The long-term goal of this workgroup is to limit the clinical, epidemiological and economical impact of virus infections by vaccination, treatment with antiviral drugs or with biological response modifiers. The research focuses on acute respiratory virus infections such as influenza viruses, human metapneumovirus (hMPV), respiratory syncytial virus (RSV), measles virus (MV), coronaviruses (CoV), as well as on chronic virus infections such as hepatitis B and C viruses and herpesviruses like herpes simplex virus (HSV), varicella zoster virus (VZV), and simian varicella virus (SVV). Knowledge of the (immuno-) pathogenesis of these virus infections, both in humans and in animals, is the fundamental basis of the work. A special element of this research is to apply  the expertise and techniques of several disciplines, including virology, immunology, pathology and genomics in a highly integrated fashion.

Studies on the natural and vaccine-induced immune response to these viruses are performed to improve existing and develop novel vaccines and other intervention strategies. Depending on the viral pathogen and what type of cellular and/or humoral immunity is most desired, subunit vaccines, whole inactivated vaccines, live-attenuated vaccines and vectored vaccines are studied. Vaccine candidates are developed using both classical and state-of-the-art techniques in biochemistry and molecular biology. Humoral and cellular immunological parameters are monitored, with special emphasis on correlates of protection and vaccine-induced immune-pathogenesis for selected viral pathogens. In addition the efficacy of novel and existing antiviral treatments and the emergence of drug-resistant virus variants are monitored in order to improve antiviral treatment regimens. Our high quality molecular diagnostics unit forms an important bridge between clinic and research. The efficacy of antiviral drugs to combat AIDS (HIV-1 and HIV-2), influenza, hepatitis (hepatitis B and C viruses) and herpes (HSV and VZV) is measured in relation to drug-resistance, host and epidemiological parameters.

An important element of the research of this workgroup is also related to studies of wild animal reservoirs of emerging viral infections: both virus discovery and studies on the epidemiology and pathology of potential emerging pathogens in wild animals are persued. The discovery of novel viruses associated with human diseases is included in this workgroup since the etiology of many human and animal diseases is still unknown; the early identification of novel viral pathogens will allow the timely development of intervention strategies using surveillance tools, vaccines, antiviral compounds and other medicines to limit their impact. It is well appreciated that interspecies transmission of pathogens may result in the emergence of new infectious diseases in humans and animals. By comparing and contrasting emerging pathogens with those of closely related pathogens in more or less related host species, the various host genomic pathways are delineated thus determining the outcome of zoonotic transmission and adaptation to the newly invaded species. It is anticipated that the outcome of this research will improve our preparedness to combat newly emerging viruses.

2.1      Influenza viruses

(Workgroup leaders: Dr. R.A.M. Fouchier; Dr. G.F. Rimmelzwaan)

• Identified of a novel influenza A virus subtype; H16.
• Identification of H7N7 viruses in poultry and humans.
• Developed candidate vaccines against H7N7 virus and preclinical evaluation.
• Identified determinants of pathogenicity of H7N7 influenza virus.
• Developed adjuvanted and (MVA-)vectored candidate vaccines against H5N1 virus and preclinical evaluation.
• Performed preclinical and clinical evaluation of vaccine candidates against H1N1v virus
• Improved reverse genetics system for influenza A virus.
• Demonstrated that influenza A virus genome packaging is a specific process.
• Identified felids and other carnivores as importants hosts for H5N1 virus.
• Developed antigenic cartography methods and used these to describe the evolutionary history of influenza A (H3N2) virus
• Provided extensive new insights on the ecology of avian influenza viruses in wild birds.
• Demonstrated that H5N1 virus predominantly targets the lower respiratory tract of humans, potentially explaining poor transmission.
• Demonstrated that H5N1 entered the African continent through several re-introductions.
• First to design animal models for new H1N1 virus, and test pathogenicity and transmission.
• Demonstrated that infection-induced heterosubtypic immunity, which correlates with cross-reactive cytotoxic T lymphocyte CTL against conserved proteins, is robust and can protect against severe disease and mortality in a mouse model.
• Demonstated that vaccination against seasonal influenza, may prevent the induction of heterosubtypic immunity against a lethal challenge with an H5N1 influenza virus
• Demonstrated that different DC  have different functions during influenza virus infection like trafficking of viral antigen, antigen presentation and initiation of T cell responses as well as formation and maintenance of iBALT structure in the lungs.
• Developed novel methods for the detection of human antibodies against influenza viruses based on GFP expressing influenza viruses.

2.2      Human metapneumovirus

(Workgroup leaders: Dr. R.A.M. Fouchier; Dr. R.L. de Swart)

• Set up animal model systems for studies on pathogenesis and vaccination.
• Set up reverse genetics systems.
• Designed diagnostic tests.
• Described antigenic and genetic variation of strains circulating around the globe.
• Described prevalence and clinical symptoms in several cohort studies.
• Designed vaccine candidates (live-attenuated HMPV, vectored live-attenuated PIV, F subunit) and performed pre-clinical studies in suitable animal models.
• Generated monoclonal antibodies for potential therapeutic use and provided proof-of-principle for their use.
•  Provided information on the evolutionary history and dynamics of human and avian metapneumoviruses.
• Identified F protein as the main determinant of metapneumovirus host range.
• Demonstrated that HMPV membrane fusion is pH-independent.
• Demonstrated that HMPV can escape from host innate immune responses.
• Demonstrated hypersensitivity to hMPV infection induced by alum-adjuvanted formalin-inactivated hMPV preparation in animal models (similar to immunopathological responses previously observed in infants and animal models with formalin-inactivated RSV and measles vaccines).
• Demonstrated safe and effective immune responses induced by candidate subunit or live-attenuated hMPV vaccines       

2.3      Respiratory syncytial virus and measles virus

(Workgroup leader: Dr. R.L. de Swart)

• Generated animal model to study immuno-pathogenesis induced by RSV vaccines.
• Provided insights in the immunopathogensis associated with formaline inactivated vaccine.
• Evaluation of the safety of BBG2Na, a recombinant RSV subunit vaccine.
• Evaluation of safety and immunogenicity of a recombinant MVA vaccine.
• Evaluation of TH-1/TH-2 responses in relation to disease severity.
• Evaluation of RSV subgroups/genetic lineages in relation to disease severity.
• Characterisation of CTL responses upon RSV infection.
• Development of Genomics technology to study respiratory virus infections.
• Evaluation of the efficacy and safety of new generation RSV vaccines in an animal model for vaccine-mediated enhancement of disease.
• Characterization of RSV-specific immune responses in atopic and non-atopic infants
• Identification of RSV-specific CD4⁺ and CD8⁺ T cell epitopes
• Development of genomics technology to study respiratory virus infections in vitro and in animal models.
• Development of model systems for studying the interaction between RSV and bacterial respiratory pathogens.
• Generated an animal model to test measles vaccines.
• Evaluated MVA vectors to be used as measles vaccine.
• Evaluated the use of ISCOMs as measles vaccine.
• Evaluated pulmonary and neurological safety of aerosol-administration of live-attenuated vaccine.
• Evaluated diagnostic and molecular epidemiologic techniques for measles virus in Sudan.
• Genotyping of measles viruses from Sudan.
• Comprehensive analysis of antibody responses to wild-type MV infections (Sudan).
• Studies on imported cases of measles in The Netherlands.
• Analysis of the contribution of protein-specific antibody responses to virus neutralization.
• Studies on imported cases of measles in The Netherlands.
• Macaque model for measles established using recombinant EGFP-expressing virus
• Demonstrated CD150-expressing lymphocytes and dendritic cells as the main target cells for MV infection in vivo.

2.4      Hepatitis- and coronaviruses

(Workgroup leader: Dr. B.L. Haagmans)    

• Identified a naturally occurring recombinant HCV
• Characterised HCV deletion mutants circulating in chronically infected patients
• Provided insight in the host factors associated with response to IFN therapy in HCV patients.
• Evaluated the pharmacokinetics of pegylated interferon-alpha in HCV patients.
• Evaluated the induction of interferon-alpha neutralising antibodies during antiviral treatment of HCV patients
• Evaluated the immunogenicity of a candidate HCV vaccine
• Set up animal models to study SARS-CoV infections.
• Evaluated the effect of interferon-alpha on SARS-CoV infection in macaques
• Evaluated immunogenicity and protective afficay of SARS-CoV vaccines.
• Evaluated humanised SARS-CoV monoclonal antibodies in ferrets
• Provided insight in the host genomic response against SARS-CoV in macaques.

2.5      Herpes viruses

(Workgroup leader: Dr. G.M.G.M. Verjans)

• Insight in the dynamics of viral populations in bone marrow transplant recipients, resulting in new intervention strategies.
• Demonstrated and detailed characterization of the systemic/intra-ocular HSV-specific B and T cell responses in patients with HSV-induced stromal keratitis and uveitis.
• Developed and applieda PCR-based assay to genotype HSV-1 isolates revealing risk factors for the development of HSV-induced keratitis and uveitis.
• Identified the putative role of corneal resident cells in the immunopathogenesis of HSV-induced stromal keratitis.
• Demonstrated and characterized the systemic/intra-ocular VZV-specific B and T cell responses in patients with VZV-induced uveitis.
• Demonstrated two variants of PHV-2 with different tropism.
• Demonstrated oncogenic properties of PHV-2.
• Ocular resident cells are involved in the pathogenesis of ocular HSV-1 and VZV infections: cytokine/chemokine secretion and local antigen presenting cell (APC) function.
• Presence of acyclovir resistant HSV-1 in corneal swaps determines the clinical outcome of HSV keratitis.
• HSV-1 DNA load in transplanted corneas determines corneal graft survival.
• Local virus-specific T-cell responses control HSV-1, but not VZV latency in human trigeminal ganglia.
• Neuron-interacting glial cells are local APC involved in controlling both HSV-1 and VZV latency.
• Experimental animal models developed to study the pathogenesis of HSV keratitis (mouse) and varicella (monkey).

2.6      Comparative pathology and wildlife disease

(Workgroup leader: Prof. Dr. T. Kuiken)

• Established comparative pathology setting at Erasmus MC by bringing expertise of human and veterinary pathology together.
• Established and coordinated wildlife disease surveillance, both at the national level (DWHC: www.DWHC.nl) and at the international level (EWDA).
• Established wild duck species as potential long-distance carriers of HPAIV H5N1
• Identified differences in pattern of attachment between human and avian influenza viruses to human lower respiratory tract tissues as important factor in determining primary disease and transmission potential
• Defined host species barriers to cross-species transmission of influenza virus and other emerging pathogens
• Identified new routes of entry and spread of HPAIV H5N1 in a mammalian model
• Established ferret and cat models for studying the pathogenesis of SARS in humans

For the respiratory virus infections, we will continue to perform surveillance studies in humans and animals in order to study genetic and antigenic differences between virus isolates, which is important for vaccine development. This work will be performed within our tasks as WHO National Influenza Center, partially also funded through the NIAID/NIH Center of Excellence in Influenza Research and Surveillance program “CRIP”, EU FP7 program Newflubird and collaborations with industry. We will continue to develop novel vaccines and test vaccines developed by others using our available animal model systems. A selection of these vaccines will be tested in clinical trials within the next 1 to 5 years, which activities are at least in part sponsored by pharmaceutical industries. Fundamental research on virus properties is essential for our understanding of viral pathogenesis, virus evolution, the immune responses required to control virus infections, identification of the viral targets most suitable for vaccine development and our understanding of immunopathogenesis associated with vaccination. Such projects are currently mostly funded by fundamental research foundations (NWO. VENI & VICI, NWO-Nivarec, Human Frontiers of Science Programme, NIAID-NIH Centers of Excellence in Influenza Research and Surveillance and EU FP7 projects). The department is integrally involved in studies on the new pandemic H1N1 virus, both in fundamental research and in applied programs to limit the impact of the pandemic. The department also remains heavily involved in similar work in the ongoing H5N1 outbreaks.

Future research will also include further analysis of the effect of vaccination on the induction of heterosubtypic immunity, in mice, ferrets and humans and novel influenza vaccine candidates will be tested in relevant animal models and the correlates of protection against infection will be assessed

for vaccine development and our understanding of immune-pathogenesis associated with vaccination. Pathogenesis studies increasingly profit from the availability of recombinant viruses expressing fluorescent proteins, enabling sensitive detection of virus infection in cell cultures, ex vivo tissue cultures or in vivo animal models. Such projects are currently mostly funded by fundamental research foundations (K.N.A.W., N.W.O., E.U.) and new projects have recently been approved (eg. EU, NIH, MRC, ZonMw) or are under evaluation.

A large genomics project (VIRGO, funded by NWO) has recently been renewed. This project will include studies on immunopathogenesis related to RSV and measles virus vaccines, pathogenic influenza virus infections, and (enhanced) disease models for RSV, measles, influenza and human metapneumovirus. In addition another major project on the development of new generations of intervention strategies for acute and chronic virus infections (“FES-Drugs”) is currently in the final stages of evalution and will probably be granted within the next month. The total subsidy of these two projects, both coordinated by A.Osterhaus will amount to approximately € 14 million.

For chronic virus infections, the current line of work on characterisation of emerging (drug-resistant) viruses will be continued, funded in part by the E.U. and pharmaceutical companies. This work is done in collaboration with clinical departments of Erasmus MC and elsewhere, and relies heavily on the molecular diagnostic unit of the department of Virology. Data gathered within this working group will be used to suggest improvements in current antiviral therapy regimens and to modify existing mathematical models. Virus-host interactions that are critical in relation to disease progression and therapy-efficacy will be studied, again sponsored by the E.U. and pharmaceutical companies. The role of CTL in controling disease, and the correlation of cellular immunity with therapy efficacies will be further evaluated. For the herpes viruses, combinatory studies (funded in part by NIH) on clinical specimens of both patients and experimentally infected animals (mice and monkeys) will be continued to unravel the virus-host interactions involved in the control of viral latency in sensory ganglia and those that initiate&perpetuate the pathology of ocular and cerebral herpesvirus infections.

Virus discovery

The department of virology continues a very successful virus discovery program that in the past 5 years has attracted significant funding from industry and the EU (most recently EU-FP7 program Emperie, coordinated by A. Osterhaus). The basis for this project was the successful identification and combat of SARS by the participants of this new program. Over the past 5 years, this long term successful research line of  the workgroup was extended by publishing several papers related to newly identified viruses including the SARS-coronavirus, human coronavirus NL63, budgerigar reovirus, and the latest influenza A virus subtype H16. Currently some new human respiratory and enteric viruses are being characterized.

Genomics

Microarray assisted mRNA profiling (affymetrix) has been implemented in various research lines to support the detailed characterization of the host response to virus infection, vaccination and/or treatment with biological response modifiers. The application of these tools provides high resolution “snapshots” not only supporting fundamental research on e.g. gene interacting networks controlling the host response to virus infection but also clinical research on diagnostic and prognostic biomarkers for specific viral infections and related disease manifestations. A standardized data storage and analysis platform has been developed to support optimal translation of genomics data into biological knowledge.

The new projects (VIRGO & FES-Drugs) will focus on the development of novel strategies for diagnosis, prognosis, and the development of novel generations of vaccines, antivirals and biological response modifiers.

Antiviral research

The workgroup, that has recently been extended with Dr. C.Boucher, an expert in antiviral HIV research, now has acquired extensive expertise in the evolution of viruses under antiviral pressure. This research line focuses on the effects of resistance development on the target enzyme functionality and viral replicative capacity as well an on the pathogenicity and transmissibility of the mutant viruses. It will now be expanded to other viruses including influenza and hepatitis B and C viruses. The current facilities and knowledge of the Department of Virology will provide the ideal setting to conduct the clinical, virological, molecular and the in vitro and in vivo studies needed for this research line.

4.1     Influenza viruses

1. Avian influenza A (H7N7) virus associated with human conjunctivitis and a fatal case of acute respiratory distress syndrome. R.A.M. Fouchier, P.M. Schneeberger, F.W. Rozendaal, J.M. Broekman, S.A.G. Kemink, V. Munster, T. Kuiken, G.F. Rimmelzwaan, M.Schutten, G.J.J. van Doornum, G. Koch, A. Bosman, M.Koopmans and A.D.M.E. Osterhaus. (2004).P.N.A.S. 101:1356-61.
2. Transmission of H7N7 avian influenza viruses to humans during a large epizootic in commercial poultry farms in The Netherlands. M. Koopmans, B. Wilbrink, M. Conyn, G. Natrop, H. van der Nat, H. Vennema, A. Meijer, M. du Ry van Beest Holle, J. van Steenbergen, R.A.M. Fouchier, A.D.M.E. Osterhaus and A. Bosman. (2004) Lancet, 363: 587-93.
3. Mapping the antigenic and genetic evolution of influenza virus. D.J. Smith, A.S. Lapedes, J.C. de Jong, T.M. Bestebroer, G.F. Rimmelzwaan, A.D.M.E. Osterhaus and R.A.M. Fouchier. (2004) Science 305:371-376.
4. Avian H5N1 influenza in cats. T. Kuiken, G.F. Rimmelzwaan, D. van Riel, G. van Amerongen, M. Baars, R.A.M. Fouchier, A.D.M.E. Osterhaus. (2004) Science. 306:241.
5. Characterization of a novel influenza A virus hemagglutinin subtype (H16) obtained from black-headed gulls.. R.A.M. Fouchier, V. Munster, A. Wallensten, T.M. Bestebroer, S. Herfst, D. Smith, G.F. Rimmelzwaan, B. Olsen and A.D.M.E. Osterhaus. (2005) J Virol. 79:2814-22.
6. Influenza A virus (H5N1) infection in domestic cats causes systemic disease with possible novel routes of virus spread within and between hosts. Rimmelzwaan GF, van Riel D, van Amerongen G, Baars M, Fouchier R and Osterhaus A, Kuiken T. (2006) Am. J. Pathol. 168:176-183.
7. H5N1 Virus Attachment to Lower Respiratory Tract. D. van Riel, V.J. Munster, E. de Wit, G.F. Rimmelzwaan, R.A.M. Fouchier, A.D.M.E. Osterhaus, and T. Kuiken. (2006) Science. 312:399.
8. Global patterns of influenza A virus in wild birds. B. Olsen, V.J. Munster, A. Wallensten, J. Waldenström, A.D.M.E. Osterhaus, R.A.M. Fouchier. (2006) Science, 312:384-8.
9. Feline friend or potential foe? T.Kuiken, R.A.M. Fouchier, G.F. Rimmelzwaan, A.D.M.E. Osterhaus, P. Roeder. (2006) Nature 440:741-2.
10. Multiple introductions of H5N1 in Nigeria. M.F. Ducatez, C.M. Ollinger, A.A. Owoade, S. De Landtsheer, W. Ammerlaan, H.G.M. Niesters, A.D.M.E. Osterhaus, R.A.M. Fouchier and C.P. Muller. (2006) Nature 442:37.
11. Spatial, temporal and species variation in prevalence of influenza A virus in wild migratory birds. V.J. Munster, C. Baas, P. Lexmond, J. Waldenstrom, A. Wallensten, T. Fransson, G.F. Rimmelzwaan, W.E.P. Beyer, M. Schutten, B. Olsen, A.D.M.E. Osterhaus and R.A.M. Fouchier. (2007) Plos pathogens 3:e61.
12. Recombinant MVA vaccines protect mice against infection with highly pathogenic H5N1 influenza viruses. Kreijtz JHCM, Süzer Y, van Amerongen G, de Mutsert G, Wood J, Kuiken T, Fouchier RAM, Osterhaus ADME, Sutter G and Rimmelzwaan GF. (2007) J. Infect. Dis.  195:1598-1606.
13. The global circulation of seasonal influenza A (H3N2) viruses. C.A. Russell, T.C. Jones, I.G. Barr, N.J. Cox, R.J. Garten, V. Gregory, I.D. Gust, A.W. Hampson, A.J. Hay, A.C. Hurt, J.C. de Jong, A. Kelso, A.I. Klimov, T. Kageyama, N. Komadina, A.S. Lapedes, Y.P. Lin, A. Mosterin, M. Obuchi, T. Odagiri, A.D.M.E. Osterhaus, G.F. Rimmelzwaan, M.W. Shaw, E. Skepner, K. Stohr, M. Tashiro, R.A.M. Fouchier, D.J. Smith. (2008) Science 320:340-6.
14. Clearance of influenza virus from the lung depends on migratory langerin+CD11b- but not plasmacytoid dendritic cells. Geurts van Kessel CH, Willart M, van Rijt LS, Kool M, Baas C, Thielemans K, Hoogsteden HC, Osterhaus ADME, Rimmelzwaan GF and Lambrecht BN. (2008) J. Exp. Med. 205(7):1621-1634
15. Recombinant modified vaccinia virus Ankara expressing HA confers protection against homologous and heterologous H5N1 influenza virus infections in macaques. Kreijtz JHCM, Süzer Y, de Mutsert G, van den Brand JMA, van Amerongen G, Schnierle BS, Kuiken T, Fouchier RAM, Löwer J, Osterhaus ADME, Sutter G and Rimmelzwaan GF. (2009) J. Infect. Dis.  199:405-413
16. Antigenic and Genetic Characteristics of Swine-Origin 2009 A(H1N1) Influenza Viruses Circulating in Humans. R. J. Garten, C.T Davis, C.A. Russell, B. Shu, S. Lindstrom, A. Balish, W.M. Sessions, X. Xu, E. Skepner, V. Deyde, M. Okomo-Adhiambo, L. Gubareva, J. Barnes, C.B. Smith, S.L. Emery, M.J. Hillman, P. Rivailler, J. Smagala, M. de Graaf, D.F. Burke, R.A.M. Fouchier, C. Pappas, C.M. Alpuche-Aranda, H. López-Gatell, H. Olivera, I. López, C.A. Myers, D. Faix, P.J. Blair, C. Yu, K.M. Keene, P.D. Dotson, Jr., D. Boxrud, A.R. Sambol, S.H. Abid, K. St. George, T. Bannerman, A.L. Moore, D.J. Stringer, P. Blevins, G.J. Demmler-Harrison, M. Ginsberg, P. Kriner, S. Waterman, S. Smole, H.F. Guevara, E.A. Belongia, P.A. Clark, S.T. Beatrice, R. Donis, J. Katz, L. Finelli, C.B. Bridges, M. Shaw, D.B. Jernigan, T.M. Uyeki, D.J. Smith, A.I. Klimov, and N.J. Cox1. (2009) Science 325:197-201
17. Pathogenesis and transmission of swine-origin 2009 A(H1N1) influenza virus in ferrets. Munster VJ, de Wit E, van den Brand JM, Herfst S, Schrauwen EJ, Bestebroer TM, van de Vijver D, Boucher CA, Koopmans M, Rimmelzwaan GF, Kuiken T, Osterhaus AD, Fouchier RA (2009).Science. Jul 24;325(5939):481-3. Epub 2009 Jul 2.
18. Vaccination of mice against influenza A/H3N2 virus infection prevents the induction of heterosubtypic immunity against lethal H5N1 influenza. Bodewes R, Kreijtz JHCM, Baas C, Geelhoed-Mieras MM, de Mutsert G, van den Brand JMA, van Amerongen G, Fouchier RAM, Osterhaus ADME and Rimmelzwaan GF. (2009) PLoS ONE 4(5) e5538:1-9.
19. Dendritic cells are necessary for maintenance of tertiary lymphoid tissue in the lung of influenza virus infected mice. Geurts van Kessel CH, Willart MAM, van Rijt LS, Bergen IM, Muskens F, Osterhaus ADME, Hendriks R, Rimmelzwaan GF and Lambrecht BN. J. Exp. Med.  In press

4.2     Human metapneumovirus

1. Experimental human metapneumovirus infection of cynomolgus macaques (Macaca fascicularis) results in virus replication in ciliated epithelial cells and pneumocytes with associated lesions throughout the respiratory tract. Kuiken T, van den Hoogen BG, van Riel DA, Laman JD, van Amerongen G, Sprong L, Fouchier RA, Osterhaus AD. Am J Pathol. 2004 Jun;164(6):1893-900.
2. Antigenic and genetic variability of human metapneumoviruses. van den Hoogen BG, Herfst S, Sprong L, Cane PA, Forleo-Neto E, de Swart RL, Osterhaus AD, Fouchier RA. Emerg Infect Dis. 2004 Apr;10(4):658-66.
3. Respiratory picornaviruses and respiratory syncytial virus as causative agents of acute expiratory wheezing in children. Jartti T, Lehtinen P, Vuorinen T, Osterback R, van den Hoogen B, Osterhaus AD, Ruuskanen O. Emerg Infect Dis. 2004 Jun;10(6):1095-101.
4. Recovery of human metapneumovirus genetic lineages a and B from cloned cDNA. Herfst S, de Graaf M, Schickli JH, Tang RS, Kaur J, Yang CF, Spaete RR, Haller AA, van den Hoogen BG, Osterhaus AD, Fouchier RA. J Virol. 2004 Aug;78(15):8264-70.
5. Newer respiratory virus infections: human metapneumovirus, avian influenza virus, and human coronaviruses.Fouchier RA, Rimmelzwaan GF, Kuiken T, Osterhaus AD. Curr Opin Infect Dis. 2005 Apr;18(2):141-6.
6. Low-pH-induced membrane fusion mediated by human metapneumovirus F protein is a rare, strain-dependent phenomenon. Herfst S, Mas V, Ver LS, Wierda RJ, Osterhaus AD, Fouchier RA, Melero JA.  J Virol. 2008 Sep;82(17):8891-5.
7. Fusion protein is the main determinant of metapneumovirus host tropism. de Graaf M, Schrauwen EJ, Herfst S, van Amerongen G, Osterhaus AD, Fouchier RA.  J Gen Virol. 2009 Jun;90(Pt 6):1408-16.

4.3     Respiratory syncytial virus and measles virus

1. De Swart RL, Ludlow M, de Witte L, Yanagi Y, van Amerongen G, McQuaid S, Yüksel S, Geijtenbeek TBH, Duprex WP, Osterhaus ADME. Predominant infection of CD150+ lymphocytes and dendritic cells during measles virus infection of macaques. PloS Pathog. 2007; 3:e178
2. Identification of a common HLA-DP4-restricted T-cell epitope in the conserved region of the respiratory syncytial virus G protein. de Waal L, Yüksel S, Brandenburg AH, Langedijk JP, Sintnicolaas K, Verjans GM, Osterhaus AD, de Swart RL. J Virol. 2004 Feb;78(4):1775-81.
3. Relative contributions of measles virus hemagglutinin- and fusion protein-specific serum antibodies to virus neutralization.   de Swart RL, Yüksel S, Osterhaus AD. J Virol. 2005 Sep;79(17):11547-51.
4. Aerosol measles vaccination in macaques: preclinical studies of immune responses and safety. de Swart RL, Kuiken T, Fernandez-de Castro J, Papania MJ, Bennett JV, Valdespino JL, Minor P, Witham CL, Yüksel S, Vos H, van Amerongen G, Osterhaus AD. Vaccine. 2006 Sep 29;24(40-41):6424-36
5. Measles vaccination of macaques by dry powder inhalation. de Swart RL, LiCalsi C, Quirk AV, van Amerongen G, Nodelman V, Alcock R, Yüksel S, Ward GH, Hardy JG, Vos H, Witham CL, Grainger CI, Kuiken T, Greenspan BJ, Gard TG, Osterhaus AD. Vaccine. 2007 Jan 26;25(7):1183-90
6. Immunization of macaques with formalin-inactivated human metapneumovirus induces hypersensitivity to hMPV infection.   de Swart RL, van den Hoogen BG, Kuiken T, Herfst S, van Amerongen G, Yüksel S, Sprong L, Osterhaus AD. Vaccine. 2007 Dec 12;25(51):8518-28. Epub 2007 Oct 26.
7. Predominant infection of CD150+ lymphocytes and dendritic cells during measles virus infection of macaques. de Swart RL, Ludlow M, de Witte L, Yanagi Y, van Amerongen G, McQuaid S, Yüksel S, Geijtenbeek TB, Duprex WP, Osterhaus AD. PLoS Pathog. 2007 Nov;3(11):e178
8. Measles vaccination: new strategies and formulations. de Vries RD, Stittelaar KJ, Osterhaus AD, de Swart RL. Expert Rev Vaccines. 2008 Oct;7(8):1215-23

4.4     Hepatitis- and coronaviruses

1. Haagmans BL, Kuiken T, Martina BE, Fouchier RA, Rimmelzwaan GF, Van Amerongen G, Van Riel D, De Jong T, Itamura S, Chan KH, Tashiro M, Osterhaus AD Pegylated interferon-alpha protects type 1 pneumocytes against SARS coronavirus infection in macaques Nat Med 2004; 10:290-293 IF 27.5
2. Human monoclonal antibody as prophylaxis for SARS coronavirus infection in ferrets. ter Meulen J, Bakker AB, van den Brink EN, Weverling GJ, Martina BE, Haagmans BL, Kuiken T, de Kruif J, Preiser W, Spaan W, Gelderblom HR, Goudsmit J, Osterhaus AD. Lancet. 2004 Jun 26;363(9427):2139-41.
3. Van der Eijk, AA, JM Vrolijk, BL Haagmans. Antibodies neutralizing peginterferon alfa during retreatment of hepatitis C. N Engl J Med 2006; 354:1323-1324 IF 50.0
4. Nonhuman primate models for SARS. Haagmans BL, Osterhaus AD. PLoS Med. 2006 May;3(5):e194
5. Identification of a naturally occurring recombinant genotype 2/6 hepatitis C virus. Noppornpanth S, Lien TX, Poovorawan Y, Smits SL, Osterhaus AD, Haagmans BL. J Virol. 2006 Aug;80(15):7569-77.
6. Interferon-gamma and interleukin-4 downregulate expression of the SARS coronavirus receptor ACE2 in Vero E6 cells. de Lang A, Osterhaus AD, Haagmans BL. Virology. 2006 Sep 30;353(2):474-81
7. The emerging role of ACE2 in physiology and disease. Hamming I, Cooper ME, Haagmans BL, Hooper NM, Korstanje R, Osterhaus AD, Timens W, Turner AJ, Navis G, van Goor H. J Pathol. 2007 May;212(1):1-11
8. Functional genomics highlights differential induction of antiviral pathways in the lungs of SARS-CoV-infected macaques. de Lang A, Baas T, Teal T, Leijten LM, Rain B, Osterhaus AD, Haagmans BL, Katze MG. PLoS Pathog. 2007 Aug 10;3(8):e112.
9. Characterization of hepatitis C virus deletion mutants circulating in chronically infected patients. Noppornpanth S, Smits SL, Lien TX, Poovorawan Y, Osterhaus AD, Haagmans BL. J Virol. 2007 Nov;81(22):12496-503
10. Pathology of experimental SARS coronavirus infection in cats and ferrets. van den Brand JM, Haagmans BL, Leijten L, van Riel D, Martina BE, Osterhaus AD, Kuiken T. Vet Pathol. 2008 Jul;45(4):551-62.
11. The Application of Genomics to Emerging Zoonotic Viral Diseases  Haagmans BL, Andeweg A, Osterhaus ADME. PLOS Pathog. 2009 in press

4.5     Herpes viruses

1. Verjans GMGM, Hintzen RQ, van Dun JM, Poot A, Milikan JCM, Laman JD, Langerak AW, Kinchington PR, Osterhaus ADME. Selective retention of herpes simplex virus specific T cells in latently infected human trigeminal ganglia. Proc Natl Acad Sci USA. 2007; 104:3496-3501. (IF year 2007: 9.6)
2. Selective retention of herpes simplex virus-specific T cells in latently infected human trigeminal ganglia. Verjans GM, Hintzen RQ, van Dun JM, Poot A, Milikan JC, Laman JD, Langerak AW, Kinchington PR, Osterhaus AD. Proc Natl Acad Sci U S A. 2007 Feb 27;104(9):3496-501.
3. Acyclovir-resistant corneal HSV-1 isolates from patients with herpetic keratitis. Duan R, de Vries RD, Osterhaus AD, Remeijer L, Verjans GM. J Infect Dis. 2008 Sep 1;198(5):659-63.
4. Neuron-interacting satellite glial cells in human trigeminal ganglia have an APC phenotype. van Velzen M, Laman JD, Kleinjan A, Poot A, Osterhaus AD, Verjans GM. J Immunol. 2009 Aug 15;183(4):2456-61.
5. Prevalence and clinical consequences of herpes simplex virus type 1 DNA in human cornea tissues. Remeijer L, Duan R, van Dun JM, Wefers Bettink MA, Osterhaus AD, Verjans GM. J Infect Dis. 2009 Jul 1;200(1):11-9.
6. High incidence of genotypic variance between sequential herpes simplex virus type 2 isolates from HIV-1-seropositive patients with recurrent genital herpes. Roest RW, Maertzdorf J, Kant M, van der Meijden WI, Osterhaus AD, Verjans GM.        J Infect Dis. 2006 Oct 15;194(8):1115-8.
7. Isopentenyl pyrophosphate-reactive Vgamma9Vdelta 2 T helper 1-like cells are the major gammadelta T cell subset recovered from lesions of patients with genital herpes. Verjans GM, Roest RW, van der Kooi A, van Dijk G, van der Meijden WI, Osterhaus AM. J Infect Dis. 2004 Aug 1;190(3):489-93.
8. Identification of a common HLA-DP4-restricted T-cell epitope in the conserved region of the respiratory syncytial virus G protein. de Waal L, Yüksel S, Brandenburg AH, Langedijk JP, Sintnicolaas K, Verjans GM, Osterhaus AD, de Swart RL. J Virol. 2004 Feb;78(4):1775-81.

4.6     Comparative pathology and wildlife disease

1. Pegylated interferon protects type 1 pneumocytes against SARS coronavirus infection in macaques  Kuiken, T. Haagmans, B.L., Chan, K.-H., Tashiro, M., Osterhaus, A.D.M.E.,  Martina, B.E., Fouchier, R.A.M., Itamura, S. (2004) Nature Medicine, 10 (3), pp. 290-293.
2. Avian influenza H5N1 in tigers and leopards Keawcharoen J, Oraveerakul K, Kuiken T, Fouchier RA, Amonsin A, Payungporn S, Noppornpanth S, Wattanodorn S, Theambooniers A, Tantilertcharoen R, Pattanarangsan R, Arya N, Ratanakorn P, Osterhaus DM, Poovorawan Y.  (2004) Emerging Infectious Diseases, 10 (12), pp. 2189-2191.
3. Avian H5N1 influenza in cats. Kuiken,T., Rimmelzwaan,G., van Riel,D., van Amerongen,G., Baars,M., Fouchier,R., and Osterhaus,A. (2004) Science, 306:241. IF 30.9
4. Pegylated interferon-alpha protects type 1 pneumocytes against SARS coronavirus infection in macaques.  Kuiken,T., Haagmans,B.L., Martina,B.E., Fouchier,R.A., Rimmelzwaan,G.F., van Amerongen,G., van Riel,D., de Jong,T., Itamura,S., Chan,K.H., Tashiro,M., and Osterhaus,A.D. (2004) Nat.Med., 10:290-293. IF 28.9
5. Experimental human metapneumovirus infection of cynomolgus macaques (Macaca fascicularis) results in virus replication in ciliated epithelial cells and pneumocytes with associated lesions throughout the respiratory tract. Kuiken,T., van den Hoogen,B.G., van Riel,D.A., Laman,J.D., van Amerongen,G., Sprong,L., Fouchier,R.A., and Osterhaus,A.D. (2004)  Am.J.Pathol., 164:1893-1900. IF 5.8
6. Experimental human metapneumovirus infection of cynomolgus macaques (Macaca fascicularis) results in virus replication in ciliated epithelial cells and pneumocytes with associated lesions throughout the respiratory tract. Kuiken, T., Van Den Hoogen, B.G., Van Riel, D.A.J., Laman, J.D., Van Amerongen, G., Sprong, L., Fouchier, R.A.M., Osterhaus, A.D.M.E.(2004) American Journal of Pathology, 164 (6), pp. 1893-1900.
7. Public health. Pathogen surveillance in animals. Kuiken,T., Leighton,F.A., Fouchier,R.A., LeDuc,J.W., Peiris,J.S., Schudel,A., Stohr,K., and Osterhaus,A.D. (2005)  Science, 309:1680-1681. IF 30.9
8. Newer respiratory virus infections: Human metapneumovirus, avian influenza virus, and human coronaviruses. Fouchier, R.A.M., Rimmelzwaan, G.F., Kuiken, T., Osterhaus, A.D.M.E. (2005) Current Opinion in Infectious Diseases, 18 (2), pp. 141-146.
9. Host species barriers to influenza virus infections.  Kuiken,T., Holmes,E.C., McCauley,J., Rimmelzwaan,G.F., Williams,C.S., and Grenfell,B.T. (2006): Science, 312:394-397. IF 30.9
10. Influenza A virus (H5N1) infection in cats causes systemic disease with potential novel routes of virus spread within and between hosts.  Rimmelzwaan,G., van Riel,D., Baars,M., Bestebroer,T.M., van Amerongen,G., Fouchier,R.A.M., Osterhaus,A.D.M.E., and Kuiken,T. (2006) Am.J.Pathol., 168:176-183. IF 5.8
11. Host species barriers to influenza virus infections. van Riel,D., Munster,V.J., de Wit,E., Rimmelzwaan,G.F., Fouchier,R.A.M., Osterhaus,A.D.M.E., and Kuiken,T. (2006) Science, 312 (5772), pp. 394-397.
12. H5N1 virus attachment to lower respiratory tract. van Riel,D., Munster,V.J., de Wit,E., Rimmelzwaan,G.F., Fouchier,R.A.M., Osterhaus,A.D.M.E., and Kuiken,T. (2006):   Science, 311:399. IF 30.9
13. Human and Avian Influenza Viruses Target Different Cells in the Lower Respiratory Tract of Humans and Other Mammals. van Riel,D., Munster,V.J., de Wit,E., Rimmelzwaan,G.F., Fouchier,R.A., Osterhaus,A.D., and Kuiken,T. (2007)  Am.J.Pathol., 171:1-9. IF 5.8
14. Wild ducks as long-distance vectors of highly pathogenic avian influenza virus (H5N1). Keawcharoen,J., van Riel,D., van Amerongen,G., Bestebroer,T., Beyer,W.E., van Lavieren,R., Osterhaus,A.D., Fouchier,R.A., Kuiken,T. (2008):  Emerg. Infect. Dis., 14:600-607.
15. Stittelaar KJ, Neyts J, Naesens L, van Amerongen G, van Lavieren RF, Holý A, De Clercq E, Niesters HG, Fries E, Maas C, Mulder PG, van der Zeijst BA, Osterhaus AD. Antiviral treatment is more effective than smallpox vaccination upon lethal monkeypox virus infection. : Nature. 2006 Feb 9;439(7077):745-8. Epub 2005 Dec 11.