Historical background

Entamoeba histolytica was first established in culture by Boeck and Drbohlav in 1925 in a diphasic egg slant medium they had developed for isolation of intestinal flagellates (2). A modification of this medium (LE) is still in use today (see below). Their success refuted the Promethean view of E. histolytica as an obligate tissue parasite. Dobell (13) introduced the use of rice starch as a carbohydrate source, and it remains a component of all media for xenic cultivation to this day. Soluble sugars, were they to be used, would be metabolized too rapidly by the bacteria and would not be available to the amebae. Several diphasic media were developed subsequently with serum, agar or egg extracts in the slants. Monophasic media that have been developed include the egg-yolk infusion medium of Balamuth (1) and TYSGM-9 of Diamond (8). Currently the most widely used media for xenic cultivation of E. histolytica are the diphasic LE and Robinson's (18) media and the monophasic TYSGM-9. These three media are detailed below. Additional recipes are given in Diamond's review (7).

Monoxenic cultivation of E. histolytica was first accomplished by Cleveland and Sanders (4) in a diphasic medium with a single species of bacterium, and the monobacterial Modified Shaffer-Frye medium (MS-F) was widely used at one time (17). Monoxenic cultivation is of limited use today except as a transitional stage between xenic and axenic cultures. Crithidia fasciculata or Trypanosoma cruzi are the associates of choice for monoxenic cultivation (6, 9). For media used in monoxenic cultivation, see Diamond's review (7).

Axenic cultivation of E. histolytica was first accomplished by Diamond in 1961 (5). The medium used was complex - a diphasic serum-enriched nutrient agar slant overlayed with a broth supplemented with chick embryo extract and vitamins. It was not until Diamond introduced the monophasic medium TP-S-1 in 1968 (9) that axenic cultures of E. histolytica started to be widely used. TP-S-1 was superseded by TYI-S-33 in 1978 (12) and this is currently the most widely used medium for axenic cultivation of E. histolytica. Diamond et al have recently described YI-S as an alternative to TYI-S-33 (10). These latter two media are described below. A partially defined medium is also available (11) but is not further considered here. Methods for obtaining cloned colonies in agar have also been published (15, 16).

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