John H. Rupnow

Behavior of hemorrhagic Escherichia coli O157:H7 during the manufacture of cottage cheese

The ability of enterohemorrhagic Escherichia coli O157:H7 to grow and survive during the manufacture of Cottage cheese was determined. Pasteurized skim milk artificially contaminated with E. coli O157:H7 was used to make Cottage cheese by the washed curd method. E. coli O157:H7 was enumerated by surface plating samples on MacConkey sorbitol agar with 5-bromo-4-chloro-3-indoxyl-β-D-glucuronic acid cyclohexylammonium salt (MSA-BCIG) and incubating at 42°C for 24 h. The heat treated samples were previously inoculated into a modified EC broth with novobiocin and incubated static at 35°C for 24 h. Sorbitol and β-glucuronidase negative colonies were picked from MSA-BCIG, spread on Levine eosin methylene blue agar plates and phenol red sorbitol agar plates with 4-methylumbelliferyl-β-D-glucuronide (PRS-MUG) added for confirmation. E. coli O157:H7 increased 100-fold in numbers during the manufacturing process, but death occurred during cooking of the curd and whey. The pH and acidity did not halt the growth of this pathogen during the manufacture of the cheese; furthermore, the values of these parameters were the same between the contaminated and control samples.

Adherence Inhibition of Enteropathogenic Escherichia coli by Chitooligosaccharides with Specific Degrees of Acetylation and Polymerization

Some oligosaccharides are known to act as molecular decoys by inhibiting pathogen adherence to epithelial cells. The present study was aimed at analyzing whether chitooligosaccharides (CHOS), that is, oligomers of D-glucosamine and N-acetyl-D-glucosamine, have such antiadherence activity. CHOS of varied degree of polymerization (DP) and fraction of acetylation (F(A)) were produced. Adherence of enteropathogenic Escherichia coli (EPEC) to the surface of a human HEp-2 cell line was determined in the absence or presence of the various CHOS fractions. Adherence was assessed by microscopic counting and image analysis of bacterial clusters and cells. The results showed that all CHOS fractions inhibited adherence of EPEC to HEp-2 cells. Hydrolysates with lower F(A) were more effective at reducing adherence. This effect is greater than that obtained with other oligosaccharides, such as galactooligosaccharides, applied at the same concentrations.

Adherence Reduction of Campylobacter jejuni and Campylobacter coli Strains to HEp-2 Cells by Mannan Oligosaccharides and a High-Molecular-Weight Component of Cranberry Extract.

Campylobacter infections are a leading cause of human bacterial gastroenteritis in the United States and are a major cause of diarrheal disease throughout the world. Colonization and subsequent infection and invasion of Campylobacter require that the bacteria adhere to the surface of host cells. Agents that inhibit adherence could be used prophylactically to reduce Campylobacter carriage and infection. Mannan oligosaccharides (MOS) have been used as a feed supplement in livestock animals to improve performance and to replace growth-promoting antibiotics. However, MOS and other nondigestible oligosaccharides may also prevent pathogen colonization by inhibiting adherence in the gastrointestinal tract. In addition, plant extracts, including those derived from cranberries, have been shown to have antiadherence activity against pathogens. The goal of this study was to assess the ability of MOS and cranberry fractions to serve as antiadherence agents against strains of Campylobacter jejuni and Campylobacter coli. Adherence experiments were performed using HEp-2 cells. Significant reductions in adherence of C. jejuni 29438, C. jejuni 700819, C. jejuni 3329, and C. coli 43485 were observed in the presence of MOS (up to 40 mg/ml) and with a high-molecular-weight fraction of cranberry extract (up to 3 mg/ml). However, none of the tested materials reduced adherence of C. coli BAA-1061. No additive effect in adherence inhibition was observed for an MOS-cranberry blend. These results suggest that both components, MOS and cranberry, could be used to reduce Campylobacter colonization and carriage in livestock animals and potentially limit human exposure to this pathogen.