Binding to and killing of human renal epithelial cells by hemolytic P-fimbriated E. coli
Related Experiment Videos
A L Trifillis1, M S Donnenberg, X Cui
1Department of Pathology, University of Maryland School of Medicine, Baltimore.
Abstract:
Acute pyelonephritis is a common invasive infection frequently caused by E. coli that possess P-fimbriae and secrete hemolysin. We have examined the role of P fimbriae and hemolysin in the killing of putative target cells of acute pyelonephritis, that is, human renal epithelial cells (HRPTEC). Cultures of HRPTEC were overlaid with (1) a prototypic pyelonephritogenic E. coli (CFT073) which expresses both P fimbriae and hemolysin; (2) its hemolysin-negative isogenic mutant (CFT073hlyD::TnphoA); or (3) a prototypic nonpyelonephritogenic fecal E. coli (FN414) which is negative for both P fimbriae and hemolysin. CFT073 and CFT073hlyD::TnphoA but not FN414 adhered to HRPTEC, as demonstrated by electron microscopy and direct counting. Adherence was diminished by antisera directed against P fimbriae and by a monoclonal antibody recognizing the epithelial receptor for P fimbriae. CFT073 was significantly more cytolethal for HRPTEC than its hemolysin-negative mutant. The bacteria-free filtrate of CFT073 was both hemolytic and cytolethal whereas that of CFT073hyD::TnphoA was not hemolytic and was significantly less cytolethal. Finally, we demonstrated that CFT073 passed through monolayers of HRPTEC at a higher rate than CFT073hlyD::TnphoA, indicating that hemolysin damages HRPTEC, facilitating passage of bacteria through the epithelial barrier. With HRPTEC and a pyelonephritogenic strain of E. coli we have reproduced in vitro bacterial attachment and toxin delivery by P fimbriae and hemolysin, factors epidemiologically associated with acute pyelonephritis in patients.
Insights
P-fimbriae mediate E. coli attachment to human renal epithelial cells (HRPTEC). Hemolysin secreted by E. coli damages HRPTEC, facilitating bacterial invasion and contributing to acute pyelonephritis pathogenesis.
Area of Science:
- Microbiology
- Urology
- Infectious Diseases
Background:
- Acute pyelonephritis is a common kidney infection.
- E. coli strains causing pyelonephritis often possess P-fimbriae and hemolysin.
- The roles of these virulence factors in kidney infection are not fully understood.
Purpose of the Study:
- To investigate the role of P-fimbriae and hemolysin in E. coli interaction with human renal epithelial cells (HRPTEC).
- To determine how these factors contribute to the pathogenesis of acute pyelonephritis.
Main Methods:
- Co-culture of HRPTEC with pyelonephritogenic E. coli (CFT073), its hemolysin-negative mutant, and non-pyelonephritogenic E. coli (FN414).
- Assessment of bacterial adherence using electron microscopy and direct counting.
- Evaluation of bacterial cytolethality and filtrate hemolytic/cytolethal activity.
- Measurement of bacterial passage through HRPTEC monolayers.
Main Results:
- P-fimbriae mediated adherence of CFT073 and its mutant to HRPTEC.
- CFT073 was significantly more cytolethal to HRPTEC than its hemolysin-negative mutant.
- Hemolysin in CFT073 filtrate caused significant cytotoxicity and hemolysis.
- Hemolysin facilitated bacterial passage through HRPTEC monolayers, indicating epithelial barrier damage.
Conclusions:
- P-fimbriae are crucial for E. coli attachment to renal epithelial cells.
- Hemolysin damages HRPTEC, promoting bacterial invasion and contributing to acute pyelonephritis.
- This study elucidates the in vitro mechanisms of bacterial attachment and toxin-mediated damage in pyelonephritis.