First molecular evidence of hybridization in endosymbiotic ciliates (Protista, Ciliophora)

Abstract

Hybridization is an important evolutionary process that can fuel diversification via formation of hybrid species or can lead to fusion of previously separated lineages by forming highly diverse species complexes. We provide here the first molecular evidence of hybridization in wild populations of ciliates, a highly diverse group of free-living and symbiotic eukaryotic microbes. The impact of hybridization was studied on the model of Plagiotoma, an obligate endosymbiont of the digestive tube of earthworms, using split decomposition analyses and species networks, 2D modeling of the nuclear rRNA molecules and compensatory base change analyses as well as multidimensional morphometrics. Gene flow slowed down and eventually hampered the diversification of Lumbricus-dwelling plagiotomids, which collapsed into a single highly variable biological entity, the P. lumbrici complex. Disruption of the species boundaries was suggested also by the continuum of morphological variability in the phenotypic space. On the other hand, hybridization conspicuously increased diversity in the nuclear rDNA cistron and somewhat weakened the host structural specificity of the P. lumbrici complex, whose members colonize a variety of phylogenetically closely related anecic and epigeic earthworms. By contrast, another recorded species, P. aporrectodeae sp. n., showed no signs of introgression, no variability in the rDNA cistron, and very high host specificity. These contrasting eco-evolutionary patterns indicate that hybridization might decrease the alpha-diversity by dissolving species boundaries, weaken the structural host specificity by broadening ecological amplitudes, and increase the nuclear rDNA variability by overcoming concerted evolution within the P. lumbrici species complex.

2D approach to reconstruct the evolutionary history of clevelandellids (Ciliophora, Armophorea) inhabiting the hindgut of the Panesthiinae cockroaches

“…A common trait shared by clevelandellids isolated from the digestive tract of cockroaches is the absence of helix 9es3b in the V2 region. Interestingly, the lack of this helix was also reported for the spirotrich genus Plagiotoma (Obert et al, 2022). On the other hand, helix 9es3b was recognized in Haptophrya, Hysterocineta Diesing, 1866, Protoptychostomum Raabe, 1949, Tetrahymena Furgason, 1940, Trichodina, and Urceolaria Stein, 1867(Lee & Gutell, 2012Obert et al, 2023;Rataj et al, 2022;, whereas the stem of this helix is shortened to only two base pairs in Trichodina .…”

“…Interestingly, the lack of this helix was also reported for the spirotrich genus Plagiotoma (Obert et al, 2022). On the other hand, helix 9es3b was recognized in Haptophrya, Hysterocineta Diesing, 1866, Protoptychostomum Raabe, 1949, Tetrahymena Furgason, 1940, Trichodina, and Urceolaria Stein, 1867(Lee & Gutell, 2012Obert et al, 2023;Rataj et al, 2022;, whereas the stem of this helix is shortened to only two base pairs in Trichodina . It is not clear whether the absence of helix 9es3b is a molecular synapomorphy of the SAL supercluster (Spirotri chea + Armophorea + Litostomatea) in which Plagiotoma and clevelandellids are classified, or whether its loss may have occurred independently in the Spirotrichea and Armophorea.…”

“…Similarly, the majority of mutations was found in helix 9 in the clevelandellid genus Nyctotheroides Grassé, 1928 as well (Li et al, 2020). Higher variability of this helix was detected also in the genera Plagiotoma and Trichodina (Obert et al, 2022; Zhang et al, 2023). The length of helix 9 is relatively unstable in Trichodina , having three to six base pairs in its stem.…”

“…As expected, mutations are situated mainly in the hypervariable regions of the 18S rRNA molecule, especially in the V2, V4, and V7–V9 regions (Li et al, 2020; present study). Similarly to clevelandellids, the majority of mutations was detected within the hypervariable regions of the 18S rRNA molecule in the ciliate genera Haptophrya Stein, 1867, Plagiotoma Dujardin, 1841, and Trichodina Ehrenberg, 1830 (Obert et al, 2022; Rataj et al, 2022; Zhang et al, 2023). A common trait shared by clevelandellids isolated from the digestive tract of cockroaches is the absence of helix 9es3b in the V2 region.…”

“…More specifically, the V9 region has gradually shortened from N. nakhoni (166 nt) and N. galerus (132 nt), the sister group of all other species included in the present analyses, through N. angustipennis (115 nt) and N. ovalis (118 nt), the sister group of the family Clevelandellidae, to the family Clevelandellidae, whose members share the shortest helix 44 (103-105 nt) (Figure 8). A relatively long hypervariable region V9 was also found in the ciliate genera Haptophrya (145-148 nt), Hysterocineta (124 nt), Tetrahymena (166-169 nt), and Trichodina (122-123 nt) (Obert et al, 2023;Rataj et al, 2022;. Remarkable structural features of the 16S rRNA molecule in ciliates include the absence/reduction of helices 26 and 26a in Trichodina as well as an extraordinarily long and branched helix 40 in Haptophrya (Rataj et al, 2022).…”