Characterization of the immune response against recombinant Tacaribe arenaviruses as new vaccine candidates to prevent argentinean hemorrhagic fever

GALLO GIOVANNA LUCRECIA; CHIALE, CAROLINA; SEPÚLVEDA, CLAUDIA; BRIGNONE, JULIA; GAMBOA GRACIELA; RIERA LAURA; SAAVEDRA MARÍA DEL CARMEN; ZUNIGA, ELINA; LOPEZ, NORA

Mendoza

Congreso; LVIII Reunión anual de la Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2022

Sociedad Argentina de Investigación en Bioquímica y Biología Molecular

Junín (JUNV) mammarenavirus is the causative agent of Argentine hemorrhagic fever (AHF), a severe human disease endemic to agricultural areas in Argentina. The current approved vaccine, live-attenuated JUNV strain Candid#1, is effective against AHF, nevertheless, part of the population at risk is excluded from vaccination. Tacaribe (TCRV) is a naturally attenuated mammarenavirus and, as it is phylogenetically and antigenically highly related to JUNV, it stands as a possible vaccine candidate against JUNV. We have previously developed a reverse genetic system that allowed us to successfully rescue infectious recombinant TCRV (rTCRV). Based on this, we generated mutations in the genomic 5’ non coding region, expected to attenuate viral propagation (rTCRV-NCR), and in the Exonuclease domain (ExoN) of the viral nucleoprotein (NP), predicted to counteract the IFN-I cell response (rTCRV-NCR-ExoN). We infected human dendritic cells (hMoDCs), the earliest mammarenavirus cell target, and observed expression of CD86 and HLA-DR activation markers at 24 and 48 hours post infection (h.p.i) with wild type TCRV and rTCRVs. Notably, rTCRV-NCR-ExoN produced the highest degree of activation, similar to that of Candid#1. No IFN-I release was observed from supernatants of infected hMoDCs with wild type TCRV, rTCRV and rTCRV-NCR; nevertheless, it was significantly increased in rTCRV-NCR-ExoN infected cells, suggesting that, as expected, the ExoN domain of NP counteracts the innate immune response. There were no differences in viral titers or in intracellular viral RNA accumulation, determined by plaque assay and RT-qPCR, respectively.We also characterized the IFN-I response in lung epithelial cell line A549 and observed that, as for hMoCDs, only rTCRV-NCR-ExoN and Candid#1 vaccine strain produced a high IFN-I peak in cell supernatants at 24 h.p.i. The IFN-I pathway was also monitored intracellularly, by using a luciferase reporter plasmid under the control of an IFN-inducible promoter. We observed high levels of the reporter gene expression only in rTCRV-NCR-ExoN and Candid#1-infected cells. These findings corroborated that rTCRV-NCR-ExoN is unable to counteract the IFN-I signaling pathway.Lastly, we evaluated virulence of these vaccine candidates in a mouse infection model. Groups of two-day old Balb/cJ mice were inoculated intracranially with serial dilutions of wild type TCRV and each of the rTCRVs, and mortality was monitored daily over a 21-day period. rTCRV and mutant rTCRV-NCR were highly virulent causing almost 100% mortality at a dose of 1 PFU. rTCRV-NCR-ExoN induced 50% mortality, indicating that the NCR and ExoN mutations associated with a considerable degree of viral attenuation in vivo. The control groups of mice inoculated with medium or left untreated survived through day 21. In conclusion, we developed a recombinant mutant TCRV that demonstrated to be attenuated both in vitro and in vivo. This work may contribute to the development of safe vaccines to prevent hemorrhagic fevers produced by mammarenaviruses.