Electronic Library of Scientific Literature - © Academic Electronic Press
Volume 45 / 2001 / number 4
E.N. PROKUDINA, N.P. SEMENOVA, I.A. RUDNEVA, V.M. CHUMAKOV, S.S. YAMNIKOVA
D.I. Ivanovsky Institute of Virology, Gamaleya St.16, Moscow 123098, Russia
Summary. – We have previously shown (Prokudina-Kantorovich EN and Semenova NP, Virology 223, 51–56, 1996) that the nucleoprotein (NP) of influenza A virus forms in infected cells oligomers which in the presence of SDS and 2-mercaptoethanol (ME) as reducing agent are stable at room temprature (RT) and dissociate at 100°C. Here we report that the efficiency of intracellular NP oligomerization depends on the host origin of influenza A virus strain. Thus, in the cells infected with avian influenza A virus strains the viral NP was almost completely oligomerized and only traces of monomeric NP were detected by polyacrylamide gel electrophoresis (PAGE) in unboiled samples. However, in the cells infected with human influenza A virus strains, besides oligomeric NP also a significant amount of non-oligomerized monomeric NP was detected in unboiled samples. In purified virions of avian and human strains the same difference in NP monomers/oligomers ratio was detected as in the infected cells. A reassortant having all internal protein genes from a human strain and the glycoprotein genes from an avian strain revealed the same intracellular pattern of NP monomers/oligomers ratio as its parental human virus. These findings suggest that the type of NP oligomerization is controlled by the NP gene. The possible connection between the accumulation of protease-sensitive monomeric NP in cells infected with a human influenza strain and the parallel accumulation of cleaved NP in these cells is discussed.
Key words: influenza
A virus; human strains; avian strains; reassortant; nucleoprotein;
glycoproteins; nucleoprotein oligomerization
Acta virologica 45: 201 – 207, 2001
Y.M. KNOX, K. HAYASHI, T. SUZUTANI, M. OGASAWARA, I. YOSHIDA, R. SHIINA, A. TSUKUI, N. TERAHARA, M. AZUMA
Department of Microbiology, Asahikawa Medical
College, 2-1, Midorigaoka-higashi, Asahikawa 078-8510, Japan;
Department of Development, Wada Sugar Refining Co. Ltd., Tokyo, Japan;
Technical Department, Nichino Kagakukogyo Co. Ltd., Saitama; Japan;
Tokyo Kasei Gakuin Junior College, Tokyo; Japan;
Department of Food Science and Technology, College of Horticulture,
Minami-Kyushu University, Miyazaki, Japan
Summary. – Earlier, we have detected antiviral activity in an extract from Ribes nigrum L. fruits (”Kurokarin”, name of the one species of black currant in Japanese) against influenza A and B viruses, and herpes simplex virus 1 (Knox et al., Food Processing 33, 21–23, 1998). In the present study, the antiviral activity of constituents of a Kurokarin extract and the mechanism of its antiviral action were examined. Kurokarin extracts were separated to fractions A to D by column chromatography. The major constituents of the fraction D were estimated as anthocyanins. The fraction D was further fractionated by thin-layer chromatography (TLC) to fractions A' to G'. The fraction E' consisted of 3-O-alpha-L-rhamnopyranosyl-beta-D-glucopyranosyl-cyanidin and 3-O-beta-D-glucopyranosyl-cyanidin, and the fraction F' consisted of 3-O-alpha-L-rhamnopyranosyl-beta-D-glucopyranosyl-delphinidin and 3-O-beta-D-glucopyranosyl-delphinidin, identified by high performance liquid chromatography (HPLC) with standards and by high resolution mass spectrometry. The fractions D' to G' showed potent antiviral activity against influenza viruses A and B. The additive antiviral effect of a combination of the fractions E' and F' was assessed. Anthocyanins in the fraction F' did not directly inactivate influenza viruses A and B, but they inhibited virus adsorption to cells and also virus release from infected cells.
Key words: Ribes
nigrum L.; Kurokarin; anthocyanins; antiviral activity; influenza
viruses A and B
Acta virologica 45: 209 – 215, 2001
L.K. CHONG, A.R. OMAR, K. YUSOFF, M. HAIR-BEJO, I. AINI
Faculty of Veterinary Medicine and
Faculty of Science and Environmental Studies, University of Putra, 43400
UPM Serdang, Selangor, Malaysia
Summary. – The complete nucleotide sequences encoding precursor polyprotein (VP2-VP3-VP4) and VP5 of a highly virulent (hv) infectious bursal disease virus (IBDV), UPM97/61 was determined. Comparison of the deduced amino acid sequences with the published ones revealed 8 common amino acid substitutions, which were found only in the hv IBDV including the UPM97/61 strain. Three of the amino acid substitutions (222 Ala, 256 Ile and 294 Ile) were used as a marker for determining hv IBDV strains. The other five substitutions (685 Asn, 715 Ser, 751 Asp, 990 Val and 1005 Ala) were also conserved in hv IBDV strains isolated in various countries. UPM97/61 strain demonstrated also 8 unique amino acid substitutions of which 3 were in VP2, 4 in VP3 and 1 in VP4. There was 1 unique amino acid substitution in VP5 at position 19 (Asp->Gly) not found in other strains. However, all the strains have a conserved 49 Arg. The amino acid sequence of UPM97/61 strain differred by 1.09% from the Japanese (OKYM) and Hong Kong (HK46) strains, and by 1.48% from the Israeli (IBDVKS) and European (UK661) strains. Hence, UPM97/61 is more closely related to the hv strains from Asia. However, phylogenetic analysis indicated that the origin of UPM97/61 might be the same as that of other hv strains isolated from other parts of the world.
Key words: infectious
bursal disease virus; A segment; nucleotide sequence; deduced amino acid
sequence
Acta virologica 45: 217 – 226, 2001
K. POLÁKOVÁ, G. RUSS
Cancer Research Institute, Slovak Academy of
Sciences, Vlárska 7, 833 91 Bratislava, Slovak Republic;
Institute of Virology, Slovak Academy of Sciences, Bratislava, Slovak
Republic
Summary. – The immunodominant peptide of human immunodeficiency virus 1 gp 160 for murine cytotoxic T cells of H-2d haplotype, has been originally identified as a 15 amino acid residue peptide P18IIIB (RIQRGPGRAFVTIGK) (Takahashi et al., 1988). Further studies have indicated that a more active form of the peptide is generated by removal of the C-terminal dipeptide by angiotensin-I-converting enzyme (ACE), and additional detailed studies have shown that the actual immunodominant peptide is a decamer P18-I10 (RGPGRAFVTI) (Kozlowski et al., 1993). The effect of proteolytic processing on the antigenicity of P18IIIB peptide and its analogs was investigated by functional T cell assays based on the ability of T cell receptor (TCR) to recognize a specific major histocompatibility complex class I (MHC-I)/peptide complex. Recently we described a new monoclonal antibody (MAb) KP15 directed against the MHC-I molecule H-2Dd complexed with the10-mer peptide P18-I10. Using this MAb, the cell surface H-2Dd/P18-I10 complex can be easily detected by flow cytometry (Polakova et al., 2000). Here we examined whether peptides longer than P18-I10 decamer form H-2Dd complexes recognized by KP15 MAb. Further we also analyzed how the ACE processing of P18IIIB-related peptides of different length affects their ability to form complexes with H-2Dd recognized by MAb KP15. These experiments confirmed that the ACE digestion of 15-mer peptide P18IIIB is the most effective in the production of a peptide capable of forming complex with H-2Dd recognized by KP15 MAb. The ACE digestion of longer peptides (16-mer to 19-mer) did not produce a significant quantity of peptides, capable of forming H-2Dd complexes recognizable with by MAb KP15. Peptides shorter than P18IIIB (13-mer to 10-mer), notably the optimally sized P18-I10 peptide lost most of their capacity to form H-2Dd complexes recognized by KP15 MAb. Our results show that the extracellular processing of MHC-I-restricted peptides, which cannot be overlooked in designing peptide-based vaccines, can be also studied by as simple and rapid assay as flow cytometry, provided MAbs specific to a particular MHC-I/peptide complex are available.
Key words: human
immunodeficiency virus 1; glycoprotein gp160; exogenous processing,
angiotensin-I-converting enzyme
Acta virologica 45: 227 – 234, 2001
A. RAUT, R. K. SINGH, M. MALIK, M. C. JOSEPH, C. S. BAKSHI, V. V. S. SURYANARAYANA, G. BUTCHAIAH
National Biotechnology Center, Indian
Veterinary Research Institute, Izatnagar 243122, India;
Rajiv Gandhi College of Veterinary and Animal Sciences, Pondicherry,
India
Summary. – The currently used Plowright’s tissue culture rinderpest vaccine (RBOK strain) gives full protection and lifelong immunity, but it is highly thermolabile and requires maintenance of cold chain from vaccine production till delivery. Keeping in view the need for a thermostabile vaccine in tropical developing countries with limited refrigeration facilities, we passaged serially the RBOK strain of rinderpestvirus (RPV) at gradually elevated temperature up to 40°C to obtain a thermoresistant RPV (TR-RPV) mutant. The thermoresistance (thermostability) and antigenicity of TR-RPV were compared with those of the vaccine virus by various methods, confirming the acquired properties. Thus, the infectivity titres of the TR-RPV mutant and vaccine virus were determined after incubation for various times at 37°C. Regression analysis indicated that TR-RPV had a half-life of 1.81 hr and a degradation constant of 0.1656, while the parent vaccine virus had a half-life of 1.11 hr and a degradation constant of 0.2686. In capture ELISA with four different monoclonal antibodies (MAbs) to the N protein of RPV, TR-RPV showed a 10-fold higher reactivity with one MAb as compared to the vaccine virus. Although TR-RPV did react also with the other three MAbs, its reactivity was only 4–5 times higher than that of the vaccine virus. A treatment of the virus with Triton X-100 resulted in 2–4 times higher reactivity with the MAbs. The 35S-methionine-labeled vaccine virus-and TR-RPV-infected Vero cell lysates showed 6 polypeptide bands with identical pattern of migration in polyacrylamide gel electrophoresis in the presence of SDS (SDS-PAGE). Radioimmunoprecipitation assay (RIPA) of the TR-RPV and vaccine virus with a rabbit anti-RPV immune serum (RHIS) and bovine anti-RPV hyperimmune serum (BHIS) showed the presence of four identical antigenic proteins, namely H, N, F and M, for both viruses. It can be concluded that TR-RPV has indeed retained the antigenic properties of the parental vaccine virus besides acquiring thermoresistance.
Key words: rinderpest
virus; RBOK strain; thermoresistant mutant; capture ELISA; SDS-PAGE;
RIPA; vaccine
Acta virologica 45: 235 – 241, 2001
Š. NĚMEČKOVÁ, P. HAINZ, P. OTÁHAL, P. GABRIEL, V. ŠROLLER, L. KUTINOVÁ
Institute of Hematology and Blood Transfusion, Department of Experimental Virology, U nemocnice 1, 128 20 Praha 2, Czech Republic
Summary. – Modified vaccinia virus Ankara strain (MVA) is a safe highly attenuated non-pathogenic virus suitable as a vector for developing various vaccines. Study of expression of a reporter ß-galactosidase gene under the control of an early vaccinia virus (VV) promoter in MVA and non-modified vaccinia virus Praha strain showed that early transcription in MVA is elevated in comparison with non modified VV. This property was demonstrated in various cell cultures including CV1 cells, human lung diploid cells, chicken primary fibroblasts but not in bone marrow-derived mouse dendritic cells. There the relationship between the elevated early transcription and the permisivity of cells for MVA was not observed.
Key words: vaccinia
virus; poxviruses; MVA; gene expression; dendritic cells; early promoter
Acta virologica 45: 243 – 247, 2001
X. WANG, S. CHANG, Z. JIN, L. LI, G. ZHOU
State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100094, China
Summary. – The nucleotide sequences of the coat protein (CP) and readthrough protein (RTP) genes of the Chinese GAV isolate of Barley yellow dwarf virus (BYDV) were determined. The CP and RTP genes of GAV isolate comprised 600 and 1374 nucleotides, respectively. When the CP and RTP gene sequences of GAV isolate were compared with those of BYDV isolates MAV-PS1, P-PAV, NY-SGV and Cereal yellow dwarf virus RPV (CYDV-RPV), the highest similarity (97.2%) between the CP genes of GAV and MAV-PS1 isolates was observed, while the RTP genes of these two isolates shared a lower similarity (87.8%). The results of the alignment of the deduced amino acid sequences of RTP showed that the sequence diversity observed was located at the C terminus.
Key words: Barley
yellow dwarf virus; GAV isolate; MAV-PS1 isolate; P-PAV isolate; NY-SGV
isolate; Cereal yellow dwarf virus RPV; coat protein gene; readthrough
protein gene; nucleotide sequences; deduced amino acid
sequences;alignment; transmission phenotype
Acta virologica 45: 249 – 252, 2001
L. KABILAN
Center for Research in Medical Entomology, Sarojini Street 4, Chinnachokkikulam, Madurai 625 002, Tamil Nadu, India
Summary. – Japanese encephalitis virus (JEV) infections are currently detected by indirect immunofluorescence (IF) assay using virus-specific antibodies on acetone-fixed smears. On few occasions, the acetone treatment was reported to damage certain epitopes on JE virus (JEV) glycoprotein. Here, we have made an attempt to adopt quick paraformaldehyde fixation followed by a short detergent treatment of cells in suspension for identification of JEV-infected brain cells of laboratory-reared Toxorhynchitis splendens mosquito larvae using virus-specific antibodies. JEV-positive cells could be scored by the presence of a well defined intracellular immunofluorescence staining against unstained uninfected antibody-treated cells. The advantage of this assay is that stained cell suspensions can be stored for up to 4 weeks, allowing analysis at convenience. Thus, the modified IF assay can be employed as an additional/alternate technique to standard IF assay for detection of JEV in cells and also to screen hybridoma cell lines for anti-JEV antibody production.
Key words: antibodies;
immunofluorescence; assay; Japanese encephalitis virus; mosquito larval
brain cells; paraformaldehyde; saponin
Acta virologica 45: 253 – 255, 2001
D. JANUSZKIEWICZ-LEWANDOWSKA, J. WYSOCKI, H. JÓŹWIAK, K. LEWANDOWSKI, J. REMBOWSKA, J. NOWAK
Institute of Human Genetics, Polish Academy of
Sciences, Strzeszynska 32, 60-479 Poznań, Poland;
Academy of Medical Sciences, Poznań;
Department of Medical Diagnostics, Poznań
Summary. – Viral etiology of hepatitis is routinely proved by standard immunological tests detecting specific antibodies. However, identification of specific antibodies cannot always be conclusive. Since specific hepatitis C virus (HCV) antibodies may appear after some months of the infection, identification of HCV RNA and/or hepatitis G virus (HGV) RNA should clarify the etiology of hepatitis. The aim of this study was to diagnose etiologically unknown hepatitis by a reverse transcription–polymerase chain reaction (RT-PCR) testing of the presence of HCV RNA and HGV RNA. The study involved 33 children with histologically proved hepatitis. The presence of HCV and any signs of autoimmune disease were not observed at the beginning of the follow-up study. During 2.5 years of the follow-up HCV-RNA was found in the blood and liver biopsies in 17 patients. Eight of them became HCV antibodies-positive during the follow-up. None of them eliminated the virus from the blood during the follow-up. In two other patients HCV-RNA was found only in the liver. HGV infection in all cryptogenic patients was excluded by PCR testing. Identification of HCV RNA RT-PCR allowed to diagnose 19 out of 33 (57.6%) patients with cryptogenic hepatitis. The etiology of the hepatitis in remaining 12 patients has to be established.
Key words: children;
cryptogenic hepatitis; hepatitis C virus; RNA
Acta virologica 45: 257 – 260, 2001
D. JANUSZKIEWICZ-LEWANDOWSKA, J. WYSOCKI, J. REMBOWSKA, K. LEWANDOWSKI, T. NOWAK, M. PERNAK, J. NOWAK
Institute of Human Genetics, Polish Academy of
Sciences, Strzeszynska 32, 60-479 Poznań, Poland;
Institute of Pediatrics, University of Medical Sciences, Poznań;
Department of Medical Diagnostics, Poznań
Summary. – Hemodialysis patients are at risk for hepatitis C virus (HCV) and hepatitis G virus (HGV) infection. The aim of this study was to investigate the possible influence of HGV co-infection on HCV RNA elimination from the peripheral blood of hemodialysis patients. The study involved 144 persons, all with HCV antibodies and HCV RNA. Among 144 patients 24 (16.7%) were positive for HGV RNA. After 2.5 years of observation 80 patients (55.6%) were still HCV RNA-positive. In the latter group 18 patients were co-infected with HGV and 62 were HGV RNA-negative. During 2.5 years of the follow-up study 64 patients eliminated HCV RNA from the serum. In this group only 6 patients were HGV co-infected. None of the HGV-positive patients eliminated HGV RNA from the serum. The higher incidence of HGV co-infection in the group of patients who remained HCV RNA-positive (18/80, 22.5%), in comparison to the group of HCV antibodies-positive patients who lost HCV in the blood (6/64, 9.4%, P <0.0001) suggests, that the co-infection with HGV may delay the spontaneous elimination of HCV RNA from the blood.
Key words: HCV;
HGV; co-infection; HCV RNA; HGV RNA; HCV RNA elimination; hemodialysis
patients
Acta virologica 45: 261 – 263, 2001
Antiviral Agents – Advances and
Problems
E. Jucker (Ed.): Antiviral Agents - Advances and Problems. A
special topic volume, the series Progress in Drug Research. Birkhäuser
Verlag, Basel-Boston-Berlin, 2001, 258 pp
11th TOMÁŠEK DAYS - Conference of Young Microbiologists with International Participation, June 5-7, 2002, Brno, Czech Republic
Electronic Library of Scientific Literature - © Academic Electronic Press