The recurring emergence of influenza virus strains that are resistant to available antiviral medications has become a global health concern, especially in light of the potential for a new influenza virus pandemic. Currently, virtually all circulating strains of influenza A virus in the United States are resistant to either of the two major classes of anti-influenza drugs (adamantanes and neuraminidase inhibitors). Thus, new therapeutic approaches that can be rapidly deployed and that will address the issue of recurring resistance should be developed. We have tested double and triple combinations of the approved anti-influenza drugs oseltamivir and amantadine together with ribavirin against three influenza virus strains using cytopathic effect inhibition assays in MDCK cells. We selected A/New Caledonia/20/99 (H1N1) and A/Sydney/05/97 (H3N2) as representatives of the wild-type versions of the predominant circulating seasonal influenza virus strains and A/Duck/ MN/1525/81 (H5N1) as a representative of avian influenza virus strains. Dose-response curves were generated for all drug combinations, and the degree of drug interaction was quantified using a model that calculates the synergy (or antagonism) between the drugs in double and triple combinations. This report demonstrates that a triple combination of antivirals was highly synergistic against influenza A virus. Importantly, the synergy of the triple combination was 2-to 13-fold greater than the synergy of any double combination depending on the influenza virus subtype. These data support the investigation of a novel combination of oseltamivir, amantadine, and ribavirin as an effective treatment for both seasonal and pandemic influenza virus, allowing the efficient use of the existing drug supplies.
SUMMARYThe sequences encoding the p24 core protein of human immunodeficiency virus type 1 were identified in a cDNA library made from infected CEM cells. The nucleotide sequence of the DNA coding for p24 was shown to be very similar but not identical to the sequences of lymphadenopathy virus and human T-cell leukaemia virus type IIIb. These sequences were expressed in Escherichia coli at the amino terminus of fl-galactosidase and used to screen a panel of monoclonal antibodies raised against virus-expressed p24. Regions containing the epitopes of five of the monoclonal antibodies were located using a series of amino-and carboxy-terminal deletion mutants of the recombinant p24 protein.
The replication and polypeptide synthesis of orbivirus isolate Mill Door/79, a member of the Kemerovo serogroup, were studied. In Vero cells cell-associated virus exceeded cell-free virus by about 2 log10 PFU/ml. Attempts to purify the virus resulted in the demonstration of five polypeptides. Thirteen virus-induced polypeptides and 10 segments of double-stranded RNA were identified in infected cells. Partial proteolysis demonstrated homology between some polypeptides.
Mill Door/79 virus, an orbivirus of the Kemerovo serogroup, Great Island Complex, was shown to induce in infected cells 10 segments of double-stranded RNA with a total molecular weight of 11.46 X 10(6) daltons. Using methyl mercuric hydroxide as a denaturing agent the double-stranded RNA was translated in a cell-free system producing 11 polypeptides, 10 of which co-migrated with those produced in Mill Door/79 virus infected Vero cells. The segments were separated and individually translated in a cell-free system allowing their coding assignments to be made. These assignments were confirmed by partial proteolysis.
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