Dr. Kassai Tibor - Dr. Murai Éva szerk.: Parasitologia Hungarica 10. (Budapest, 1977)
(homogenisation, filtration, sporulation, aeration, etc. ) to increase the efficiency of recovery. The optimal procedure has been described in detail by RYLEY et al. (1976). These methods are generally impracticable with fish coccidia. Flotation usually gives rise to deformation of the oocysts, and is therefore out of question. Filtration of the oocysts through a fine mesh and subsequent sedimentation is a valuable complementary procedure to faecal examination. Sedimentation is a relatively rapid procedure, oocysts can be recovered from the sediment, but although the soluble and coarse fibrous components are removed by previous processing, masses of faecal debris are present in the sediment together with oocysts. Digestion: An oocyst population free of debris can be obtained exclusively by digestion. The mucosal lining of the infected segment of intestine is scraped off after through washing, and is placed in an 0.25-0.5% trypsin solution. Trypsin can be changed repeatedly by cautious centrifugation and décantation of the supernatant. Tissue scraps gradually become digested until finally only oocysts are found at the bottom of the centrifuge tube . The oocysts so collected can be stored in water for a relatively long time. (In our experiments trypsin did not injured the oocysts, nor the sporocysts released from disrupted fully sporulated oocysts, not even when bile or 0. 75% desoxycholate was added to the system. ) Examination for oocysts localizing in tissues is made as follows: a small piece of tissue, the size of a pea, is impressed between two slides, or between two glass plates if is larger, and examined under the microscope. Since such oocysts usually form groups, the aggregations may be recognized already at a low power of magnification. With small fish hosts the entire gut can in toto be examined in this manner. With larger organs the digestion procedure should be employed, as described above. Storage of oocysts Oocysts purified by digestion or sedimentation could be stored in water over periods ranging from two weeks to five months without notable morphological change. Oocysts recovered from the faeces ( E. carpelli , E. sinensis ) soon became disrupted in the course of storage, however, the contours of sporocysts and residual body could distinctly be recognized inside the resistant sporocysts for several months. Oocysts deteriorated rapidly in 4% formaline or 2% potassium bichromate solution, but addition of a few drops of 4% formaline to the faecal sample did not affect them. In view of this, a practicable method was elaborated for use in field studies. Samples of mucus were taken from the gut of the examined fish host, preferentially from the anterior intestine. The host had been starved for one or two preceding days whenewer possible, to ensure complete evacuation of the gut. If there was no other choice faeces was used instead of intestinal mucus. A small amount of mucus or faeces was transferred to a slide, not more than sufficient to cover three quarters of a coverslip, surface when compressed. A few drops of 4% formaline or 2. 5% glutaraldehyde solution were subsequently instilled under the coverslip, which was then fixed in position with Canada balsam. The shape and structure of the oocysts remained preserved in such preparations for periods ranging from 4 weeks to 1 year, in the worst case for a few days at least, and the preparations were thus available for microscopic study (depiction and measurements) in the laboratory, after the conclusion of sampling in the field. The same procedure can also be used for obtaining preparations of Eimeriae localizing in tissues. Discussion This paper was written with the aim to encourage studies of fish coccidia by methodical instructions derived from the present writer's own experience. According to LOM (1970) up to now about 40 species of Eimerian parasites have been described from marine fish hosts. Of some 60 Eimerian species known from freshwater fishes most
