Abstract
BackgroundThe RGD motif in the mosquito-borne flaviviruses envelope protein domain III (EDIII) FG loop was shown to bind negatively charged cellular molecules and mediate virus entry in mammals. However, its importance in virus entry in the mosquito has not yet been defined. The sequences of RGD motifs are conserved in JEV-serocomplex members primarily transmitted by Culex mosquitoes but absent from members of the DENV serocomplex, which utilize Aedes mosquitoes as vectors. Interestingly, the RGD sequence is present in the attenuated 17D strain of yellow fever virus as a result of the T380R mutation in the EDIII of Asibi strain following extensive in vitro passage in mice and chicken embryos and was found to contribute to the more rapid clearance in mice challenged with 17D. However, viral infectivity and dissemination in mosquitoes had not been evaluated for this mutant.FindingsThe study utilized the reverse genetics system of YFV and Ae. aegypti RexD WE mosquitoes to assess the impact of a T380R mutation in YFV Asibi and 17D/Asibi M-E chimera. The T380R mutation led to higher infection rates but similar dissemination rates when introduced into the YFV Asibi strain and 17D/Asibi M-E chimera.ConclusionsWhile the increase of the positive charge in EDIII may reduce the virulence of YFV in mice, this mutation favored the establishment of the viral infection in Ae. aegypti. However, such gain in viral infectivity did not increase dissemination in infected mosquitoes.
Highlights
Introduction ofT380R mutation in E of infectious clones of YFV Asibi strain and 17D/Asibi M-E chimera was achieved by subcloning the structural genes into the pGEM®-T Easy vector (Promega, Madison, WI) followed by PCR site-directed mutagenesis with Quikchange®II XL kit (Stratagene, La Jolla, CA) and ligation of BspEI (New England Biolabs, Ipswich, MA)- and MluI (New England Biolabs)- digested plasmids as previously described [12]
While the increase of the positive charge in envelope protein domain III (EDIII) may reduce the virulence of YFV in mice, this mutation favored the establishment of the viral infection in Ae. aegypti
Several molecules of mammalian origin have been proposed as putative receptors for mosquitoborne flaviviruses, knowledge about arthropod receptors remains limited [1]
Summary
Introduction ofT380R mutation in E of infectious clones of YFV Asibi strain and 17D/Asibi M-E chimera was achieved by subcloning the structural genes into the pGEM®-T Easy vector (Promega, Madison, WI) followed by PCR site-directed mutagenesis with Quikchange®II XL kit (Stratagene, La Jolla, CA) and ligation of BspEI (New England Biolabs, Ipswich, MA)- and MluI (New England Biolabs)- digested plasmids as previously described [12]. Conclusions: While the increase of the positive charge in EDIII may reduce the virulence of YFV in mice, this mutation favored the establishment of the viral infection in Ae. aegypti. Such gain in viral infectivity did not increase dissemination in infected mosquitoes.
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