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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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Phylogeny and Poisson-Fitter analysis of DG21 macaque env sequences.We analyzed 19 full-length envelope sequences from SIVsmE660 infected DG21 macaque. (A) phylogenetic comparison of DG21 env sequences to the generated consensus DG21 envelope sequence. (B) Highlighter plot exhibiting the nucleotide differences in DG21 env sequences compared to DG21 consensus sequence. (C) Poisson-Fitter plots of DG21 env sequences including APOBEC mutations. (D) Poisson-Fitter plots of DG21 env sequences excluding the APOBEC mutations.
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pone-0109678-g005: Phylogeny and Poisson-Fitter analysis of DG21 macaque env sequences.We analyzed 19 full-length envelope sequences from SIVsmE660 infected DG21 macaque. (A) phylogenetic comparison of DG21 env sequences to the generated consensus DG21 envelope sequence. (B) Highlighter plot exhibiting the nucleotide differences in DG21 env sequences compared to DG21 consensus sequence. (C) Poisson-Fitter plots of DG21 env sequences including APOBEC mutations. (D) Poisson-Fitter plots of DG21 env sequences excluding the APOBEC mutations.

Mentions: In gag-env vaccinated animals, four out of six macaques exhibited apparent sterilizing immunity to the challenge, while the other two had transient viremia. SGA was performed on the day 14 (peak viremia) plasma samples from both macaques. We obtained in total 21 full-length envelope sequences from DF38 and the sequences were used to construct a phylogenetic tree. A consensus envelope sequence was generated and individual envelope nucleotide sequences were compared to the consensus. Highlighter plots were made to enumerate the number of T/F variants and identify the putative env sequence that initiated infection. (Fig. 4A). Nine out of twenty one (43%) sequences showed a significant number of APOBEC mutations (red star in Fig. 4A). These mutations were created by host restriction of the retrovirus and often obscure enumeration of T/F viruses from infected animals (Fig. 4B) [36]. After, accounting for these mutated sequences, a single low diversity variant was observed in DF38 representing infection with a single T/F variant. Of the remaining 12 sequences, 7 were identical to the consensus sequence and to themselves and represent the T/F sequence. Sequences that differed from consensus sequence contained only one or at most two mutations randomly distributed in the genome. Further, to confirm the infection with a single variant, we performed Poisson-Fitter analysis of SGA envelope sequences [37], which determines if frequency of mutations accumulate are consistent with Poisson distribution. This analysis distinguishes between the presence of one or more founder viruses from infected animals. The DF38 macaque putative founder virus fit the Poisson-Fitter analysis (after eliminating APOBEC mutations) confirming that this macaque was infected with only one founder virus (Fig. 4C and D). In case of DG21, the plasma viral load was low (<40,000 copies/ml) and hence we obtained viral RNA after pelleting the plasma sample as described in the methods. In total, 19 full-length envelope sequences were obtained and were analyzed phylogenetically (Fig. 5A) of which 4 sequences showed APOBEC mutations on the highlighter plot (Fig. 5B). Similar to DF38, after subjecting the sequences for Poisson-Fitter analysis this macaque was also infected with only one founder virus (Fig. 5C and D).


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)

Phylogeny and Poisson-Fitter analysis of DG21 macaque env sequences.We analyzed 19 full-length envelope sequences from SIVsmE660 infected DG21 macaque. (A) phylogenetic comparison of DG21 env sequences to the generated consensus DG21 envelope sequence. (B) Highlighter plot exhibiting the nucleotide differences in DG21 env sequences compared to DG21 consensus sequence. (C) Poisson-Fitter plots of DG21 env sequences including APOBEC mutations. (D) Poisson-Fitter plots of DG21 env sequences excluding the APOBEC mutations.
© Copyright Policy
Related In: Results  -  Collection

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

pone-0109678-g005: Phylogeny and Poisson-Fitter analysis of DG21 macaque env sequences.We analyzed 19 full-length envelope sequences from SIVsmE660 infected DG21 macaque. (A) phylogenetic comparison of DG21 env sequences to the generated consensus DG21 envelope sequence. (B) Highlighter plot exhibiting the nucleotide differences in DG21 env sequences compared to DG21 consensus sequence. (C) Poisson-Fitter plots of DG21 env sequences including APOBEC mutations. (D) Poisson-Fitter plots of DG21 env sequences excluding the APOBEC mutations.
Mentions: In gag-env vaccinated animals, four out of six macaques exhibited apparent sterilizing immunity to the challenge, while the other two had transient viremia. SGA was performed on the day 14 (peak viremia) plasma samples from both macaques. We obtained in total 21 full-length envelope sequences from DF38 and the sequences were used to construct a phylogenetic tree. A consensus envelope sequence was generated and individual envelope nucleotide sequences were compared to the consensus. Highlighter plots were made to enumerate the number of T/F variants and identify the putative env sequence that initiated infection. (Fig. 4A). Nine out of twenty one (43%) sequences showed a significant number of APOBEC mutations (red star in Fig. 4A). These mutations were created by host restriction of the retrovirus and often obscure enumeration of T/F viruses from infected animals (Fig. 4B) [36]. After, accounting for these mutated sequences, a single low diversity variant was observed in DF38 representing infection with a single T/F variant. Of the remaining 12 sequences, 7 were identical to the consensus sequence and to themselves and represent the T/F sequence. Sequences that differed from consensus sequence contained only one or at most two mutations randomly distributed in the genome. Further, to confirm the infection with a single variant, we performed Poisson-Fitter analysis of SGA envelope sequences [37], which determines if frequency of mutations accumulate are consistent with Poisson distribution. This analysis distinguishes between the presence of one or more founder viruses from infected animals. The DF38 macaque putative founder virus fit the Poisson-Fitter analysis (after eliminating APOBEC mutations) confirming that this macaque was infected with only one founder virus (Fig. 4C and D). In case of DG21, the plasma viral load was low (<40,000 copies/ml) and hence we obtained viral RNA after pelleting the plasma sample as described in the methods. In total, 19 full-length envelope sequences were obtained and were analyzed phylogenetically (Fig. 5A) of which 4 sequences showed APOBEC mutations on the highlighter plot (Fig. 5B). Similar to DF38, after subjecting the sequences for Poisson-Fitter analysis this macaque was also infected with only one founder virus (Fig. 5C and D).

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