Fig. 1. Typical ensemble workflow for alignment and phylogeny assessment.

An ensemble of MSAs is generated and assessed for accuracy using Muscle5. Gray rectangles are processing steps made by an algorithm or software package. First, Muscle5 (step 1) generates an ensemble of MSAs (step 2), each alignment is generated by a different combination of a perturbed HMM and permuted guide tree. The accuracy of the MSAs can be assessed by Muscle5 (step 3) using accuracy metrics such as Column Confidence (CC). A phylogeny algorithm (step 4), e.g. maximum likelihood (ML), is used to predict a tree from each MSA (step 5). Finally, accuracy metrics, e.g. Ensemble Confidence (EC), are calculated from the resulting ensemble of trees (step 6). The Newick package (https://github.com/rcedgar/newick) was used to calculate the novel metrics described in this paper.