Integrative taxonomy resolves species identities within the Macrobiotus pallarii complex (Eutardigrada: Macrobiotidae)

Abstract

The taxonomy of many groups of meiofauna is challenging due to their low number of diagnostic morphological characters and their small body size. Therefore, with the advent of molecular techniques that provide a new source of traits, many cryptic species have started to be discovered. Tardigrades are not an exception, and many once thought to be cosmopolitan taxa are being found to be complexes of phenotypically similar species. Macrobiotus pallarii Maucci, 1954 was originally described in South Italy and has been subsequently recorded in Europe, America, and Asia. This allegedly wide geographic range suggests that multiple species may be hidden under this name. Moreover, recently, genetic evidence to support this was put forward, and the Macrobiotus pallarii complex has been proposed to accommodate putative species related to M. pallarii. Here, we describe three new pseudocryptic species based on populations that would have been all classified as Macrobiotus pallarii if molecular methods were not employed. Using an integrative taxonomy approach, we analyzed animals and eggs from the topotypic population of Macrobiotus pallarii, together with four other populations of the complex. We recovered four distinct phylogenetic lineages that, despite the overlap of morphometric traits, can be separated phenotypically by subtle but discrete morphological characters. One lineage corresponds to Macrobiotus pallarii, whereas the other three are newly described as Macrobiotus margoae Stec, Vecchi & Bartels, sp. nov. from the USA, Macrobiotus ripperi Stec, Vecchi & Michalczyk, sp. nov. from Poland and Finland, and Macrobiotus pseudopallarii Stec, Vecchi & Michalczyk, sp. nov. from Montenegro. To facilitate species identification, we provide a dichotomous key for species of the M. pallarii complex. Delimitation of these pseudocryptic taxa highlights the need for an integrative approach to uncover the phylum’s diversity in full.

First insights into female sperm storage duration in tardigrades

“…A similar time threshold can be expected in other closely related macrobiotids with known or suspected presence of spermatheca (i.e. taxa clustered in subclade B in Stec, Vecchi, Calhim, et al, 2021 , Stec, Vecchi, Dudziak, et al, 2021 ). Monitoring oviposition after the experimentally controlled mating encounters beyond the first batch of eggs (see what has been previously done e.g., Sugiura et al, 2019 ; Sugiura & Matsumoto, 2021a ) would provide key data for interspecific variation in sperm storage ability in Macrobiotidae.…”

“…Furthermore, the presence of sperm in the spermatheca of stock population individuals with an immature ovary (e.g., using Orcein staining) is suggestive, albeit not conclusive, evidence that sperm could have been retained from the previous oviposition cycle, and/or that they might be stored for the full duration of the current oocyte development. A recent phylogenetic analyses of the genus Macrobiotus places most species with indications of the presence of spermatheca in a subclade B sensu Stec, Vecchi, Calhim, et al ( 2021 ) and Stec, Vecchi, Dudziak, et al ( 2021 ). None of the three macrobiotid taxa with detailed descriptions of sexual interactions (Sugiura & Matsumoto, 2021c ) belong to this subclade.…”

“…Storage structures (organs) 1 This study species (Figure 2a-f and Poprawa et al, 2015) and other members of the subclade B of the genus Macrobiotus (Rebecchi, 1997;Stec, Vecchi, Dudziak, et al, 2021) have a spermatheca.…”

Reaching the Monophyly: Re-Evaluation of the Enigmatic Species Tenuibiotus hyperonyx (Maucci, 1983) and the Genus Tenuibiotus (Eutardigrada)

“…Over the last ten years, the implementation of molecular techniques and taxonomic analyses by means of an integrative approach have accelerated the emergence of new species being characterized morphologically and genetically in detail, e.g., [5][6][7][8][9][10][11][12]. The acquisition and accumulation of genetic data tightly associated with phenotypic information enhanced studies on two major subjects in tardigrade taxonomy: (i) The recognition and disentanglement of cryptic/pseudocryptic diversity, e.g., [13][14][15][16][17][18][19] and (ii) the construction of comprehensive molecular phylogenies at the family level or higher that considerably impacted tardigrade systematics, e.g., [20][21][22][23][24][25][26][27][28][29][30]. Notably, there were not only integrative descriptions of new species for science that have contributed greatly to these subjects' investigation, but also revisional notes with updated diagnoses and new information on already-known taxa, e.g., [31][32][33][34][35][36][37].…”

Sisubiotus hakaiensis sp. nov. (Tardigrada, Macrobiotidae), a new tardigrade species from Calvert Island (British Columbia, Canada)

“…Sisubiotus wuyishanensis (Zhang & Sun, 2014), described from China (Zhang & Sun 2014) was deemed to be insufficiently characterized to be differentiated from S. spectabilis and S. grandis and thus considered as species inquirenda by Stec et al (2021a). Integrative taxonomy integrates multiple lines of evidence from multiple technique (morphological, molecular) to solve taxonomic issues, and it has been providing to be extremely useful in tardigrades taxonomy (see for example Kiosya et al 2021;Stec et al 2021b;Stec & Morek 2022). Thus, using an integrative taxonomy approach we here describe Sisubiotus hakaiensis sp.…”