Plasmodium falciparum:Heterologous Synthesis of the Transmission-Blocking Vaccine Candidate Pfs48/45 in Recombinant Vaccinia Virus-Infected Cells

Abstract After assembly in the cytosol, some Vaccinia virus particles go through a complex process that leads to virus egress and eventually cell-to-cell transmission. Intracellular particles are fully infectious, and therefore virus mutants lacking essential functions in the exit pathway are unable to form plaques but can multiply intracellularly. We isolated virus mutants in which two of the genes required for virus spread (F13L and A27L) were deleted independently or concurrently. The phenotypes of the mutant viruses were consistent with the need of A27L and F13L for intercellular virus transmission, the effect of the ΔA27L mutation being more severe than that of ΔF13L. Despite their defect in spread, ΔA27L mutant viruses could be expanded by infecting cell cultures at high multiplicity of infection, followed by the release of virions from infected cells by physical means. We developed a novel system for the isolation of recombinant Vaccinia virus in which selection is efficiently achieved by recovering plaque formation capacity after re-introduction of A27L into a ΔA27L virus. This system allowed the insertion of foreign DNA into the viral genome without the use of additional genetic markers. Furthermore, starting with a double mutant (ΔA27L-ΔF13L) virus, A27L selection was used in conjunction with F13L selection to mediate simultaneous dual insertions in the viral genome. This selection system facilitates combined expression of multiple foreign proteins from a single recombinant virus.

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Green fluorescent protein expressed by a recombinant vaccinia virus permits early detection of infected cells by flow cytometry

Journal of Immunological Methods ◽

10.1016/s0022-1759(98)00156-2 ◽

1998 ◽

Vol 220 (1-2) ◽

pp. 115-121 ◽

Cited By ~ 22

Author(s):

Javier Domı́nguez ◽

Marı́a del Mar Lorenzo ◽

Rafael Blasco

Keyword(s):

Flow Cytometry ◽

Green Fluorescent Protein ◽

Early Detection ◽

Vaccinia Virus ◽

Fluorescent Protein ◽

Recombinant Vaccinia Virus ◽

Recombinant Vaccinia ◽

Infected Cells ◽

Green Fluorescent

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Long-Term Sterilizing Immunity to Rinderpest in Cattle Vaccinated with a Recombinant Vaccinia Virus Expressing High Levels of the Fusion and Hemagglutinin Glycoproteins

Journal of Virology ◽

10.1128/jvi.76.2.484-491.2002 ◽

2002 ◽

Vol 76 (2) ◽

pp. 484-491 ◽

Cited By ~ 25

Author(s):

Paulo H. Verardi ◽

Fatema H. Aziz ◽

Shabbir Ahmad ◽

Leslie A. Jones ◽

Berhanu Beyene ◽

...

Keyword(s):

Vaccinia Virus ◽

Lethal Dose ◽

Syncytium Formation ◽

Viral Disease ◽

Recombinant Vaccinia Virus ◽

Recombinant Vaccinia ◽

Heat Stable ◽

Infected Cells ◽

Sterilizing Immunity

ABSTRACT Rinderpest is an acute and highly contagious viral disease of ruminants, often resulting in greater than 90% mortality. We have constructed a recombinant vaccinia virus vaccine (v2RVFH) that expresses both the fusion (F) and hemagglutinin (H) genes of rinderpest virus (RPV) under strong synthetic vaccinia virus promoters. v2RVFH-infected cells express high levels of the F and H glycoproteins and show extensive syncytium formation. Cattle vaccinated intramuscularly with as little as 103 PFU of v2RVFH and challenged 1 month later with a lethal dose of RPV were completely protected from clinical disease; the 50% protective dose was determined to be 102 PFU. Animals vaccinated with v2RVFH did not develop pock lesions and did not transmit the recombinant vaccinia virus to contact animals. Intramuscular vaccination of cattle with 108 PFU of v2RVFH provided long-term sterilizing immunity against rinderpest. In addition to being highly safe and efficacious, v2RVFH is a heat-stable, inexpensive, and easily administered vaccine that allows the serological differentiation between vaccinated and naturally infected animals. Consequently, mass vaccination of cattle with v2RVFH could eradicate rinderpest.

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