Reversible Inhibition of Development and Hatching of Infective Eggs of Ascaris lumbricoides var. hominis

Abstract

The inhibition of hatching of Ascaris suum eggs by iodine and its reversal by a reducing agent, hydrogen-sulphide, has been described (Hurley and Sommerville, 1982, International Journal of Parasitology 12: 463-465). Attempts have been made to explain the inhibition of hatching of A. suum eggs using carbon dioxide-nitrogen gas mixture as a hatching stimulus. Rogers (1966, Comparative Biochemistry and Physiology 17: 1103-1110) proposed that reactive thiol groups, acting as receptors for CO2, are present on the egg surface. These receptors are oxidized by iodine, which blocks hatching. Treatment with a reducing agent would restore the reactive groups. An alternate hypothesis is that hydrated CO2 may act as a hatching stimulus, and the hydration of CO2 is catalyzed by carbonic anhydrase (Hurley and Sommerville, 1982, loc. cit.). Iodine inhibits carbonic anhydrase (Riese and Hastings, 1940, Journal of Biological Chemistry 132: 281-292); this inhibition is reversed by reducing agents, which explains the reversible inhibition of hatching infective eggs. In our experiments, we produced a similar reversible inhibition of hatching, as well as development of infective eggs of Ascaris lumbricoides var. hominis, without using a CO2-N2 gas mixture as a hatching stimulus. Fresh fertile eggs, recovered from the stool samples of patients suffering from ascariasis, were exposed to iodine (0.04 M) solution for 120 min before allowing them to embryonate at 25 C. Some of the iodine-treated eggs were also exposed to 1% sodium-thiosulphate for 60 min prior to embryonation. Fresh untreated eggs were treated with 1% sodium-thiosulphate and allowed to embryonate. Percentage embryonation at the end of 28 days of incubation was compared with the control eggs (Fairbrain, 1961, Canadian Journal of Zoology 39: 153-162). To study the effect of iodine on hatching of infective eggs, fresh fertile eggs, after 28 days incubation at 25 C, were exposed to iodine (0.04 M) solution from 120 min before exposing them to the hatching stimulus. Hatching is defined as the emergence of the larvae from the egg shell with the activation of the embryo. Some of the embryonated eggs were subsequently treated with 1% sodium-thiosulphate and then subjected to the hatching stimulus. The hatching stimulus used was the change in the incubation temperature. Eggs that were incubated at 25 C until the development of larva, were, after treatment with iodine and sodium-thiosulphate, incubated at 37 C in Eagle's Minimum Essential Medium for 45 min. Percentage hatch was compared with the control percentage hatch. Iodine inhibited development as well as hatching of the infective eggs of A. lumbricoides. This inhibition could be reversed by subsequent treatment of iodine-treated eggs with sodium-thiosulphate. Eggs that were treated with iodine alone did not develop beyond the 2-celled stage, whereas the percentage embryonation of eggs that were treated with iodine and later with sodium-thiosulphate approached the percentage embryonation of the control group. The eggs that were treated with sodium-thiosulphate alone, developed larva 7 days earlier than the control eggs. The mechanism of this inhibition is not known. Possibly iodine penetrates the egg shell and affects its metabolism. Inhibition of development does not appear to be total since a 2-celled stage developed in some of the iodine-treated eggs. A similar effect was seen on hatching of the infective eggs. A C02-N2 gas mixture was not used as the hatching stimulus, so a mechanism involving the blocking of egg surface receptors for CO2 or inhibition of carbonic anhydrase by iodine does not appear to be possible. Moreover, the medium used for embryonation (0.1 M formalin) and hatching, did not contain any significant amount of dissolved CO2, which could act as a hatching stimulus, together with the change in temperature. How the reduced conditions enhance development, as seen by the treatment of eggs with sodium-thiosulphate alone, is not known.