Does the yicJI operon contribute to uropathogenic Escherichia coli fitness when cultured in human urine
Kalinka, Christoper P.
MetadataShow full item record
Escherichia coli is a common bacterium found in the intestinal tracts of many mammals. A medically relevant type of E. coli is known as uropathogenic E. coli (UPEC). One of the most critical research areas involves genes that are responsible for bacterial fitness, like the yicJ gene, which encodes the sodium:galactoside symporter; yicI, which encodes the α-glycosidase YicI; and frzR, which encodes FrzR that is the activator of the frz operon and a putative activator of the yicJI operon. To examine what impacts yicJ, yicI, the yicJI operon, and frzR had on fitness, each gene was cloned into the pMMB91 plasmid and growth curves in urine and buffered Luria broth were done for each recombinant strain in 96-well plates from 0 h-72 h. The growth curve studies demonstrated that overexpression of yicJI and frzR significantly enhanced growth in shaken human urine, whereas overexpression of yicJI attenuated growth in Luria-Bertani broth. Additionally, a portion of the promoter region of frzR was also cloned into the pCRISPathBrick plasmid to act as a gRNA scaffold for dCas9 to knock down the gene expression of the frzR gene. When frzR expression was knocked down, fitness was enhanced in shaken human urine, but decreased growth in static pooled human urine. Quantitative reverse transcriptase polymerase chain reaction analysis showed that overexpression of the frzR gene led to the transcriptional activation of both the frz and yicJI operons. These results demonstrate the potential importance of the yicJI operon and frzR gene in UPEC pathogenesis.
Urine -- Microbiology