(In)validating experimentallyderived knowledge about influenza A defective interfering particles

A defective interfering particle (DIP) in the context of influenza A is a virion with a significantly shortened RNA segment substituting one of eight fulllength parent RNA segments, such that it is preferentially amplified. Hence, a cell coinfected with DIPs will produce mainly DIPs, suppressing infectious virus yields, and affecting infection kinetics. Unfortunately, the quantification of DIPs contained in a sample is difficult since they are indistinguishable from standard virus. Using a mathematical model, we investigated the standard experimental method for counting DIPs based on the reduction in infectious virus yield (Bellett and Cooper, 1959). We found the method is valid for counting DIPs provided that: (1) a cell’s coinfection window is approximately half its eclipse phase (it blocks infection by other virions before it begins producing progeny virions); (2) a cell coinfected by virus and DIP produces less than 1 infectious virus per 1,000 DIPs; and (3) highly concentrated virus stock (>4 PFU/cell) is used to perform the assay. Prior work makes no mention of these criteria such that the method has been applied incorrectly in several publications discussed herein. We determined influenza A virus meets these criteria, making the method suitable for counting influenza A DIPs. 
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