A live attenuated recombinant dengue-4 virus vaccine candidate with restricted capacity for dissemination in mosquitoes and lack of transmission from vaccinees to mosquitoes.

Abstract

Abstract. 2A⌬30 is a live dengue-4 virus vaccine candidate with a 30-nucleotide deletion in its 3Ј-untranslated region. To assess the transmissibility of 2A⌬30 by mosquitoes, we compared its in vivo replication in mosquitoes with that of its wild type DEN-4 parent. Both the vaccine candidate and wild type virus were equally able to infect the mosquito Toxorhynchites splendens after intrathoracic inoculation. Relative to its wild type parent, 2A⌬30 was slightly restricted in its ability to infect the midgut of Aedes aegypti mosquitoes fed on an artificial blood meal and was even more restricted in its ability to disseminate from the midgut to the salivary glands. Thus, the 30-nucleotide deletion rendered the vaccine candidate more sensitive than its wild type parent to the mosquito midgut escape barrier. Most significantly, 2A⌬30 was not transmitted to 352 Ae. albopictus mosquitoes fed on 10 vaccinees, all of whom were infected with the vaccine candidate.

Dengue virus type 3 vaccine candidates generated by introduction of deletions in the 3′ untranslated region (3′-UTR) or by exchange of the DENV-3 3′-UTR with that of DENV-4

“…When the Δ30 mutation was introduced into the DENV-4 3'-UTR (rDEN3-3'D4Δ30), significant attenuation was observed in C6/36 cells, Vero cells, SCID-HuH-7 mice, and rhesus monkeys. The impact of Δ30 in rDEN3-3'D4 is generally consistent with its impact in rDEN4, save that in the latter case the addition of Δ30 does not cause attenuation in Vero cells [27]. This finding further indicates that the context of the Δ30 deletion dictates whether the mutation will confer attenuation upon a given virus.…”

“…In the case of rDEN3Δ86 and rDEN3Δ30/31, the lack of detectable viremia in rhesus monkeys suggests that they would indeed replicate to low levels in humans similar to other DENV vaccine candidates studied in our laboratory [18]. Second, live attenuated DENV vaccine candidates should have an inherent restriction for growth in mosquitoes, as observed for the rDEN4Δ30 virus [27]. Therefore, the DENV-3 described here were assayed for replication in C6/36 cells which are derived from Aedes albopictus.…”

“…Intrathoracic inoculation of serial ten-fold dilutions of test virus was performed as described previously [27]. After a 14 day incubation, heads were separated and homogenized in diluent.…”

Replacement of the 3' untranslated variable region of mosquito-borne dengue virus with that of tick-borne Langat virus does not alter vector specificity

“…Ae. aegypti were maintained on sucrose and fed on bloodmeals containing designated quantities of virus as described previously (Troyer et al, 2001), incubated for 14 days and frozen. Viral titre in body and head homogenates from mosquito bodies that tested positive for virus was determined in Vero cells as described previously (Hanley et al, 2008).…”

Size Heterogeneity in the 3′ Noncoding Region of South American Isolates of Yellow Fever Virus

“…Experimental studies indicate that variations in the 3Ј NCR are well tolerated but may sometimes lead to altered growth characteristics in cell culture (3,6,19), changes in virulence and immunogenicity in vertebrate animals (10,17,21,22), or reduced infectivity in mosquitoes (1,44). Molecular epidemiological studies of naturally occurring flaviviruses also suggest that 3Ј NCR structural differences correlate with patterns of virus distribution and transmission (39,50).…”