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Artificial cross between two genetically different populations of Japanese Misgurnus loach was made to examine the reproductive capacity of the artificial inter-populational hybrid females. Ploidy status and microsatellite genotypes of the eggs laid by these hybrids were inferred from those determined in progenies developed by normal fertilization with haploid loach sperm, induced gynogenesis with UV-irradiated goldfish sperm and/or hybridization with intact goldfish sperm. Some hybrid females laid unreduced diploid eggs genetically identical to the mother. However, these diploid eggs could not develop by spontaneous gynogenesis, but grow to triploid by incorporation of a sperm nucleus. Other hybrid females laid haploid eggs together with diploid eggs and/or various aneuploid and polyploid eggs. Thus, a disruption of normal meiosis occurred in inter-populational hybrid females. The results suggested that the two populations should be so distant as to give rise to atypical formation of unreduced and other unusual eggs in their hybrids.

Here, we propose a simple and inexpensive method for fish sperm cryopreservation. Sperm samples of the loach Misgurnus anguillicaudatus (Teleostei: Cobitidae) were diluted 7-fold by an extender containing 63.5 mM NaCl, 114 mM KCl, 20 mM Tris and 10% methanol. The cryogenic straws were placed in three kinds of self-made tubes which diameter was changed by commercially available materials and then immersed into powdered dry ice for 2 min and plunged into liquid nitrogen. This procedure resulted in a cooling rate at -421.4 +/- 119.84 (control), -55.8 +/- 4.32 (tube 1), -40.2 +/- 3.43 (tube 2) and -33.3 +/- 2.09 C/min (tube 3). In the slowest cooling rate by the tube 3, total motility (72 +/- 3 %), duration (146 +/- 12 s) and hatching rates (29 +/- 04 %) were higher than those by other rates. Progressive motility (83 +/- 5 %) did not differ significantly from fresh samples.