Colpodean ciliate phylogeny and reference alignments for phylogenetic placements.

Area of Science:

• Microbiology
• Evolutionary Biology
• Bioinformatics

Background:

• Colpodea are a major ciliate clade frequently detected in environmental DNA (eDNA) sequencing.
• They share somatic ciliature but possess diverse oral structures, complicating classification.
• Existing molecular phylogenies show disagreement on the branching order of four well-supported subclades.

Purpose of the Study:

• To resolve the phylogenetic position of bursariomorphids and platyophryids within the Colpodea.
• To evaluate the branching order of early diverging colpodean lineages using nuclear SSU-rRNA sequences.
• To provide reference alignments for phylogenetic placement of eDNA data.

Main Methods:

• Phylogenetic inference using nuclear small subunit ribosomal RNA (SSU-rRNA) gene sequences.
• Application of various sequence alignment and masking strategies.
• Phylogenetic network analyses to assess relationships with outgroups.
• Constrained phylogenetic analyses to test specific branching orders.

Main Results:

• Inferred "platyophryids-early" topologies using multiple methods.
• Constrained analyses could not reject the "bursariomorphids-early" topology.
• Both bursariomorphid and platyophryid clades exhibit similar nucleotide positions shared with the outgroup, with interconnected phylogenetic networks.
• Mitochondrial SSU-rRNA data also support the "platyophryids-early" topology, but discordant results complicate definitive placement.

Conclusions:

• The precise branching order of early diverging Colpodea lineages remains ambiguous.
• Both "platyophryids-early" and "bursariomorphids-early" topologies are plausible based on current nuclear SSU-rRNA data.
• Reference alignments are provided to aid phylogenetic placement of eDNA sequences, facilitating evolutionary and ecological interpretations.