Molecular variability in the Celleporella hyalina (Bryozoa; Cheilostomata) species complex: evidence for cryptic speciation from complete mitochondrial genomes.

The bryozoan Celleporella has been shown to be composed of multiple, often cryptic, lineages. We sequenced two complete mitochondrial (mt) genomes of the Celleporella hyalina species complex from Wales, UK and Norway (i) to determine genetic divergence at the complete mt genome level, and (ii) to design new molecular markers for examining the interrelationships amongst the major lineages. In addressing (i), we estimated genetic divergence at three levels: (a) nucleotide diversity (π), (b) genome size, and (c) gene order. Genes nad4L, nad6, and atp8 showed the highest levels of divergence, and rrnL, rrnS, and cox1 showed the lowest levels. Inter-genome nucleotide divergence of protein-coding and ribosomal RNA genes, measured as π, was 0.21. The two genomes differed substantially in size, with the Norwegian genome being 2,573 base pairs (bp) longer than the Welsh genome, 17,265 and 14,692 bp, respectively. This difference in size is attributable to long non-coding regions present in the Norwegian genome. Both genomes exhibit similar gene orders, except for the translocation of one transfer RNA (trnA). Considering the high nucleotide diversity, genome size difference and change in gene order, these mt genomes are considered sufficiently divergent to have originated from two distinct species. In addressing (ii) we designed PCR primers that flank the most conserved regions of the genome: 1,300 bp of cox1 and a contiguous 2,000 bp fragment of rrnL + rrnS. The primers have yielded products for tissue from Wales, Norway, New Zealand, Alaska and Chile and should provide useful tools in establishing species- and population-level diversity within the Celleporella complex.