Isolation and separation of physicochemically distinct fimbrial types expressed on a single culture of Escherichia coli O7:K1:H6

Abstract

The fimbrial (pili) profile of a single strain of Escherichia coli 07:K1:H6 (WF96) was evaluated. Fimbriae were isolated by sucrose density gradient ultracentrifugation, purified from flagellae by the use of 0.4% sodium dodecyl sulfate (SDS), and separated into distinct fimbrial types. Analysis of the purified WF96 fimbriae by SDS-polyacrylamide gel electrophoresis revealed two polypeptide bands with molecular weights of 16,000 and 21,000. Treatment of the fimbrial mixture with saturated guanidine hydrochloride resulted in the appearance of a third band with a molecular weight of 19,500. The relative susceptibilities of the WF96 fimbrial types to disrupting chemicals (octyl-glucoside, urea, SDS, and guanidine hydrochloride) were assessed by exposure of the fimbrial mixture to each agent, separation of the depolymerized fimbriae from intact fimbriae by gel filtration on Sepharose CL-4B, and identification of the disaggregated fimbrial types by SDS-polyacrylamide gel electrophoresis of column fractions. The physicochemical heterogeneity of the three fimbrial types coexpressed on WF96 was exploited to develop a method for separation of individual fimbriae.

Three-dimensional structure of fimbriae determines specificity of immune response

“…Even among fimbriae that belong to the same group, such as K88, antigenic diversity exists. Moreover, a single culture of a strain of E. coli can express up to three or four fimbrial types as demonstrated by 0rskov et al (23) and Korhonen et al (16) and in our laboratory (12). The complexity of analysis of fimbrial heterogeneity is compounded by the finding that several E. coli fimbrial types, e.g., type 1 fimbriae (2, 4) and P-fimbriae (25), undergo phase variation.…”

“…E. coli D182 (not typable) and D539 (not typable) were originally obtained from J. P. Duguid, Dundee University. Strain B875 is a K-12 strain, and strain WF96 (07:K1:H6) has been previously described (12). The hemagglutination types of these strains, according to the Evans hemagglutination scheme (8), were as follows: D182, type V; D539, type VII; B875, type IIIa; and WF95, type VIIc.…”

A plasmid of enterohemorrhagic Escherichia coli O157:H7 is required for expression of a new fimbrial antigen and for adhesion to epithelial cells

“…The protein content of fimbriae was determined as described by Markwell et al (24). Fimbrial samples were pretreated with hot acid as described previously (15). SDS-PAGE.…”

“…SA), whereas the nonfimbriate, plasmid-cured strain did not (Fig. SB) (15) were run on this gel for comparison (lane 4). The positions of molecular weight markers purchased as a mixture from Pharmacia (low-molecular-weight protein kit) containing phosphorylase b (94K), bovine serum albumin (67K), ovalbumin (43K), carbonic anhydrase (30K), soybean trypsin inhibitor (20.1K), and a lactalbumin (14.4K) are indicated on the left.…”

Specific and non-specific interactions in bacterial adhesion to solid substrata

“…Several of the surface components have been recognized as molecular probes reacting stereochemically with molecules on the opposing surface and are therefore termed adhesins, whereas other components significantly affect the macroscopic properties of the bacterial cell. Both gram-negative and gram-positive bacteria have been shown to possess a variety of surface layers and surface appendages of different structural and chemical nature [13] and having various physico-chemical properties [22,24,25]. Surface appendages, by virtue of their small radius, are believed to bridge the distance between the cell and the adhesion substratum and to counteract electrostatic repulsion [16].…”