tetano
Editor, Senior Moderator
Virus Res. 2014 May 12. pii: S0168-1702(14)00184-1. doi: 10.1016/j.virusres.2014.05.003. [Epub ahead of print]
Genome rearrangement of influenza virus for anti-viral drug screening.
Sutton TC1, Obadan A1, Lavigne J1, Chen H1, Li W1, Perez DR2.
Author information
Abstract
Rearrangement of the influenza A genome such that NS2 is expressed downstream of PB1 permits the insertion of a foreign gene in the NS gene segment. In this report, the genome rearranged strategy was extended to A/California/04/2009 (pH1N1), and Gaussia luciferase (GLuc) or GFP was expressed downstream of the full-length NS1 gene (designated GLucCa04 and GFPCa04, respectively). In growth kinetics studies, culture of amantadine sensitive GLucCa04 (Sens/GlucCa04) in the presence of amantadine significantly decreased GLuc expression and viral titers for 48h post-infection (hpi). When Sens/GlucCa04 was subsequently used in an in vitro anti-viral screening assay, amantadine treatment significantly decreased GLuc expression from amantadine sensitive compared to amantadine resistant GLucCa04 (Res/GlucCa04) as early as 16 hpi. In in vivo screening studies, DBA mice were treated daily with amantadine from 1 day prior to infection and inoculated with either Sens/GlucCa04 or Res/GlucCa04 alone or as a co-infection with the parental strain. On days 3 and 5 post-infection, lung samples were collected and amantadine treatment was shown to decrease GLuc expression by two orders of magnitude (p<0.05) in Sens/GlucCa04 infected mice. Furthermore, while both Sens and Res/GlucCa04 were highly attenuated, addition of the parental strain to the inoculum yielded clinical disease indicative of GLuc expression and pulmonary viral titers. These findings indicate that the use of GLucCa04 can potentially accelerate in vitro and in vivo anti-viral screening by shortening the time required for virus detection.
Copyright ? 2014. Published by Elsevier B.V.
KEYWORDS:
Amantadine, Anti-viral screening, Gaussia luciferase, Influenza
PMID:
24833536
[PubMed - as supplied by publisher]
http://www.ncbi.nlm.nih.gov/pubmed/24833536
Genome rearrangement of influenza virus for anti-viral drug screening.
Sutton TC1, Obadan A1, Lavigne J1, Chen H1, Li W1, Perez DR2.
Author information
Abstract
Rearrangement of the influenza A genome such that NS2 is expressed downstream of PB1 permits the insertion of a foreign gene in the NS gene segment. In this report, the genome rearranged strategy was extended to A/California/04/2009 (pH1N1), and Gaussia luciferase (GLuc) or GFP was expressed downstream of the full-length NS1 gene (designated GLucCa04 and GFPCa04, respectively). In growth kinetics studies, culture of amantadine sensitive GLucCa04 (Sens/GlucCa04) in the presence of amantadine significantly decreased GLuc expression and viral titers for 48h post-infection (hpi). When Sens/GlucCa04 was subsequently used in an in vitro anti-viral screening assay, amantadine treatment significantly decreased GLuc expression from amantadine sensitive compared to amantadine resistant GLucCa04 (Res/GlucCa04) as early as 16 hpi. In in vivo screening studies, DBA mice were treated daily with amantadine from 1 day prior to infection and inoculated with either Sens/GlucCa04 or Res/GlucCa04 alone or as a co-infection with the parental strain. On days 3 and 5 post-infection, lung samples were collected and amantadine treatment was shown to decrease GLuc expression by two orders of magnitude (p<0.05) in Sens/GlucCa04 infected mice. Furthermore, while both Sens and Res/GlucCa04 were highly attenuated, addition of the parental strain to the inoculum yielded clinical disease indicative of GLuc expression and pulmonary viral titers. These findings indicate that the use of GLucCa04 can potentially accelerate in vitro and in vivo anti-viral screening by shortening the time required for virus detection.
Copyright ? 2014. Published by Elsevier B.V.
KEYWORDS:
Amantadine, Anti-viral screening, Gaussia luciferase, Influenza
PMID:
24833536
[PubMed - as supplied by publisher]
http://www.ncbi.nlm.nih.gov/pubmed/24833536