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6 International Geminivirus Symposium Poster Session Virus Family Poster Session Anelloviridae Epidemiology Virus Family: Anelloviridae Category: Epidemiology/Transmission Title: Genogroup distribution and viral load of Torquetenovirus in fecal samples Authors: C. A. PINHO-NASCIMENTO1, J. P. G. LEITE 2, C. NIEL1, L. DINIZ- MENDES1 Address: 1Laboratório de Virología Molecular; 2Laboratório de Virología Comparada e Ambiental, Instituto Oswaldo Cruz, Fiocruz, Avenida Brasil 4365, 21040-360 Rio de Janeiro, RJ, Brasil. E-mail:[email protected] Abstract: Torquetenovirus (TTV, floating genus Anellovirus) is a non enveloped DNA virus with circular, single-stranded genome with size of 3.8 kb. TTV is highly prevalent in populations from around the world. TTV isolates have been classified into five main phylogenetic groups (1-5) showing a large genetic distance between them. The presence of TTV has been detected in feces. However, whether all five TTV genogroups are excreted in feces and the frequency of these events are presently unknown. The presence of TTV DNA was assessed in feces from 135 Brazilian (0-90 years old) patients with gastroenteritis by using three PCR methods, including real-time PCR. One hundred and twenty one (91.1%) samples were positive with at least one method. Using a genogroup-specific assay, it was shown that all genogroups were present. Thirty-seven (27.4%), 27 (20.0%), 57 (42.2%), 29 (21.5%) and 33 (24.4%) fecal samples contained TTV isolates belonging to genogroups 1 to 5, respectively. Coinfections with two, three, four and five TTV genogroups were found in 23 (17.0%), 15 (11.1%), 7 (5.2%) and 7 (5.2%) fecal samples, respectively. Thus, 52 (38.5%) samples contained more than one TTV genogroup. Viral loads ranged from 2.6 to 6.5 log genome equivalents per gram of feces. However, only moderate variations of viral load were noted depending on genogroup and number of coinfecting TTV genogroups. These results are the first to show high prevalence and the diversity of TTV isolates in feces. th 6 International Geminivirus Symposium Poster Session Anelloviridae Replication Virus Family: Anelloviridae Category: Replication/Gene expression Title: Transcription of TTV-HD types during in vitro virus replication Authors: JIAN-WEI FEI, ROMANA KIMMEL AND ETHEL-MICHELE DE VILLIERS Address: Division for the Characterization of Tumorviruses, German Cancer Research Centre, Im Neuenheimer Feld 242, 69120 Heidelberg, Germany Abstract: Transcription of single TTV types has previously been reported. We replicated full-length genomes and propagated virus of 12 isolates of 7 TTV-HD types in vitro. Transcripts from all cultures were analyzed using two different approaches. Single- as well as double stranded cDNA served as template for 5’- and 3’-RACE–PCR. We report a large range of transcripts resulting from varying splicing events. Our data indicate possibilities of additional proteins being formed during the replication of TT viruses. These data will facilitate future attempts to associate TTV infection with the pathogenesis of human disease. th 6 International Geminivirus Symposium Poster Session Anelloviridae Replication Virus Family: Anelloviridae Category: Replication/Gene expression Title: Stimulation of TT replication by EBV in cells of lymphatic origin. Authors: SILVIA S. BORKOSKY, CORINNA WHITLEY, ANNETTE KOPP- SCHNEIDER, HARALD ZUR HAUSEN AND ETHEL-MICHELE DE VILLIERS Address: Division for the Characterization of Tumorviruses, German Cancer Research Center, Im Neuenheimer Feld 242, 69120 Heidelberg, Germany. Abstract: Viral infections have been implicated in the pathogenesis of multiple sclerosis. Epstein-Barr virus (EBV) has frequently been investigated as a possible candidate. Torque teno virus (TTV) has also been discussed in this context. Nevertheless, mechanistic aspects remain unresolved. We report viral replication of two TTV-HD isolates obtained from multiple sclerosis brain tissue in a series of EBV- positive and -negative hematopoietic cell lines. Our results demonstrate the replication of both transfected TTV genomes in all the evaluated cell lines. Quantitative amplification indicates an enhanced TTV replication in the EBV- positive cell lines in comparison to the EBV-negative Burkitt’s lymphoma cell line BJAB, suggesting a helper effect of EBV infections in the replication of TTV. The present study provides information on a possible interaction of EBV and TTV in the etiology and progression of multiple sclerosis. th 6 International Geminivirus Symposium Poster Session Anelloviridae Other Virus Family: Anelloviridae Category: Other Title: Assessment of human torque teno virus removal from two wastewater treatment plants Authors: L. DINIZ-MENDES1, C.A. PINHO-NASCIMENTO1, L.R. BERGSTEN- TORRALBA1, V.S. DE PAULA3, M.P. MIAGOSTOVICH2 AND C. NIEL1 Address: 1Laboratório de Virologia Molecular; 2Laboratório de Virologia Comparada e Ambiental; 3Laboratório de Desenvolvimento Tecnológico em Virologia, Instituto Oswaldo Cruz, FIOCRUZ – Avenida Brasil, 4365 – CEP21040-360 Rio de Janeiro, RJ, Brasil. E-mail: [email protected] Abstract: Sewage is one of the primary sources of human viruses in environment. It is thus important to investigate the effectiveness of viral removal in sewage plants. Human torque teno virus (TTV) has been suggested as a possible indicator of viral contamination in environment, due to its long-term excretion in faeces, high persistence and elevated concentrations in sewage and polluted aquatic environments. The ability of two sewage treatment plants (STP1 and STP2) to remove TTV from effluents was assessed by real time PCR, and compared to removal profile of other common viruses found in wastewaters, namely enteric adenovirus (AdV) and hepatitis A virus (HAV). In STP1, the presence of viruses was evaluated monthly, for one year, in raw and treated sewage. In STP2, samples from raw sewage, as well as after primary and secondary treatment, were assessed over 20 collections which were done twice a week. Viruses were concentrated 1000-fold from two liters of water by adsorption-elution to a negatively charged membrane. The loads of both TTV and AdV reached about 4.3 log copies per 100 mL of raw sewage in STP1, and the treatment used in this plant removed approximately 68% of the input amount of these viruses. Furthermore, 4.6 log copies of TTV and AdV, and 2.7 log copies of HAV were detected per 100 mL of raw sewage in STP2. This plant was able to eliminate about 98% of the input amount of both TTV and AdV, and 82% of HAV content. STP1 effluents contained about 3.7 log copies of TTV and AdV. Viral amounts in STP2 effluents were 2.6 log copies of both TTV and AdV and 2 log copies of HAV per 100 mL. We conclude that, despite an appreciable reduction of viral loads, notable amounts were still discharged in the environment after treatment. These results reinforce the necessity of surveillance for viruses in sewage and the establishment of a viral indicator for water quality. th 6 International Geminivirus Symposium Poster Session Circoviridae G enetic Diversity/Evolution Virus Family: Circoviridae Category: Genetic Diversity/Evolution Title: Origin and evolution of porcine circovirus type 2 in Cuba Authors: Lester Josué Pérez, Heidy Díaz de Arce, Martí Cortey, Patricia Domínguez, Maria I. Percedo, Carmen Laura Perera, Joan Tarradas, Maria T. Frías, Joaquim Segalés, LLilianne Ganges, José Ignacio Núñez. Address: Centro Nacional de Sanidad Agropecuria (CENSA), La Habana, Cuba Abstract: Porcine circovirus type 2 (PCV2) is the essential etiological infectious agent of postweaning multisystemic wasting syndrome (PMWS), which is considered one of the most economically important swine diseases worldwide. In this study, a comparison between methodologies based on classical phylogenetic trees and networks to infer the origin of PCV2 in Cuba was performed. In addition, the mechanisms supporting the genetic variability of Cuban PCV2 populations were investigated. A retrospective study, using pig sera collected in Cuba from 1993 to 2004, to evaluate the presence of PCV2 genome and PCV2-specific antibodies was also conducted. A total of 24 complete Cuban PCV2 sequences collected between 2005 and 2009 from different regions of the country were analyzed. Three classical methods of phylogenetic analysis, namely Neighbour- Joining, Maximum Parsimony and Bayesian Inference, as well as haplotype network construction, were used. Whereas the classical phylogenetic trees suggested different origins for the Cuban PCV2 strains, the haplotype network revealed a direct connection between all the Cuban sequences in agreement with the obtained epidemiological and experimental data. Moreover, the importation of pigs carried out in 2005 from the Quebec-Ontario region, Canada, seems to be the most likely origin of PCV2 in Cuba. Likewise, the genetic variability of Cuban PCV2 sequences was supported by geographic segregation and positive selection pressure with estimated rates of nucleotide substitution on the order of 3.12 × 10−3 and 6.57 × 10−3 substitutions/site/year, which are closer to those reported for RNA viruses. th 6 International Geminivirus Symposium Poster Session Circoviridae G enetic Diversity/Evolution Virus Family: Circoviridae Category: Genetic Diversity/Evolution Title: Discovery of a novel single-stranded DNA virus in dragonflies Authors: K. Rosario1, M. Marinov2, D. Stainton2, S. Kraberger2, D. Martin3, M. Breitbart1, and A. Varsani2 Address: 1College of Marine Science, University of South Florida, St. Petersburg, FL, USA 2School of Biological Sciences, University
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