A cytological study of the Gregarine Parasites of Tenebrio molitor, using the Ultra-centrifuge
Authors: Daniels Margaret I. 
Published: April 2021

ABSTRACT
A. Three gregarine species are found to inhabit the mid-gut of the mealworm larvae used: Gregarina cuneata Stein, Gregarina polymorpha Hamm, and Gregarina steini Berndt. The often described Steinina ovalis is probably seldom or never found.
They live only in the mid-gut of larvae. They are never found in pupal or adult forms.
Gregarines have been seen moving when in a stratified condition.
B. The gregarine cytoplasm has five important inclusions, each having a characteristic position in a centrifuged animal (Text-fig. 2).
Paraglycogen.—This gives a dark brown colour with iodine, a pinkish general colour with the acid fuchsin of the Feulgen technique, and often a red colour with Bauer’s reaction. It occupies the centrifugal pole of the centrifuged cell and is in the form of disc-like granules of varying size. In young centrifuged Gregarina steini chromidial granules are seen in the paraglycogen area, and have, therefore, approximately the same specific gravity. They arise by karyosomic budding with the subsequent extrusion of these buds into the cytoplasm. They stain with iron alum haematoxylin, like the karyosome, and both give a negative result with the Feulgen test for thymonucleic acid. They probably correspond to Joyet-Lavergne’s ‘albuminoid reserves’, but do not have the mitochondrial ‘cap’ he describes. Mitochondria.—These are usually granular, but sometimes rod-like. They are seen between the ‘alveoli’ formed by the paraglycogen granules. They lie distally to the paraglycogen in a centrifuged parasite ; they stain by the iron alum haematoxylin long method, after Benoit, Champy, or Altmann fixation, also with Altmann’s fuchsin picric acid stain and the Bensley Cowdry modification of it. The Nucleus is karyosomic, and the karyosome is moved to the centrifugal pole by pressure as is the nucleolus of metazoan cells. The nucleus shows budding of the karyosome. There is plasmatic as well as chromatic material in the karyosome, as shown by centrifuging. The nucleus gives a negative result with Feulgen’s nuclear reaction, but chromatin may exist in a very dispersed condition. Golgi Material.—This lies at the centripetal end of the nucleus. It is hest shown by Weigl fixation. The large and regular Golgi elements are slightly heavier than the granular Golgi material, which may be compared with that of young oocytes. Fatty Material lies at the extreme centripetal pole of the cell, in globules of varying size. It becomes brown or black after treatment with osmium tetroxide, and vivid cherry red with Sudan IV. It gives a negative result with the Schultz reaction for cholestrol.
C. Large globules are seen in the protomerite of Gregarina steini, eosinophile, sometimes fuchsinophile, and also staining with methylene blue. These move towards the centrifugal pole.
Methylene blue preparations show blue granules among the paraglycogen granules in the centrifuged animal. They are remarkably resistant to dilute sulphuric acid. They are possibly allied to volutin or chromidia.
Tests for the presence of Vitamin C yielded negative results.
Only the inclusions of the gregarines in the gut lumen were studied, and the complete life-cycles of the species were not followed out.