Example 5 



Smear Preparation of Monocystis from the Seminal 

 Vesicle of an Earthworm 



Very few Sporozoa are available for class demonstration purposes, and one's 

 choice is practically limited to the inhabitants of the intestines of a cockroach 

 or to the specimen under discussion. 



The advantage of using Monocystis is that all the forms from the sporozoite 

 to the trophozoite occur in the seminal vesicle of the earthworm and, there- 

 fore, may be made available on a single smear. The degree of infection among 

 earthworms varies greatly, but it has been the author's experience that the 

 larger the earthworm, the more likely the chance of a heavy infestation. At the 

 same time, there is no use making a great number of smears for class demon- 

 stration purposes until a preliminary survey of a single smear has shown that 

 the material will be satisfactory. 



There is no need to kill or anesthetize the earthworm, which is simply 

 pinned down in a dissecting tray and slit from the anterior end to about the 

 sixteenth or seventeenth segment. The edges of this slit are pulled back and 

 pinned into place, disclosing the large white seminal vesicles. 



One should have available, before making the smear, a petri dish in which 

 are a couple of short lengths of glass rod, a supply of the fixative selected, an 

 adequate supply of clean glass slides, an eye-dropper type pipette, some 0.8 

 per cent sodium chloride, and some coplin jars of distilled water. Enough 

 fixative is poured into the petri dish so that when a slide is laid on the pair 

 of glass rods its lower but not its upper surface will be in contact with the 

 fixative. This level should be established with a plain glass slide before the 

 smears are made. 



Now the seminal vesicle of the earthworm is slit, and a drop of the con- 

 tained fluid is removed with the eye-dropper type pipette. This pipette is used 

 to smear a relatively thick layer of the material on the center of one of the 

 clean slides. Before it has time to dry, this slide is laid face down in the fixa- 

 tive for a period of about two minutes. Then the slide is removed, rinsed 

 under the tap, and examined under a high power of the microscope after a 

 coverslip has been placed over the smear. It is rather difficult to see the tro- 

 phozoite stages in an unstained preparation, but no difficulty will be experi- 

 enced in picking out the spore cases (pseudonavicellae) because of their 

 relatively high index of refraction. It may be assumed that adequate numbers 

 of the parasites are present if not less than three or four of these spore cases 

 occur within the field of a four-millimeter objective in a thick smear of this nature. 



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