Interaction of Salmonella enterica with Bovine Epithelial Cells Demonstrates Serovar-Specific Association and Invasion Patterns.

Dairy cows are known reservoirs of Salmonella enterica and human salmonellosis has been attributed to the consumption of contaminated dairy and beef products as well as poultry meat and eggs. Although many S. enterica serovars are known to colonize the gastrointestinal tract of cattle, the interactions between dairy commensal (or persistent) and transient Salmonella serovars with bovine epithelial cells are not well understood. Association-invasion assays were used to characterize the interactions of 26 S. enterica strains from bovine origins, comprising serovars Anatum, Cerro, Dublin, Give, Kentucky, Mbandaka, Meleagridis, Montevideo, Muenster, Newport, Oranienburg, Senftenberg, and Typhimurium, with cultured bovine epithelial cells. There were significant differences in the association with and invasion of bovine epithelial cells within and across Salmonella serovars (Tukey's Honestly Significant Difference test, p < 0.05). Salmonella enterica serovar Dublin strains were the most invasive, whereas Kentucky, Mbandaka, Cerro, and Give strains were the least invasive (p < 0.05). Significant differences in motility on semisolid medium were also observed between strains from different serovars. Findings from this study demonstrate an underappreciated level of phenotypic diversity among Salmonella strains within and across serovars and serve as a baseline for future studies that may identify the molecular mechanisms of asymptomatic Salmonella carriage and bovine salmonellosis.

Diversity of Extended-Spectrum Cephalosporin-Resistant Escherichia coli in Feces from Calves and Cows on Pennsylvania Dairy Farms.

The global incidence of human infections associated with extended-spectrum ß-lactamase (ESBL)-producing Escherichia coli is increasing. Dairy animals are reservoirs of ESBL-producing E. coli, especially, third-generation cephalosporin (3GC)-resistant strains. To further understand the diversity of 3GC-resistant E. coli across animals of different age groups (e.g., pre- and postweaned calves, lactating cows, and dry cows) and farms, we used pulsed-field gel electrophoresis (PFGE) to characterize 70 fecal isolates from 14 dairy farms located in nine Pennsylvania counties. Results of this analysis indicated that 3GC-resistant E. coli were highly diverse and grouped into 27 PFGE clades (80% similarity cutoff) and 24 unique antimicrobial resistance patterns were observed among the isolates. For eight farms, clonal E. coli with the same resistance patterns were isolated from two or more age groups, indicating that strains were carried in both the calves and adult cows within the same herd. However, there were also several isolates with the same resistance pattern that were distributed to different clades, including isolates from different animal age groups on the same farm, suggesting different strains of E. coli within a farm harbored the same resistance-conferring elements. Results of this analysis indicated that 3GC-resistant E. coli were highly diverse, associated with multidrug resistance, and circulated through different (noncommingled) animal groups on individual farms.

Prevalence of Salmonella enterica, Listeria monocytogenes, and pathogenic Escherichia coli in bulk tank milk and milk filters from US dairy operations in the National Animal Health Monitoring System Dairy 2014 study.

The dairy farm environment is a well-documented reservoir for zoonotic pathogens such as Salmonella enterica, Shiga-toxigenic Escherichia coli, and Listeria monocytogenes, and humans may be exposed to these pathogens via consumption of unpasteurized milk and dairy products. As part of the National Animal Health Monitoring System Dairy 2014 study, bulk tank milk (BTM, n = 234) and milk filters (n = 254) were collected from a total of 234 dairy operations in 17 major dairy states and analyzed for the presence of these pathogens. The invA gene was detected in samples from 18.5% of operations and Salmonella enterica was isolated from 18.0% of operations. Salmonella Dublin was detected in 0.7% of operations. Sixteen Salmonella serotypes were isolated, and the most common serotypes were Cerro, Montevideo, and Newport. Representative Salmonella isolates (n = 137) were tested against a panel of 14 antimicrobials. Most (85%) were pansusceptible; the remaining were resistant to 1 to 9 antimicrobials, and within the resistant strains the most common profile was resistance to ampicillin/clavulanic acid, ampicillin, cefoxitin, ceftiofur, ceftriaxone, chloramphenicol, streptomycin, sulfisoxazole, and tetracycline. Listeria spp. were isolated from 19.9% of operations, and L. monocytogenes was isolated from 3.0% of operations. Serogroups 1/2a and 1/2b were the most common, followed by 4b and 4a. One or more E. coli virulence genes were detected in the BTM from 30.5% of operations and in the filters from 75.3% of operations. A combination of stx2, eaeA, and γ-tir genes was detected in the BTM from 0.5% of operations and in the filters from 6.6% of operations. The results of this study indicate an appreciable prevalence of bacterial pathogens in BTM and filters, including serovars known to infect humans.

Diversity of Listeria monocytogenes within a U.S. dairy herd, 2004-2010.

Listeria monocytogenes, the causative agent of listeriosis, is frequently isolated from the environment. Dairy cows and dairy farm environments are reservoirs of this pathogen, where fecal shedding contributes to its environmental dispersal and contamination of milk, dairy products, and meat. The molecular diversity of 40 L. monocytogenes isolates representing 3 serogroups (1/2a, 1/2b, and 4b) collected between 2004 and 2010 from the feces of dairy cattle on a single dairy farm was assessed using a multivirulence locus sequence typing (MVLST) assay. The dairy farm L. monocytogenes MVLST patterns were compared to those from 138 strains isolated globally from clinical cases, foods, and the environment. Results of the study demonstrated that several distantly related L. monocytogenes strains persisted among members of the herd over the course of the study while other strains were transient. Furthermore, some strains isolated during this study appear to be distantly related to previously isolated L. monocytogenes while others are closely related to Epidemic Clones associated with human illness. This work demonstrates that dairy cows can be reservoirs of a diverse population of potentially human pathogenic L. monocytogenes that represents a risk to consumers of milk, dairy products, and meat.

Inactivation of Escherichia coli, Salmonella enterica, and Listeria monocytogenes using the Contamination Sanitization Inspection and Disinfection (CSI-D) device.

The Contamination Sanitization Inspection and Disinfection (CSI-D) device is a handheld fluorescence-based imaging system designed to disinfect food contact surfaces using ultraviolet-C (UVC) illumination. This study aimed to determine the optimal CSI-D parameters (i.e., UVC exposure time and intensity) for the inactivation of the following foodborne bacteria plated on non-selective media: generic Escherichia coli (indicator organism) and the pathogens enterohemorrhagic E. coli, enterotoxigenic E. coli, Salmonella enterica, and Listeria monocytogenes. Each bacterial strain was spread-plated on non-selective agar and exposed to high-intensity (10 mW/cm2) or low-intensity (5 mW/cm2) UVC for 1-5 s. Control plates were not exposed to UVC. The plates were incubated overnight at 37 °C and then enumerated. Three trials for each bacterial strain were conducted. Statistical analysis was carried out to determine if there were significant differences in bacterial growth between UVC intensities and exposure times. Overall, exposure to low or high intensity for 3-5 s resulted in consistent inhibition of bacterial growth, with reductions of 99.9-100 % for E. coli, 96.8-100 % for S. enterica, and 99.2-100 % for L. monocytogenes. The 1 s exposure time showed inconsistent results, with a 66.0-100 % reduction in growth depending on the intensity and bacterial strain. When the results for all strains within each species were combined, the 3-5 s exposure times showed significantly greater (p < 0.05) growth inhibition than the 1 s exposure time. However, there were no significant differences (p > 0.05) in growth inhibition between the high and low UVC intensities. The results of this study show that, in pure culture conditions, exposure to UVC with the CSI-D device for ≥3 s is required to achieve consistent reduction of E. coli, S. enterica, and L. monocytogenes.