Multiscale model of defective interfering particle replication for influenza A virus infection in animal cell culture

Cell culture-derived defective interfering particles (DIPs) are considered for antiviral therapy due to their ability to inhibit influenza A virus (IAV) production. DIPs contain a large internal deletion in one of their eight viral RNAs (vRNAs) rendering them replication-incompetent. However, they can propagate alongside their homologous standard virus (STV) during infection in a competition for cellular and viral resources. So far, experimental and modeling studies for IAV have focused on either the intracellular or the cell population level when investigating the interaction of STVs and DIPs. To examine these levels simultaneously, we conducted a series of experiments using highly different multiplicities of infections for STVs and DIPs to characterize virus replication in Madin-Darby Canine Kidney suspension cells. At several time points post infection, we quantified virus titers, viable cell concentration, virus-induced apoptosis using imaging flow cytometry, and intracellular levels of vRNA and viral mRNA using real-time reverse transcription qPCR. Based on the obtained data, we developed a mathematical multiscale model of STV and DIP co-infection that describes dynamics closely for all scenarios with a single set of parameters. We show that applying high DIP concentrations can shut down STV propagation completely and prevent virus-induced apoptosis. Interestingly, the three observed viral mRNAs (full-length segment 1 and 5, defective interfering segment 1) accumulated to vastly different levels suggesting the interplay between an internal regulation mechanism and a growth advantage for shorter viral RNAs. Furthermore, model simulations predict that the concentration of DIPs should be at least 10000 times higher than that of STVs to prevent the spread of IAV. Ultimately, the model presented here supports a comprehensive understanding of the interactions between STVs and DIPs during co-infection providing an ideal platform for the prediction and optimization of vaccine manufacturing as well as DIP production for therapeutic use. Author summary Influenza viruses replicate inside their hosts after infection. Along with the release of wild-type standard virus (STV), they can also generate specific kinds of particles that have deletions in their genome. These so-called defective interfering particles (DIPs) are unable to replicate on their own. However, during co-infection with STV they can interfere with the viral life cycle suppressing STV production and instead release a large number of progeny DIPs. Recent studies have shown promising results regarding their potential as antiviral agents. To characterize the interactions between STVs and DIPs during co-infections, we infected animal cell cultures using 12 different multiplicities of infection for STVs and DIPs, respectively. We measured intra- and extracellular infection dynamics and show that STV replication can be suppressed completely using high amounts of DIPs. Then, we developed a mathematical model that describes co-infection dynamics closely using a single set of parameters. We used this model to predict optimal dosing ratios for DIPs to suppress STV infections and for cell culture-derived production of DIPs for antiviral therapy.

Automated summary

Cell culture-derived defective interfering particles (DIPs) have shown potential as antiviral agents as they can inhibit the production of standard virus (STV) during influenza A virus (IAV) infection. High concentrations of DIPs can completely shut down STV propagation, and a mathematical model developed in this study can accurately predict optimal dosing ratios for DIPs to suppress STV infections. This research provides insight into the interplay between STVs and DIPs during co-infection, which can be important for vaccine manufacturing and therapeutic DIP production.