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Acta virologica

Volume 47 / 2004 / number 1

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Short Communications

Letters to the Editor


F.L. Xu1,2, Z.Q Yang1, C.C. Yang1,2*, S.Y. Xiao3, H. Xiao1, L. Wen1

*Corresponding author. Visiting professor of Institute of Virology, Medical School, Wuhan University, Wuhan, Hubei, P.R.China. E-mail: cyang@csmu.edu.tw; fax: +8864-23767469, +8627-87307966

1Institute of Virology, Medical School, Wuhan University, 115, Dong-Hu Road, Wuhan, Hubei 430071, P.R. China;
2School of Medical Technology, Chung Shan Medical University, 110, Section 1, Chien-Kuo North Road, Taichung, Taiwan 402, R.O.C.;
3Departments of Pathology and Internal Medicine, and Center for Biodefense and Emerging Infectious Diseases, University of Texas Medical Branch, Texas, USA

Received September 2, 2003; accepted January 22, 2004

Summary. – Hantavirus HV114, isolated from urine of a patient during epidemic of hemorrhagic fever with renal syndrome (HFRS) in China, was subjected to a detailed serological characterization using enzyme-linked immunosorbent assay (ELISA), neutralization test and indirect immunofluorescence antibody assay (IFA). It has been found that HV114 is antigenically similar to the hantavirus A9 strain isolated in China and to the Hantaan 76-118 virus (HTNV 76-118), but different from the hantaviruses isolated from Apodemus agrarius in the region endemic for HFRS.
Key words: hantavirus; HV114 virus; serology; ELISA; neutralization test; immunofluorescent assay

Acta virologica 48: 5 – 8, 2004



*Corresponding author. E-mail: olive40@uol.com.br; fax: +5521-30883753

1Instituto Adolfo Lutz, Sao Paulo, Brasil; 
2Faculdade de Saúde Pública, Universidade de Sao Paulo, Sao Paulo, Brasil; 
3Instituto de Ciencias Biomédicas, Universidade de Sao Paulo, Sao Paulo, Brasil

Received October 14, 2004; accepted February 18, 2004

Summary. – This study reports on molecular analysis of a Measles virus (MV) isolate from a patient who was infected in Japan but showed symptoms after arriving to Brazil. This patient had typical clinical measles infection symptoms: fever, rash, cough and coryza. After isolating the virus in B95a cells, a fragment of the nucleoprotein (N) gene was amplified by reverse transcription–polymerase chain reaction (RT-PCR) and subjected to direct nucleotide sequencing. The sequence data showed that the MV isolate of concern is of the D5 genotype.
Key words:
 Measles virus; nucleotide sequence; molecular epidemiology; nucleoprotein; Brazil

Acta virologica 48: 9 – 14, 2004

A Vero cell-attenuated goatpox virus provides protection against virulent virus challenge

M. Hosamani*, S. Nandi1, B. Mondal, R.K. Singh, T.J. Rasool, S.K. Bandyopadhyay1

*E-mail: m_hosa@email.com; fax: +91-5942286347
1Present address: Indian Veterinary Research Institute, Izatanagar, Bareilly 243 122, India.

Division of Virology, Indian Veterinary Research Institute, Mukteswar, Uttaranchal 263138, India

Received October 3, 2003; accepted February 23, 2004

Summary. – An Indian isolate of Goatpox virus (GTPV) was adapted and propagated in Vero cells for development of an attenuated virus. The virus was initially passaged in primary lamb testes cells and subsequently in Vero cells. At the 55th passage, the virus showed evidence of attenuation when tested for safety in sero-negative goats. At this stage, the virus was found to be completely non-pathogenic. The virus was passaged further and the 60th passage was usedfor testing its immunogenicity in goats. The latter were inoculated with 10, 100 and 1000 TCID50 of the attenuated virus by intradermal (i.d.) route and challenged after 28 days with virulent GTPV. The attenuated virus produced no adverse reaction even at the highest dose and conferred complete protection even at the lowest dose against challenge with a high dose (2 x 106 of 50% skin-reactive dose SRD50) of virulent virus. Increased levels of virus-specific serum antibodies could be demonstrated by both indirect enzyme-linked immunosorbent assay (ELISA) and virus neutralization (VN) test in all the immunized goats. No horizontal transmission of the virus from the immunized to in-contact animals took place. Our results suggest that this attenuated virus could be a safe, immunogenic and potent candidate for developing a vaccine against goatpox.
Key words: attenuated Goatpox virus; immunogenicity; Vero cells

Acta virologica 48: 15 – 21, 2004

Detection of Newcastle disease virus using A SYBR green I Real time POLYMERASE Chain Reaction

S.W. Tan1, A.R. Omar1*, I. Aini1, K. Yusoff2, W.S. Tan2

*Corresponding author. Fax: +603-89486317; E-mail: aro@vet.upm.edu.my

1Faculty of Veterinary Medicine, 
2Faculty of Science and Environmental Studies, University Putra Malaysia, 43400 UPM, Serdang, Selangor, Malaysia

Received September 17, 2003; accepted February 25, 2004

Summary. – A two-step SYBR Green I real time polymerase chain reaction (PCR, real time PCR) for the detection of Newcastle disease virus (NDV) was developed. A melting curve analysis was performed to distinguish specific from non-specific products and primer dimers. Regardless of different virus pathotypes the melting temperature (Tm) ranged from 86oC to 87oC. The sensitivity of the real time PCR was compared with the reverse transcription (RT)-nested PCR enzyme-linked immunosorbent assay (ELISA, RT-nested PCR ELISA). Whereas the detection limit of the real time PCR was 10 pg DNA, the RT-nested PCR ELISA and conventional PCR could only detect up to 1 ng and 10 ng DNA, respectively. Thus the real time PCR offers a sensitive, rapid and convenient method for screening large number of NDV specimens.
Key words: ELISA; Newcastle disease virus; SYBR Green I; real time PCR; RT-nested PCR ELISA

Acta virologica 48: 23 – 28, 2004

initial peptide mass fingerprinting analysis OF PROTEINS obtained by lysis of Coxiella burnetii cells

Ľ. Škultéty1, L. Hernychová2, R. Toman1, M. Kroča3, J. Stulík2, A. Macela2*

*Corresponding autor. E-mail: amacela@pmfhk.cz, fax: +420 49-5513018

1Institute of Virology, Slovak Academy of Sciences, 845 05 Bratislava, Slovak Republic; 
2Institute of Molecular Pathology and Proteome Centre for the Study of Intracellular Parasitism of Bacteria, Purkynì Military Medical Academy, 500 01 Hradec Králové, Czech Republic; 
3Central Military Medical Institute, 561 66 Techonin, Czech Republic

Received December 12, 2003; accepted March 3, 2004

Summary. – Whereas the complete genome of Coxiella burnetii (C.b.), the etiological agent of Q-fever, has recently been published (Seshadri et al., Proc. Natl. Acad. Sci. USA. 100, 5455–5460, 2003), the C.b. proteome is still under study. Using the bioinformatic approach, we found in total 309 proteins on two dimensional electroctrophoretic images of C.b. whole cell lysates. Eighteen major protein species were subjected to peptide mass fingerprinting and identified as the products of 6 known open reading frames (ORFs): the chaperone DnaK (heat shock 70 K protein), chaperonin 60 K (GroEL protein, heat shock protein B), DnaJ-like protein djlA (mucoidy activation protein mucZ), elongation factor Ts (EF-Ts), ribosomal protein L7/L12, and chaperonin 10 K (GroES protein, heat shock protein A).
Key words: Coxiella burnetii; proteins; peptide mass fingerprinting; 2-D electrophoresis; MALDI-ToF

Acta virologica 48: 29 – 34, 2004



*Corresponding author. E-mail: phusa@med.muni.cz; fax: +4205-32232221

1Department of Infectious Diseases, University Hospital Brno, Jihlavska 20, 625 00 Brno, Czech Republic; 
2Department of Clinical Haematology, University Hospital Brno, Brno, Czech Republic; 
3Medical Gastroenterological Department, University Hospital Brno, Brno, Czech Republic

Received January 16, 2004; accepted March 1, 2004

Summary. – Chronic hepatitis C infection is common among hemophiliacs in all the developed countries. Since 1996, only alpha-interferon (alpha-IFN) in monotherapy has been used for the treatment of chronic hepatitis C in hemophiliacs (6 patients). In Czech Republic a combination therapy with alpha-IFN and ribavirin has been used since 1999 (13 patients). Finally, a combination therapy with pegylated alpha-IFN (PEG-alpha-IFN) and ribavirin is being used since 2001 (still 3 patients). In all cases, the treatment lasted 48 weeks. A sustained virological response (SVR, defined as an undetectable serum HCV RNA level 24 weeks after the treatment was completed) was not achieved in any of 6 patients treated with alpha-IFN alone. A combination therapy with alpha-IFN and ribavirin yielded better results: four of eight patients still untreated with alpha-IFN (naive patients), one of two relapsers, and one of three non-responders to previous alpha-IFN monotherapy achieved SVR. So far the combination therapy with PEG-alpha-IFN and ribavirin has been used only in 3 patients. SVR was achieved in one patient who had relapsed after the combination therapy with IFN-alpha and ribavirin, and in 1 of 2 non-responders to this therapy. We conclude that the efficacy and tolerability of the treatment of chronic hepatitis C in hemophiliacs did not differ from that of chronic hepatitis C in other patients.
Key words: chronic hepatitis C; hemophilia; alpha-interferon; pegylated alpha-interferon; ribavirin

Acta virologica 48: 35 – 38, 2004

Detection of infectious bursal disease virus by ELISA using an antipeptide antibody raised against VP3 region

P. Saravanan1*, SatishKumar2, J.M. Kataria3, T.J. Rasool1

This study is a part of PhD thesis of the first author.
*E-mail: drsaravana72@dr.com; drsaravana72@epatra.com; fax: +91-581-286347

1Division of Virology, Indian Veterinary Research Institute, Mukteswar, Nainital, Uttaranchal, 263 138 India; 
2National Biotechnology Centre, Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh, India; 
3Division of Avian Diseases, Indian Veterinary Research Institute, Izatnagar, Uttar Pradesh, India

Received August 12, 2003; accepted March 2,2004

Summary. – Antigenic determinant analysis was carried out on VP3, one of major immunogenic proteins of Infectious bursal disease virus (IBDV) using computer algorithms. Altogether 17 peptides were synthesized for predicted putative regions and were tested for their reactivity with IBDV-positive polyclonal sera as well as with antisera to other common avian viruses to confirm specificity and to rule out cross reactivity. Of 17 peptides tested, three were selected and synthesized in multiple antigenic peptide (MAP) format. The immunization of rabbits with the three MAPs resulted in high humoral immune response. The purified antipeptide antibodies were screened against native IBDV antigen and the respective titers were determined. Out of the three antisera to MAPs that raised against the MAP3, spanning the amino acids (aa) 974–995 region on the VP3 protein had a very high titer (2048) and reacted specifically with IBDV. Thus, the antiserum to MAP3 detected native virus in enzyme-linked immunosorbent assay (ELISA), revealing the presence of a potential antigenic determinant on the C-terminus of the protein. This study proved that an antipeptide antibody could be used as a safe and specific tool for the diagnosis of IBD in chickens.
Key words: antipeptide antibodies; Infectious bursal disease virus; multiple antigenic peptide; antigenic determinants; diagnosis

Acta virologica 48: 39 – 45, 2004



*E-mail: rjkumar48@yahoo.com; fax: +25362787/+253889445

Department of Animal Biotechnology, Madras Veterinary College, Tamil Nadu Veterinary and Animal Sciences University, Chennai-600 0007, India

Received October 17, 2003; accepted February 18, 2004

Summary. – Rabies occurs in all parts of Indian sub-continent except Andaman and Nicobar and Lakshadweep group of islands. The full-length nucleoprotein (N) gene sequence of a rabies virus isolate from India is reported for the first time and the same has been compared with available N gene sequences from the database. A central domain of 230 amino acids (aa) from aa 141 to aa 370 exhibited more than 95% similarity. There were 8 amino acid positions (aa 29, 32, 38, 84, 119, 379, 438, and 439) at which substitution was unique for Indian isolates but common for laboratory strains. In antigenic epitopes, except for a single amino acid difference at the antigenic site IV, the amino acids were conserved. The Indian isolate also possessed two Bam HI sites (aa 247 and 278), while the other Asian isolates had only one site at aa 278 or were not digested with Bam HI at all. Phylogenetic analysis also demonstrated that the Indian isolate was closely related to the Sri Lankan isolate and grouped in the cluster that comprised of the isolates from other Asian countries namely China and Pakistan.
Key words: Rabies virus; Indian isolate; nucleoprotein; nucleotide sequencing; phylogenetic analysis

Acta virologica 48: 47 – 50, 2004

Ganciclovir resistance mutations in UL97 and UL54 genes of human cytomegalovirus isolates resistant to ganciclovir


*Corresponding author. E-mail: m-segondy@chu-montpellier.fr; fax: +33467-337793
1Present address: Département des Maladies Infectieuses et Tropicales, Hôpital Tenon, Paris, France
2Present address: Laboratoire de Virologie, Hôpital Bichat-Claude Bernard, Paris, France.

Laboratoire de Virologie, Hôpital Saint-Eloi, Centre Hospitalier Universitaire (CHU) de Montpellier, 80 Avenue Augustin Fliche, 34295 Montpellier Cedex 5, France

Received November 18, 2003; accepted February 24, 2004

Summary. – Human cytomegalovirus (HCMV) resistance to ganciclovir results from mutations in viral phosphotransferase (UL97) and/or DNA polymerase (UL54) genes. The HCMV isolates from the blood of immunocompromised patients with persisting presence of the pp65 antigen in the blood in spite of ganciclovir therapy were tested for ganciclovir susceptibility by an immediate-early antigen plaque reduction assay, and the UL54 and UL97 genes were sequenced. Nine isolates from eight patients (six patients with acquired immune deficiency syndrome (AIDS), one liver transplant recipient and one renal transplant recipient) showed phenotypic resistance to ganciclovir. All these ganciclovir-resistant HCMV isolates harbored one or more of the following UL97 mutations: M460V, A594V, A594T, L595S, C603W, and M615V. Two isolates harbored the P522S mutation in the UL54 gene. The M615V mutation in the UL97 gene has not been reported earlier and its role in ganciclovir resistance remains to be elucidated. In ganciclovir-resistant HCMV isolates the UL54 gene was less frequently mutated than the UL97 gene. The P522S mutation was relatively frequent in UL54-mutated HCMV isolates.
Key words: Human cytomegalovirus; resistance; ganciclovir; UL54; UL97

Acta virologica 48: 51 – 55, 2004

bovine respiratory syncytial virus strains currently circulating in the Czech Republic are most closely related to Danish strains from 1995

K. Kovařčík, V. Valentová*

*Corresponding author. E-mail: valentova@vri.cz; fax: +420541-211229.

Veterinary Research Institute, Hudcova 70, Brno 621 32, Czech Republic

Received January 23, 2004; accepted March 4, 2004

Summary. – In this study we showed a high degree of genetic homogeneity among recently (2002–2003) circulating Bovine respiratory syncytial virus (BRSV) strains in cattle population in the Czech Republic. These strains are in a phylogenetic tree more closely related to the Danish strains from 1995 than to the Czech strain VS97 from 1997 that shares the highest similarity with the French strain F1 and the Belgian strain P10. From the sequence analysis we deduce that the revealed high diversity between BRSV strains from 2002–2003 and those from 1997, at both nucleotide (0–11.4%) and amino acid (0–21%) level, is more likely due to distinct sources of the virus strains than to the sequence evolution.
Key words: Bovine respiratory syncytial virus; sequence analysis; phylogenetic analysis

Acta virologica 48: 57 – 62, 2004

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