Computational protocol: Dissecting HIV Virulence: Heritability of Setpoint Viral Load, CD4+ T-Cell Decline, and Per-Parasite Pathogenicity

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Protocol publication

[…] The transmission pairs were identified as monophyletic clusters on a previous HIV transmission tree (). Of these previously established transmission pairs, 196 were present in our study population. The direction of transmission cannot be inferred in these pairs.The reconstruction of the phylogenetic tree relies on pol gene sequencing of the virus carried by the study subjects. In particular, we had sequences of pol extending over the HXB2 positions 2253–3870, comprising the protease and the reverse transcriptase. All sequences were initially aligned to an HXB2 reference genome (; last accessed September 24, 2017) using MUSCLE (). We selected the earliest sequence if more than one sequence was available for a person.To reconstruct the evolutionary history, we first removed insertions relatively to HXB2. To exclude signatures of parallel evolution due to drug pressure that can distort the inferred evolutionary history, we further removed drug resistance mutations according to the databases of Stanford (; last accessed September 24, 2017) and the International Antiviral Society (; last accessed September 24, 2017). We used Gblocks to refine the alignment. The final number of positions was 1106.We constructed the phylogenetic tree using FastTree (Version 2.1.8 SSE3, OpenMP; ). We used a maximum-likelihood-based inference using a Generalized Time-Reversible evolutionary model and a CAT model () with 20 discrete evolutionary rate categories. We use the most rigorous and time-consuming FastTree parameters (FastTreeMP -pseudo -spr 4 -mlacc 2 -slownni -gtr -nt). We rooted the tree with 10 Subtype C sequences as an outgroup, using the R package APE. The branch lengths in our tree correspond to genetic distances between the sequences, and not to time.We also compared the results obtained from this tree with those of two trees reconstructed with RaxML (). These trees were reconstructed assuming a Generalized Time-Reversible evolutionary model. One assumed CAT model with 25 discrete evolutionary rate categories, the other Γ-distributed evolutionary rates. , online, shows that there is generally good agreement between the results. […]

Pipeline specifications

Software tools MUSCLE, Gblocks, FastTree, RAxML
Applications Phylogenetics, Nucleotide sequence alignment
Organisms Human immunodeficiency virus 2, Homo sapiens
Diseases HIV Infections