EPIDEMIOLOGICAL AND BIOLOGICAL PROBLEMS 23 



the firemen who drank polluted water than among those who did not, 

 serves as good evidence that many of those cyst passers among the higher 

 percentage group were infected by the polluted water they drank. 



C. Discovery of Cysts of E. histolytica in Sewage and Sewage Effluent. 

 Recently, Gordon (1941) reported that the effluent of the Moscow sewage 

 discharging into a river had been found to contain E. histolytica-\ike cysts 

 constantly during a period of 3 years of examination, and that stor- 

 age of the effluent for 6% hours in a tank failed to remove all cysts from 

 the supernatant. Cram (1943) , in this country, reported that E. histolytica- 

 like cysts were occasionally found in the sewage sludge both from munici- 

 palities and from army camps, and that storage and sewage treatment 

 failed to remove all the cysts from the sewage effluent. In a previous 

 report, the present author (1945a) showed that the cysts of E. histolytica 

 have a specific gravity of about 1.06 and that even in perfectly quiet, pure 

 water, it would take 2 or more days for the cysts to settle through a 5-foot 

 depth at 10-28° C. These results imply that in places where carriers 

 exist, the sewage will contain the cysts, and some of these cysts are likely 

 to remain in the effluent after treatment. Pollution of water with this 

 sewage or sewage effluent will render the former a source of amoebic infec- 

 tion. 



III. Biological Problems in Water-Borne Amoebiasis 



Having shown that polluted water supply may constitute one of the 

 most important sources of amoebic infection, this section of the paper 

 attempts to bring out some of the biological problems related to the cysts 

 of E. histolytica in order to furnish information on the purification or dis- 

 infection of water for removal or destruction of the amoebic cysts that may 

 be present. 



A. Cultivation of E. histolytica in Artificial Media. To detect the 

 presence of cysts in possible agents of transmission, or to determine the 

 viability of cysts in survival tests requires cultivation of the cysts in arti- 

 ficial media. Excystation of the mature cysts and multiplication of the 

 trophozoites result in a positive culture. Although several good entamoeba 

 media have been successfully used by protozoologists, obtaining positive 

 culture of the organism is not simple. As shown in a previous report by 

 the author (1946), E. histolytica requires a moderately anaerobic condition 

 for excystation and a profound anaerobic condition for perpetuation, and 

 these anaerobic conditions are furnished by the growth activity of an 

 accompanying bacterial flora. This bacterial flora must also not produce 

 unfavorable pH range in the culture. These growth requirements of the 

 amoebae are not provided by every kind of bacterial flora. Therefore, 

 unless the cultures are handled by experienced persons, cultural results 

 obtained in tests for the presence or survival of amoebic cysts may be 

 misleading. 



B. Survival of Cysts of E. histolytica in Water and Sewage. In a 

 previous report by the author (1943), it was shown that culture-induced 



