lentiviruses, including human immunodeficiency virus type 1 (hiv-1) and feline immunodeficiency virus (fiv), two cysteine residues, conserved in most retroviruses, are thought to form a loop containing five to seven amino acids. these elements make up a b-cell epitope recognized by nearly 100% of sera from infected patients or animals, designated the principal immunodominant domain (pid). the pid amino acid sequences are highly conserved between ...19957884857
delineating minimal protein domains and promoter elements for transcriptional activation by lentivirus tat proteins.lentivirus tat proteins comprise a novel class of rna-binding transcriptional activators that are essential for viral replication. in this study, we performed a series of protein fusion experiments to delineate the minimal protein domains and promoter elements required for tat action. we show that a 15-amino-acid region of equine infectious anemia virus (eiav) tat protein, when fused to the gal4 or lexa dna binding domain, can activate transcription in appropriate promoter contexts. in the natur ...19957884911
trifluoroethanol stabilizes a helix-turn-helix motif in equine infectious-anemia-virus trans-activator protein.the solution structure of the 75-amino-acid trans-activator (tat) protein of the equine infectious-anemia virus in trifluoroethanol-containing solution was determined by two-dimensional and three-dimensional nuclear magnetic resonance spectroscopy, resulting in a total of 838 nuclear-over-hauser-enhancement distance restraints, and restrained molecular-dynamics simulations. in contrast to the recently determined structure of this protein in trifluoroethanol-free ph 6.3 solution, the hydrophobic ...19947957222
major histocompatibility complex-restricted cd8+ cytotoxic t lymphocytes from horses with equine infectious anemia virus recognize env and gag/pr proteins.cytotoxic t lymphocytes (ctl) can control some viral infections and may be important in the control of lentiviruses, including human immunodeficiency virus type 1. since there is limited evidence for an in vivo role of ctl in control of lentiviruses, dissection of immune mechanisms in animal lentiviral infections may provide needed information. horses infected with equine infectious anemia virus (eiav) a lentivirus, have acute plasma viremia which is terminated in immunocompetent horses. viremic ...19948107209
enhancement of eiav replication and disease by immunization with a baculovirus-expressed recombinant envelope surface glycoprotein.the potential for antibody-dependent enhancement of replication of macrophage/monocyte tropic viruses has posed a significant problem in the development of vaccines for several animal and human viruses and has raised significant concern in the design of potential aids vaccines. using the previously described equine infectious anemia virus/shetland pony system as a model for hiv-1 vaccine development, we have evaluated the efficacy of a recombinant subunit vaccine containing a baculovirus-express ...19948116252
expression of functional protease and subviral particles by vaccinia virus containing equine infectious anaemia virus gag and 5' pol genes.cells infected with vaccinia viruses expressing the equine infectious anaemia virus (eiav) gag gene (vgag) or gag plus the 5' pol encoding protease (vgag/pr) were evaluated with monoclonal antibody to a p26 capsid protein linear epitope (qeiskfltd). both recombinant viruses expressed gag precursor protein (55k) whereas only vgag/pr expressed a detectable gag-pol fusion protein (82k) with a functional protease, shown by subviral particles containing processed p26. horses inoculated with vgag/pr p ...19948151302
translation of equine infectious anemia virus bicistronic tat-rev mrna requires leaky ribosome scanning of the tat ctg initiation codon.we have examined the translational regulation of the equine infectious anemia virus (eiav) bicistronic tat-rev mrna. site-directed mutagenesis of the tat leader region followed by expression of the tat-rev cdna both in vitro and in transiently transfected cells established that tat translation is initiated exclusively at a ctg codon. increasing the efficiency of tat translation by altering the ctg initiator to atg resulted in a dramatic decrease in translation of the downstream (rev) cistron, in ...19938382305
sequence-specific resonance assignments of the 1h-nmr spectra of a synthetic, biologically active eiav tat protein.the equine infectious anemia virus (eiav) trans-activating (tat) protein is a close homologue of the human immunodeficiency virus (hiv) tat protein. both of these proteins bind to an rna trans-activation responsive element (tar). we synthesized chemically a protein with the sequence of the 75 amino acid tat protein from eiav. the chemically synthesized protein was shown to be biologically active. circular dichroism (cd) and 1h nuclear magnetic resonance (nmr) spectroscopy were used to structural ...19938395203
equine infectious anemia.the ability of eiav to persistently infect horses in the face of a profound immune response by the host makes it a potentially devastating disease for the horse population of the united states. its ability to evade host immune defenses by lying dormant in apparently healthy animals and by rapidly changing its antigenic determinants is proving to be a major obstacle to vaccine development. because most infected horses appear clinically normal and a large proportion of horses in this country remai ...19938395326
expression and mutational analysis of the reverse transcriptase of the lentivirus equine infectious anemia virus.the reverse transcriptase of equine infectious anemia virus (eiav) shows sequence similarity with the reverse transcriptases of other lentiviruses, particularly with those of human immunodeficiency viruses types 1 and 2 (hiv-1 and hiv-2). we have constructed a plasmid that when introduced into e. coli induces the synthesis of substantial quantities of the nearly authentic eiav reverse transcriptase. the viral and bacterially expressed reverse transcriptases are similar in their molecular weights ...19937694581
distinct subsets of retroviruses encode dutpase.the nonprimate lentiviruses feline immunodeficiency virus, equine infectious anemia virus, visna virus, and caprine encephalitis virus contain a gene segment in the polymerase gene that is lacking in the primate lentiviruses. a related sequence has been noted in other retroviruses, most notably the type d retroviruses. computer searches have indicated a relatedness between this unique gene segment, termed proteaselike element and elements of both the aspartate proteinase and the dutpase enzyme f ...19921310783
inhibition of human immunodeficiency virus type 1 tat activity by coexpression of heterologous trans activators.we examined the mechanism of tat-mediated trans activation through competition experiments employing tat proteins of human immunodeficiency virus type 1 (hiv-1) and equine infectious anemia virus (eiav). eiav tat, as well as chimeric eiav/hiv-1 tat proteins, inhibited hiv-1 tat-mediated trans activation in a cell-type-dependent fashion. furthermore, these proteins inhibited trans activation by tat-bacteriophage r17 coat protein chimeras. inhibition resulted from competition between activation do ...19921312617
a soluble recombinant fusion protein of the transmembrane envelope protein of equine infectious anaemia virus for elisa.the use of the bacterial expression vector, pgex, to produce an abundant, soluble fusion protein of gp45 from equine infectious anaemia virus is described. purification of the recombinant protein was achieved by one step affinity chromatography on immobilized glutathione using competitive elution so no harsh conditions were required. this provides a readily available antigen that is defined, plentiful and cheap. yields of 3.5 mg of purified soluble protein/litre of bacterial culture were obtaine ...19921320787
equine lentivirus, comparative studies on four serological tests for the diagnosis of equine infectious anaemia.serological diagnosis of equine infectious anemia is of necessity group-reactive, i.e. based on viral core protein p26, because viral envelope components as well as the host's immune response to them undergo rapid antigenic change. since 1970 the agar gel-immunodiffusion test ("coggins-test") has been the diagnostic method of choice. recently, elisa tests have been introduced for faster and theoretically more sensitive serodiagnosis, while western blots have been used to clarify doubtful results ...19921336247
african horse sickness and equine infectious anaemia serology in the gambia. 19921339038
detailed mapping of the antigenicity of the surface unit glycoprotein of equine infectious anemia virus by using synthetic peptide strategies.we describe here a detailed analysis of the antigenic determinants of the surface unit glycoprotein (gp90) of equine infectious anemia virus (eiav), using a comprehensive panel of synthetic peptides in enzyme-linked immunosorbent assays with immune serum from naturally and experimentally infected horses and with a panel of gp90-specific neutralizing and nonneutralizing monoclonal antibodies. the results of these studies identify immunoreactive segments throughout the conserved and variable domai ...19921370556
detecting single base substitutions as heteroduplex polymorphisms.we have developed a sensitive technique for detecting single base substitutions in polymerase chain reaction (pcr) products from individuals heterozygous for polymorphisms or new mutations. this technique takes advantage of the formation of heteroduplexes in the pcr between different alleles from heterozygous individuals. these heteroduplexes can be detected on polyacrylamide gels because they migrate slower than their corresponding homoduplexes. using pcr, we have generated a series of point mu ...19921740339
identification of a hypervariable region in the long terminal repeat of equine infectious anemia virus.an avirulent, field-derived isolate of equine infectious anemia virus (eiav), designated ma-1, was molecularly cloned, and the complete nucleotide sequence was determined for the 3' half of the viral genome. comparisons between ma-1 and the prototype wyoming strain of eiav identified a 66-nucleotide stretch between caat (-91) and tataa (-25) in the u3 region of the long terminal repeat, where sequence divergence was as high as 39.3%. the polymerase chain reaction was used to amplify and clone lo ...19911847479
proviral sequences detected by polymerase chain reaction in peripheral blood cells of horses with equine infectious anemia lentivirus.proviral sequences in the peripheral blood mononuclear cells of 3 horses with acute equine infectious anemia virus were monitored using the polymerase chain reaction. provirus was detected during the initial viremic episode in each horse and during each of 3 relapsing viremic cycles, although the appearance of provirus lagged behind the onset of viremia. following each viremic episode, provirus levels in the peripheral monocytes decreased to less than 1 copy in 5 x 10(6) cells.19911848747
mutational analysis of the equine infectious anemia virus tat-responsive element.a hairpinlike structure is predicted to exist at the 5' end of equine infectious anemia virus (eiav) rna which is similar in many ways to the human immunodeficiency type 1 (hiv-1) tat-responsive element (tar). in eiav, this structure has a shorter stem than in hiv-1 and lacks the uridine bulge. primer extension analysis of eiav rna was used to identify the transcriptional start site in the viral long terminal repeat. premature termination of primer elongation at the predicted double-stranded rna ...19911645778
transient suppression of equine immune responses by equine infectious anemia virus (eiav).suppression of the immune system is a common aspect of the disease pathogenesis associated with retroviral infections in both man and animals. we have measured transient suppression of the equine immune system as a loss or decrease in antigen-specific and polyclonal lymphocyte proliferation following experimental infection of ponies with three variants of equine infectious anemia virus (eiav) with difference virulence characteristics. the transient suppression of proliferative responses was temp ...19911651604
immune-mediated thrombocytopenia in horses infected with equine infectious anemia virus.an adult horse infected with a virulent, cell culture-adapted strain of equine infectious anemia virus (eiav) developed cyclical thrombocytopenia in which the nadir of platelet counts coincided with peak febrile responses. in order to investigate the mechanism of thrombocytopenia during acute febrile episodes, four adult horses were experimentally infected with the wild-type wyoming strain of eiav. platelet counts decreased from baseline as rectal temperature increased. serum reverse transcripta ...19911717720
antiretroviral activity of synthetic hypericin and related analogs.hypericin and pseudohypericin are naturally occurring polycyclic quinones which have recently been shown to inhibit the infectivity of several retroviruses, including human immunodeficiency virus. to better understand the antiviral mechanisms of these compounds, hypericin and a series of analogous quinones were synthesized and tested for anti-retroviral activity against equine infectious anemia virus (eiav). treatment of eiav-infected cells with hypericin reduced the production of infectious vir ...19901699534
equine monoclonal antibodies recognize common epitopes on variants of equine infectious anaemia virus.equine-murine xenohybridoma cells were produced using sp2/0 murine myeloma cells and splenic lymph node cells obtained from horses infected with 10(6) tcid50 of single cloned variants of equine infectious anaemia virus (eiav). the xenohybridomas secreted equine igg monoclonal antibodies reactive with eiav in enzyme immunoassays employing purified virus. seven antibodies were studied in detail. they bound to viral glycoproteins (gp90 or gp45) in radioimmunoprecipitation assays, and reacted with h ...19901703988
characterization of eiav immunogenicity during persistent infections: humoral responses and antigen targets. 19901704323
proviral dna integration and transcriptional patterns of equine infectious anemia virus during persistent and cytopathic infections.the structure and integration patterns of equine infectious anemia virus (eiav) proviral dna and the patterns of viral transcription were examined in persistent and cytopathic infections of cultured cells. the results of southern blot analyses indicated that, in persistently infected cells, about 30% of the eiav provirus exists as randomly integrated dna, while the remaining 70% is equally divided between unintegrated linear and closed circular forms. the cytopathic infection, in contrast, is ch ...19902152836
analysis of antibody reactivities in elisa using protein blots as antigen substrates: s-elisa.we describe here a novel immunoassay procedure, designated strip-elisa (s-elisa), in which specific antigens are purified by sds-page, transferred to support membranes, and utilized in situ as substrate in routine elisa procedures. using two different lentivirus systems, simian immunodeficiency virus and equine infectious anemia virus, we demonstrate the utility of s-elisa for screening hybridoma supernatants during production of monoclonal antibodies and for the dissection of polyclonal antibod ...19902157766
equine infectious anemia virus (eiav) humoral responses of recipient ponies and antigenic variation during persistent infection.three ponies were inoculated with plasma containing 10(4.8) tcid50 of equine infectious anemia virus (eiav) and observed for 165 to 440 days. each pony developed a febrile response within 3 weeks of infection during which a plasma viremia greater than or equal to 10(3.5) tcid50/ml was observed. analyses of four isolates from sequential febrile episodes in a single pony were conducted by two-dimensional tryptic peptide maps and with monoclonal antibodies in immunoblots. structural and antigenic a ...19902162160
cloning and characterization of cdnas encoding equine infectious anemia virus tat and putative rev proteins.we isolated and characterized six cdna clones from an equine infectious anemia virus-infected cell line that displays a rev-defective phenotype. with the exception of one splice site in one of the clones, all six cdnas exhibited the same splicing pattern and consisted of four exons. exon 1 contained the 5' end of the genome; exon 2 contained the tat gene from mid-genome; exon 3 consisted of a small section of env, near the 5' end of the env gene; and exon 4 contained the putative rev open readin ...19902164593
selection against cpg dinucleotides in lentiviral genes: a possible role of methylation in regulation of viral expression.extremely low frequencies of cpg dinucleotides are found in the genomes of the lentivirus subfamily of retroviruses, including the human, simian and feline immunodeficiency viruses (hiv1, hiv2, siv, and fiv, respectively), equine infectious anemia virus (eiav), and the ovine lentivirus, visna. the occurrence of cpg dinucleotides is greater in the 2-3 (ncg) than in the 1-2 (cgn) codon-defined frame, as well as in the gag and env genes, compared to the more conserved pol gene. these differences su ...19902170945
topoisomerase i activity associated with human immunodeficiency virus (hiv) particles and equine infectious anemia virus core.in the present study, we found a topoisomerase i (topo i) activity in two strains of human immunodeficiency virus type 1 (hiv-1) and equine infectious anemia virus (eiav) particles. the topo i activity was located in the eiav cores and differed from the cellular topo i in its ionic requirements and response to atp, indicating that these were two distinct forms of this enzyme. topo i activity was removed from the viral lysates and viral cores by anti-topo i antiserum. the only protein recognized ...19902174357
immunopathogenesis of equine infectious anemia lentivirus disease.virus replication and subsequent viremia are clearly correlated with clinical disease in eiav infected horses. termination of viremia is the result of specific immune responses. recurrences of viremia are associated with antigenic variation of neutralization-sensitive epitopes. immunosuppression experiments indicate that the eventual control of eiav and development of carriers is mediated by the immune system. even though the immune response to eiav has a protective effect, immune responses also ...19902178127
change in host cell tropism associated with in vitro replication of equine infectious anemia virus.similar to other human and animal lentiviruses, equine infectious anemia virus (eiav) is detectable in vivo in cells of the monocyte-macrophage lineage. owing to their short-lived nature, horse peripheral blood macrophage cultures (hmc) are rarely used for in vitro propagation of eiav, and equine dermal (ed) or kidney cell cultures, which can be repeatedly passed in vitro, are used in most studies. however, wild-type isolates of eiav will not grow in these cell types without extensive adaptation ...19892470916
animal lentivirus replication and reverse transcriptase inhibitors.we summarize the pathogenesis of animal lentiviruses (visna-maedi virus, caprine arthritis and encephalitis virus, and equine infectious anemia virus), which have raised considerable interest since the discovery of human lentiviruses. the human lentiviruses possess structural, genetic, and clinical properties similar to those of animal lentiviruses. we describe the different mechanisms of and the principal work on reverse transcriptase inhibitors of animal lentiviruses, such as hpa-23, phosphono ...19892479076
the preparation and biochemical characterization of intact capsids of equine infectious anemia virus.capsids of equine infectious anemia virus have been isolated as cone-shaped particles 60 x 120 nm in size. detergent treatment of whole virus followed by two cycles of rate-zonal centrifugation in ficoll produces these capsids in a yield of approximately 10%. the major protein components are the gag-encoded p11 nucleocapsid protein and p26 capsid protein, which are present in equimolar amounts. substantial cleavage of p11 to p6 and p4 can be observed under conditions where the viral protease pac ...19892541703
occurrence of equine infectious anaemia in india. 19892547265
analysis of regulatory elements of the equine infectious anemia virus and caprine arthritis-encephalitis virus long terminal repeats.we analyzed the equine infectious anemia virus (eiav) long terminal repeat (ltr) for sequences that influence its promoter activity and ability to be trans-activated by the eiav tat gene product. a series of ltr deletion mutants and recombinants between ltr and simian virus 40 (sv40) regulatory sequences were used for these studies. we were able to identify the eiav promoter region and showed that sequences within the u3 region significantly inhibited ltr-directed transcription. however, when pl ...19892552171
localization of conserved and variable antigenic domains of equine infectious anemia virus envelope glycoproteins using recombinant env-encoded protein fragments produced in escherichia coli.previous characterizations of equine infectious anemia virus (eiav) glycoprotein variation by dna sequence analysis and epitope mapping using monoclonal antibodies (mabs) have revealed the presence of conserved and variable regions within the eiav env gene. to extend these studies, fragments of the eiav envelope proteins gp90 and gp45 were expressed in escherichia coli and used in western blot analysis with a diverse panel of equine immune sera to identify antigenic segments. all sera from eiav- ...19892552661
cross-neutralizing and subclass characteristics of antibody from horses with equine infectious anemia virus.antibody responses in horses with equine infectious anemia virus (eiav) were examined to determine their cross-neutralizing capacity. antibodies induced by infection with any of six biologically cloned variants of eiav cross-neutralized multiple variants from the group. anti-eiav antibody was found in both the igg and igg(t) subclasses in plasmas with virus-neutralizing activity and the majority of antiviral antibody was of the igg(t) subclass. depletion of igg(t) did not increase the neutraliza ...19892559537
comparison of diagnostic tests for the detection of equine infectious anemia antibody.two diagnostic tests are approved for detecting antibody to equine infectious anemia virus: the agar-gel immunodiffusion (agid) test and the competitive enzyme-linked immunosorbent assay (elisa). a total of 420 sera from national veterinary services laboratories check sets were tested with the agid and competitive elisa. a 100% correlation was obtained. the agid and competitive elisa were further used to test difficult samples with low levels of equine infectious anemia antibody (weak positives) ...19892562211
antigenic variation in lentiviral diseases. 19882838047
eiav genomic organization: further characterization by sequencing of purified glycoproteins and cdna.nucleotide sequence analyses of two different proviral clones of equine infectious anemia virus (eiav), designated lambda 12 (k. rushlow et al., 1986, virology 155, 309-321) and 1369 (t. kawakami et al., 1987, virology 158, 300-312), indicate significant differences in the organization of two critical regions of the viral genome, i.e., in the short open reading frames in the pol-env intergenic region and in the 5'-end of the env gene. to determine the correct structure of the eiav genome, we hav ...19882841805
the lentiviruses: maedi/visna, caprine arthritis-encephalitis, and equine infectious anemia. 19882843016
a perspective on equine infectious anemia with an emphasis on vector transmission and genetic analysis. 19882847392
a propagating epizootic of equine infectious anemia on a horse farm.an epizootic of equine infectious anemia (eia) involved 35 horses on a farm in south georgia. during a 126-day period, 21 of these horses became seropositive for eia. after the initial diagnosis in july, the horses were tested every 7 to 10 days. at least one additional horse was found to be seropositive on each testing day. as soon as they were determined to be seropositive, the horses were removed from the herd and sent to slaughter. the removal of the seropositive horses, however, did not sto ...19882848789
studies on viral-induced anemia in horses infected with equine infectious anemia virus. 19883386089
hybridoma cell lines secreting monoclonal antibodies against equine infectious anemia virus.a monoclonal anti-equine infectious anemia virus (anti-eiav) antibody (1b15) has been generated by fusion of x63 ag 8.653 myeloma cells and spleen cells from mice hypersensitized with viral antigen p29. ouchterlony double-diffusion analysis indicated that antibody 1b15 is of the igg class. the specificity of the immune reaction for p29 was confirmed by cross-over immunoelectrophoresis and disc-gel electrophoresis. mab 1b15 was used to devise a solid-phase 'capture' ria for eiav-p29 antigen. the ...19872435751
characterization of equine infectious anemia virus long terminal repeat.the long terminal repeats (ltrs) of equine infectious anemia virus (eiav) were examined with respect to their ability to function as transcriptional promoters in various cellular environments. nucleotide sequence analyses of the ltrs derived from two unique proviral clones revealed the requisite consensus transcription and processing signals. one of the proviruses possessed a duplication of a 16-base-pair sequence in the ccaat box region of the ltr which was absent in the other provirus. to asse ...19873027401
phagocytosis of horse erythrocytes treated with equine infectious anemia virus by cultivated horse leukocytes.horse erythrocytes treated with equine infectious anemia virus hemagglutinin were phagocytized by cultivated horse leukocytes (mainly macrophage-like cells and partly polymorphonuclear cells) after incubation with fresh horse serum but not with inactivated horse serum. the phagocytosis began as soon as the erythrocytes were added to the leukocyte cultures, and the majority of the reaction proceeded within 30 minutes. addition of antiserum showed a slightly suppressing but no enhancing effect on ...19873036046
a review of antigenic variation by the equine infectious anemia virus. 19873040337
the visna virus genome: evidence for a hypervariable site in the env gene and sequence homology among lentivirus envelope proteins.the complete nucleotide sequence of the visna virus 1514 genome was determined. our sequence confirms the relationship of visna virus and other lentiviruses to human immunodeficiency virus (hiv) both at the level of sequence homology and of genomic organization. sequence homology is shown to extend to the transmembrane proteins of lentivirus env genes; this homology is strongest in the extracellular domain, suggesting that close structural and functional similarities may also exist among these e ...19872824836
antigenic variation and lentivirus persistence: variations in envelope gene sequences during eiav infection resemble changes reported for sequential isolates of hiv.the extent and nature of genomic variation among nine antigenically distinct eiav isolates recovered during sequential clinical episodes from two experimentally infected ponies were examined by restriction fragment analysis and nucleotide sequencing. only minor variations in restriction enzyme patterns were observed among the viral genomes. in contrast, env gene sequences of four isolates from one pony revealed numerous clustered base substitutions. divergence in env gene nucleotide and deduced ...19872825406
rapid detection of viral-specific antibodies by enzyme-linked immunosorbent assay (elisa).the development of three separate rapid elisas for detecting antibodies in host serum to three different viruses is described. these include: 1. a direct antigen assay using enzyme labelled anti-canine ig for detecting antibodies to canine parvovirus, 2. a competitive elisa using a feline infectious peritonitis virus-specific monoclonal antibody labelled with enzyme, and 3. a competitive elisa using an equine infectious anemia virus-specific monoclonal antibody and enzyme labelled antigen, p. 26 ...19872829416
lav/htlv-iii gag gene product p24 shares antigenic determinants with equine infectious anemia virus but not with visna virus or caprine arthritis encephalitis virus.antigenic cross-reactivity of the human acquired immunodeficiency disease syndrome virus lav/htlv-iii with the lentiviruses visna virus, caprine arthritis encephalitis virus (caev), and equine infectious anemia virus (eiav) was determined with indirect enzyme-linked immunosorbent assays, immunoblot analysis, and virus-specific polyclonal antisera. nonreciprocal cross-reactivity was seen between the gag gene products p24 of lav/htlv-iii and p28 of eiav. reciprocal cross-reactivity was seen betwee ...19862438251
shedding and interspecies type sero-reactivity of the envelope glycopolypeptide gp120 of the human immunodeficiency virus.two glycopolypeptides with molecular weights 160,000 and 120,000 (gp120) are regularly recognized by human immunodeficiency virus (hiv)-specific antisera in lysates of cells persistently infected with hiv. in the present study, gp120 was characterized as the major envelope glycopolypeptide of hiv. gp120 was identified as the external viral glycoprotein by radiosequencing and by its presence in purified virus. however gp120 was predominantly shed as a soluble protein into the culture fluid. furth ...19862431105
rapid emergence of novel antigenic and genetic variants of equine infectious anemia virus during persistent infection.previous results from our laboratory have demonstrated that equine infectious anemia virus displays structural variations in its surface glycoproteins and rna genome during passage and chronic infections in experimentally infected shetland ponies (montelaro et al., j. biol. chem. 259:10539-10544, 1984; payne et al., j. gen. virol. 65:1395-1399, 1984). the present study was undertaken to obtain an antigenic and biochemical characterization of equine infectious anemia virus isolates recovered from ...19863001367
equine infectious anemia virus: immunopathogenesis and persistence.equine infectious anemia (eia) is a chronic, relapsing infectious disease of horses caused by a nononcogenic retrovirus. virus persists in infected animals for life and can be reliably detected by serologic tests that measure levels of antibody to the major structural protein of the virus. periodic virus replication in macrophages leads to an immunologically mediated acute disease characterized primarily by severe anemia. recrudescence of acute eia is the result of antigenic variation of the sur ...19852984759
rapid solid-phase radioimmunoassay for detection of equine infectious anemia viral antigen and antibodies: parameters involved in standardization.solid-phase radioimmunoassays (spria) are described for the detection of equine infectious anemia (eia) viral antigen and antibodies. protein-antigen p29 currently used in the agar-gel immunodiffusion (agid) test was used as antigen in the spria. rabbit sera selected from positive agid test data were used to standardize the method. briefly, wells of flexible microtitre plates coated with antigen were incubated with antiserum followed by a secondary labelled antibody. the radioactivity remaining ...19853001113
human t-cell lymphotropic virus type iii: immunologic characterization and primary structure analysis of the major internal protein, p24.the major internal structural protein of human t-cell lymphotropic virus type iii (htlv-iii), a virus etiologically implicated in acquired immunodeficiency syndrome (aids), was purified to homogeneity. this 24,000-molecular-weight protein (p24) was shown to lack immunologic cross-reacting antigenic determinants shared by other known retroviruses, including htlv-i and htlv-ii, with the exception of equine infectious anemia virus (eiav). a broadly reactive competition immunoassay was developed in ...19852410630
studies on equine infectious anemia virus transmission by insects.there are several factors involved in the mechanical transmission of equine infectious anemia (eia) virus by insects. large hematophagous insects, especially tabanids, which feed from extravascular sites (ie, pool feeding) appear to be the most efficient vectors. the biology of the host-seeking and blood-feeding behavior of the vectors are important variables that have been overlooked in the mechanical transmission of pathogens like eia virus. the biology, population levels, and diversity of the ...19846321420
structural proteins of equine infectious anemia virus and their antigenic activity.using purified equine infectious anemia (eia) virus labeled with 3h-glucosamine or 14c-protein hydrolysate, structural proteins were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. as a result, 2 glycoproteins and 10 proteins with molecular weights (mol wt) ranging from 12,000 to 115,000 daltons were demonstrated. of 12 structural proteins, 3 proteins, namely a glycoprotein with mol wt of 76,000 (gp76) and 2 proteins with mol wt of 25,000 (p25) and 12,000 (p12), respective ...19846322625
effects of common radioiodination procedures on the binding of glycoproteins to immobilized lectins.representative glycoproteins including fetuin, protein a, ovalbumin, alpha 1 acid glycoprotein, and the major glycoprotein of equine infectious anemia virus were labelled with 125i by the chloramine-t or bolton-hunter procedure and their binding to immobilized con a or lentil lectin compared to untreated samples of each glycoprotein. glycoprotein modification was no greater than one substituted residue per protein molecule. yet the radioiodinated glycoproteins typically displayed only 0-50% of t ...19836838504
mechanical transmission of equine infectious anemia virus by deer flies (chrysops flavidus) and stable flies (stomoxys calcitrans). 19836297339
transmission of equine infectious anemia virus from horses without clinical signs of disease.twenty seven adult horses positive to the agar gel immunodiffusion (agid) test for equine infectious anemia (eia), but with no history of clinical eia, were used in transfusion studies to determine whether infectious eia virus was present in 1 to 5 ml of their blood. of 27 recipients, 21 (78%) became agid test-positive at an average of 24 days after inoculation. two horses that were initially negative when screened were retested and found to carry infectious virus in 5-300 ml of whole blood; the ...19826276353
antigenic stimulation of t lymphocytes in chronic nononcogenic retrovirus infection: equine infectious anemia.equine infectious anemia is a chronic disease of horses caused by a nononcogenic retrovirus. studies were undertaken to determine the types of cells involved in the in vitro lymphoproliferative response to viral antigens and the dynamics of this reaction. it was observed that reactive lymphocytes were present at unpredictable times in the peripheral blood of infected horses. this reaction was shown to be specific for the interaction of equine infectious anemia virus and t lymphocytes. enriched b ...19826281191
equine infectious anaemia: detection of antibodies using an immunofluorescence test.an indirect immunofluorescence test for detecting antibodies to equine infectious anaemia virus is presented. using monolayers of equine dermal cells within a defined period after infection, discrete fluorescent spots were observed in the cytoplasm of as many as 95 per cent of the cells. these inclusions appeared as ring-like structures when high titred sera were employed but became spots when the sera were diluted. cells showing optimal antigen fluorescence were used immediately or after storag ...19826296954
hemagglutination of several strains of equine infectious anemia virus.six strains of equine infectious anemia (eia) virus propagated in equine leukocyte cultures were found to agglutinate horse erythrocytes. concentrated virus material containing about 20 units of complement fixation (cf) titer showed hemagglutinating (ha) titers ranging from 4 to 8 units. the ha activity remained stable after ether treatment and was reduced by trypsin, formaldehyde and kio4. cesium chloride equilibrium density gradient centrifugation revealed two populations of hemagglutinin, one ...19816263225
transmission of equine infectious anaemia virus from a horse negative to agar gel immunodiffusion testing. 19816265206
studies with equine infectious anemia virus: transmission attempts by mosquitoes and survival of virus on vector mouthparts and hypodermic needles, and in mosquito tissue culture.biological and mechanical transmission trials with psorophora columbiae (dyar and knab) and aedes sollicitans (walker) and ponies acutely infected with equine infectious anemia virus (eiav) were negative. the eiav antigen was detected by radioimmunoassay in ae sollicitans immediately after the mosquitoes had fed on an acutely ill pony, but not 14 days after feeding. psorophora columbiae mosquitoes had detectable eiav antigen as determined by radioimmunoassay 24 hours after they fed on an acutely ...19816119953
isolation and characterization of low-molecular-weight dna-binding proteins from retroviruses. 19806249039
the role of stable flies and mosquitoes in the transmission of equine infectious anemia virus. 19806118874
synthesis of long complementary dna in the endogenous reaction by equine infectious anemia virus.in the endogenous reverse transcriptase reaction, equine infectious anemia virus is able to synthesize complementary dna (cdna) of 8,000 nucleotides in high yield. after 2 h in 50 mum dntp, about 2.8 mug of cdna per mg of protein is produced, almost 30% of which is long cdna. the system thus compares favorably with the other two well-characterized endogenous reaction systems, moloney murine leukemia virus and avian sarcoma virus. elongation rates of 100 to 150 nucleotides per min have been obser ...197987522
equine infectious anemia: current knowledge. 1979218920
prevalence of antibodies to equine viruses in the netherlands.the prevalence of antibodies to various viruses was investigated in a series of serum samples collected from horses in the netherlands between 1963 and 1966 and from 1972 onwards. neutralizing antibodies to equine rhinopneumonitis virus, equine arteritis virus and to equine rhinovirus types 1 and 2 were detected in respectively 76%, 14%, 66% and 59% of the equine serum samples tested. the observed incidence of serum samples positive to equine adenovirus in the complement fixation test was 39%. p ...1979219560
serologic survey for equine infectious anemia virus in louisiana.in 1975, a survey was conducted in east baton rouge parish, louisiana, to determine the prevalence of equine infectious anemia. using the agar gel immunodiffusion test, 94 of 1,398 horses (6.7%) were found to be infected. infection rates were especially high in areas where clinical cases of equine infectious anemia had been diagnosed. clinical signs compatible with the disease were noted in 1 of the 94 seropositive horses. the sample set of 1,398 horses represented 22% of the census population o ...1979221447
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