Plasma viral loads. Viral RNA copies per ml of plasma are shown for macaques inoculated orally with low doses (A) or high doses (B) of SIV.

Phylogenetic tree. Neighbor-joining phylogenetic trees were constructed from the sequences of clones obtained after PCR amplification of the SIV env V1-V2 region from day 7 or 14 PBMCs of low dose inoculated macaques (earliest SIV-positive sample). Bootstrap values from 500 replicates are indicated on each node if greater than 75%. Dots of the same shape and shade represent clones from the same animal. SIV Env V1-V2 sequences are depicted from all six low dose macaques as well as the SIV inoculum (A). Sequences from macaques 18984 and 18412 with the SIV inoculum are shown separately to demonstrate the higher diversity of transmitted virions in these two macaques (B and C respectively).

Heteroduplex Mobility Assay (HMA). Viral diversity in macaques administered low or high doses of SIV. Genomic DNA was obtained from PBMCs from the first SIV-positive sample of all orally inoculated macaques (day 14 for low dose macaques, and day 7 or 14 for high dose macaques) and the V1-V2 region of SIVenv was PCR amplified. Lanes 1 to 6 show the six macaques that were inoculated with low doses of SIVmac251 (lane 1: 17742, 2: 18984, 3: 18412, 4: 18414, 5: 18981, 6: 19147). Lanes 7 to 11 show the five macaques inoculated with high doses of SIVmac251 (lane 7: RM11, 8: RM12, 9: RM13, 10: RM14, 11: RM15). Lane 12 shows HMA for V1-V2 region of a SIVmac239 plasmid, to indicate the pattern when only one clonal sequence is present. HD indicates the homoduplex bands which is the brightest and fastest migrating band on the gel.

Fold changes in mRNA expression of immune modulating genes in LN and PBMCs. Fold changes are shown for mRNA expression of six immune response genes (from left: OAS, CXCL9, CXCL10, IFNγ, TNFα, IL-10) in PBMCs (A, B) as well as eight immune response genes (from left: IFN-α, OAS, CXCL9, CXCL10, IFN-γ, IL-12, TNF-α, IL-10) in LN (C, D) of macaques infected orally with low doses (A, C) or high doses (B, D) of SIV. Symbols of the same shape represent the same macaque and each symbol represents a different time point of sampling. Symbols within the black boxes represent mRNA expression levels that are increased more than two standard deviations away from the baseline gene expression. For PBMC, gene expression in uninfected macaques was determined by average gene expression levels of preinfection timepoint from both high and low dose macaques. For lymph nodes, the average of gene expression levels of preinfection time point from low dose macaques and 4 uninfected macaques from California primate center was used to determine baseline gene expression level at peripheral lymph nodes. For low dosed macaques, PBMCs were obtained at four time points between days 7 and 14, 28 and 35, 70 and 85, and at day 112; LN biopsies were obtained at three time points for the low dose macaques between days 28 and 35, day 70 and 85 and also at day 112. Immune gene expression in LN biopsies and PBMCs shown here for high dose macaques include PBMCs obtained at days 7 or 15, 21 or 28, 45 or 56 as well as 85 dpi and lymph node biopsies taken at three time points on days 21 or 28, 45 or 56 as well as 85 dpi as previous described [28].

OAS and CXCL10 mRNA expression changes in peripheral blood. OAS (A, B, C) and CXCL10 (D, E, F) expression from acute to chronic SIV infection time points are shown for PBMCs of Rhesus macaques orally inoculated with low (A, D) or high doses (B, E) of SIVmac251. Also, OAS (C) and CXCL10 (F) expression in PBMCs are shown for macaques 7 days after i.v. infection with high doses of SIVmac239. Each bar represents the fold change of expression at one time point compared to uninfected macaques. The shaded area across the x-axis represents two standard deviations of the average expression of the corresponding gene in uninfected macaques. ND – not determined.

OAS (A, B), CXCL9 (C, D) and CXCL10 (E, F) expression during chronic SIV infection at lymph node are shown from Rhesus macaques orally inoculated with low (A, C, E) or high doses (B, D, F) of SIVmac251. Each bar represents the fold change of gene expression at one time point compared to uninfected macaques. The shaded area across the x-axis represents two standard deviations of the average expression of the corresponding gene in uninfected macaques.

Fold changes of expression of OAS (A, B, C) and CXCL10 (D, E, F) are shown in isolated CD3+ T cells (A, D), CD14+ monocytes (B, E) and cells that do not have either of these markers (CD3−CD14−) (C, F). PBMCs were taken from macaque RM14 (84 dpi, left bar) and RM11 (14 dpi, right bar). Expression of OAS or CXCL10 in each cell subset is compared to gene expression in the same cell subset of uninfected Rhesus macaques. The fold change is considered increased or decreased in SIV infected compared to uninfected macaques when it is greater than two standard deviations (shaded area) away from the average expression in uninfected animals.