18S rRNA variability maps reveal three highly divergent, conserved motifs within Rotifera

Abstract

Background: 18S rRNA is a major component of the small subunit of the eukaryotic ribosome and an important phylogenetic marker for many groups, often to the point of being the only marker available for some. A core structure across eukaryotes exists for this molecule that can help to inform about its evolution in different groups. Using an alignment of 18S rDNA for Rotifera as traditionally recognized (=Bdelloidea, Monogononta, and Seisonacea, but not Acanthocephala), I fitted sequences for three exemplar species (Adineta vaga, Brachionus plicatilis, and Seison nebaliae, respectively) to the core structure and used these maps to reveal patterns of evolution for the remainder of this diverse group of microscopic animals.

Results: The obtained variability maps of the 18S rRNA molecule revealed a pattern of high diversity among the three major rotifer clades coupled with strong conservation within each of bdelloids and monogononts. A majority of individual sites (ca. 60%) were constant even across rotifers as a whole with variable sites showing only intermediate rates of evolution. Although the three structural maps each showed good agreement with the inferred core structure for eukaryotic 18S rRNA and so were highly similar to one another at the secondary and tertiary levels, the overall pattern is of three highly distinct, but conserved motifs within the group at the primary sequence level. A novel finding was that of a variably expressed deletion at the 3' end of the V3 hypervariable region among some bdelloid species that occasionally extended into and included the pseudoknot structure following this region as well as the central "square" of the 18S rRNA molecule. Compared to other groups, levels of variation and rates of evolution for 18S rRNA in Rotifera roughly matched those for Gastropoda and Acanthocephala, despite increasing evidence for the latter being a clade within Rotifera.

Conclusions: The lack of comparative data for comparable groups makes interpretation of the results (i.e., very low variation within each of the three major rotifer clades, but high variation between them) and their potential novelty difficult. However, these findings in combination with the high morphological diversity within rotifers potentially help to explain why no clear consensus has been reached to date with regard to the phylogenetic relationships among the major groups.

Keywords: Acanthocephala; Bdelloidea; Conservation; Deletion; Evolution; Hypervariable; Monogononta; Phylogeny; Rate of evolution; Seisonacea.

Fig. 1

The three exemplar rotifer 18S rRNA molecules: A Adineta vaga (Bdelloidea), b Brachionus plicatilis (Monogononta), and c Seison nebaliae (Seisonacea). Individual nucleotides are coloured according to their relative rate of evolution (for Bdelloidea, Monogononta, and all Rotifera, respectively) as determined using TIGER (see inline legends) and hypervariable regions V1–V9 are labeled and outlined in blue. In addition, the maximal extent of the variable deletion at the 3'-end of the V3 region in bdelloids (see Fig. 2) is highlighted using square parentheses. Each map was created initially using VARNA v3.93 [59] and modified using Adobe® Illustrator® 2020 to match the traditional topology of the eukaryotic core structure as closely as possible. Empty circles were added at the terminal ends of the sequences as needed to present the presumed full length of the molecule for each species

Fig. 1

The three exemplar rotifer 18S rRNA molecules: A Adineta vaga (Bdelloidea), b Brachionus plicatilis (Monogononta), and c Seison nebaliae (Seisonacea). Individual nucleotides are coloured according to their relative rate of evolution (for Bdelloidea, Monogononta, and all Rotifera, respectively) as determined using TIGER (see inline legends) and hypervariable regions V1–V9 are labeled and outlined in blue. In addition, the maximal extent of the variable deletion at the 3'-end of the V3 region in bdelloids (see Fig. 2) is highlighted using square parentheses. Each map was created initially using VARNA v3.93 [59] and modified using Adobe® Illustrator® 2020 to match the traditional topology of the eukaryotic core structure as closely as possible. Empty circles were added at the terminal ends of the sequences as needed to present the presumed full length of the molecule for each species

Fig. 1

The three exemplar rotifer 18S rRNA molecules: A Adineta vaga (Bdelloidea), b Brachionus plicatilis (Monogononta), and c Seison nebaliae (Seisonacea). Individual nucleotides are coloured according to their relative rate of evolution (for Bdelloidea, Monogononta, and all Rotifera, respectively) as determined using TIGER (see inline legends) and hypervariable regions V1–V9 are labeled and outlined in blue. In addition, the maximal extent of the variable deletion at the 3'-end of the V3 region in bdelloids (see Fig. 2) is highlighted using square parentheses. Each map was created initially using VARNA v3.93 [59] and modified using Adobe® Illustrator® 2020 to match the traditional topology of the eukaryotic core structure as closely as possible. Empty circles were added at the terminal ends of the sequences as needed to present the presumed full length of the molecule for each species

Fig. 2

Partial alignment of the 18S rDNA gene showing the variably expressed deletion present at and extending beyond the 3' end of the V3 hypervariable region in bdelloids and two species in the monogonont genus Lindia. The locations of the entire V3 region as well as the pseudoknot following it are indicated at the top of the alignment. The taxonomic groups are numbered as (1) Calicophoron calicophorum (Platyhelminthes, outgroup), (2) Bdelloidea, (3) Seisonacea, and (4) Monogononta (selected species)

Fig. 3

Histogram of relative TIGER rates of evolution across the three rotifer data sets (green, Bdelloidea; red, Monogononta; and blue, Rotifera). n/a stands for sites represented by less than 15% of the species in a given data set and so with insufficient coverage for the TIGER analyses

Fig. 4

Sizes and relative rates of evolution (as determined using TIGER) of each of the hypervariable and pooled non-hypervariable regions for the three rotifer data sets (green, Bdelloidea; red, Monogononta; and blue, Rotifera). Error bars represent standard errors and are subsumed by the data point when not visible

Fig. 5

Relative TIGER rates of evolution presented as a rolling average of the 35 positions centred on the focal position for each of the three rotifer data sets (green, Bdelloidea; red, Monogononta; and blue, Rotifera). The locations of the hypervariable regions are indicated by bars at the top of the graph

Fig. 6

Maximum-likelihood phylogeny of the 18S rDNA sequences examined in this study using RAxML v8.2.12 [60]. The analysis consisted of a fast bootstrap search followed by a thorough search for the maximum-likelihood topology [61] under a GTR + Γ model, with the gamma distribution being approximated initially through a CAT model [62]. The tree was rooted on Calicophoron calicophorum. Values above selected nodes represent bootstrap support [63] and the scale bar represents the average number of substitutions per site per unit time. Species names have been removed for clarity and the major clades are labeled as well as colour-coded (green, Bdelloidea; red, Monogononta; blue, Seisonacea; black, Acanthocephala)