Article Figures & Data
Figures
- FIG. 1.
Light microscopy micrographs of ileal explants inoculated with the wild-type O45 strain 86-1390 (A) or with the LEE-negative strain 862 (B). Strain 86-1390 showed a typical intimate-adherence pattern (arrowheads) with irregularity of the associated epithelial cells, whereas a loose association of bacteria with the intestinal mucosa of some villi with no obvious change in associated epithelial cells (arrow) was observed for negative-control strain 862. Magnification, ×400.
- FIG. 2.
Adherence of wild-type strains and their paa mutant strains. (A) paa mutant strain M155 (n = 18) showed a decreased number of intact ileal villi with bacterial adherence to epithelial cells, compared to wild-type PEPEC strain 86-1390 (n = 12) and to the complemented mutant strain M155c (n = 20). The porcine strain 862 (n = 15), which does not possess the LEE, was used as a negative control. (B) paa mutant strain E22Δpaa (n = 19) showed a decreased number of intact ileal villi compared to wild-type REPEC strain E22 (n = 19) and to the complemented mutant strain E22c (n = 10). Error bars, standard deviations of the means. Asterisk, statistically significant difference (P < 0.0001, when compared by Kruskal-Wallis test) from wild-type strains 86-1390 (A) and E22 (B).
- FIG. 3.
Transmission electron micrographs of ileal explants inoculated with the wild-type O45 strain 86-1390 (A; magnification, ×20,664), the complemented mutant strain M155c (B; magnification, ×20,702), or TnphoA mutant M155 (C; magnification, ×13,500). Typical A/E lesions were observed for both wild-type and complemented-mutant strains, whereas bacteria in the lumen without any direct contact with the epithelium were observed for the mutant M155.
- FIG. 4.
The nucleotide sequence of paa and its flanking sequences and the deduced amino acid sequences. The putative −10 and −35 promoter sites, the ribosome binding site (RBS), and the putative transcription terminator are underlined. The translation initiation codon and the TAG translation termination codon are in boldface. Vertical arrow, potential peptide signal cleavage site; arrowhead, insertion site of TnphoA.
- FIG. 5.
Alignment of the deduced amino acid sequences of the Paa of the 86-1390 strain and the Paa proteins of the O157:H7 EDL933 and Sakai E. coli strains, the AcfC protein of V. cholerae, and the PEB3 protein of C. jejuni. ∗, identical or conserved residues in all sequences in the alignment; colons, conserved substitutions; periods, semiconserved substitutions. The amino acid sequence alignment was performed with the Clustal W program.
- FIG. 6.
Reduction of the percentage of intact villi showing intimate adherence when pig ileal explants are inoculated with PEPEC strain 86-1390 following treatment with anti-Paa antibodies, compared to percentages for explants inoculated with strain 86-1390 following treatment with antibodies from hens immunized with a sonicate preparation from host strain M15(pREP4) (T −). The porcine strain 862, which does not have the LEE, was used as a negative control. Asterisk, statistically significant difference (P < 0.0001, when compared by Kruskal-Wallis tests) from the T (−) treatment.
- FIG. 7.
Transmission electron micrographs showing the uniform distribution of immunogold-labeled Paa protein (arrow) over the bacterial surface of the complemented strain M155c (A) following overnight growth at 37°C in TSB. When anti-Paa serum was adsorbed against the Paa protein, only a few gold beads were observed for strain M155c, mostly in the background (B). Bars = 300 nm.
Tables
- TABLE 2.
Clinical and histopathological findings in piglets inoculated with porcine E. coli O45 isolates
Straine No. of pigs with diarrhea/no. inoculated Onset of diarrhea (h)b Extent of A/E lesionsd Presence of paa gene 81-4420 2/2 24 ++++ + 86-1390 2/2 30 ++++ + 91-19-172 2/2 35 ++++ + 90-2061 2/2 39 +++ + 90-1513 2/2 41 +++ + 88-4299 1/1a 44 ++ + 86-4733 2/2 70 + + 83-2315 2/2 83 ++ − 88-1861 1/1a 91 − − 89-56-196 1/2 96 − − 82-4378 0/2 NMc + − 81-1786 1/2 85 − − ↵ a One piglet died of causes unrelated to the infection within 20 h after birth.
↵ b Mean time of onset of diarrhea p.i.
↵ c NM, nonmeasurable data.
↵ d ++++, extensive bacterial colonization and severe effacement of microvilli; +++, large areas of bacterial colonization and heavy effacement; ++, focal lesions; +, small scattered focal lesions; −, no lesions observed.
↵ e Strains were eae and LEE positive, except for 81-1786, which was a control eae-negative strain.
