The method devised by Boeck and Drbohlav for the cultivation of E. histolytica has been successfully used for cultivating this species and various other entozoic amoebae of man and monkeys.Various modifications of the original technique have been tried, and several improvements are here described. Among these are methods of cultivation (involving the addition of solid rice-starch to the medium) whereby (1) more luxuriant and prolonged growth of the amoebae, and (2) all stages—including encystation and excystation—in the life-cycle of E. histolytica and other species can be obtained in vitro.Methods by which cultures of entozoic amoebae can be initiated from encysted forms have been discovered and are also described, and some indications are given for the isolation of pure strains from mixed cultures.The influence of the accompanying bacterial flora upon the various amoebae cultivated has been studied, and is briefly discussed in the light of certain experimental findings.Loss of infectivity to kittens has been observed in E. histolytica as an apparent result of cultivation in media containing solid starch.

No abstract

A pure strain of Entamoeba histolytica has been isolated and cultivated, and an attempt has been made to study and describe its whole life-history in detail.This strain (K. 28 c) was derived from the dysenteric dejecta of a kitten experimentally infected per os by means of typical cysts from the faeces of a monkey (Macacus sinicus). It has now been under continuous cultivation for about 20 months (220 serial subcultures), and its development in vitro has been uniform throughout.Methods have been devised, and are here described, whereby any desired stage in the life-history of this strain—amoebae, cysts, and all intermediate stages (including encystation and excystation)—can be readily procured in vitro at will.Detailed study has shown that the trophic amoebae multiply in cultures by simple binary fission only, as they do in their natural hosts. Their mode of division is briefly described.Encystation also occurs in vitro just as it does in the bowel, with formation of characteristic precystic amoebae and the final production of typical quadrinucleate cysts.Excystation has been carefully studied, and it has been found that a single quadrinucleate, amoeba escapes from each cyst through a minute perforation in its wall. An account is given of this remarkable process, which has not been described previously.The 4-nucleate excysted (metacystic) amoeba has been found to produce a new generation of trophic forms by a complicated series of nuclear and cytoplasmic divisions, which are described in detail for the first time. The final result of this subdivision is the production of eight uninucleate amoebulae by each quadrinucleate amoeba hatched from a cyst.These amoebulae are young trophic amoebae, and not gametes or conjugants. No sexual phenomena of any sort have been observed during the metacystic itages: and the life-history of E. histolytica, as visible in vitro, is thus wholly sexual.A development similar to that here described in the case of Strain K. 28 c has been found to occur in many other cultivated strains of E. histolytica— including a strain isolated directly from man, and a human strain experiaentally implanted in a monkey (M. sinicus) and recovered therefrom in pure culture. There are therefore good reasons for concluding that the development were described is not abnormal, and that it is probably closely parallel to that which occurs naturally inside man.

It is now becoming generally recognized that there are three different amoebae which may be present in the human bowel—the harmless Entamoeba coli(Lösch) Schaudinn and “Entamoeba” nana Wenyon and O'Connor, and the pathogenic E. histolytica Schaudinn. We have discussed these three species in an earlier paper (1917), and there noted their chief distinctive characters. In the present paper we shall describe a fourth and much less common species which we have recently encountered. It differs in some respects so conspicuously from the others that it appears to us necessary to place it not only in a new species, but even in a new genus. We propose therefore the new name Dientamoeb fragilis for our amoeba, but we can discuss the question of nomenclature most conveniently after we have described the organism itself. This, therefore, we shall now do.

See Dobell (1920), where these observations are considered in detail. * The discovery was reported by Virchow in i860. See Dobell (1919) for further details.f This discovery is usually incorrectly attributed to Lambl (i860). Cf. Dobell (1919 a, pp. 8-9, and 71 et seq. ) where additional details will be found.(2) The Phylum Mastigophora consists of all those Protozoa which move, in their fully developed and typical condition, by means of whip-like filaments or flagella-familiar to all who have studied Euglena, or any other common flagellate.(3) The Phylum SPOROZOA contains a number of exclusively parasitic forms, which in their motile stages-when present-move without the aid of any special external locomotory organs. The several common species of Monocystis, parasitic in earthworms, supply familiar examples-with their slow, worm-like motions, performed by the body as a whole.(4) The Phylum Ciliophora contains all the Protozoa which move, in their typical active stages, by the agency of many little hair-like threads or cilia -exemplified in the familiar Paramecium and other common ciliates.Each of these Phyla contains a vast array of species, variously collected into genera, families, orders, and higher groups. It will be unnecessary, however, to discuss their classification in detail here, and we shall limit ourselves to a consideration of the systematic position of those species alone with which the present work is concerned. It will suffice to note the general grouping of our forms, and their more obvious relations to one another.The human intestine harbours protozoa belonging to all the four Phyla just enumerated. As these groups contain organisms as different * See especially Yorke, Carter, Mackinnon, Matthews, and Smith (1917), Smith (1919, 1919a), Dobell (1921).tSee Dobell (1921). % Among more recent contributions to this subject the reader may be referred to the following : Galliard and Brumpt (1912), Paviot and Garin (1913), Landouzy and Debrd (1914), Bloch (1916) -French cases; Kuenen (1918) -Dutch cases; Fischer (1920) -German cases; Yakimoff (1917)-Russian cases; Kofoid, Kornhauser, and Plate (1919), Cort and McDonald (1919) -United States cases. There are also numerous other works dealing with the occurrence of intestinal protozoa in the inhabitants of temperate climates, but it would lead us too far to discuss-or even to attempt to citeall of them. See also the papers on French cases of Balantidiosis cited on p. 119 infra.* Numerous references will be found in the Tropical Diseases Bulletin. The reader interested in this subject may be referred to the following recent works, which will also supply him with numerous further references to the immense literature dealing with the incidence of intestinal protozoal infections :