Mutations in the csgD Promoter Associated with Variations in Curli Expression in Certain Strains of Escherichia coli O157:H7

Abstract

Single-base-pair csgD promoter mutations in human outbreak Escherichia coli O157:H7 strains ATCC 43894 and ATCC 43895 coincided with differential Congo red dye binding from curli fiber expression. Red phenotype csgD::lacZ promoter fusions had fourfold-greater expression than white promoter fusions. Cloning the red variant csgDEFG operon into white variants induced the red phenotype. Substrate utilization differed between red and white variants.Many Escherichia coli organisms and salmonellae express coiled surface appendages, known as curli fibers and thin, aggregegative fimbriae, respectively, that are typically produced under stressful environmental conditions, such as low temperature, low osmolarity, and stationary growth (3, 9, 10). Curli fibers bind fibronectin, laminin, certain serum proteins, and Congo red dye (4,8,9,18). Two divergently transcribed operons are required for curli expression: csgBA encodes the curli subunit protein (CsgA) and a nucleator protein (CsgB); csgDEFG encodes a transcriptional regulator (CsgD), an outer membrane lipoprotein (CsgG), and two putative curli assembly factors (CsgE and CsgF). Transcription from the csgBA promoter requires csgD expression; both operons require stationary-phase sigma factor ( s ) for expression (1, 4, 10). Expression of thin, aggregative fimbriae in Salmonella enterica serovar Typhimurium is regulated by a similar agf operon (13).Curli expression has not been reported for enterohemorrhagic E. coli (EHEC) O157:H7, the most common Shigatoxigenic serotype associated with human disease (11). In order to identify potential factors involved in this pathogen's survival and persistence outside of the mammalian host, we screened 49 diverse bovine and human E. coli O157:H7 isolates for curli expression on Congo red indicator (CRI) plates after 48 h at 28°C (5). The 41 bovine isolates were from infected beef calves in five states (6). The eight human-associated isolates were American Type Culture Collection (ATCC, Rockville, Md.) strains ATCC 35150, ATCC 43888, ATCC 43889, ATCC 43890, ATCC 43894, and ATCC 43895 and Washington state strains Tarr4A and Tarr1A (2, 19). All of the bovine and six of the human isolates displayed smooth, moist, white colonies typical of the curli-deficient E. coli strain HB101 on CRI plates (9). However, strains ATCC 43894 and ATCC 43895 displayed mixed red and white colonies. Red colonies were dry, rough, and curliated as verified by electron microscopy (results not shown). Red and white colonies retained their parental phenotypes when subcultured on CRI plates with or without 1% NaCl and at either 28 or 37°C, suggesting that curli expression was neither low-temperature nor low-osmolarity dependent. Red variants passaged daily (1:100) in Luria-Bertani broth (Difco Laboratories, Detroit, Mich.) at 37°C generated mixed red and white phenotypes in as few as 3 passages, with white variants persisting at 40 to 60% of total colonies over 10 passages. White variants were stable under all growth conditions tested except for one, strain ATCC 4389...

Comparative Genetics of the rdar Morphotype inSalmonella

“…For Sarc16, the change was in the Ϫ35 region and shifted the PagfD sequence closer to the 70 consensus (39). In E. coli, enhanced Tafi (curli) production has been linked to PagfD (PcsgD) mutations in the Ϫ10 region that move the sequence closer to the 70 consensus (62). The changes that occurred in Sarc4 and Sarc16 were also predicted to allow for 70 -based transcription of agfD.…”

Differential Binding of Escherichia coli O157:H7 to Alfalfa, Human Epithelial Cells, and Plastic Is Mediated by a Variety of Surface Structures

“…(40,49). Both genera have curli genes; however, it has been reported that single-base-pair csgD promoter mutations leave Ն95% of E. coli O157:H7 cells unable to express curli (47). These mutations also cause E. coli O157:H7 to display increased virulence in mice and increased invasion of HEp-2 cells (48).…”

“…coli O157:H7 expresses multiple fimbrial and nonfimbrial adhesins which have no obvious role in pathogenesis and may be involved in adhesion to other surfaces (reviewed in reference 46). For example, curli-expressing thin aggregative fim-briae, which are rarely reported in E. coli O157:H7 compared with other pathogenic E. coli strains (47,48), have been reported to bind eukaryotic extracellular matrix proteins as well as to enhance the formation of E. coli O157:H7 biofilms on inert surfaces (11,38). Biofilm formation may increase E. coli O157:H7 survival and would likely result in protection against many environmental conditions.…”

Expression of cellulose and curli fimbriae by Escherichia coli isolated from the gastrointestinal tract

“…In contrast, enteroinvasive E. coli (EIEC), Shigella spp. and enterohaemorrhagic E. coli (EHEC) regularly do not express curli fimbriae in vitro (Bian et al, 2000;Cookson et al, 2002;Patri et al, 2000;Sakellaris et al, 2000;Sjöbring et al, 1994;Uhlich et al, 2001).…”