1. Quantitative analysis of particles, genomes and infectious particles in supernatants of haemorrhagic fever virus cell cultures
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Faye Fatou Traoré, Gunnel Lindegren, Ali Mirazimi, Jean-Claude Manuguerra, Amadou A. Sall, Aime Adjami, Manfred Weidmann, Lamine Koivogui, Kjell-Olof Hedlund, University Medical Center Göttingen (UMG), Institut Pasteur de Dakar, Réseau International des Instituts Pasteur (RIIP), Cellule d'Intervention Biologique d'Urgence - Laboratory for Urgent Response to Biological Threats (CIBU), Institut Pasteur [Paris] (IP), Université Gamal Abdel Nasser de Conakry, World Health Organizatin [Ouagadougou, Burkina Faso] (WHO Burkina Faso), Fondation Mérieux, Smittskydds Institutet - Swedish Institute for Infectious Disease Control [Solna, Sweden] (SMI), Public Health Agency of Sweden, Department of Microbiology, Tumor and Cell Biology, Karolinska Institutet [Stockholm], and This work was supported by EU grant FP6-INCO-032180 'Development of rapid field diagnostics for identification, control and management of haemorrhagic fever outbreaks (VHF Diagnostics).
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Hemorrhagic Fevers, Viral ,Virus Cultivation ,Arenaviridae ,[SDV]Life Sciences [q-bio] ,viruses ,Cell Culture Techniques ,Filoviridae ,Genome, Viral ,Dengue virus ,medicine.disease_cause ,Virus ,lcsh:Infectious and parasitic diseases ,Microbiology ,Cell Line ,MESH: Virus Cultivation ,03 medical and health sciences ,Virology ,Veterinary virology ,medicine ,Humans ,RNA Viruses ,lcsh:RC109-216 ,Virus classification ,030304 developmental biology ,0303 health sciences ,MESH: Cell Culture Techniques ,MESH: Humans ,biology ,030306 microbiology ,Research ,Virion ,MESH: Hemorrhagic Fevers, Viral ,biology.organism_classification ,3. Good health ,MESH: Cell Line ,Lassa virus ,Infectious Diseases ,MESH: Virion ,Bunyaviridae ,MESH: RNA Viruses - Abstract
Information on the replication of viral haemorrhagic fever viruses is not readily available and has never been analysed in a comparative approach. Here, we compared the cell culture growth characteristics of haemorrhagic fever viruses (HFV), of the Arenaviridae, Filoviridae, Bunyaviridae, and Flavivridae virus families by performing quantitative analysis of cell culture supernatants by (i) electron microscopy for the quantification of virus particles, (ii) quantitative real time PCR for the quantification of genomes, and (iii) determination of focus forming units by coating fluorescent antibodies to infected cell monolayers for the quantification of virus infectivity. The comparative analysis revealed that filovirus and RVFV replication results in a surplus of genomes but varying degrees of packaging efficiency and infectious particles. More efficient replication and packaging was observed for Lassa virus, and Dengue virus resulting in a better yield of infectious particles while, YFV turned out to be most efficient with only 4 particles inducing one FFU. For Crimean-Congo haemorrhagic fever virus (CCHFV) a surplus of empty shells was observed with only one in 24 particles equipped with a genome. The complete particles turned out to be extraordinarily infectious. Background Viral haemorrhagic fevers (VHF) are caused by various haemorrhagic fever viruses (HFV), of the Arenaviridae, Filoviridae, Bunyaviridae ,a ndFlavivridae virus families. Only few laboratories specialize in the research on these agents. Basic virological info rmation on these viruses is scant and described exclusively in the frame of case reports and pathological animal models. Although some progress has been achieved concerning the interaction of these viruses with mechanisms of innate immunity [1-9] and nitrite oxide pathways (CCHFV) [7] concise information on their basic virological characteristics is limited. This type of data however is important for biosafety risk assessment purposes. Here, we present a comparative analysis of quantities of HFV in cell culture determined by electron microscopic counting, quantitative real time RT-PCR and focus forming units, which reveal some features of the replication of these viruses that have not been described before. Methods Virus propagation
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