Standardized phylogenetic and molecular evolutionary analysis applied to species across the microbial tree of life

Abstract

chien-chi Lo & patrick S. G. chain * there is growing interest in reconstructing phylogenies from the copious amounts of genome sequencing projects that target related viral, bacterial or eukaryotic organisms. to facilitate the construction of standardized and robust phylogenies for disparate types of projects, we have developed a complete bioinformatic workflow, with a web-based component to perform phylogenetic and molecular evolutionary (phaMe) analysis from sequencing reads, draft assemblies or completed genomes of closely related organisms. furthermore, the ability to incorporate raw data, including some metagenomic samples containing a target organism (e.g. from clinical samples with suspected infectious agents), shows promise for the rapid phylogenetic characterization of organisms within complex samples without the need for prior assembly. The reconstruction of organismal evolutionary history using phylogenetics is a fundamental method applied to many areas of biology. Single nucleotide polymorphisms (SNPs), one of the dominant forms of evolutionary change, have become an indispensable tool for phylogenetic analyses 1-4. Phylogenies in the pre-genomic era relied on SNPs and conserved sites within a single locus, and was later extended to multiple loci, such as in multiple locus sequence typing (MLST). Although still valuable, these methods only consider evolutionary signals originating within a small fraction of the genome, are unable to capture the complete variation within species, and generally provide a weak phylogenetic signal, particularly within a species, and do not always reflect the true evolutionary history of species 5. While phylogenetic analyses that use many conserved genes (orthologs) are a great improvement, these methods require annotated coding regions, whose predictions are not always accurate or available 6. Furthermore, they are impacted by horizontal gene transfer (HGT) 7 , recombination 8 , rate heterogeneity 9 , and incomplete lineage sorting. Genome-wide SNPs are one of the best measures of phylogenetic diversity as they can discriminate among closely related organisms and help resolve both short and long branches in a tree 10,11. Since selectively neutral SNPs accumulate at a uniform rate, they can be used to measure divergence between species as well as strains 12,13. Furthermore, due to the large number of SNPs found along the length of entire genomes, the use of whole-genome SNPs minimizes the impact of random sequencing and assembly errors that can impact individual loci, as well as biases due to individual genes under strong selective pressure. Some inherent biases remain with whole genome SNP approaches that are similar to loci-based phylogenies such as HGT, recombination, and rate heterogeneity. Although genome-wide sequencing now allows examination of the full complement of genomic variation, the number of completed and finished genomes are increasingly falling behind the generation of new draft genomes, due to the lack of computational or other resources. For example, o...

Mycoplasma bovis in Nordic European Countries: Emergence and Dominance of a New Clone

“…All branches and sub-branches contained a mix of strains isolated from different sites in the body, with no evidence to suggest anatomical specificity of strains. As an alternative to cgMLST, which took into consideration the genes conserved in all genomes (n = 500 here), a genome-wide SNP phylogeny [29] was also generated on our set of genomes using the methodology already validated for M. bovis [11]. The resulting phylogenetic tree corresponded well to that of the cgMLST ( Figure S1), showing the same phylogenetic signals, albeit with a lower discriminatory power.…”

“…However, the dnaA allele alone was poorly discriminatory for the isolates included in our study. Because it considers evolutionary signals originating from the whole genome, either the whole set of CDS (cgMLST) or genome-wide polymorphisms, the WGS-typing approaches are a more appropriate way forward [29]. However, they remain relatively expensive, time-consuming, and require expertise for interpretation.…”

Genomic Analysis of Diverse Members of the Fungal Genus Monosporascus Reveals Novel Lineages, Unique Genome Content and a Potential Bacterial Associate

“…Three other assemblies contained a small (≤ 5) number of Ralstonia contigs, while the remaining four assemblies contained zero Ralstonia contigs. Phylogenetic analyses generated with PhaME ( Shakya et al 2020 ) using both raw Illumina reads and assembled contigs confirmed these bacterial sequences belong to distinct R. pickettii lineages, which is evidence against local contamination ( Figure 5 ).…”

“…Subsequent to the acquisition of genome sequence data, evolutionary relationships among Monosporascus isolates were evaluated using whole-genome phylogenetic analyses performed with PhaME ( Shakya et al 2020 ). PhaME analyses were conducted with the reads used in the assembly of the ten Monosporascus genomes, employing the smallest (GIB2) assembly as the reference and 100 bootstrap replicates.…”

Subtelomeric regions and a repeat-rich chromosome harbor multicopy effector gene clusters with variable conservation in multiple plant pathogenicColletotrichumspecies

“…For each GCASP orthogroup, the best scoring tree of 500 parsimonious and 427 500 random trees was used with support values from 1,000 bootstrap replicates. For maximum 428 likelihood phylogeny of all Colletotrichum members, whole genome SNPs were identified from 429 nucmer whole genome alignments by PhaME (Shakya et al 2020). Maximum likelihood 430 phylogenetic analysis was carried out using RAxML-ng on the 199,953 SNP positions identified with 431 the GTR+G4 model of substitution, which was determined as the best model according to 432 modeltest-ng (Darriba et al 2020).…”