"Everything is not everywhere": Time-calibrated phylogeography of the genus Milnesium (Tardigrada)

Abstract

There is ample evidence that macroscopic animals form geographic clusters termed as zoogeographic realms, whereas distributions of species of microscopic animals are still poorly understood. The common view has been that micrometazoans, thanks to their putatively excellent dispersal abilities, are subject to the "Everything is everywhere but environment selects" hypothesis (EiE). One of such groups, <1 mm in length, are limnoterrestrial water bears (Tardigrada), which can additionally enter cryptobiosis that should further enhance their potential for long distance dispersion (e.g., by wind). However, an increasing number of studies, including the most recent phylogeny of the eutardigrade genus Milnesium, seem to question the general applicability of the EiE hypothesis to tardigrade species. Nevertheless, all Milnesium phylogenies published to date were based on a limited number of populations, which are likely to falsely suggest limited geographic ranges. Thus, in order to test the EiE hypothesis more confidently, we considerably enlarged the Milnesium data set both taxonomically and geographically, and analysed it in tandem with climate type and reproductive mode. Additionally, we time-calibrated our phylogeny to align it with major geological events. Our results show that, although cases of long distance dispersal are present, they seem to be rare and mostly ancient. Overall, Milnesium species are restricted to single zoogeographic realms, which suggests that these tardigrades have limited dispersal abilities. Finally, our results also suggest that the breakdown of Gondwana may have influenced the evolutionary history of Milnesium. In conclusion, phylogenetic relationships within the genus seem to be determined mainly by paleogeography.

Keywords: Apochela; EiE; climate type; long-distance dispersal (LDD); molecular clock; reproductive mode.

FIGURE 1

Approximate localities of the 127 Milnesium populations analysed in this study (see Tables S1 and S2 for details). Yellow circles indicate the newly analysed populations (N = 83), whereas red circles stand for data retrieved from the literature (N = 44)

FIGURE 2

Simplified Bayesian Inference time calibrated phylogenetic tree based on concatenated 18S rRNA+28S rRNA+ITS‐2+COI nucleotide sequences obtained with BEAST (left) and its relation to the most recent published phylogeny of Milnesium by Morek and Michalczyk (2020) (right). The left side tree: the upper values above nodes indicate the 95% highest posterior densities, which are graphically presented as horizontal blue bars, whereas the values under the nodes and in italics represent the posterior probability (PP) supports. Yellow circles superimposed on nodes indicate the two calibration points used as tree priors. The dashed branches indicate phylogenetic uncertainty. The main geological eras as well as the estimated time of the Gondwana breakup (vertical grey bar) are marked. Mya, million years ago. The six main clades within Milnesium (A–F) are collapsed and their sizes indicate the number of populations within each clade in both phylogenetic trees. The percentages within the collapsed clades indicate the support for the origin of the ancestor in given zoogeographic realm (see Results for more details). See Figure 3a–c for detailed relationships within each of the main clades uncovered in this study. The right side tree: values at nodes represent PP supports and the scale bar shows the number of substitutions per site. The arrows between both trees show the relationships between clades obtained in this contribution and in the phylogeny by Morek and Michalczyk (2020). The pie chart below indicates the number of species detected in the genus Milnesium to date divided into three categories (blue – species described under the integrative taxonomy framework; green – classically described species, that is, of which phylogenetic position is unknown; yellow – undescribed putative new species detected in this study)

FIGURE 3

Milnesium phylogeny reconstructed using the time calibrated Bayesian Inference based on concatenated 18S rRNA +28S rRNA +ITS‐2 + COI nucleotide sequences obtained with BEAST: A – clades A and B, B – clades C and D, C – clades E and F (see Figure 2 for the simplified tree). Values at nodes represent posterior probability (PP) supports and the scale indicates the time before present in Millions of years. Black vertical bars to the right of the population codes encompass species delineated using both phylogeny and morphology, whereas thin grey vertical lines indicate putative species suggested by the bPTP analysis alone and that did not agree with the integrative species delineation; numbers within vertical black bars are running species numbers (see Results for more details on species delineation). Further right, zoogeographic origin, climate type and reproductive mode are indicated by numbered and differently coloured squares. Colour and number coding are explained in the legend at the bottom of the last tree

FIGURE 4

The summary and interpretation of distribution patterns of Milnesium species recovered in this study. Species with limited ranges =species found only in a single zoogeographic realm; widespread species =species found in more than one zoogeographic realm (i.e., suggesting recent long distance dispersal, LDD); “inclusion species” species that were found in a different zoogeographic realm than the majority of species in a given clade (i.e., suggesting ancient dispersal); SSD, short distance dispersal (refers to species found in localities close to the borders of adjacent zoogeographic realms). Overall, there were 86% species showing no evidence for natural LDD and 14% species suggesting natural LDD (mostly ancient, only rarely recent)