Glycogen Metabolism in Larval Moniliformis dubius (Acanthocephala)

Abstract

Study of glycogen metabolism in larval Moniliformis dubius showed significant increase in the rate of metabolism as a result of activation of the cystacanths by bile salts. Anaerobiosis with increased levels of C02 also enhanced the rate of metabolism, a possible explanation of apparent enhancement of activation of the cystacanths by this gas phase. From the results of this study it appears that the role of the gas phase can be explained in terms of the nature of metabolic end products formed. The mechanism by which bile salts elicit the increase in rate of glycogen metabolism could not be determined. Graff and Kitzman (1965) reported that bile salts in vitro or bile of the host in vivo activate the cystacanth of Moniliformis dubius. They also noted that anaerobiosis and increased levels of CO2 enhanced the rate of bile salt activation of this larval parasite. In the rat host, rapid growth of the parasite follows activation and establishment of the cystacanth in the gut. Since Read and Rothman (1958) showed that M. dubius requires carbohydrate from the host's diet as a factor for growth and Fisher (unpublished) quoted by Graff and Kitzman (loc. cit.) stated that the cystacanth of M. dubius contains only small amounts of stored carbohydrate, a study was undertaken to determine the exact amount of glycogen present in the cystacanth as it reaches infective stage and to determine some of the parameters of its metabolism following activation. MATERIALS AND METHODS Maintenance of hosts and the life cycle The definitive host, Holtzman white rats (Holtzman Rat Co.), were maintained in groups of 10 to 20 in 12by 15by 18-inch wire mesh cages in air-conditioned quarters. They were provided Laboratory Chow (Ralston Purina Co.) and tap water ad lib. Only male rats were used. Periplaneta americana, the intermediate hosts, were maintained in painted 5to 15-gal aquaria on a diet of fresh apples, powdered rat chow, and water. Received for publication 26 May 1970. * From a dissertation submitted in partial satisfaction of the requirements for the degree of Doctor of Philosophy. t Supported in part by grants from the USPHS (GM 12263 and 5-T01-AI 00106) and by a predoctoral fellowship to the author (GM 37755). t Present address: Department of Biology, Texas A&M University, College Station, Texas 77843. Eggs of Moniliformis dubius were obtained from gravid female worms and concentrated by sedimentation in tap water. After removal of excess moisture by blotting on filter paper, the eggs were mixed in a ratio approximately 1:1 with fresh minced apple and the mixture was presented to cockroaches which had been denied food and water for the previous week. Complete development to the cystacanth stage in the hemocoel of the cockroaches required approximately 90 days at 24 to 28 C. To complete the life cycle, male rats of 60 to 90 g weight were infected with 25 cystacanths. Adult M. dubius were recovered 40 to 45 days later upon necropsy of the hosts. Handling of the cystacanths Infective cystacanths were removed from the cockroaches by flushing the hemocoel with KrebsRinger saline buffered at pH 7.4 with 0.025 M tris(hydroxymethylamino) methanemaleate, hereafter referred to as KRT. In experiments involving the use of cystacanths, the cyst wall was removed by expressing the intact cystacanths through a narrow tipped pipette which allowed passage of the larvae but impeded the passage of the cysts. The larvae were examined under a dissecting microscope for possible anatomical damage and/or abnormalities and o ly anatomically perfect larvae were selected. The use of a short glass cylinder having a stainless steel mesh bottom facilitated the rapid handling of the cystacanths during experiments and permitted he extraction of larvae by immersion of the cylinders nd the parasites in a given volume of 70% ethanol. Wet and ethanol-extracted dry weight ratios were determined using groups of 50 cystacanths, which were extracted for 18 hr in 70% ethanol and dried at 95 C for 24 hr. All incubations were performed in KRT. The gas phases during incubations were established by equilibrating the media for 5 min with either air, ir-CO2 (95:5), N2 (100), or N2-C02 (95:5). Bicarbonate was added to all CO2-equilibrated preparations in concentrations to maintain pH 7.4 at 37 C following gas phase equilibration.