Studies on the egg stage of Heterobilharzia americana (Trematoda: Schistosomatidae)
The purpose of the present investigation is to describe the hatching process of the egg of Heterobilharzia americana (one of two mammalian schistosomes in North America), and to assess the various factors which may influence the ability of this egg to hatch The hatching of the egg in dechlorinated water was followed under low power of the compound microscope, and it was noted that the egg underwent a dramatic prehatch expansion just prior to eclosion. Measurements of hatching eggs with an ocular micrometer showed an average 47% increase in volume, an expansion due to the rapid influx of water during this period, according to vital stain experiments. At the time of hatching, the egg was seen to burst and the miracidium extruded. Phase contrast examination of egg shells cleared in glycerol showed no prevalent shell rupture patterns, indicating that no preformed line of weakness in the shell was present in this non-operculate egg Miracidial participation in the hatch process did not appear to be necessary, at least in the final stages of hatching. A small percentage of eggs containing a dead miracidium was shown to rupture when placed in dechlorinated water. The viable miracidium within the egg, however, did become active in a series of documentable steps of unvarying sequence prior to hatching. It was shown that these steps are reversible upon reimmersing the egg in 0.85% NaCl, and that the egg/miracidium complex can be activated and deactivated repeatedly in this manner The conditions under which the egg of H. americana would hatch were explored utilizing a variety of conditions of light, temperature, pH, oxygen concentration, and NaCl concentration of the hatching medium. It was shown unequivocally that these eggs do not require light for hatching, as eggs hatched equally well in light or darkness Eggs were allowed to hatch in water in temperatures of 10(DEGREES)C to 37(DEGREES)C, and hatched well within this range, with the smaller percentages hatching at the temperature extremes. Optimum temperature for hatching appears to be 13(DEGREES) - 30(DEGREES)C. No requirement for oxygen in the hatching water was evident as eggs hatched equally well in both oxygenated and deoxygenated water Tris-maleate buffers of pH 6.7, 7.2, and 8.5 were used to assess hatching at these pH values; and sodium acetate/acetic acid buffers of pH 4 and pH 5 provided the hatching solution for lower pH values. The optimum pH for hatching appears to be pH 7.2, but eggs will hatch at pH 6.7 and pH 8.5. Acetate media at pH 4 and pH 5 are lethal for miracidia, and eggs did not hatch in these solutions Solutions of NaCl--0%, 0.5%, 0.85%, and 1.0%--were formulated and standardized with a solu bridge. Eggs were hatched in these solutions, and it was demonstrated that salinity has a marked effect on the hatching rate of eggs, with the optimum hatching occurring at 0% NaCl, and no hatching at 1.0% NaCl. In order to separate the effects of osmolarity from the possible effects of ions in the above experiment, hatching solutions of NaCl, CaCl, sucrose and glycerol were formulated for osmolarities of 50, 100, 200, 300, and 400 mOsmols. In both ionic and nonionic solutions, increasing osmolarity yielded lower hatch rates than did low osmolarity solutions. To further define the need, if any, for a specific ion component, artificial pond water containing calcium, magnesium, sodium, and potassium was formulated, and the hatch percentage compared with those eggs hatching in artificial pond water lacking one of these ions. No requirement for any of the specific ions tested was demonstrable