Development of a reverse genetics system to generate recombinant Marburg virus derived from a bat isolate.
Recent investigations have shown the Egyptian fruit bat (Rousettus aegyptiacus) to be a natural reservoir for marburgviruses. To better understand the life cycle of these viruses in the natural host, a new reverse genetics system was developed for the reliable rescue of a Marburg virus (MARV) originally isolated directly from a R. aegyptiacus bat (371Bat). To develop this system, the exact terminal sequences were first determined by 5' and 3' RACE, followed by the cloning of viral proteins NP, VP35, VP30 and L into expression plasmids. Novel conditions were then developed to efficiently replicate virus mini-genomes followed by the construction of full-length genomic clones from which recombinant wild type and GFP-containing MARVs were rescued. Surprisingly, when these recombinant MARVs were propagated in primary human macrophages, a dramatic difference was found in their ability to grow and to elicit anti-viral cytokine responses.
Centers for Disease Control and Prevention, Atlanta, GA 30333, USA.
César G
CG
Albariño
Luke S
LS
Uebelhoer
Joel P
JP
Vincent
Marina L
ML
Khristova
Ayan K
AK
Chakrabarti
Anita
A
McElroy
Stuart T
ST
Nichol
Jonathan S
JS
Towner
D000818:Animals; D002478:Cells, Cultured; D002685:Chiroptera / Q000821:virology*; D003001:Cloning, Molecular; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008264:Macrophages / Q000821:virology; D029024:Marburgvirus / Q000235:genetics* / Q000302:isolation & purification; D010957:Plasmids; D011995:Recombination, Genetic; D059386:Reverse Genetics / Q000379:methods*; D013194:Staining and Labeling / Q000379:methods; D014764:Viral Proteins / Q000235:genetics; D014773:Virology / Q000379:methods*
D014764:Viral Proteins; D049452:Green Fluorescent Proteins
Bat isolate; Cytokines; Filovirus; Full-length; GFP; Macrophages; Marburg virus; Marburgvirus; Minigenome; Recombinant