Establishment and stability of the latent HIV-1 DNA reservoir

  1. Johanna Brodin
  2. Fabio Zanini
  3. Lina Thebo
  4. Christa Lanz
  5. Göran Bratt
  6. Richard A Neher
  7. Jan Albert  Is a corresponding author
  1. Karolinska Institute, Sweden
  2. Max Planck Institute for Developmental Biology, Germany
  3. Stockholm South General Hospital, Sweden
  4. Karolinska University Hospital, Sweden
4 figures, 2 tables and 2 additional files

Figures

Sampling times before and after start of suppressive antiretroviral therapy.

For each study participant, the thick grey bar indicates the period of untreated HIV-1 replication. Circles indicate the collection times of the plasma samples used for whole genome deep sequencing …

https://doi.org/10.7554/eLife.18889.002
Figure 2 with 3 supplements
Reconstructed phylogenetic trees of plasma HIV-1 RNA sequences (circles) and PBMC HIV-1 DNA (triangles and squares) from two patients.

The RNA sequences were obtained from plasma samples collected before the start of suppressive antiretroviral therapy (ART). DNA sequences 1, 2, and 3 were obtained from PBMCs collected after many …

https://doi.org/10.7554/eLife.18889.004
Figure 2—figure supplement 1
Phylogenetic trees of plasma HIV-1 RNA and PBMC HIV-1 DNA sequences from all patients included in the study.
https://doi.org/10.7554/eLife.18889.005
Figure 2—figure supplement 2
Distributions of frequencies of haplotypes.

We found a wide variation in haplotype frequencies (measured by fraction of reads), from <0.001 to >0.5. Most of the haplotypes were present at frequencies of approximately 0.01 (note the …

https://doi.org/10.7554/eLife.18889.006
Figure 2—figure supplement 3
Distributions of mutations in reads classified as hypermutanted or as non-hypermutated.
https://doi.org/10.7554/eLife.18889.007
Figure 3 with 3 supplements
Root-to-tip distances.

The plasma HIV-1 RNA sequences evolved steadily before the start of antiretroviral therapy (ART), while no evidence of evolution was found in the PBMC HIV-1 DNA sequences obtained after the start of …

https://doi.org/10.7554/eLife.18889.008
Figure 3—figure supplement 1
Mean root-to-tip distances for plasma HIV-1 RNA sequences obtained before the start of ART and PBMC HIV-1 DNA sequences obtained after the start of ART.

This figure is analogous to Figure 3 in the main text, but presents root-to-tip distance values for DNA sequences classified as hypermutants. The root-to-tip distances of the hypermutant HIV-1 DNA …

https://doi.org/10.7554/eLife.18889.009
Figure 3—figure supplement 2
Mean root-to-tip distances for plasma HIV-1 RNA sequences obtained before the start of antiretroviral therapy (ART) and PBMC HIV-1 DNA sequences obtained after the start of ART.

This figure is analogous to Figure 3 in the main text. Instead of weighing each sequence by its number of reads, each sequence was counted only once. The two approaches yielded similar results. The …

https://doi.org/10.7554/eLife.18889.010
Figure 3—figure supplement 3
Short lived cells can generate a false signal of evolution.

HIV-1 DNA from cell samples taken at the initiation of antiretroviral therapy (ART) will be dominated by virus variants that have very recently infected short-lived cells (blue). The short-lived …

https://doi.org/10.7554/eLife.18889.011
Figure 4 with 2 supplements
Probable origin of sequences in the DNA reservoir.

For each HIV-1 DNA read obtained from the PBMCs, we determined the pre-treatment plasma sample and HIV-1 RNA variant that was the most likely origin of the read. Panel A presents the results for the …

https://doi.org/10.7554/eLife.18889.013
Figure 4—figure supplement 1
The distribution of plausible seeding times of reservoir HIV-1 DNA sequences for all 10 patients.

For each HIV-1 DNA read obtained from the PBMCs, the plasma sample and HIV-1 RNA variant from which the read was most likely derived was determined. The plots present the distributions of these most …

https://doi.org/10.7554/eLife.18889.014
Figure 4—figure supplement 2
Same as Figure 4—figure supplement 1, but counting each sequence once instead of weighted by the number of reads it represents.
https://doi.org/10.7554/eLife.18889.015

Tables

Table 1

Summary of patient characteristics.

https://doi.org/10.7554/eLife.18889.003
PatientGenderTransmissionSubtypeAge*

HIV RNA from plasma

HIV DNA from PBMCs

# samples

First/last since EDI

Time on ART# templates

p1

F

HET

01_AE

37

12

0.3

8.2

7.9/9.9/10.4

820/148/38

p2

M

MSM

B

32

6

0.2

5.5

6.9

75

p3

M

MSM

B

52

10

0.4

8.4

4.6/6.7/7.2

243/102/108

p5

M

MSM

B

38

7

0.4

5.9

4.0/6.3

180/72

p6

M

HET

C

31

7

0.2

7.0

3.0/5.0/5.5

115/15/nd

p7

M

MSM

B

31

11

6.3

16.1

6.3/8.4/8.8

88/279/108

p8

M

MSM

B

35

7

0.2

6.0

8.4/10.6/10.9

180/55/175

p9

M

MSM

B

32

8

0.3

8.1

7.7/9.7/10.2

60/72/72

p10

M

MSM

B

34

9

0.1

6.2

16.2/18.3/18.6

249/116/51

p11

M

MSM

B

53

7

0.6

5.6

6.4/8.4/8.8

124/120/123

  1. *at diagnosis;

  2. EDI: estimated date of infection; all times are given in years;

  3. sequencing failed in earlier samples due to low plasma HIV-1 RNA levels.

Table 2

Rates of evolution in plasma HIV-1 RNA and PBMC HIV-1 DNA sequences obtained before the start and after the start of suppressive antiretroviral therapy, respectively.

https://doi.org/10.7554/eLife.18889.012
Patient

RNA rate

DNA rate

[Year−1]p-value[Year−1]p-value
p1

4.4 × 10−3

<10−6

−6 × 10−4

0.22

p2

3.7 × 10−3

<10−2

−8 × 10−4

p3

4.1 × 10−3

<10−6

−2 × 10−4

0.39

p5

4.8 × 10−3

<10−3

4 × 10−4

0.45

p6

1.4 × 10−3

<10−3

−9 × 10−4

0.22

p7

1.3 × 10−3

<10−2

−7 × 10−4

0.14

p8

2.9 × 10−3

<10−5

8 × 10−5

0.22

p9

2.6 × 10−3

<10−4

1 × 10−4

0.12

p10

3.6 × 10−3

<10−5

−1 × 10−4

0.20

p11

1.2 × 10−3

<10−2

2 × 10−4

0.16

Additional files

Supplementary file 1

Sequencing and hypermutation statistic for all samples.

'good' refers to proviral sequences that are not obviously defective, 'hyper' refers to those with an excess of G→A mutations. The column '% recaptured' indicates the fraction of sequences >1% frequency that were present in other samples from the same patient.

https://doi.org/10.7554/eLife.18889.016
Supplementary file 2

PCR primers used for HIV-1 p17gag amplification.

https://doi.org/10.7554/eLife.18889.017

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