Identification and Characterization of Defective Viral Genomes in Ebola Virus-Infected Rhesus Macaques

Abstract

Ebola virus (EBOV), of the family Filoviridae, is an RNA virus that can cause hemorrhagic fever with a high mortality rate. Defective viral genomes (DVGs) are truncated genomes that have been observed during multiple RNA virus infections, including in vitro EBOV infection, and have previously been associated with viral persistence and immunostimulatory activity. As DVGs have been detected in cells persistently infected with EBOV, we hypothesized that DVGs may also accumulate during viral replication in filovirus-infected hosts. Therefore, we interrogated sequence data from serum and tissues using a bioinformatics tool in order to identify the presence of DVGs in nonhuman primates (NHPs) infected with EBOV, Sudan virus (SUDV) or Marburg virus (MARV). Multiple 5’ copy-back DVGs (cbDVGs) were detected in NHP serum during the acute phase of filovirus infection. While the relative abundance of total DVGs in most animals was low, serum collected during acute EBOV and SUDV infections, but not MARV infection, contained a higher proportion of short trailer sequence cbDVGs than the challenge stock. This indicated an accumulation of these DVGs throughout infection, potentially due to the preferential replication of short DVGs over the longer viral genome. Using RT-PCR and deep sequencing, we also confirmed the presence of 5’ cbDVGs in EBOV-infected NHP testes, which is of interest due to EBOV persistence in semen of male survivors of infection. This work suggests that DVGs play a role in EBOV infection in vivo and further study will lead to a better understanding of EBOV pathogenesis. Importance The study of filovirus pathogenesis is critical for understanding the consequences of infection and the development of strategies to ameliorate future outbreaks. Defective viral genomes (DVGs) have been detected during EBOV infections in vitro , however their presence in in vivo infections remains unknown. In this study, DVGs were detected in samples collected from EBOV- and SUDV-infected nonhuman primates (NHPs). The accumulation of these DVGs in the trailer region of the genome during infection indicates a potential role in EBOV and SUDV pathogenesis. In particular, the presence of DVGs in the testes of infected NHPs requires further investigation as it may be linked to the establishment of persistence.

Natural History of Sudan ebolavirus to Support Medical Countermeasure Development

“…The approach of sequence analysis used in this study is an effective means of determining the consensus sequence but is not as accurate in describing the RNA molecules late in infection. Analysis of the RNA genomes during the course of Ebola infection has shown that defective truncated panhandle genomes accumulate late in infection [33]. In another SUDV natural history study performed at another BSL-4 facility, viral RNA was enriched by selecting out ribosomal and messenger RNA and analyzing the resulting cDNA library for evidence of defective genomes [34].…”

Why does viral RNA sometimes persist after recovery from acute infections?

“…For example, in lung infection with RSV, early production of DVGs activates RIG-I-like receptors to stimulate the activation of IRF3 and IRF1, leading to production of TNFα, IFNλ, and IFIT1, suppression of virus replication, and survival of persistently infected cells [ 136 , 139 , 140 ]. DVGs have been demonstrated in the testes during filovirus infection of nonhuman primates [ 141 ] and in the lungs of children with RSV infection [ 140 ].…”

Characterization of an air-liquid interface primary human vaginal epithelium to study Ebola virus infection and testing of antivirals

“…Regarding laboratory experiments, there is still no published in vivo study demonstrating sexual transmission. However, several studies in macaques and guinea pigs have found virus genome or antigen in tissues, and glands of the male but also female genital system following an intra-muscular or peritoneal challenge ( Cooper et al, 2018 ; Liu et al, 2022 ; Perry et al, 2018 ; Baskerville et al, 1985 ; Johnson et al, 2021 ; Zeng et al, 2017 ). This suggests that both genders can be infectious and ultimately infect their partners by the sexual route.…”