Central circadian clock, located in the brain, synchronizes peripheral clocks found in almost every cell of the body and with its effect on metabolism, physiology and behavior of the organism maintains homeostasis. An important factor, when it comes to establishing homeostasis in the host, is also the intestinal microbiota. It is well known that microbiota and host immune system are tightly connected, the latest research showed that dysbiosis results in altered circadian clock. In our study we focused on the bacteria Escherichia coli (E. coli) as an important representative of microbiota and investigated its effect on the host circadian rhythms and immune system in the established stable coculture with human liver cells HepG2. For the assays two different E. coli strains from the collection of fecal E. coli strains from healthy individuals were selected: the strain BJ23 with many and strain BJ17 with only a few genes encoding virulence associated factors. After a 4 hour incubation of the BJ strain with the HepG2 cell line, the exposed HepG2 cells were sampled for 72 hours and the relative quantity of mRNA of the circadian and immune genes was analyzed with qPCR. We found that after 72 hours the bacterial strain BJ23 had a greater impact on the circadian clock genes of HepG2 cells, while there was no difference in activation of immune system genes between both employed strains. Our research therefore contributed to a better understanding of the E. coli impact on the host’s circadian clock.