From bat coronaviruses to SARS and MERS

Institute of Biochemistry, University of Lübeck, Germany

In recent years, it has been shown that bats constitute a rich reservoir of RNA viruses. Occasionally, these viruses are transmitted into the human population by zoonosis, often via an intermediate species. Thus, the outbreak of Severe Acute Respiratory Syndrome (SARS) in 2002/2003 was caused by a coronavirus (SARS-CoV) that probably crossed species barriers from bats to the civet cat and other animals treated at markets in Southern China, and from there to humans. A very recent event is the emergence of a new human coronavirus in countries on and surrounding the Arab peninsula. Designated Middle-East Respiratory Syndrome coronavirus (MERS-CoV), this virus appears to have a high case-fatality ratio (57 documented cases, 27 deaths at the time of writing this abstract), although at the moment, we probably only see the "tip of the iceberg. Similar to SARS, MERS is characterized by severe atypical pneumonia, but in addition, renal failure is observed in most cases.

Through functional and structural characterization of components of the huge replicase/transcriptase complex of the coronaviruses, we try to understand the evolution of these viruses. How do they adapt to the new host after entering the human population? We also examine interactions between viral proteins and host proteins, with a focus on viral components that seem to influence protein synthesis in the host cell. The structural findings are also used to design antiviral compounds. In preparation against future zoonotic transmissions, we develop and synthesize antivirals on the basis of our structures of bat-CoV proteins; these compounds can immediately enter preclinical development should another epidemic caused by a novel human coronavirus strike. Using this approach, we already had an antiviral in our hands when MERS-CoV was first described in September, 2012. 

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Genetic diversity and dilution effects during virus emergence from pristine to modified landscapes

Institute of Virology, University of Bonn Medical Centre, Germany

Tropical rainforests show the highest level of terrestrial biodiversity and may be an important contributor to microbial diversity. Exploitation of these ecosystems may foster the emergence of novel pathogens. At present, however, we lack both an understanding of pathogen prevalence in remote rainforest areas and reliable detection systems for novel pathogens. Monitoring of pathogen prevalence and circulation at the interface of pristine rainforests and disturbed landscapes is crucial for emerging disease surveillance and forecasting.

We investigated the variation in mosquito distribution and mosquito-associated viruses along anthropogenic disturbance gradients in Africa and the Neotropics. Mosquito species composition and diversity greatly differed between natural and modified habitat types. Investigating concomitant viral infections revealed an extremely high diversity of numerous previously unknown RNA viruses belonging to the families Bunyaviridae, Flaviviridae, Reoviridae, and Rhabdoviridae, as well as to the discovery of the first insect-associated nidovirus (Cavally virus, CAVV) that is likely to represent a novel family within the order Nidovirales. General virus abundance and diversity was examined along the gradient, yielding a decrease in diversity and an increase in prevalence, from natural to modified habitat types. Phylogenetic analyses indicated ancestral relationships of the newly discovered viruses to established virus groups suggesting that tropical ecosystems may contain a larger spectrum of viruses than currently known from epidemic isolates. Knowledge on the biological mechanisms behind ecosystem modification and arbovirus emergence could provide innovative approaches for epidemic risk assessment and intervention strategies. 

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Prof. Ian Goodfellow will provide you with up-to-date information about the work of his Calicivirus Research Group. (web page)

Division of Virology, Department of Pathology, University of Cambridge, UK

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From virus entry to replication: A hitchhiker guide to the cell

Laboratory of Viral Infection and Innate Immune Sensing Dynamics, Heidelberg University, Germany

Viruses are intracellular parasites. They must enter the cells to deliver their genetic information to replication-competent compartments within the host cell. The first barrier that a virus must face to establish an infection in the host is breaching the cellular membrane. Viruses have developed efficient ways to hijack existing cellular processes to breach this barrier. They either directly penetrate the plasma membrane or hijack endocytic pathways to gain access to the host cells. For viruses that enter cells by endocytosis, after uptake, they traffic to the endosomal pathway. In the endosomal compartments, viruses will encounter acidic environments and proteases that are often required for the viruses to undergo fusion or membrane penetration, which will allow either the release of the viral particles or of the viral genome into the cytoplasm so that infection/replication can occur.

In this lecture, the students will have an overview of the various strategies used by viruses to enter and to replicate in the host. By using examples from non-enveloped and enveloped viruses, the participants will have an introduction to the methodologies used in laboratories to study virus entry and replication. This seminar will mainly focus on imaging methods (live-cell confocal microscopy and super-resolution microscopy) that allow researchers to visualize and track individual virus particles as they infect the host. Finally, we will examine the advantages and disadvantages of these methods and discuss how molecular mechanisms could be concluded from these microscopy approaches.

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Prof. Stephan Urban will provide you with up-to-date information about the work of his HBV Research Group. (web page)

University Hospital Heidelberg, Germany

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A new concept of influenza treatment by a combination of antiviral drugs and inhibitors targeting hemagglutinin activating host proteases

Institute for Molecular Virology, Philipps-University of Marburg, Germany

Human and animal influenza virus infections are caused by numerous genetically variable influenza virus strains with an outcome of severe illnesses, high burden on the healthcare system, and serious economic losses. A development of a more efficacious anti-influenza therapy on the basis of existed drugs and newly created agents is considered to be needed, since the limitations and usefulness of vaccination against influenza and the emergence of drug-resistant influenza viruses. Host proteases activating viral hemagglutinin spikes were discovered in recent years. Moreover, highly efficient protease inhibitors were designed and chemically synthesized by my colleague Prof. Dr. Torsten Steinmetzer, Pharmaceutical Chemistry, University of Marburg. Now we examined influenza virus propagation in presence of these inhibitors, alone in double-combination with neuraminidase inhibitors or in triple-combination with virus replication inhibitors. Combinatorial treatment of virus infected cells showed a more significant inhibitory effect compared with that treated with a single drug. Such a combination therapy may reduce the emergence of resistant influenza viruses. Our data of combined therapies will be presented, proposing a viable strategy for treatment of humans afflicted with seasonal influenza or with a highly pathogenic avian influenza virus infection.

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Design of novel antiviral drugs on the basis of cellular targets

Institute for Clinical and Molecular Virology, University of Erlangen-Nuremberg, Germany

Infections with human cytomegalovirus (HCMV) represent a serious, sometimes life-threatening danger for immunosuppressed individuals, including transplant recipients and patients under antitumoral chemotherapy. Currently approved antiviral drugs have the disadvantage of inducing side effects and drug-resistant virus mutants. These limitations create an increasing need for new antiviral drugs, particularly for drugs that exhibit low levels of toxicity and activity against HCMV variants that are resistant to conventional drugs. Viral replication is a complex process regulated on a network of interacting viral and cellular proteins that are expressed during infection. Here, we describe innovative anticytomegaloviral approaches based on novel, potent and selective small-molecule inhibitors targeting cellular host factors crucial for the efficiency of virus replication. These antiviral principles are based on the targets dihydroorotate dehydrogenase (DHODH, an enzyme that mediates de novo biosynthesis of pyrimidine ribonucleotides) or cyclin-dependent kinase 7 (CDK7, a CDK-activating kinase and a co-activator of RNA polymerase II). Our findings demonstrate that both targets play important roles during viral gene expression and replication and may be efficiently exploited for novel antiviral strategies. Thus, we conclude that DHODH- and CDK7-inhibitors represent highly interesting candidates for the development of novel antiviral drugs.

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Parvovirus infections: prospects for cancer treatment

German Cancer Research Center, Heidelberg, Germany

Rodent parvoviruses (PV) are recognized for their intrinsic oncotropism and oncolytic activity. These features contribute to the natural capacity of PV for tumor suppression, for which human cancer cells can be targets in animal models. Although PV uptake occurs in most host cells, some of the subsequent steps leading to expression and amplification of the viral genome and production of progeny particles are upregulated in malignantly transformed cells. By usurping cellular processes such as DNA replication, DNA damage response, and gene expression, and/or by interfering with cellular signaling cascades involved in cytoskeleton dynamics and cell integrity, PVs can induce cytostasis and cytotoxicity. Furthermore, there is growing evidence that parvoviral oncosuppression involves an immune component. Besides exerting direct oncolytic effects, PV can indeed serve as adjuvants to hand further tumor destruction to the immune system in animal models. Current and planned parvovirotherapy clinical trials should indicate whether PV oncolysis can be translated into long-term protection by the immune system with improved tumor destruction and patient survival.

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Towards an HIV Cure by Provirus Excision

Heinrich Pette Institute – Leibniz Institute for Experimental Virology,  Hamburg, Germany

HIV-1 integrates into the host chromosome and persists as a provirus flanked by long terminal repeats (LTR). To date, treatment regimens primarily target the virus enzymes or virus entry, but not the integrated provirus. Therefore, current antiviral therapy (HAART) requires lifelong treatment which, unfortunately, may be accompanied by the occurrence of substantial drug-related toxicities and/or the development of drug-resistant viruses. Moreover, HAART cannot cure HIV infection.

Previously, we engineered a LTR-specific recombinase (Tre-recombinase) that effectively excises integrated HIV-1 proviral DNA from infected human cell cultures, suggesting that customized enzymes might someday help to eradicate HIV-1 from the body. Here we analyzed potential cytopathic effects in Tre-expressing cells and demonstrate pronounced antiviral Tre-activity in HIV-1 infected Rag2-/-γc-/- mice, which were either engrafted with Tre-transduced human CD4+ T cells or with Tretransduced human CD34+ hematopoietic stem cells (HSC). The presented data suggest that Tre-recombinase may be a valuable component of future antiretroviral therapies of the post HAART era that aim at virus eradication.

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