Donnenberg Lab Images
Fig. 1.  Scanning electron micrograph showing microcolonies
of EPEC displaying the characteristic localized adherence
pattern of adherence to HEp-2 cells.

 
Fig. 2.  High power scanning electron micrograph of EPEC displaying localized adherence to
HEp-2 cells.  Note the elongated microvilli to which the bacteria appear to attach.

 
Fig. 3.  Transmission electron micrograph of negatively-
stained EPEC.  Arrow shows aggregates of bundle-forming pili.

 
Fig. 4.  Map of the bfp gene cluster encoding the bundle-forming pilus.
Putative functions that have been assigned to gene products are shown.

 
 Prepilin model


Fig. 5.  Model for the early steps in the biogenesis of the bundle-forming pilus.  The nascent pre-pilin (pre-bundlin) exists as a cytoplasmic transmembrane protein that is processed simultaneously and independently by the BfpP pre-pilin peptidase, which cleaves off the signal sequence and N-methylates the mature amino terminus (triangle), and by the periplasmic oxidoreductase DsbA, which catalyzes the formation of a disulfide bond that is necessary for bundlin stability.

 

 

 

 

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Fig. 6.  Time-lapse photography of auto-aggregates of wild type (left) or bfpF mutant (right) EPEC bacteria.  The wild type bacteria disaggregate, while the bfpF mutant auto-aggregates remain fixed.  It is believed that BfpF generates the energy required for pilus retraction, which leads to disaggregation.
 
BfpB EM
Fig. 7.  Electron micrograph of purified BfpB showing ring-shaped mutimers.  BfpB is an outer membrane lipoprotein that is believed to form the channel through which the bundle-forming pilus protrudes.


















 
LEE
Fig. 8.  The Locus of Enterocyte Effacement (LEE) pathogenicity island, which consists of 41 open reading frames, is necessary and sufficient for the attaching and effacing effect.  Genes shown in black have unknown function and are designated either as ORFs or rORFs depending on orientation.  Genes shown in dark blue are esc genes, which encode homologues of the Yersinia type III secretion system.  Genes shown in light blue are the sep genes, which encode components of the type III secretion system that lack Yersinia homologues.  The genes in red encodes the outer membrane adhesin intimin.  Genes shown in teal blue encode chaperones of secreted proteins.  Genes shown in green encode esp or other secreted proteins, as indicated.  The positive regulator of the LEE is shown in olive.

 
 Fig. 9.  Laser-scanning confocal micrograph of EPEC interacting with HeLa cells.  Bacteria and host cell nuclei are labeled with DAPI and appear blue.  Actin is labeled with FITC-phalloidin and appears green.  Not the large cluster of bacteria toward the left (localized adherence) and the individual bacteria that are located at the tip of actin rich pedestals.
Fig. 10.  Transmission electron micrograph showing the attaching and effacing effect.  EPEC have effaced microvilli and are intimately attached to the surface of the HEp-2 cell which responds to the bacteria by forming the typical cup shaped pedestals that embrace the bacteria.
 Fig. 11.  Laser-scanning confocal micrograph of EPEC interacting with HeLa cells.  The image is a composite of a Nomarsky optical image in which the cells and bacteria are visible, an FITC-phalloidin stain, in which actin is labeled green, and an affinity-purified EspB antibody visualized with rhodamine secondary antibody,which labels the EspB protein red.  EspB is visible within the host cell adjacent to bacteria involved in attaching and effacing.
Fig. 12.  A composite image derived from 18 optical sections taken at 0.5 micron intervals through a HeLa cell that had been infected with EPEC and stained with affinity-purifed EspB antibodies.  The images were reassembled and are color coded so that those furthest from the camera are red, those closest are violet and intermediate sections are orange, yellow, green and blue.  These images demonstrate that the EspB protein is distributed throughout the cytoplasm of the host cell.

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