Establishment of Myotis myotis cell lines--model for investigation of host-pathogen interaction in a natural host for emerging viruses.

Xiaocui He,Jiří Pikula,Tomáš Korytář,Jan Zukal,Bernd Köllner,Hana Bandouchova,Yaqing Zhu,Matthias Johannes Schnell

doi:10.1371/journal.pone.0109795

Abstract

Bats are found to be the natural reservoirs for many emerging viruses. In most cases, severe clinical signs caused by such virus infections are normally not seen in bats. This indicates differences in the virus-host interactions and underlines the necessity to develop natural host related models to study these phenomena. Due to the strict protection of European bat species, immortalized cell lines are the only alternative to investigate the innate anti-virus immune mechanisms. Here, we report about the establishment and functional characterization of Myotis myotis derived cell lines from different tissues: brain (MmBr), tonsil (MmTo), peritoneal cavity (MmPca), nasal epithelium (MmNep) and nervus olfactorius (MmNol) after immortalization by SV 40 large T antigen. The usefulness of these cell lines to study antiviral responses has been confirmed by analysis of their susceptibility to lyssavirus infection and the mRNA patterns of immune-relevant genes after poly I:C stimulation. Performed experiments indicated varying susceptibility to lyssavirus infection with MmBr being considerably less susceptible than the other cell lines. Further investigation demonstrated a strong activation of interferon mediated antiviral response in MmBr contributing to its resistance. The pattern recognition receptors: RIG-I and MDA5 were highly up-regulated during rabies virus infection in MmBr, suggesting their involvement in promotion of antiviral responses. The presence of CD14 and CD68 in MmBr suggested MmBr cells are microglia-like cells which play a key role in host defense against infections in the central nervous system (CNS). Thus the expression pattern of MmBr combined with the observed limitation of lyssavirus replication underpin a protective mechanism of the CNS controlling the lyssavirus infection. Overall, the established cell lines are important tools to analyze antiviral innate immunity in M. myotis against neurotropic virus infections and present a valuable tool for a broad spectrum of future investigations in cellular biology of M. myotis.

Highlights

Bats belong to one of the most abundant, diverse and widely distributed mammalian groups
Permanent cell lines of different origin could be established after immortalisation Five M. myotis cell lines brain (MmBr), tonsil (MmTo), peritoneal cavity (MmPca), nasal epithelium (MmNep) and nervus olfactorius (MmNol) were successfully established by transformation with Simian Vacuolating Virus 40 large T antigen (SV40T) gene integrating into the chromosomal DNA
Varying cell morphologies were observed in the cell lines, with MmBr, MmTo, MmNep and MmNol being fibroblastic-like, and MmPca being epithelial-like (Fig. 1A)

Summary

Introduction

Bats belong to one of the most abundant, diverse and widely distributed mammalian groups. Bats evolved early and changed very little over the past 52 million years [2]. Their wide distribution and migratory behaviour favour bats as vectors for viruses and raise concerns over their role in zoonotic diseases [3,4,5]. 13 of the 15 lyssaviruses, except Mokola virus and Ikoma lyssavirus, were detected in bats. In North America bats host RABV, whereas in Europe European Bat lyssavirus type 1 and 2 (EBLV-1 and EBLV-2) are found in different bat species [12,13].

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Conclusion