Establishment, immortalisation and characterisation of pteropid bat cell lines.
BACKGROUND
Bats are the suspected natural reservoir hosts for a number of new and emerging zoonotic viruses including Nipah virus, Hendra virus, severe acute respiratory syndrome coronavirus and Ebola virus. Since the discovery of SARS-like coronaviruses in Chinese horseshoe bats, attempts to isolate a SL-CoV from bats have failed and attempts to isolate other bat-borne viruses in various mammalian cell lines have been similarly unsuccessful. New stable bat cell lines are needed to help with these investigations and as tools to assist in the study of bat immunology and virus-host interactions.


RESULTS
Black flying foxes (Pteropus alecto) were captured from the wild and transported live to the laboratory for primary cell culture preparation using a variety of different methods and culture media. Primary cells were successfully cultured from 20 different organs. Cell immortalisation can occur spontaneously, however we used a retroviral system to immortalise cells via the transfer and stable production of the Simian virus 40 Large T antigen and the human telomerase reverse transcriptase protein. Initial infection experiments with both cloned and uncloned cell lines using Hendra and Nipah viruses demonstrated varying degrees of infection efficiency between the different cell lines, although it was possible to infect cells in all tissue types.


CONCLUSIONS
The approaches developed and optimised in this study should be applicable to bats of other species. We are in the process of generating further cell lines from a number of different bat species using the methodology established in this study.
Crameri
Gary
G
CSIRO Livestock Industries, Australian Animal Health Laboratory, Geelong, Australia.
Todd
Shawn
S
Grimley
Samantha
S
McEachern
Jennifer A
JA
Marsh
Glenn A
GA
Smith
Craig
C
Tachedjian
Mary
M
De Jong
Carol
C
Virtue
Elena R
ER
Yu
Meng
M
Bulach
Dieter
D
Liu
Jun-Ping
JP
Michalski
Wojtek P
WP
Middleton
Deborah
D
Field
Hume E
HE
Wang
Lin-Fa
LF
D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods*; D002461:Cell Line, Transformed / Q000166:cytology*; D048430:Cell Shape / Q000187:drug effects; D002685:Chiroptera; D003001:Cloning, Molecular; D045404:Hendra Virus / Q000187:drug effects / Q000502:physiology; D045464:Henipavirus Infections / Q000821:virology; D006801:Humans; D007113:Immunity, Innate / Q000187:drug effects / Q000276:immunology; D007372:Interferons / Q000235:genetics; D045405:Nipah Virus / Q000187:drug effects / Q000502:physiology; D011070:Poly I-C / Q000494:pharmacology; D013539:Simian virus 40 / Q000235:genetics
D007372:Interferons; D011070:Poly I-C