DOI: 10.5061/dryad.1c59zw42s
title: Dating the bacterial tree of life based on ancient symbiosis

description: Obtaining a timescale for bacterial evolution is crucial to understanding
 early life evolution but is difficult owing to the scarcity of bacterial
 fossils and the absence of maximum age constraints of the available
 fossils. Here, we introduce multiple new time constraints to calibrate
 bacterial evolution based on ancient symbiosis. This idea is implemented
 using a bacterial tree constructed with mitochondria-originated genes
 where the mitochondrial lineage representing eukaryotes is embedded within
 Proteobacteria, such that the date constraints of eukaryotes established
 by their abundant fossils are propagated to ancient co-evolving bacterial
 symbionts and across the bacterial tree of life. Importantly, we formulate
 a new probabilistic framework that considers uncertainty in inference of
 the ancestral lifestyle of modern symbionts to apply 19 relative time
 constraints (RTC) each informed by host-symbiont association to constrain
 bacterial symbionts no older than their eukaryotic host. Moreover, we
 develop an approach to incorporating substitution mixture models that
 better accommodate substitutional saturation and compositional
 heterogeneity for dating deep phylogenies. Our analysis estimates that the
 last bacterial common ancestor (LBCA) occurred approximately 4.0-3.5
 billion years ago (Ga), followed by rapid divergence of major bacterial
 clades. It is generally robust to alternative root ages, root positions,
 tree topologies, fossil ages, ancestral lifestyle reconstruction, and gene
 sets, among other factors. The obtained time tree serves as a foundation
 for testing hypotheses regarding bacterial diversification and its
 correlation with geobiological events across different timescales.
description: # Dating the bacterial tree of life based on ancient symbiosis
 [https://doi.org/10.5061/dryad.1c59zw42s](https://doi.org/10.5061/dryad.1c59zw42s) ``` More detailed introductions for each folder and data set are given in a_more_detailed_readme.zip. ``` ## Description of the data and file structure Original data and results for the study "Dating the bacterial tree of life based on ancient symbiosis". Organized into the following archives: Note that **Supplemental data** are available at [https://zenodo.org/records/14348151](https://zenodo.org/records/14348151) (see **Related works** on the right-hand side) #### **DATING** *Relevant figures and tables:* Figs. 2-3, Figs. S15-S19. Included in this folder are the **results of the MCMCtree analysis**. * bootstrap/: using the focal strategy (some alternatives included) * bootstrap.root2/: with different root placements * date-alt/: analyses under various alternative dating schemes to explore the uncertainty associated with divergence time estimation * date-alt.dup/: run2 of the MCMC analysis included in date-alt/ * date-fg/: the full gene set instead of 19 “high-quality” genes * deltaLL/: using 5, 10, and 15 (best) top-ranking genes out of the 19 mitochondrial genes according to relative rate difference between mitochondrial and bacterial lineages or △LL which measures the the degree of species- and gene-tree incongruence (see Data S3) The structures of the MCMCtree results are similar. This is detailed in the “Readme” file in the directory “bootstrap/”. The result of the **focal analysis can be found in “bootstrap\1-pf\dating\C60\mcmctree-rate\rrtc\official-mcmc987\marginal”.** #### **PTL** *Relevant figures and tables:* Figs. S2, S10, S11; Note S3. **ASR of the hosts/lifestyles** of seven selected bacterial symbiont lineages. * full/: main analysis #### **Coleman60** *Relevant figures and tables:* Fig. S6, Table S2. This folder is about **IQ-Tree phylogenomic tree construction**. All 265 bacterial genomes used in the prior study Coleman et al., 2021 are included. Individual alignments of the 60 genes used in the study Coleman et al., 2021 can be found in **./pep**/. See Table S2 for the details of each scheme. Files used and generated in IQ-Tree analysis are given in **tree-CxxPMSF-RML/** where Cxx represents C20, C40, or C60. * combined.aln: concatenated alignment (after trimming) * aln/: original sequence alignments * trimal/: trimmed sequence alignments * phylo/iqtree/: IQ-Tree analysis. Specifically, the command can be found in the ".log" file, and the ml tree is indicated in the "iqtree.treefile" file. For NONREV model analysis, you may see the file "REV.treefile" which is the guide tree constructed using a reversible model. #### **hessian\_in\_BV** *Relevant figures and tables:* Fig. 1B, Figs. S12-S13, Note S4. The simulation analysis of the **bootstrap-based Hessian approximation** in MCMCtree analysis using bs_inBV ([https://github.com/evolbeginner/bs_inBV](https://github.com/evolbeginner/bs_inBV)) or CODEML (see the GitHub online repository for more details). Briefly, a [timetree]{.underline} is simulated, and then a [phylogeny]{.underline} where branch lengths correspond to the number of expected substitutions/sites is simulated under a relaxed clock model. Last, [sequence alignment]{.underline} is simulated under a given model. For more details, please see **Note S4** in the original paper. The three folders included correspond to different models used in [sequence evolution]{.underline} simulation. i) LG+G{1.0}: LG substitution model and across-site rate variation under a four-category discrete gamma distribution with the shape parameter α=1.0, thus Gamma(1.0,1.0), meaning that the mean and variance of the relative rates across site are 1/1 = 1 and 1/1^2=1 respectively (Yang 1994). ii) LG+G{1.0}+C40: LG substitution model plus a mixture of amino acid site frequency profiles C40 and across-site rate variation under a four-category discrete gamma distribution with α=1.0. iii) LG+G{0.5}+C40: LG substitution model plus a mixture of amino acid site frequency profiles C40 and across-site rate variation under a four-category discrete gamma distribution with α=0.5, which reflects more among-site rate variation than using α=1.0. For each of the above models, you will see 30 independent simulations/comparisons based on which many further analyses are performed. #### **mcmc3r** *Relevant figures and tables*: Fig. S5. Data for clock model selection using **mcmc3r**. MCMC3R.tar.xz: used for eukaryote divergence time estimation as the 1st step of the Bayesian sequential molecular clock analysis. bac_20genome/: used for the bacterial timetree analysis (2nd step of the Bayesian sequence molecular clock analysis), with 20 genomes. bac_40genome/: used for the bacterial timetree analysis (2nd step of the Bayesian sequence molecular clock analysis), with 40 genomes. In the folder of each gene, there are four folders: * date/: MCMCtree analysis results * nucl/: recoded nucleotide sequences * pep/: original amino acid sequences * tree.sp:/ original and trimmed alignment ## Sharing/Access information The sources of genomes and their accessions are well indicated in Table S1. ## Code/Software We have made the following two scripts freely available on GitHub. bs_inBV: A tool to help generate the file in.BV when using "complex" substitution models for MCMCTree. [https://github.com/evolbeginner/bs_inBV](https://github.com/evolbeginner/bs_inBV) pRTC: A script to do probability-based relaxed time constraint **(pRTC)**-based dating. [https://github.com/evolbeginner/rrtc.](https://github.com/evolbeginner/rrtc.) ## Change log **10-Dec-2024** - A description of the state information contained in the file traits.txt for PTL/ is added to the file "PTL_readme.docx" in a_more_detailed_readme.zip. The Zeonodo link to all **Supplemental Data** is added in the Related Works section.