Repeatable methods to establish bdelloid rotifer cultures of cold-dwelling origins

In this study, we investigated the influence of several culturing conditions on seven species (eight strains) of endemic Antarctic bdelloid rotifers including Adineta grandis, Adineta sp., Philodina gregaria, Philodina sp. (Bd25), Habrotrocha devetteri, Habrotrocha sp. (Bd14), and Macrotrachela donneri (two strains: CL-VH strain, MR-VH strain). A multifactorial design was used to assess temperature, media type, and food concentration. In early development of cultures, individuals of P. gregaria, A. grandis, and Habrotrocha sp. (Bd14) were monitored daily and behaviour scored (active, feeding, slightly active, immobile, tun, dead, offspring presence). This data is contained in the pilot trial file. The final optimization trial tested three temperatures (5, 10, and 15 °C), two food volumes (low = 5 µL, high = 25 µL; dispensed once at the beginning of the test), and two media (1 mL per well of either DMW or MilliQ) for a total of 12 different treatment conditions. One optimization test was completed for each of the seven species. Counts of the total number of rotifers (i.e. the alive original animals and any active offspring) and eggs in each well were completed periodically for up to 44 days. Data for culture optimization trials are contained in the optimisation trial file.