What to do when ontogenetic tracking is unavailable: a morphometric method to classify instars in Milnesium (Tardigrada)

Abstract

Abstract Identification of tardigrade species is still reliant mostly on morphological and morphometric traits. Given that the number of taxonomically significant traits available in the genus Milnesium is exceptionally low, ontogenetic variability may provide additional methods for species delineation. However, it is notoriously difficult to establish whether both immature and sexually mature instars are present in examined populations. Here, we developed a simple analytical morphometric method that helps in testing whether both immature and mature life stages are present in a given set of individuals. In order to achieve this, we analysed 21 populations representing 11 Milnesium species, with developmental tracking data available for four species. Using cluster analysis, individuals were assigned accurately to three groups of instars: hatchlings, juveniles and adults (third instar onwards). Our analyses revealed that the most useful character in life-stage classification is buccal tube length. The method of classification of instars presented herein could be used as a simple tool for identification of the number of life stages present in a population. Finally, using our algorithm, we analysed morphometric datasets underlying the descriptions and redescriptions of all known Milnesium species to date and concluded that the majority of them did not include all three life-stage groups.