7 th NATIONAL CONGRESS OF THE ITALIAN SOCIETY OF VIROLOGY

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1 7 th NATIONAL CONGRESS OF THE ITALIAN SOCIETY OF VIROLOGY Orvieto (TR) Palazzo del Capitano del Popolo June 24-26,

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3 COMMITEES President of the Congress Palù Giorgio Scientific Committee Franco Buonaguro Canio Buonavoglia Gabriella Campadelli-Fiume Francesco Maria Cancellotti Maurizio Conti Carlo De Giuli Morghen Antonina Dolei Giuseppe Gerna Colomba Giorgi Giorgio Gribaudo Maria Paola Landini Roberto Manservigi Giorgio Palù Maria Cristina Parolin Carlo Federico Perno Mauro Pistello Luisa Rubino Franco Maria Ruggeri Paola Verani Borgucci Maurizio Zazzi Organizing Committee Elisabetta Affabris Alberta Azzi Francesco Maria Cancellotti Colomba Giorni Anna Maria Iorio Franco Maria Ruggeri Cristiano Salata Eurovirology Travel Grant and poster-prize Committee Canio Buonavoglia Gabriella Campadelli-Fiume Antonina Dolei Giovanni Martelli Giorgio Palù 3

4 SECRETARIATS SCIENTIFIC SECRETARIAT SIV - Italian Society of Virology c/o Department of Histology, Microbiology and Medical Biotechnologies University of Padova - Italy The Italian Society of Virology is supported by ORGANIZING SECRETARIAT MZ CONGRESSI Member of the MZ International Group (Milano, Torino, Barcelona, London) Via Carlo Farini, Milano - Italy Phone Fax Speakers, chairs and companies: Patrizia Sirtori - Registrations: ACKNOWLEDGMENTS We thank the Comune di Orvieto for its continuous support. GENERAL INFORMATION OFFICIAL LANGUAGE The official language of the symposium is English CME Italian CME Credits have been assigned by the Italian Ministry of Health for the following categories: - Medical Doctor: 10 credits (Specialist Areas: Microbiology and Virology, Infectious Diseases, Pharmacology and Clinical Toxicology) - Biologist: 10 credits (Specialist Areas: Microbiology and Virology) - Biomedical Laboratory Technician: 11 credits Italian CME certificates will be sent after the Congress directly to the address indicated on the registration form. 4

5 PROGRAMME Sunday, 24 June : Registration 17:30 Opening and welcome Chair: Antonina Dolei (Sassari) 17:45 GB Rossi Lecture New aspects of interferon systems, immune evasion and new potential applications Filippo Belardelli (Roma) GENERAL VIROLOGY AND VIRAL GENETICS Chairs: Maurizio Conti (Torino), Maria Cristina Parolin (Padova) Selected oral presentations Increasing circulation and diversification of HIV-1 non-b subtypes in Italy over a 10- year observation period F. Razzolini Detection and quantification of noroviruses in environmental samples by newly designed nested PCR assays and Taqman Real-Time RT-PCR M. Muscillo UL products are required for the activation of HCMV fusion complex during endothelial cell infection M. Patrone Prevalence and Persistence of human papillomavirus genotypes and their variants in Human Immunodeficiency Virus (HIV) positive women from south-italy F.M. Buonaguro Polyomavirus BK and anti-dsdna antibodies in renal transplant recipients C. Costa Inconsistent responses of cytomegalovirus-specific T-cells to pp65 and IE-1 vs infected dendritic cells in organ transplant recipients D. Lilleri 5

6 Monday, 25 June 2007 MEDICAL VIROLOGY AND ANTIVIRAL THERAPY Chairs: Carlo Federico Perno (Roma), Maurizio Zazzi (Siena) KeyNote Lecture Antiviral strategies, mechanism, and clinical efficacy Massimo Puti (Brescia) 09: Selected oral presentations Human respiratory syncytial virus (hrsv) RNA quantification in nasopharyngeal secretions identifies the hrsv etiologic role in acute respiratory tract infections of hospitalized infants G. Campanini The human bocavirus role in acute respiratory tract infections of pediatric patients as defined by viral load quantification G. Gerna Antiviral activity of Retinoic Acid derivatives against HHV-8 E. Caselli Pre-transplant analysis of preservation and washing solutions for rapid detection of viruses in the kidney graft M. Pacenti Antiviral activity of WC5, a new 6-Aminoquinolone compound, against human herpesviruses B. Mercorelli BREAK VIRAL BIOTECHNOLOGIES AND GENE THERAPY Chairs: Giorgio Gribaudo (Torino), Giorgio Palù (Padova) Selected oral presentations Persistent viral detection in the kidney allograft is a risk factor for chronic allograft injury L. Barzon Viral clearance evaluation model using 4 different viruses for GMP-Production of biopharmaceutical drugs or vaccines JL. McDermott Effects of adenoviral vector infection on human adrenocortical cells U. Matkovic Role of recombinant poxvirus vectors expressing HIV/SIVgenes in modulating Th1/Th2 cytokine profiles in human dendritic cells P. Beggio 6

7 Human placenta derived mesenchymal stem cells are fully permissive to human cytomegalovirus infection L. Luconi Expression of the HPV-16 L1 Antigen in Transplastomic Tobacco Plants P. Lenzi 12: LUNCH VIRAL ONCOGENESIS AND VACCINES Chairs: Franco Buonaguro (Napoli), Colomba Giorgi (Roma) KeyNote Lecture New strategies in Vaccine Development The HIV experience Hans Wolf (Regensburg Germany) Selected oral presentations Anti-cancer activity of plant-produded HPV16 E7 vaccine S. Massa Development of a vaccine against Toscana Virus G. Gori Savellini Development of avipoxvirus recombinant vaccines for the control of HPV16-induced cervical cancer E. Pozzi Impact of the seropositivity status on the pattern of DC activation by Virus-Like Particles L. Buonaguro Distribution of human papillomavirus type 16 variants in penile keratinizing squamous cell carcinoma ML. Tornesello Expression, processing and assembly of the HIV-1 Pr55gag protein in transgenic tobacco chloroplasts N. Scotti BREAK 7

8 HPV INFECTION AND PREVENTION Chairs: Franco Buonaguro (Napoli), Colomba Giorni (Roma) HPV and HPV-associated diseases worldwide burden Kate Soldan (London UK) Characterisation of the interaction of HPV E6 with the Discs Large Tumour Suppressor Lawrence Banks (Trieste) VLPs: past and future Reinhard Kirnbauer (Vienna Austria) Gardasil Franco Scaglione (Milano) Long-term impact of prophylactic ant-hpv 16/18 vaccine Davis Jenkins (Rixensart Belgium) New therapeutic strategies Aldo Venuti (Roma) Tavola rotonda Panel discussion: issues and guidelines (in italiano) Coordinator Sergio Pecorelli (Brescia) La vaccinazione in Italia Maria Grazia Pompa (Roma) Metodi diagnostici e screening nell'era del vaccino Francesca Carozzi (Firenze) Il punto di vista del virologo Giorgio Palù (Padova), Carlo Federico Perno (Roma), MariaLina Tornesello (Napoli) Il punto di vista del ginecologo Aldo Vecchione (Napoli) Come controllare l'efficacia del vaccino, chi lo farà e come lo farà Marta Ciofi degli Atti (Roma) 8

9 Tuesday, 26 June KeyNote Lecture Virological monitoring of environmental matrices and food: importance for the risk assessment Annalaura Carducci (Pisa) VIRUS HOST INTERACTIONS AND PATHOGENESIS Chairs: Mauro Pistello (Pisa), Luisa Rubino (Bari) Selected oral presentations Down-regulation of proteolytic complexes following Epstein Barr Virus activation in Burkitt s lymphoma cells G. Matusali Analysis of HCMV ppul44 homodimerization, intra-nuclear mobility and phosphorylation-regulated nuclear transport using live cells imaging G. Alvisi Human cytomegalovirus replicates in adrenocortical cells and stimulates steroid hormone production M. Trevisan HSV-1 induces dysregulation of monocyte anticandida functions C. Cermelli What s the clue in the progression of liver damage during HCV-infection? Analysis of the role of HCV NS5A and Core proteins M. Marcolongo Involvement of the E62EEE65 Nef Acidic Domain and TRAF Family Members on Signaling Events Induced by treatment of Monocytes/Macrophages with HIV-1 Nef G. Mangino HPV infection in HIV positive subjects: 10 years experience of follow-up F. Lillo 10: BREAK 11: Guidelines for human Cytomegalovirus infections Giuseppe Gerna (Pavia) 11:45-12:45 Poster Session 12: LUNCH 9

10 EMERGING AND ZOONOTIC VIRAL INFECTIONS Chairs: Canio Buonavoglia (Bari), Franco Ruggeri (Roma) KeyNote Lecture An emerging zoonosis: European bat lyssaviruses Franco Mutinelli (Legnaro PD) 14: Selected oral presentations Emerging respiratory infections in hematopoietic stem cell-transplanted children in Germany V. Falcone Characterization of the biological properties of human and rhesus VP8* proteins F. Cappuccini Detection of norovirus in a captive lion cub with haemorrhagic enteritis (Panthera leo) V. Martella Virologic Characterization of a Poxvirus Zoonosis in Northern Italy F. Carletti 15:30-17:00 SOCIETY MEETING 17:00 Best Poster Award and Travel Awards for Eurovirology 2007 Closing remarks 10

11 GB Rossi & Keynote Lectures 11

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13 New aspects of interferon systems, immune evasion and new potential applications Filippo Belardelli Department of Cell Biology and Neurosciences, Istituto Superiore di Sanità, Rome, Italy. Half a century has passed since the discovery of interferons (IFNs) as antiviral factors released by virus-infected cells. The first two decades of IFN research were characterized by major difficulties in convincing the scientific community about the specificity of the emerging biological effects. In the early 1980s, when recombinant IFNs became available, it was recognized that this family of cytokines, comprising the type I or viral IFNs (mainly α and β) and the type II or immune IFN (γ), could also exert multiple activities on cell growth, differentiation and immune response. Today, in revisiting 50 years of history of IFN research, the impact of early studies on the IFN system appears to be impressive. IFNs-α are the cytokines with a longest record of clinical use and are still used in patients with certain malignancies and viral diseases, while IFN-β is currently used in patients with multiple sclerosis. Notably, IFN research has been instrumental for understanding cytokine signal transduction pathways and important mechanisms regulating antiviral, antitumor and immune responses. Recently, multiple mechanisms by which viruses can undermine the potent IFN-mediated host defence system have been described. These mechanisms are important for evading the direct IFNmediated control on viral replication as well as for allowing evasion from an effective immune response, which is also regulated by the IFN system. Likewise, immune suppression, linked to impaired IFN production or defective cytokine response, has been observed in cancer patients. These findings are consistent with data in mouse models, demonstrating: i) the importance of the integrity of the IFN system in the host immune surveillance against tumors; ii) the role of endogenous IFN in chemotherapy-induced reversion of immune suppression in tumor-bearing animals. Notably, recent studies have underscored new effects of IFNs (especially type I IFN) on cells of the immune system (including T cells and dendritic cells), which are important in linking innate and adaptive immunity. Thus, in mouse models, type I IFN can act as a powerful vaccine adjuvant by acting on dendritic cells (DCs). Likewise, DCs rapidly generated from human monocytes after exposure to IFN-α (IFN-DCs) act as powerful cellular adjuvants for the generation of MHC class I restricted CD8+ T cell responses against viral (HIV and EBV) and tumor (melanoma) antigens and efficiently crossprime CD8+ T cells against exogenous antigens. On the whole, the preclinical data suggest that these IFN-DCs exhibit some unique characteristics particularly suitable for the generation of effective DC-based vaccines. New therapeutic strategies can now be based on a novel rationale for using IFN-α, including the direct in vivo use of these cytokines as vaccine adjuvants and their in vitro use to generate highly active patient s monocyte-derived DCs to be utilized in therapeutic vaccination protocols. Such immunotherapy strategies are particularly suitable in patients showing immune suppression features (induced either by virus infection of by tumor progression) which can be effectively restored by IFN-induced immune interventions targeted to DCs and by additional combination treatments. Moreover, a detailed knowledge on the IFN system in patients with cancer and HCV-infection would be instrumental in selecting categories of patients responding to IFN therapy. Planned clinical studies are expected to provide evidence on the importance of IFN-DC interactions in patients and on the perspectives of this novel use of IFN-α in immunotherapy protocols. 13

14 Antiviral strategies, mechanism and clinical efficacy Massimo Puoti Le terapie antivirali negli ultimi tempi hanno assunto una crescente importanza in Clinica in virtù del loro impatto sulla sopravvivenza di soggetti con malattie virali croniche legato all'immediato efetto sulle loro sequele. L'impato dei farmaci antiretrovirali sulla sopravvivenza e sulla qualità di vita dei soggetti con AIDS è ben noto, ma recentemente l'impiego delle terapie anti HBV ed anti HCV ha consentito di ridurre la mortalità per cirrosi ed epatocarcinoma e l'impiego delle terapie anti CMV quella per le infezioni da CMV in soggetti immunodepressi. NEW STRATEGIES IN VACCINE DEVELOPMENT: THE HIV EXPERIENCE Prof. Dr. Hans Wolf University of Regensburg, Institute for Medical Microbiology and Hygiene In response to the growing problems related with HIV-infections in poor economics, industrialized countries responded with strategies to intensify the development of HIV-vaccines. INCO-projects and the formation of sizeable research clusters in the EU allowed strategies starting from molecular epidemiology to evaluation of vaccine candidates in primates and humans. Together with the Chinese Center for Disease Control early at onset of the HIV-epidemic in China relevant HIV-strains could be selected (B-clade RL42 and C-clade 97CN54, 97CN54 is a B -C recombinant and the most prevalent strain in Western and North-Western provinces of China). Using sequence modified genes from 97CN54 a set of immunogens comprising gag pol nef and env was developed which was found to be highly immunogenic but inactive in their original enzymatic functions. In context of the EUROVAC-cluster and the INCO-programme different presentation systems based amongst others DNA-plasmids (COBRA) and vaccinia viruses (NYVAC, MVA, TienTan) have been developed into GMP-manufactured products. These have been evaluated in parallel to the human trials in Rhesus monkeys. The combination of DNA prime vaccinia boost gave strong immune response in almost all animals in multiple HIV reading frames and on the basis of IFNα, IL-2 and IL-4 Elispots. Parallel experiments using the SHIV 89.6p-model showed protection from disease and rapid clearance of viremia to set point in challenge experiments. Data from human trials with NYVAC-C alone looked encouraging with 45 % responders, DNA alone did, upon first preliminary evaluation, not induce significant immune response jugged by CTL with about 10 % of the vaccinees. A trial with the combination of DNA-C prime and NYVAC-C boost parallels data in primates and achieved rigorous (Elispots above 400 up to 1200) broad (multiple antigens and epitopes), rigorous (multiple cytokines) and long lasting immunogenicity on almost all vaccinees (above 90 %). Further trials are up-coming in Regensburg and other European sites and in Beijing/China. 14

15 Virological monitoring of environmental matrices and food: importance for the risk assessment Annalaura Carducci, Dip. Biologia dell'università di Pisa The virological risk coming from the exposure to environmental contaminated matrices have characteristics that impose its specific consideration: viruses cannot be efficacy represented by the classic microbial indicators (i.e. E. Coli, enterococci and coliphages) because they have a lower infectious dose, multiple ways of transmission, different ecology in comparison with bacteria and a higher genetic variability often associated with the possibility of recombination. The recent examples of SARS and H5N1 avian influenza as well as studies on HEV, rotavirus and calicivirus, showed the importance of animal viruses in human pathology and the possible role of foods and environment as vehicle between infected and susceptible organisms: although viruses cannot multiply outside living cells, they can persist in the environment for a very long time because of their high resistance to natural and artificial disinfection agents and the protective action of environmental factors. The role of viruses as environmental biological risk agents has been proven by epidemiological data for different matrices: potable and recreational waters, food, arosol and surfaces. Unfortunately, in most cases, the epidemiological evidence has not been confirmed by the virus detection in the suspected matrices owing to the difficulties in applying to the environment analytical techniques developped for clinical materials. In fact, environmental samples have a highly variable composition often interfeering with the detection procedures, viruses are extremely diluted and multiple contamination of human and animal origin are frequent. Nevertheless, progress in clinical virology, in particular biomolecular techniques, and the development of methods for sample concentration and purification, have greatly improved the efficiency of virological analysis of environmental matrices and increased the amount of data on dectected viruses in waters, foods, aerosol, surfaces, etc. Although the majority of methods are still produced in house and need standardization, the virological environmental and food monitoring is now a real possibility and it can be useful for the risk analysis (assessment, control and communication) in different ways: to identify the routes of virus spreading, to verify the efficacy of control measures, to assess the level of sanitation thus helping the operators education, to monitor the environmental pollution, to carry out epidemiological surveillance of viral circulation, allowing molecular epidemiology and phylogenetic studies. The virological monitoring for the routine risk assessment of food and enviroment needs, besides the standardization of techniques, the selection of significant viral species and types or representative indicators and the clarification of the real meaning of the nucleic acid detection in the environment. Furtherly, monitoring data should be related to the clinical and surveillance ones to obtain quantitative information in terms of a dose-response relationship and probability of adverse outcomes. In conclusion, to use the environmental and food virology in the risk assessment, sharing knowledge and data with clinical human and veterinarian virologists will be an essential evolution. An emerging zoonosis: European bat lyssaviruses Franco Mutinelli, NRL for Rabies, Istituto Zooprofilattico Sperimentale delle Venezie, Legnaro (PD) In Europe, two bat lyssaviruses referred to as European bat lyssaviruses (EBLVs) types 1 and 2 (genotypes 5 and 6 respectively) which are closely related to classical rabies virus are responsible for an emerging zoonosis. EBLVs are host restricted to bats, and have been known to infect not only their primary hosts but also in rare circumstances, induce spillover infections to terrestrial mammals including domestic livestock, wildlife and man. Although spillover infections have occurred, there has been no evidence that the virus adapted to a new host. Since 1977, five human deaths from EBLVs have been reported. None of them had a record of prophylactic rabies immunization. Only fragmentary data exist about the effectiveness of current vaccines in cross-protection against EBLVs. EBLV in bats cannot be eliminated using conventional strategies similar to the control programmes based on vaccine baits used for fox rabies in Europe. Due to the protected status of bats in Europe, our knowledge of EBLV prevalence and epidemiology is limited. It is possible that EBLV is under-reported and that the recorded cases of EBLV represent only a small proportion of the actual number of infected bats. Human exposure through biting incidents, especially unprovoked attacks, should be treated immediately with rabies post-exposure treatment and the bat, where possible, retained for laboratory analysis and species identification. Preventative measures include educating all bat handlers of the risks posed by rabies-infected animals and advising them to be immunized. 15

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17 General Virology and Viral Genetics 17

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19 SELECTED ORAL COMMUNICATIONS Increasing circulation and diversification of HIV-1 non-b subtypes in Italy over a 10-year observation period Razzolini F, Saladini F, Vicenti I, Marconi A, Balestrieri M, Romano L, Zazzi M Department of Molecular Biology, University of Siena, Italy HIV-1 isolates belonging to the M (major) group are classified into subtypes (A, B, C, D, F, G, H, J, K), sub-subtypes (A1, A2, A3, F1, F2) and more than 30 circulating recombinant forms (CRFs). Geographic distribution of the different clades is subject to evolution over time as a result of human migration and settlement as well as possible preferential routes of transmission. HIV-1 pol sequence databases maintained at genotypic antiretroviral resistance testing reference laboratories can provide useful insights into the temporal changes in the circulation of HIV-1 clades. We analyzed clade assignment data stored at the HIV Monitoring Service of the University of Siena, Italy over a 10-year period ( ). More than 8000 HIV-1 sequences obtained from a total of 3485 patients were available. The first sequence of each patient was considered and assigned to the year of sampling. Subtyping was carried out by phylogenetic analysis using the Kimura distance model and neighbor-joining method with 1000-replicate bootstrap analysis. The overall prevalence of non-b subtypes in the data set was 10.4% and increased significantly over time reaching 29.8% in 2006 (p < ). A wide variety of subtypes and CRFs was represented, with subtypes A1, C, G, F1 and CRFs 01_AE and 02_AG each accounting for >5% of the pooled non-b sequences. Interestingly, as many as 9.4% (n = 34) of the non-b viruses were unique recombinant forms (URFs), particularly involving recombination between B and F subtypes. Significant association between individual highly represented subtypes and patient demographics were detected (e. g. CRF02_AG and African origin, decreased male-to-female ratio and clades A1, C, e CRF02_AG). Sequencing of additional HIV-1 regions (gag, env and in some cases full-length genome) for select strains allowed to characterize several complex and novel recombinant forms. The diversity of the HIV-1 sequence pool has increased significantly in recent years in Italy and will likely contribute to further exploration of HIV-1 genetic space at the population level. Detection and quantification of noroviruses in environmental samples by newly designed nested PCR assays and Taqman Real-Time RT-PCR. Muscillo, M., Pourshaban M., Fontana S., Di Grazia A., Iaconelli M., and La Rosa, G. Noroviruses have received increased attention in recent years because their role as etiologic agents in acute gastroenteritis outbreaks is now clearly established. In the present study, environmental samples from sea water, estuarine water, and effluents of sewage treatment plants were analyzed in order to evaluate the role of environmental surface contamination as a possible vehicle for transmission of norovirus genogroups I (GI) and II (GII). We used a previously published RT-PCR assay and also one that was developed in this study. The two assays resulted in similar detection, however with the new assay longer amplicons were obtained that provided more accurate phylogenetic assessment of the norovirus strains detected. To complete qualitative PCR assays, we performed quantitative RT-PCR (RT-Q-PCR) assays to detect norovirus concentrations in environmental samples by broadly reactive one-step TaqMan reverse transcription (RT)-PCR assays. Transcribed norovirus RNA from GI and GII plasmids were used as calibration curves. Quantitative data on the concentrations of noroviruses present in recreational water are indispensable for assessment of the public health risks caused by norovirus infections. While river, estuarine, and seawater samples were scarcely contaminated (0.9% of samples), all sewage samples were positive for norovirus GII; in 60% of samples both genogroup I and II were detected. This is the first study in Italy reporting the presence of Noroviruses in environmental samples not associated to evident outbreaks and outline the possible role played by water as a vehicle for transmission. 19

20 UL products are required for the activation of HCMV fusion complex during endothelial cell infection PATRONE Marco, MILANESI Gabriele and GALLINA Andrea University of Milano School of Medicine, Department of Medicine Surgery and Dentistry, Milano, Italy UL locus products are determinants of HCMV tropism for a number of cell types including leukocytes, DC and endothelial cells (EC) other than fibroblasts. The encoded proteins have been detected in the virion, complexed to envelope glycoproteins gh-gl, which suggests that UL polypeptides play role in viral entry. We have directly addressed this issue by comparing the fusion capability of an EC-tropic isolate (VR1814) with that of fibroblast-adapted strains. Indeed, all non-ec-tropic strains failed to fuse. The role of UL proteins in fusion was further proven by the ability of a recombinant pul128 (rpul128) to specifically inhibit VR1814 fusion with, and infection of, EC. Virion fusion was executed at the plasma membrane, as only fusion-deficient viral particles were found to be endocytosed. What is more, in VR1814 infections of EC we have characterized a post-attachment conformational intermediate of gb, the appearance of which is followed by a transient gb-gh interaction, detected in proximity of fusion. Both the gb intermediate and the gb-gh complex were inhibited by rpul128, and were absent in infections with non-ec-tropic strains. Altogether these findings indicate that UL products are required for the initiation of a chain of events triggering HCMV multi-component fusion machinery. Prevalence and Persistence of human papillomavirus genotypes and their variants in Human Immunodeficiency Virus (HIV) positive women from south-italy Maria Lina Tornesello 1, Maria Luisa Duraturo 1, Matilde Sansone 2, Roberto Piccoli 2, Luigi Buonaguro 1, Franco Maria Buonaguro 1,*, 1 Ist. Naz. Tumori "Fond. G. Pascale", Cappella Cangiani, I-80131, Naples, Italy, 2 Gynecology Department, University of Medicine, Naples, Italy Human immunodeficiency virus (HIV) positive women have high rates of infection with a broad range of human papillomavirus genotypes (HPV) whose oncogenic risk is not well known. Furthermore revalence and persistence of mucosal HPV genotypes have been analysed in 112 HIV-positive and 115 HIV-negative women during a 3-year followup period. Compared with HIV-negative, HIV-positive women were at the higher risk of HPV infection with a prevalence rate of 39.3% at the first visit and 43.7% after at least three visits. Among the twenty different viral genotypes identified the high risk HPVs (16, 18, 31, 33, 35, 45, 52, 58, and 66) were detected in 33% of HIV-positive and in 13% of HIV negative women, with the HPV16 as the most represented (16.1% and 6.9%) in both groups. Persistent infections with less characterized HPVs (45, 53, 54, 55, 61, 62, 70, 72, 81 and 83) were frequently found in HIV-positive women (within a range of 0.9%-16.3%) and significantly associated with low and high grade intraepithelial lesions. Phylogenetic analysis of HPV16 isolates allowed the identification of HPV16 African 2 and African 1 variants differently distributed in HIV-positive and HIV-negative women, suggesting different sexual mixing behaviours. The association of uncommon genotypes and variants with cytologic abnormalities in HIV-positive women underscores the need for targeting a wide range of HPV types in the screening analysis and in the design of future vaccines. 20

21 Polyomavirus BK and anti-dsdna antibodies in renal transplant recipients 1 Cristina Costa, 1 Massimiliano Bergallo, 2 Giovanni Antonio Touscoz, 1 Francesca Sidoti, 1 Maria Elena Terlizzi, 1 Sara Astegiano, 1 Chiara Merlino, 3 Giuseppe P. Segoloni, 1 Rossana Cavallo 1 Dept. of Public Health and Microbiology, Virology Unit, University of Turin, Italy 2 Gastro-Hepatology Unity, Laboratory of Digestive and Hepatic Physiopathology, and 3 Dept. of Internal Medicine, Renal Transplant Unit, Molinette Hospital, Turin, Italy Background. Polyomavirus BK reactivation is common in renal transplant recipients and may cause nephropathy with significant graft dysfunction. The induction of anti-dsdna antibodies by BKV has been described in experimental animals and during primary infection, and a role in the pathogenesis of systemic lupus erythematosus has been suggested. Methods. In this study we evaluated the occurrence of anti-dsdna antibodies and non-organ-specific autoantibodies by indirect immunofluorescence in 90 renal transplant recipients and the association with BKV reactivation as determined by BKV-DNA positivity on urine and/or serum samples. Results. Forty-four of 90 patients experienced a BKV reactivation; 17/90 developed anti-dsdna antibodies. Considering the 44 patients with BK viruria and/or viremia, anti-dsdna antibodies were present in 15 patients vs 29 who did not developed anti-dsdna antibodies, while in BKV DNA-negative patients 2 were anti-dsdna-positive vs 44 anti-dsdna-negative (p < 0.001). Considering the 22 patients with BK viremia (with or without viruria), anti-dsdna antibodies were present in 9 patients vs 13 who did not developed anti-dsdna antibodies, while in BK viremia-negative patients 8 were anti-dsdna positive vs 60 anti-dsdna negative (p < 0.01). No significant difference in terms of clinical parameters of renal function was found between anti-dsdna positive and negative patients. Conclusions. There is a significant association between BKV and production of anti-dsdna antibodies in renal transplant patients, however there is no evidence that these virus-induced autoreactive responses are themselves pathogenic. Inconsistent responses of cytomegalovirus-specific T-cells to pp65 and IE-1 vs infected dendritic cells in organ transplant recipients Daniele Lilleri 1, Paola Zelini 1, Chiara Fornara 1, Giuditta Comolli 1,2, Giuseppe Gerna 1 1 Servizio di Virologia, 2 Laboratori Sperimentali di Ricerca-Area Biotecnologie, Fondazione IRCCS Policlinico San Matteo, Pavia, Italy CD4 + and CD8 + T-cells specific for human cytomegalovirus (HCMV) and two immunodominant HCMV antigens (pp65 and IE-1) were monitored in 20 solid organ transplant recipients undergoing primary (n=4) or reactivated (n=16) HCMV infection during the first year after transplantation by using as a stimulator either HCMV-infected autologous dendritic cells (DCs) or pp65- or IE-1 peptide mixtures. Turnaround times for test performance were 7 days for infected DCs and 24h for peptides. Using infected DCs, HCMV-specific T-cell restoration occurred in all patients for CD8 + and in 18/20 (90%) for CD4 + T-cell subpopulations, resulting in virus clearance from blood. Using peptide mixtures, T-cell responses were less frequently detected. In detail, 14 (70%) patients showed pp65-specific CD8 + T-cells and 10 (50%) patients IE-1-specific CD8 + T-cells, whereas pp65-specific CD4 + T-cells were detected in 14 (70%) patients, and IE-1- specific CD4 + T-cells in 3 (15%) patients only. Protection from HCMV infection was associated with the presence of a HCMV-specific T-cell response directed against multiple viral proteins, but not against pp65 or IE-1 only. In conclusion, the use of pp65 and IE-1 peptide mixtures for rapid monitoring of HCMV-specific T-cell responses in solid organ transplant recipients underestimates the actual level of protection against HCMV. 21

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