## 4 Discussion
condylurus_ after experimental infection implies that respiratory or oral spread of infection could occur in the confined spaces of bats roosts [22]. Future experiments may reveal mechanisms that allow EBOV persistence in _M. condylurus_ primary cells and whether ebolaviruses can persist in vivo in certain organs of this microbat species. Another mechanism contributing to the establishment of virus persistence is the accumulation of defective interfering (DI) particles, which may dampen virus replication [64; 99; 100; 101; 102; 103]. Ten undiluted passages of EBOV with high MOI (>1) resulted in production of DI particles and persistently infected Vero cells [104]. In contrast, in our experiment the amount of virus used to infect MoLu Prim cells was low and the amount of infectious virus in the supernatant increased over time. Therefore, the establishment of EBOV persistence in lung primary cells of _M. condylurus_ was not enforced through accumulation of DI particles as described by [PERSON] et al. but seemed to be facilitated by specific characteristics of the cells. Additionally, the selection of virus variants with reduced cytopathogenicity may be a mechanism during the establishment of persistent infections [37; 64; 105; 106; 107; 108]. As MoLu Prim cells tolerated EBOV in several experiments directly after infection, such selection processes seemed to be unlikely. Comparing the full EBOV genome sequences of the inoculum and in the supernatant of MoLu Prim_EBOV cells on day 150, only one nucleotide change from T to G in position 10,590 in the VP24 CDS was detected, resulting in an amino acid change from Arginine to Lysine (both are basic amino acids with similar characteristics). Therefore, no selective pressure for virus mutation and selection of virus variants was observed. Repeated establishment of persistent EBOV infections in primary cells from _M. condylurus_ might reflect the intrinsic ability, that ebolaviruses may persist and be permanently maintained in this bat species and in the bat population, being an essential criterion for a natural reservoir [36; 56; 109]. Difficulties in isolating infectious Nipah virus (NiV) from wild-caught or experimentally infected bats may be, due to near elimination by immune mechanisms, but this virus likely persists at low levels in specific organs [110]. Additionally for MARV, long-term viral persistence in _R. aegyptiacus_ bats has been indirectly observed, due to late transmissions from experimentally infected to naive contact bats 7 months post-infection [55; 56]. In vivo studies will elucidate whether EBOV is also tolerated in _M. condylurus_ bats and can persist without being eliminated by innate and adaptive immune responses. The potential persistence of ebolaviruses in lungs of _M. condylurus_ bats with intermittent viral shedding may explain why no infectious virus has been detected in this or any other bat species, and why spillovers occur rarely. In summary, low NPC1 receptor expression levels in most primary cells from _M. condylurus_, compared to human or monkey cell lines or cells from a European microbat, might contribute to low level virus replication rates in all tested primary cells from _M. condylurus_ and might reflect a potential adaptation between EBOV and its natural reservoir. Most cells derived from _M. condylurus_ showed high tolerance and no cell damage upon infection with EBOV, while cell destruction was observed in control cells. We noticed the repeated establishment of EBOV persistence in primary lung cells from _M. condylurus_, potentially reflecting the intrinsic ability of in vivo persistence in this bat species. With lower NPC1 receptor expression levels, lower EBOV replication rates, high tolerance to EBOV infections, and repeated establishment of persistent infection of primary cells from _M. condylurus_ with slow virus spread in the monolayer, these in vitro experiments provide additional evidence that this bat species might develop asymptomatic infections in vivo and is a likely reservoir of ebolaviruses.