Until now, very little is known about the ability of adult and deutonymph water mites (Acari, Hydrachnidia) to survive in sub-zero temperatures. Information concerns mainly water mites from vernal astatic waters, and the knowledge has never been experimentally verified. To determine the sensitivity of water mites to freezing, experiments were conducted on (1) the impact of acclimatization, (2) temperature, and (3) duration of freezing on survival, (4) the survival rate of water mites from various types of water bodies, and (5) the survival rate of water mites from different climatic zones. The experiments were carried out in a phytotron chamber, and water mites were placed in containers (10 × 10 × 5 cm) filled with 4/5 of water for 10 specimens each. Water mites were identified to the species level after finishing the experiments. The temperature was lowered 1 °C every hour until the target temperature was reached. After a certain period of freezing (depending on the treatment) the temperature was raised by 1 °C every hour until it reached 4 °C. The time of the experiment was measured from the moment the desired temperature was reached (below 0 °C) until the ice thawed and the temperature of 4 °C was reached again. The highest survival rates had Limnochares aquatica, Piona nodata, Sperchon clupeifer and Lebertia porosa, followed by L. insignis, Hygrobates longipalpis, H. setosus, Limnesia undulatoides, Piona pusilla, Arrenurus globator, Hydrodroma despiciens, Piona longipalpis, Sperchonopsis verrucosa, Unionicola crassipes and Mideopsis crassipes; no specimens of Torrenticola amplexa survived. The following conclusions were drawn: (1) water mites can survive freezing to −2 °C, lower temperatures are lethal for them; (2) they survived better the short period of freezing (24–48 h) than the long period (168 h); (3) resistance to freezing seems to be an evolutionary trait of individual species, only partly related to the living environment; and (4) freezing survival rates are linked to the region of Europe and are much lower in Southern than in Central Europe.