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Transmitted/founder simian immunodeficiency virus envelope sequences in vesicular stomatitis and Semliki forest virus vector immunized rhesus macaques.

Gambhira R, Keele BF, Schell JB, Hunter MJ, Dufour JP, Montefiori DC, Tang H, Rose JK, Rose N, Marx PA - PLoS ONE (2014)

Bottom Line: We used single genome amplification and phylogenetic analyses to characterize transmitted/founder SIVs both in the inoculum and in immunized-infected rhesus monkeys.We obtained 246 full-length envelope sequences from SIVsmE660 infected macaques at the peak of infection and determined the number of transmitted/founder variants per animal.Because there were only 2 infected animals in the gag-env vaccinated rhesus compared to 10 infected rhesus in the other 2 groups, the significance of finding single env variants in the gag-env vaccinated group could not be established.

View Article: PubMed Central - PubMed

Affiliation: Division of Microbiology, Tulane National Primate Research Center, Tulane University, Covington, Louisiana, United States of America.

ABSTRACT
Identification of transmitted/founder simian immunodeficiency virus (SIV) envelope sequences responsible for infection may prove critical for understanding HIV/SIV mucosal transmission. We used single genome amplification and phylogenetic analyses to characterize transmitted/founder SIVs both in the inoculum and in immunized-infected rhesus monkeys. Single genome amplification of the SIVsmE660 inoculum revealed a maximum diversity of 1.4%. We also noted that the consensus sequence of the challenge stock differed from the vaccine construct in 10 amino acids including 3 changes in the V4 loop. Viral env was prepared from rhesus plasma in 3 groups of 6 immunized with vesicular stomatitis virus (VSV) vectors and boosted with Semliki forest virus (SFV) replicons expressing (a) SIVsmE660 gag-env (b) SIVsmE660 gag-env plus rhesus GM-CSF and (c) control influenza hemagglutinin protein. Macaques were immunized twice with VSV-vectors and once with SFV vector and challenged intrarectally with 4000 TCID50. Single genome amplification characterized the infections of 2 unprotected animals in the gag-env immunized group, both of which had reduced acute plasma viral loads that ended as transient infections indicating partial immune control. Four of 6 rhesus were infected in the gag-env + GM-CSF group which demonstrated that GM-CSF abrogated protection. All 6 animals from the control group were infected having high plasma viral loads. We obtained 246 full-length envelope sequences from SIVsmE660 infected macaques at the peak of infection and determined the number of transmitted/founder variants per animal. Our analysis found that 2 of 2 gag-env vaccinated but infected macaques exhibited single but distinct virus envelope lineages whereas rhesus vaccinated with gag-env-GM-CSF or HA control exhibited both single and multiple env lineages. Because there were only 2 infected animals in the gag-env vaccinated rhesus compared to 10 infected rhesus in the other 2 groups, the significance of finding single env variants in the gag-env vaccinated group could not be established.

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Comparison of T/F env sequences to SIVsmE660 stock sequence and to T/F env sequences from different infected macaques.We phylogenetically compared the T/F env sequences obtained from all the infected macaques to the sequences obtained from challenge virus from groups (A) Gag-Env (B) Gag-Env-GM-CSF and (C) VSV-HA macaques. (D) Representative env sequences (5) from each infected macaque from each lineage of T/F viruses from all of the infected animals were compared to each other.
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pone-0109678-g008: Comparison of T/F env sequences to SIVsmE660 stock sequence and to T/F env sequences from different infected macaques.We phylogenetically compared the T/F env sequences obtained from all the infected macaques to the sequences obtained from challenge virus from groups (A) Gag-Env (B) Gag-Env-GM-CSF and (C) VSV-HA macaques. (D) Representative env sequences (5) from each infected macaque from each lineage of T/F viruses from all of the infected animals were compared to each other.

Mentions: We phylogenetically analyzed the combined full-length envelope sequences obtained from all SIVsmE660 infected animals. The envelope sequences from different founder viruses had overlap with challenge stock virus sequence even though they were not identical to the stock envs (Fig. 8A). The two founder lineages from the gag-env immunized (Group I) animals were separated into two different regions of the tree with none of the envelope sequences overlapping with gag-env-GM-CSF (Group II) or control HA (Group III). In the case of gag-env-GM-CSF macaques, full-length envelope sequences from EF28, EA28 overlapped demonstrating that there was common founder virus envelope infecting both the RhM. Furthermore; sequences from EF28 and EJ74 were found in multiple sites on the phylogenetic tree confirming that these animals were infected with more than one lineage. Sequences from the control HA vaccinated animals DD04, FT80 and N288 showed evidence of a single founder variant each whereas sequences from EN82, EP84 and FG05 exhibited multiple lineages. The sequences from EJ74 were the most variable of all the envelope sequences and were distributed along the entire phylogenetic tree. Interestingly, a few of the envelope sequences from FG05, EF28 and EA28 were identical suggesting a potential fitness advantage for this particular linage. In VSV-HA immunized animals many T/F sequences were identical in several animals (N288, EP84, EN82, FG05 and EJ74) again suggesting a selective advantage to this particular lineage in transmission. Finally, we analyzed 5 T/F envelope sequences (representing majority lineages) from all the infected macaques and compared them phylogenetically (Fig. 8D). We observed that there is not only an overlap of envelope sequences between different animals of the same immunized group but also between different groups of macaques. The T/F lineage from both the gag-env group infected macaques grouped separately and didn't overlap with any other lineages. The envelope sequences from gag-env-GM-CSF and HA RhM did overlap with each other. The T/F viral envelope sequences from all the infected macaques will be deposited in the Genbank. We further analyzed the glyocsylation status of founder viruses by using the N-Glycosite software from HIV database web site (http://www.hiv.lanl.gov/). HIV contains two heavily glycosylated proteins gp120 and gp41, which mediate attachment of virions to CD4 on the cell surface. Further, protein N-glycosylation is critical to the pathogenesis of HIV at the levels of viral infectivity and cytopathicity but not at the level of virus replication or host-cell infectability (38–40). We wanted to examine if there were any differences in the glycosylation profiles of envelopes derived from gag-env vaccinated macaques to those in the control HA group. We failed to observe any differences in the glycosylation profiles between these two groups and on an average each sequence had 25–27 N-glycosylation sites (data not shown).


Transmitted/founder simian immunodeficiency virus envelope sequences in vesicular stomatitis and Semliki forest virus vector immunized rhesus macaques.

Gambhira R, Keele BF, Schell JB, Hunter MJ, Dufour JP, Montefiori DC, Tang H, Rose JK, Rose N, Marx PA - PLoS ONE (2014)

Comparison of T/F env sequences to SIVsmE660 stock sequence and to T/F env sequences from different infected macaques.We phylogenetically compared the T/F env sequences obtained from all the infected macaques to the sequences obtained from challenge virus from groups (A) Gag-Env (B) Gag-Env-GM-CSF and (C) VSV-HA macaques. (D) Representative env sequences (5) from each infected macaque from each lineage of T/F viruses from all of the infected animals were compared to each other.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215841&req=5

pone-0109678-g008: Comparison of T/F env sequences to SIVsmE660 stock sequence and to T/F env sequences from different infected macaques.We phylogenetically compared the T/F env sequences obtained from all the infected macaques to the sequences obtained from challenge virus from groups (A) Gag-Env (B) Gag-Env-GM-CSF and (C) VSV-HA macaques. (D) Representative env sequences (5) from each infected macaque from each lineage of T/F viruses from all of the infected animals were compared to each other.
Mentions: We phylogenetically analyzed the combined full-length envelope sequences obtained from all SIVsmE660 infected animals. The envelope sequences from different founder viruses had overlap with challenge stock virus sequence even though they were not identical to the stock envs (Fig. 8A). The two founder lineages from the gag-env immunized (Group I) animals were separated into two different regions of the tree with none of the envelope sequences overlapping with gag-env-GM-CSF (Group II) or control HA (Group III). In the case of gag-env-GM-CSF macaques, full-length envelope sequences from EF28, EA28 overlapped demonstrating that there was common founder virus envelope infecting both the RhM. Furthermore; sequences from EF28 and EJ74 were found in multiple sites on the phylogenetic tree confirming that these animals were infected with more than one lineage. Sequences from the control HA vaccinated animals DD04, FT80 and N288 showed evidence of a single founder variant each whereas sequences from EN82, EP84 and FG05 exhibited multiple lineages. The sequences from EJ74 were the most variable of all the envelope sequences and were distributed along the entire phylogenetic tree. Interestingly, a few of the envelope sequences from FG05, EF28 and EA28 were identical suggesting a potential fitness advantage for this particular linage. In VSV-HA immunized animals many T/F sequences were identical in several animals (N288, EP84, EN82, FG05 and EJ74) again suggesting a selective advantage to this particular lineage in transmission. Finally, we analyzed 5 T/F envelope sequences (representing majority lineages) from all the infected macaques and compared them phylogenetically (Fig. 8D). We observed that there is not only an overlap of envelope sequences between different animals of the same immunized group but also between different groups of macaques. The T/F lineage from both the gag-env group infected macaques grouped separately and didn't overlap with any other lineages. The envelope sequences from gag-env-GM-CSF and HA RhM did overlap with each other. The T/F viral envelope sequences from all the infected macaques will be deposited in the Genbank. We further analyzed the glyocsylation status of founder viruses by using the N-Glycosite software from HIV database web site (http://www.hiv.lanl.gov/). HIV contains two heavily glycosylated proteins gp120 and gp41, which mediate attachment of virions to CD4 on the cell surface. Further, protein N-glycosylation is critical to the pathogenesis of HIV at the levels of viral infectivity and cytopathicity but not at the level of virus replication or host-cell infectability (38–40). We wanted to examine if there were any differences in the glycosylation profiles of envelopes derived from gag-env vaccinated macaques to those in the control HA group. We failed to observe any differences in the glycosylation profiles between these two groups and on an average each sequence had 25–27 N-glycosylation sites (data not shown).

Bottom Line: We used single genome amplification and phylogenetic analyses to characterize transmitted/founder SIVs both in the inoculum and in immunized-infected rhesus monkeys.We obtained 246 full-length envelope sequences from SIVsmE660 infected macaques at the peak of infection and determined the number of transmitted/founder variants per animal.Because there were only 2 infected animals in the gag-env vaccinated rhesus compared to 10 infected rhesus in the other 2 groups, the significance of finding single env variants in the gag-env vaccinated group could not be established.

View Article: PubMed Central - PubMed

Affiliation: Division of Microbiology, Tulane National Primate Research Center, Tulane University, Covington, Louisiana, United States of America.

ABSTRACT
Identification of transmitted/founder simian immunodeficiency virus (SIV) envelope sequences responsible for infection may prove critical for understanding HIV/SIV mucosal transmission. We used single genome amplification and phylogenetic analyses to characterize transmitted/founder SIVs both in the inoculum and in immunized-infected rhesus monkeys. Single genome amplification of the SIVsmE660 inoculum revealed a maximum diversity of 1.4%. We also noted that the consensus sequence of the challenge stock differed from the vaccine construct in 10 amino acids including 3 changes in the V4 loop. Viral env was prepared from rhesus plasma in 3 groups of 6 immunized with vesicular stomatitis virus (VSV) vectors and boosted with Semliki forest virus (SFV) replicons expressing (a) SIVsmE660 gag-env (b) SIVsmE660 gag-env plus rhesus GM-CSF and (c) control influenza hemagglutinin protein. Macaques were immunized twice with VSV-vectors and once with SFV vector and challenged intrarectally with 4000 TCID50. Single genome amplification characterized the infections of 2 unprotected animals in the gag-env immunized group, both of which had reduced acute plasma viral loads that ended as transient infections indicating partial immune control. Four of 6 rhesus were infected in the gag-env + GM-CSF group which demonstrated that GM-CSF abrogated protection. All 6 animals from the control group were infected having high plasma viral loads. We obtained 246 full-length envelope sequences from SIVsmE660 infected macaques at the peak of infection and determined the number of transmitted/founder variants per animal. Our analysis found that 2 of 2 gag-env vaccinated but infected macaques exhibited single but distinct virus envelope lineages whereas rhesus vaccinated with gag-env-GM-CSF or HA control exhibited both single and multiple env lineages. Because there were only 2 infected animals in the gag-env vaccinated rhesus compared to 10 infected rhesus in the other 2 groups, the significance of finding single env variants in the gag-env vaccinated group could not be established.

Show MeSH
Related in: MedlinePlus