Electronic Library of Scientific Literature Electronic Library of Scientific Literature
Volume 44 / February 2000 / number 1
Y.A. Smirnov, A.S. Lipatov, R. van Beek, A.K. Gitelman, A.D.M.E. Osterhaus, E.C.J. Claas
The D.I. Ivanovsky Institute of Virology, Russian Academy of Medical Sciences, Gamaleya
16, 123 098 Moscov, Russsia;
National Influenza Center, Department of Virology, Erasmus University, Rotterdam, the
Netherlands;
Department of Virology, CKVL, Leiden University Medical Center, Leiden, the Netherlands
Summary. - We have used the mouse model to monitor the acquisition of virulence of a non-pathogenic influenza A virus upon adaptation to a new mammalian host. An avian strain, A/Mallard duck/Pennsylvania/10218/84 (H5N2) (Mld/PA/84) was adapted to mice by 23 serial lung-to-lung passages until a highly virulent mouse-adapted (MA) variant (Mld/PA/84-MA) emerged. This MA variant was characterized and compared to the parental strain as well as some of its intermediate passage variants. MA variant caused bronchopneumonia in mice with a high mortality rate (the virulence of Mld/PA/84-MA measured as log (EID50/LD50) was 1.75), while the parental, avirulent strain Mld/PA/84 did not cause illness and mortality in mice (log (EID50/LD50) was 7.25). Hemagglutination-inhibition (HAI) test with a set of hemagglutinin- (HA) specific monoclonal antibodies (MAbs) revealed antigenic differences between the parental strain and MA variant. Mld/PA/84-MA reacted with HA-specific MAbs in higher titers than the parental strain. The HA genes of the parental strain Mld/PA/84, the 1st, 3rd, 8th, and 15th intermediate passage variants, and Mld/PA/84-MA were sequenced. Three amino acid changes at positions 203, 273 and 320 were determined in the HA of MA variant. The first of them, Leu -> Pro (320), appeared in the HA stem region at the 8th passage. Two other in the HA1 globular region (Ser -> Phe (203) and Glu -> Gly (273)) appeared at the 15th passage. All of these substitutions were associated with the increase of viral infectivity for mouse lungs and changes in the HA antigenicity. The potential role of these changes in HA with respect to the process of viral interspecies transmission and acquisition of virulence for new host is discussed.
Key words: avian influenza virus; H5N2; mammalian host; adaptation
Acta virologica 44: 1 - 8, 2000
I.D. GARG, S.M.P. KHURANA
Central Potato Research Institute, Shimla 171 001, India
Summary. - During isolation of strains of potato viruses X (PVX) and A (PVA) from indigenously collected potato germplasm, an inseparable association between these viruses was discovered. As a result, all the hosts of PVX, used to free PVX from PVA, also showed infection of PVA along with PVX. Furthermore, Nicandra physaloides, which is a host of PVA but not PVX, also did not free PVA from PVX. These results suggested a reciprocal complementation of movement function of these viruses due to which they together infected various hosts sensitive to PVX or PVA. Relative concentration of PVX, in all the hosts tested, was much higher than that of PVA.
Key words: potato virus A; potato virus X; movement function;
complementation
Acta virologica 44: 9 - 13, 2000
S.S. KIM, E.Y. KIM, K.Y. PARK, S.D. SUH, H.K. PARK, Y.O. SHIN, M. BAE, J.S. LEE
Center for AIDS Research, Department of Virology, National Institute of Health, 5
Nokbun-dong Eunpyung-gu, Seoul, 122-701 South Korea;
Department of Microbiology, Kangwon National University College of Medicine, Chunchon,
South Korea;
Department of Biological Science, Ewha Womans University, South Korea
Summary. - Although human immunodeficiency viruses 1 and 2 (HIV-1 and HIV-2) share mode of transmission, their epidemiologic characteristics differ and international spread of HIV-2 has been limited. To investigate the extent of HIV-2 infection in South Korea and to clarify the characteristic of HIV-2 isolates, we describe epidemiological, serological and genetic analyses of five HIV-2 isolates from South Korea. Five of 964 HIV antibody-positive serum specimens showed positive reactivity by HIV1/2 enzyme immunoassay (EIA), HIV-2 Western blot analysis, HIV-2 particle agglutination (PA) test and line immunoassay (LIA) but negative or indeterminate one by HIV-1 PA test and HIV-1 Western blot analysis. To confirm HIV-2 infection by genetic analysis, reverse transcription-polymerase chain reaction (RT-PCR) was performed on five HIV-2 seropositive samples. PCR products from gag (197 bp) and env gene regions (137 bp) were obtained with three of the five samples with HIV-2 specific gag primers and with all the five samples with env primers. To obtain larger sequences for a more comprehensive phylogenetic analysis, we performed PCR for a 1191 bp env region of HIV-2 but only two such products were obtained. For the phylogenetic analysis, three 197 bp gag and two 1191 bp env PCR products were cloned and sequenced. Based on the gag and env sequences alignments, three isolates (KR4063, KR7051 and KR8091) were clustered phylogenetically within HIV-2 subtype A. In conclusion, HIV-2 virus is present in South Korea and was detected in five subjects. Furthermore, the prevalence of HIV-2 infection should be monitored continuously in South Korea to assess the spread of this virus and to assist in the diagnosis of HIV infection.
Key words: HIV-2; PCR; nucleotide sequencing; serological
diagnosis; South Korea
Acta virologica 44: 15 - 22, 2000
J. Stránský, L. Malina, B. Cieslarová, J. Stříteský, I. Půtová, J. Horák
Department of Internal Medicine I and Department of Dermatology and Pathology, Third Faculty of Medicine, Charles University, Šrobárova 50, 100 34 Prague 10, Czech Republic
Summary. - The aim of this study was to assess the rate of hepatitis B virus (HBV) and hepatitis C virus (HCV) coinfection (“the coinfection”) in chronic liver disease (CLD) and to reveal overt and hidden HBV infection in patients with antibodies to HCV (anti-HCV). A total of 209 untreated patients (64 with chronic hepatitis B, 79 with chronic hepatitis C and 66 with porphyria cutanea tarda (PCT)) were screened for serological markers of HBV and HCV infection in serum by third generation enzyme-linked immunosorbent assay (ELISA) methods and for HBV DNA and HCV RNA in serum by polymerase chain reaction (PCR). The rate of the overt coinfection in chronic hepatitis B was very low (2/64, 3%). However, in chronic hepatitis C, the rate of the hidden coinfection with HBV was relatively high (19/79, 24%); these patients had higher alanine transaminase (ALT) and asparagine transaminase (AST) levels in serum and a more advanced liver disease. In PCT patients, the rates of HBV and HCV infections were the same, 21% (14/66). In the PCT patients infected with HBV or HCV, the rate of the coinfection was 33% (7/21). The PCT patients with the coinfection had a high serum ALT level and the worst histological picture in the liver. The hidden HBV infection was more frequent than the overt one. The possibility of the overt or hidden coinfection in CLD renders a detailed analysis of all serum samples for both viruses mandatory. Vaccination against HBV infection should be offered to anti-HCV-positive individuals as well as to PCT patients not showing antibodies to HBV (anti-HBV).
Key words: hepatitis B virus; hepatitis C virus; antibodies;
antigens; coinfection; porphyria cutanea tarda; chronic liver disease; vaccination
Acta virologica 44: 23 - 28, 2000
C. EROGLU, E. YILDIZ, M. OZTURK, E. PINARBASI
Department of Molecular Biology and Genetics, Science Faculty, Bilkent University,
Bilkent Ankara, Turkey;
Department of Medical Biology and Genetics, Medicine Faculty, Cumhuriyet University, 58140
Sivas, Turkey
Summary. - In this study, a 178 amino acids long portion of the hepatitis C virus (HCV) core gene was cloned, sequenced, expressed in Escherichia coli, and purified. The resulting antigen (C178) was tested with human sera enzyme-linked immunosorbent assay (ELISA) in order to assess its ability to diagnose HCV. It was shown by ELISA that 92% of the patients sera, diagnosed previously by a 3rd generation enzyme immunoassay (EIA) as HCV-positive, had antibodies against the C178 antigen. This antigen gave no false positive results when tested with anti-HCV-negative sera.
Key words: hepatitis C virus; core protein gene; cloning;
nucleotide sequencing; expression; E. coli; purification; ELISA; RT-PCR
Acta virologica 44: 29 - 33, 2000
E. Cánepa, M.M. Siqueira, M. Hortal, F. Friedrich
Departamento de Laboratórios de Salud Pública, Ministério de Salud Pública,
Montevideo, Uruguai;
Departamento de Virologia, Instituto Oswaldo Cruz, FIOCRUZ, 21040-360 Rio de Janeiro, RJ,
Brazil
Summary. - In the summer 1999, a measles outbreak occurred in Uruguai. During this outbreak 58 cases were recorded, 36 of which were laboratory confirmed as positive for measles virus (MV) IgM. The cases occurred in touristic places (Montevideo and Maldonado) predominantly among health facilities and tourist service personnel. Urine specimens collected between days 1 and 4 after the onset of the rash from seven cases were analyzed by reverse transcription-polymerase chain reaction (RT-PCR) and nested PCR with primers specific for the carboxyl-terminal region of the nucleoprotein (N) gene. Three of these specimens/cases were positive for MV. Sequencing of 300 nucleotides (nt) of PCR products corresponding to a part of the carboxyl-terminal region of the MV N gene detected in these specimens MV of D6 genotype. The same nucleotide sequences and the same genotype were also previously observed for MV isolates from the 1997 epidemic in Brazil and the 1998 epidemic in Argentina, demonstrating that the D6 genotype was, and may be still circulating in South America.
Key words: measles virus; outbreak; RT-PCR; nucleotide sequence;
molecular epidemiology; Uruguai
Acta virologica 44: 35 - 39, 2000
J. MATOUŠEK, J. PTÁČEK, P. DĚDIČ, J. SCHUBERT
Department of Molecular Genetics, Institute of Plant Molecular Biology, Czech Academy
of Sciences, Branišovská 31, 370 05 České Budějovice; Czech Republic;
Institute for Potato Research, Havlíčkův Brod, Czech Republic;
Federal Centre for Breeding Research, Institute for Resistance Research and Pathogen
Diagnostics, Aschersleben, Germany
Summary. - Reaction conditions specific for reverse taranscription-polymerase chain reaction (RT-PCR) of potato virus Y strain NTN (PVYNTN) were used to amplify a 394 bp fragment of the P1 gene from selected PVY isolates with the aim to study the PVY variability within this genomic region. The P1 gene fragment from the Nicola isolate (Nicola P1/1 clone) was sequenced and characterized by temperature-gradient gel electrophoresis (TGGE). The Nicola P1/1 clone differed from that from the Hungarian isolate by double point mutation resulting in two changes at the deduced amino acid level. The clone showed simple transition from double-stranded to single-stranded form with two characteristic melting end points of about 41°C and 48°C. A more complicated TGGE pattern was, however, found for the whole P1 cDNA library of the Nicola isolate, suggesting accumulation of some minor sequence variants of PVY in this isolate. Based on the TGGE pattern, 46°C was selected as the standard temperature for electrophoretic analysis of heteroduplex DNAs formed with the Nicola P1/1 DNA as reference. More than 40 other PVY isolates from PVYN group were analysed using this method. In most cases only minor fractions of electrophoretically distinguishable DNA heteroduplexes were found, however, in most isolates of PVYN-Wilga type, mixtures of the major sequence variants were observed. Two of these variants from the hybrid 220-5 (Czech Republic) were sequenced. Both of them differed from the Nicola P1/1 clone by 6 point mutations, which led to several changes at the amino acid level.
Key words: potato virus Y; P1 gene; Solanum tuberosum; virus
quasispecies; mixed virus infection; RT-PCR; cloning; nucleotide sequencing; TGGE;
heteroduplex DNA
Acta virologica 44: 40 - 46, 2000
J. FRÁNOVÁ
Department of Plant Virology, Institute of Plant Molecular Biology, Academy of Sciences of the Czech Republic, Branišovská 31, 370 05 České Budějovice, Czech Republic
Summary. - Deadly nightshade plants showing severe necrotic lesions on leaves were observed in southern Bohemia. In negatively stained preparations of spontaneously infected deadly nightshade, artificially inoculated host plants and purified preparations two types of virus particles, isometric ones of about 26 nm in diameter and flexuous ones with length of 765 nm were seen by electron microscopy. The virus with isometric particles was identified as belladonna mottle virus (BMV), indistinguishable serologically from the Hungarian isolate of this virus. Identification of the virus with flexuous particles is discussed. Observations of the ultrastructure revealed the presence of filamentous virus particle aggregates and chloroplasts with peripheral vesicles bounded by double membranes, a feature typical for tymoviruses.
Key words: Atropa belladonna; belladonna mottle virus; electron
microscopy; flexuous virus particles; ultrastructure; virus aggregates
Acta virologica 44: 47 - 51, 2000
R. OTSU, A. ISHIKAWA, K. MUKAE
Faculty of Health Science, Kyushu University of Health and Welfare, Yoshino, Nobeoka,
Miyazaki, 882-8508 Japan;
Kyushu Sangyou University, Fukuoka, Japan
Summary. - In testing 60 stool specimens small round structured virus (SRSV) particles were detected in 35 (58%) specimens from all 17 outbreaks of gastroenteritis by electron microscopy (EM), while SRSV genes were found in 36 (60%) specimens from 15 outbreaks by reverse transcription-polymerase chain reaction (RT-PCR) by use of 2 primer pairs. Specimens from 2 outbreaks were found SRSV particles-positive by EM but SRSV genes-negative by RT-PCR. EM remains the basic examination method for diagnosis of SRSV agents.
Key words: small round structured virus; gastroenteritis;
detection; electron microscopy; RT-PCR
Acta virologica 44: 53 - 55, 2000
J.J. Paxman, A.M. Enriquez, J.T. May
Department of Microbiology, Faculty of Science, Technology and Engineering, La Trobe University, Bundoora, Victoria 3083, Australia
Acta virologica 44: 57 - 59, 2000