Coordinate expression of fimbriae in uropathogenic Escherichia coli

Abstract

Uropathogenic Escherichia coli is the most common etiological agent of urinary tract infections. Bacteria can often express multiple adhesins during infection in order to favor attachment to specific niches within the urinary tract. We have recently demonstrated that type 1 fimbria, a phase-variable virulence factor involved in adherence, was the most highly expressed adhesin during urinary tract infection. Here, we examine whether the expression of type 1 fimbriae can affect the expression of other adhesins. Type 1 fimbrial phase-locked mutants of E. coli strain CFT073, which harbors genes for numerous adhesins, were employed in this study. CFT073-specific DNA microarray analysis of these strains demonstrates that the expression of type 1 fimbriae coordinately affects the expression of P fimbriae in an inverse manner. This represents evidence for direct communication between genes relating to pathogenesis, perhaps to aid the sequential occupation of different urinary tract tissues. While the role of type 1 fimbriae during infection has been clear, the role of P fimbriae must be further defined to assert the relevance of coordinated regulation in vivo. Therefore, we examined the ability of P fimbrial isogenic mutants, constructed in a type 1 fimbrial-negative background, to compete in the murine urinary tract over a period of 168 h. No differences in the colonization of these mutants were observed. However, comparison of these results with previous studies suggests that inversely coordinated expression of adhesin gene clusters does occur in vivo. Interestingly, the mutant that was incapable of expressing either type 1 or P fimbriae compensated by synthesizing F1C fimbriae.

FIG. 1.

Microarray analysis of the fim and pap gene clusters in E. coli CFT073 type 1 fimbrial phase-locked mutants. The signal intensity, corresponding to the relative expression of a gene, is shown for genes encoding type 1 fimbriae (fim) and both copies of P fimbriae (pap). Genes upregulated (⊕) or downregulated (⊖) in E. coli CFT073-ON relative to CFT073-OFF are indicated.

FIG. 2.

qRT-PCR analysis of papA_2 expression in E. coli CFT073 and type 1 fimbrial phase-locked mutants. The black bars represent the change (n-fold) in gene expression of papA_2 between E. coli CFT073-OFF, CFT073-ON, and wild-type CFT073. Changes (n-fold) were calculated using CFT073-OFF as the relative measure of comparison.

FIG. 3.

Transmission electron microscopy of E. coli CFT073 fim pap. A. E. coli CFT073 fim pap, a mutant which does not express type 1 or P fimbriae, remains capable of other fimbrial production. B. This strain was most often observed in large clusters of fimbriated bacteria when grown in vitro.

FIG. 4.

Coinoculation of E. coli CFT073 fim pap and CFT073-OFF in the murine model of ascending UTI. E. coli CFT073 fim pap and CFT073-OFF were inoculated together into the murine urinary tract. At the indicated times postinoculation, mice (n = 8) were sacrificed and the level of colonization was determined for each strain in the urine (A), in the bladder (B), and in the kidneys (C). Individual counts of strain CFT073-OFF are represented by squares (□), and individual counts of strain CFT073 fim pap are represented by circles (○). Bars, representing the median counts, are connected by a solid line in CFT073-OFF, and a dotted line in CFT073 fim pap.

FIG. 5.

F1C fimbriae expressed in static cultures of E. coli CFT073 fim pap. SDS-polyacrylamide gel electrophoresis was used to analyze the protein profiles of fimbrial preparations from concentrated static and aerated (exponential phase) Luria broth cultures of E. coli strain CFT073, UPEC76, and CFT073 fim pap. The protein ladder is shown on the left (kDa). The result of BLASTP analysis of the N-terminal sequence of the protein of small size (∼10 kDa) observed in the fimbrial preparation from a static culture of CFT073 fim pap is shown below the gel. This protein was absent in the fimbrial preparations from static cultures of wild-type CFT073 and UPEC76.