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Flies are vectors of disease because flies can transfer bacteria from dirty places to food or the human body. The purpose of this study was to determine the differences in the number of Escherichia coli carried by three types of flies in different location (settlement and garbage) dump. This factorial study involved three types of flies, i.e. the house fly (Musca domestica), green fly (Chrysomya megacephala), and fruit flies (Drosophila sp.). Catching flies is carried out at 02.00 to 05.00 pm for four times a week. Calculation of the number of E. coli using the colony counter in CFU / ml units and two-way ANOVA was chosen to analyze the data that has been obtained. The result, differences in location did not give a significant effect (p = 0.533), while the type of fly had a significant effect on the number of E. coli (p < 0.001). The location and the type of fly also did not show a significant interaction (p = 0.816). Interestingly, Drosophila is the type of fly that carries the least amount of E. coli when compared to the other two types of flies.