Mammarenaviral Infection Is Dependent on Directional Exposure to and Release from Polarized Intestinal Epithelia

Warner, Nikole L.; Jokinen, Jenny D.; Beier, Juliane I.; Sokoloski, Kevin J.; Lukashevich, Igor S.

Apical exposure of polarized Caco-2 cells to both strains of LCMV resulted in robust infection and virus release from primarily the apical cell surface. Nonetheless, basolateral release of infectious virus particles was observed at later times post infection. Therefore, while infectious particles were released from both surfaces, the release was more efficient from the apical surface, with a~2-log difference between the two supernatants ( Figure 3A ). In contrast, when polarized Caco-2 cells were exposed to LCMV-Arm and LCMV-WE via the basolateral side, infection resulted in roughly the equivalent release of infectious particles from both cell surfaces ( Figure 3B ). during infections of LCMV strains with different pathogenic potential for NHPs, the polarized Caco-2 cells were exposed either apically or basolaterally to the aforementioned OW arenaviruses. To verify the integrity of the polarized monolayer during the experiment, TEER was measured regularly, and the apical and basolateral supernatants were collected every 24 h for a period of 5 days. Apical exposure of polarized Caco-2 cells to both strains of LCMV resulted in robust infection and virus release from primarily the apical cell surface. Nonetheless, basolateral release of infectious virus particles was observed at later times post infection. Therefore, while infectious particles were released from both surfaces, the release was more efficient from the apical surface, with a ~2-log difference between the two supernatants ( Figure 3A ). In contrast, when polarized Caco-2 cells were exposed to LCMV-Arm and LCMV-WE via the basolateral side, infection resulted in roughly the equivalent release of infectious particles from both cell surfaces ( Figure 3B ). Supernatants were collected from both the insert, and well of the Transwells, to determine viral release from the apical or basolateral surfaces independent from one another. Viral titer was measured using standard plaque assay. Release from the Apical surface (black) and the Basolateral surface (red) is plotted with respect to time, with initial viral load subtracted. Values shown are the means of 3 biological replicates with the error bar representing the standard deviation of the mean. If viral plaque forming units (PFUs) were not observed, data received a place-holder value to signify samples were tested, but no data (ND) was collected. # indicates that one or more biological replicates was below limit of detection. * p-value ≤ 0.05.