An in-vitro model for studying the interaction of Escherichia coli O157:H7 and other enteropathogens with bovine primary cell cultures

Abstract

Sections of kidney, trachea, ileum, colon, rectum and rumen were removed at post mortem from a neonatal calf and, with the exception of the rumen, primary cell lines were established for each of the cell types. The adherence of enterohaemorrhagic Escherichia coli (EHEC) serotype O157:H7, enteropathogenic E. coli (EPEC) serotype O111, E. coli K12 (a laboratory adapted non-pathogenic strain) and Salmonella enterica serotype Typhimurium was assayed on each cell type. For all adherence assays on all cell lines, EHEC O157:H7 adhered to a signi®cantly greater extent than the other bacteria. S. Typhimurium and EPEC O111 adhered to a similar extent to one another, whereas E. coli K12 was signi®cantly less adherent by 100-fold. In all cell types, >10% of adherent S. Typhimurium bacteria invaded, whereas c. 0.01±0.1% of adherent EHEC O157:H7 and EPEC O111 bacteria invaded, although they are regarded as non-invasive. EHEC O157 generated actin re-arrangements in all cell types as demonstrated by¯uorescent actin staining (FAS) under densely packed bacterial micro-colonies. EPEC O111 readily generated the localised adherent phenotype on bovine cells but generated only densely packed micro-colonies on HEp-2 cells. The intensity of actin re-arrangements induced in bovine cells by EPEC O111 was less than that induced by EHEC O157:H7. The intimate attachment on all cell types by both EHEC O157:H7 and EPEC O111 was clearly demonstrated by scanning electron microscopy.

Preliminary characterization of jejunocyte and colonocyte cell lines isolated by enzymatic digestion from adult and young cattle

“…The cells were derived following the adaptation of an enzymatic digestion approach (Dibb-Fuller et al, 2001;Föllmann et al, 2000) coupled to a mechanical dissection, which had been previously successful in yielding primocultures of both jejunocytes and colonocytes from adult cattle (Rusu et al, 2005). Immortalization was achieved by transfection with the tumour large T antigen of the simian leukaemia virus 40 (SV40 LT).…”

A rapid, sensitive method for testing the activity of Escherichia coli 0157:H7 secreted cytotoxins against epithelial cells from the jejunum and descending colon of cattle

“…A number of studies have shown that specific type III effector molecules are required for the formation of classic attaching and effacing lesions in calves (Cookson and Woodward 2003); however, not all colonization is associated with the formation of actin pedestals (Vlisidou et al 2006). In addition, EHEC O157:H7 actin pedestals do not have the same appearance in cattle as those in disease-susceptible animals (Dibb-Fuller et al 2001), and a recent study suggests that this is attributable to lower expression of type III effector molecules in cattle (Rashid et al 2006). In contrast, Rashid et al (2006) determined that Shiga toxin expression is not different for human and cattle systems suggesting that cytotoxins could play a similar role in both systems.…”

Clearance of Escherichia coli O157:H7 Infection in Calves by Rectal Administration of Bovine Lactoferrin

“…E. coli O157:H7 strain NCTC12900, a well-characterized Shiga toxin-negative EHEC strain of human origin (15), was used for experimental infections in calves. We used this Shiga toxin-negative strain for biosafety reasons.…”