Herpesvirus saimiri-induced proteins in lytically infected cells. I. Time-ordered synthesis

The addition of TPA (phorbol-12-myristate-13-acetate) to cultures during the lytic infection with herpesvirus saimiri led to an enhanced and accelerated production of polypeptides induced by H. saimiri and to a rapid shut-down of host cell protein synthesis and allowed a detailed analysis of the protein patterns. Analysis of sequential protein synthesis in owl monkey kidney cells lytically infected with H. saimiri 11 permitted the identification of 31 virus-induced polypeptides. The use of the amino acid analogues canavanine (for arginine) and azetidine (for proline) in parallel allowed experiments on the identification of proteins synthesized early and late during lytic infection

The spontaneous and induced synthesis of Epstein-Barr virus antigens in Raji cells immobilized on surface coated with anti-lymphocyte globulin

Immobilization of Raji cells on surface coated with anti-lymphocyte globulin (ALG) at low cell densities lead to the synthesis of Epstein-Barr virus (EBV) early antigen (EA) in up to 5% of the cells. At higher cell densities the percentage of antigen-positive cells decreased and at confluency no antigen synthesis was observed. Addition of iododeoxyuridine (IdUrd) to low density cultures increased the expression of EA to 20%, whereas in confluent cultures the cells could not be induced to synthesize EA. Treatment of cells in suspension with ALG failed to induced EA synthesis and did not potentiate the effect of IdUrd. Immobilized Raji cells proved to be suitable targets for superinfection with EBV derived from P3HR1 cultures

The regulated expression of Epstein-Barr virus. III. Proteins specified by EBV during the lytic cycle

The experiments show that 30 virus-induced or virus-specified proteins were synthesized in Raji cells after superinfection with Epstein-Barr virus (EBV) derived from P3HR1 cells. Using a combination of pulse labelling, application of cycloheximide blocks at different times post-infection, treatment with amino acid analogues and inhibition of DNA synthesis it was shown that three groups of proteins appear in Raji cells after superinfection; the synthesis of the proteins in any one group appears to be coordinately regulated. Amongst the six virus-induced proteins which were synthesized immediately after release from an early cycloheximide block one would expect to find those proteins essential for the transition from EBNA to EA synthesis. Using human sera with differing specificities for the various antigen groups 11 proteins were identified as being specifically precipitated by sera having high titres against the EBV-induced early antigen complex

Identification of type A and B isolates of Epstein-Barr virus by polymerase chain reaction

A method is described for the identification of type A and type B isolates of Epstein-Barr virus (EBV) by means of the polymerase chain reaction. The use of three pairs of primers specific for genomic sequences coding for the two forms of EBV nuclear antigen (EBNA), 2A and 2B, and for a DNA sequence from the BamZ/BamR region allows the reliable and rapid detection of type A and B viruses in as little as 1000 EBV positive cells

Der Nachweis von Epstein-Barr-Virus-Genomen in der Ohrspeicheldrüse = Evidence for the persistence of Epstein-Barr-virus in the parotid gland (author's transl.)

EBV is associated with B-lymphocytes. It is, however, not clear whether spontaneous activation of EBV genomes in carrier lymphocytes, which are present in the lymphocyte rich area of the oropharynx, is responsible for lifelong persistence of antibody-titers directed against EBV related antigens and the shedding of virus into the oropharynx. Alternatively EBV could reside in specific sites of the body and be produced there resembling somehow the situation of Marek's disease virus of chicken. That virus is found in T-lymphocytes and produced and spread from the epithelium of featherfollicles. In situ hybridisations with frozen sections of tonsils and parotid glands reveiled that EBV genomes are present in the tissue of the parotid gland of healthy seropositive persons. The data from in situ hybridisations could be confirmed using stringent reassociationkinetics. It is suggested that spontaneous activation of the lymphocytes is a very rare event and neither the source of virus in the saliva nor the reason for the lifelong persistence of antibodies to EBV related antigens

Fatal lymphoproliferation and acute monocytic leukemia-like disease following infectious mononucleosis in the elderly

Three elderly patients are reported, in whom serologically confirmed recent infectious mononucleosis is followed by fatal lymphoproliferation (case 1), by acute monocytic leukemia (case 2), and by acute probably monocytic leukemia (case 3)

Purification and quantification of recombinant Epstein-Barr viral glycoproteins gp350/220 from Chinese hamster ovary cells

Truncated Epstein-Barr virus (EBV) membrane antigen gp350/220 (EBV-MA) lacking the membrane anchor was expressed and secreted into the medium of recombinant Chinese hamster ovary cells that had been cultured in Plasmapur hollow-fibre modules using defined serum-free medium. The EBV-MA in the medium was concentrated by 70% (w/v) ammonium sulphate precipitation and subsequently purified by immunoaffinity chromatography using an anti-EBV-MA (EBV.0T6) monoclonal antibody (mAb) column. Adsorbed antigen was eluted with 3 M MgCl2 in phosphate-buffered saline, concentrated by Mono Q anion-exchange chromatography and analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis, silver staining and Western blotting using EBV-positive serum and anti-EBV-MA specific mAbs. Monospecific polyclonal rabbit antibodies against the purified EBV-MA were raised and purified by protein G affinity chromatography. For the measurement of EBV-MA antigen levels a sandwich enzyme-linked immunosorbent assay using rabbit polyclonal antibodies and a horseradish peroxidase-conjugated anti-MA mAb was developed having a detection level of 10 ng/ml

BIOMECHANICAL ASPECTS OF THE POLE VAULT - ANALYSlS OF THE 4th IAAF WORLD CHAMPIONSHIP

World class pole vaulters were analysed at the 4th IAAF World Championships in Stuttgart, 1993 Performance relevant parameters were obtained using standard APAS procedures. DLT for non-panning cameras provided spatial coordinates for the 3-d analysis Graphical and numerical postprocessing of the data was done using Excel. In addition to the video graphic recording, the event was filmed with two panned 16mm Locam cameras operating at 100 Hz to determine temporal parameters. Furthermore approach velocities were measured using doubled IR photocells. Vertical CM accelerations in the free flight phase verified, that the digitally filtered data are reliable despite the large object space There was little lateral motion of the CM. Thus, for many applications, a 2-d analysis can be justified. Top vaulters clear heights 1,Zm above their grip height. Huffman cleared 5,Sm using a new 'straddle like' clearance. This was 1,17m above grip height. Compared to major competitions in the last decade approach speed stagnated or even decreased while performance improved. However, better vaulters tend to have higher speed and acceleration into the last approach section. High approach speed is a necessary but non-sufficient prerequisite for high vaults. Analysis of the time course of potential and kinetic energy reveals the importance of the enery at takeoff and the work done on the pole. These factors are not independent of each other. Thus, there is an optimum relationship between energy produced in the respective phases of the vault. This optimum depends on the current technical and conditional standard of the athlete. The inter- and intraindividual variations in the measured parameters support the notion, that compensation does take place and is indeed a common strategy on the world class level. Optimum technique remains a time variant, individual and situation specific motor pattern

Zur Virusätiologie der idiopathischen Fazialisparese (Enzymimmunserologische Untersuchungen) = On the viral etiology of Bell's palsy (An enzyme-linked immunosorbent assay study) [author's transl.]

In a prospective study paired sera of 14 patients suffering from Bell's palsy were examined for antibodies against Varicella-zoster Virus (VZV), Herpes-simplex Virus (HSV), Cytomegalovirus (CMV) and Epstein-Barr Virus (EBV). For determination of antibodies against VZV, HSV and CMV an enzyme-linked immunosorbent assay was carried out. Indirect immunofluorescence was carried out to determine antibodies against EBV. Serum samples were taken within 5 days of Bell's palsy having been diagnosed and were compared with further serum samples taken 4 weeks later. No evidence of significant differences between the antibody titers of the paired sera was found. The viral etiology of Bell's palsy due to an exogenous infection or by activation of a latent infection seems unlikely

Immunological reactivity of a human immunodeficiency virus type I derived peptide representing a consensus sequence of the GP120 major neutralizing region V3

To reduce the opportunities for human immunodeficiency virus type 1 (HIV-1) to evade vaccine induced immunity, the development of subunit vaccines must focus on the characterization of immunogenic epitopes, which are major targets for the immune system. The most dominant site for elicitation of neutralising immune response is located on the external envelope glycoprotein gp120 within the third variable domain (V3). To overcome virus type specificity of antibodies directed to the V3-domain we designed a 36 amino acids long gp120/V3-consensus peptide (V3-C36) based on published biological data and sequence comparisons of various HIV-1 virus isolates. This peptide contains a conserved core sequence which is suggested to form a surface-exposed beta-turn. This peptide also includes T-cell epitopes defined in mice and humans, an ADCC-epitope and two highly conserved cysteine residues which were oxidized to form a cystine derivate, thus allowing correct peptide folding. In ELISA-tests, this peptide reacts with at least 90% of randomly selected sera of European and African patients infected with HIV-1 and is recognized by three different HIV-1/V3 "type-specific" antisera (MN, RF, IIIB-strain). Using this peptide as immunogen in rabbits, antisera could be raised with highly cross-reactive and HIV-1/IIIB strain neutralizing properties. Moreover, HTLV/HIV-1/IIIB specific cytotoxic T-lymphocytes (CTLs) of BALB/c mice infected with a gp120 recombinant vaccinia virus recognized the central 16- and 12-mer peptides of the V3-C36 consensus peptide in cytolytic assays, indicating perfect compatibility of the consensus peptide with the IIIB-primed CTLs. The DNA-sequence encoding the V3-consensus loop region might be an important component in newly designed recombinant subunit vaccines. In addition, due to its broad serological reactivity, the V3-consensus peptide might play an important role in special diagnostic purposes