WGS can provide comprehensive resistance genotypes and is capable of accurately predicting resistance phenotypes, making it a valuable tool for surveillance. Moreover, the data presented here showing the ability to accurately predict resistance suggest that WGS may be used as a screening tool in selecting anti-infective therapy, especially as costs drop and methods improve.

Bacteriophage CEV1 was isolated from sheep resistant to Escherichia coli O157:H7 colonization. In vitro, CEV1 efficiently infected E. coli O157:H7 grown both aerobically and anaerobically. In vivo, sheep receiving a single oral dose of CEV1 showed a 2-log-unit reduction in intestinal E. coli O157:H7 levels within 2 days compared to levels in the controls.

Escherichia coli O157:H7 can live undetected in the gut of food animals and be spread to humans directly and indirectly. Bacteriophages are viruses that prey on bacteria, offering a natural, nonantibiotic method to reduce pathogens from the food supply. Here we show that a cocktail of phages isolated from commercial cattle feces reduced E. coli O157:H7 populations in the gut of experimentally inoculated sheep. A cocktail of phages was used in order to prevent the development of resistance to the phages. In our first in vivo study we found that our cocktail of phages reduced E. coli O157:H7 populations in the feces of sheep (p < 0.05) by 24 hours after phage treatment. Upon necropsy, populations of inoculated E. coli O157:H7 were reduced by phage treatment in both the cecum (p < 0.05) and rectum (p < 0.1). In our second in vivo study, several ratios of phage plaque-forming units (PFU) to E. coli O157:H7 colony-forming units (CFU) were used (0:1, 1:1, 10:1, and 100:1 PFU/CFU) to determine the most efficacious phage dose. A 1:1 ratio of phage to bacteria was found to be more effective (p < 0.05) than either of the higher ratios used (10:1 or 100:1). Ruminal levels of E. coli O157:H7 were not significantly reduced (p > 0.10) in any of the studies due to relatively low inoculated E. coli O157:H7 ruminal populations. Our results demonstrate that phage can be used as a preharvest intervention as part of an integrated pathogen reduction scheme.

Aims:  To examine the variability in faecal shedding of Salmonella and Escherichia coli O157:H7 in healthy lactating dairy cattle and to evaluate the genetic relatedness of Salmonella isolates. Methods:  Faecal samples were obtained from lactating Holstein dairy cattle on four commercial farms in the southwestern US. All farms were within an 8‐km radius and were sampled in August 2001, January 2002 and August 2002 (60 cows per farm per sampling; n = 720 total samples). Samples were cultured for E. coli O157:H7 and Salmonella and a portion of the recovered Salmonella isolates were examined for genetic relatedness using pulsed‐field gel electrophoresis (PFGE). Results:  Faecal shedding of E. coli O157:H7 and Salmonella varied considerably between farms and at the different sampling times. Large fluctuations in the percentage of positive animals were observed from summer to summer for both of these pathogens. Similarly, Salmonella serotype and serotype prevalence varied from farm to farm and within farm from one sampling time to another. Multiple Salmonella genotypes were detected for a number of serotypes and identical genotypes were found on different farms with one genotype of Salmonella Senftenberg identified on three of the four farms. Significance and Impact of the Study:  This study demonstrated the wide variability in pathogen shedding within and among dairy farms all located in a small geographical region and highlights the complexity of pathogen control at the farm level.

Aims: To examine factors affecting faecal shedding of the foodborne pathogens Escherichia coli O157:H7 and Salmonella in dairy cattle and evaluate antimicrobial susceptibility of these isolates. Methods: Faecal samples were obtained in replicate from lactating (LAC; n ¼ 60) and non-lactating (NLAC; n ¼ 60) Holstein cattle to determine influence of heat stress, parity, lactation status (LAC vs NLAC) and stage of lactation [£60 or >60 days in milk (DIM)] and cultured for E. coli O157:H7 and Salmonella. A portion of the recovered isolates were examined for antimicrobial susceptibility using the broth microdilution technique. Results: No effects of heat stress were observed. Lactating cows shed more (P < 0AE01) E. coli O157:H7 than NLAC cows (43% vs 32%, respectively). Multiparous LAC cows tended to shed more (P ¼ 0AE06) Salmonella than primiparous LAC cows (39% vs 27%, respectively). Parity did not influence (P > 0AE10) bacterial shedding in NLAC cows. Cows £60 DIM shed more (P < 0AE05) Salmonella than cows >60 DIM. Seventeen Salmonella serotypes were identified with the most prevalent being Senftenberg (18%), Newport (17%) and Anatum (15%). Seventy-nine of the Salmonella isolates were resistant to at least one of the seven antibiotics. Escherichia coli O157:H7 isolates were resistant to 11 different antibiotics with multiple resistance to nine or more antibiotics observed in five isolates. Significance and Impact of the Study: This study demonstrated differences in the shedding patterns of foodborne pathogens due to the stage of the milk production cycle and may help identify times when on-farm pathogen control would be the most effective.

The objective of this study was to understand the conjugative transmissibility of resistance plasmids present in 205 Salmonella enterica isolates from bovine sources. Polymerase chain reaction (PCR)-based replicon typing was used to type plasmid replicons. Conjugation experiments were preformed in triplicate at 30 degrees C and 37 degrees C on solid medium. PCR mapping of the A/C transfer gene operon was done on 17 Salmonella Newport isolates that were only positive for A/C. Eighty-six percent (n = 177) of the Salmonella isolates were multidrug resistant (MDR) with resistance to 3-12 antimicrobial agents. Of these, 82% (n = 146) were resistant to extended-spectrum cephalosporins and possessed a bla(CMY) gene. A/C was the predominant replicon detected, present in 90% (n = 160) of the MDR isolates. Twenty-three percent (n = 37) of the A/C-positive strains were positive for a second replicon. Replicons coresident with A/C included I1, N, B/O, HI1, and HI2. Only 31% (n = 54) of the MDR isolates produced transconjugants, and most of these donors carried multiple replicons. A/C cotransferred with B/O, N, and I1 at both 30 degrees C and 37 degrees C and with HI2 at 30 degrees C. Seven Salmonella Newport isolates that produced transconjugants possessed only the single A/C replicon and lacked bla(CMY). PCR mapping of the A/C transfer gene operon in ten Salmonella Newport isolates that carried bla(CMY) revealed a bla(CMY) inverted repeat element integrated between the traA and traC genes. These results suggest that A/C may have been a conjugative plasmid before the integration of bla(CMY) into the transfer gene operon. Additionally, transfer deficient A/C replicons may be mobilized in the presence of certain compatible conjugative plasmids.

Salmonella is one of the most serious foodborne pathogenic bacteria in the United States, causing an estimated 1.3 million human illnesses each year. Dairy cows can be reservoirs of foodborne pathogenic bacteria, including Salmonella spp.; it is estimated that from 27 to 31% of dairy herds across the United States are colonized by Salmonella. The present study was designed to examine the occurrence of Salmonella spp. on dairies and to examine the serotypic diversity of Salmonella isolates on sampled dairies from across the United States. Fecal samples (n = 60 per dairy) were collected from 4 dairies in each of 4 states for a total of 960 fecal samples representing a total population of 13,200 dairy cattle. In the present study, 93 of 960 samples (9.96%) collected were culture-positive for Salmonella enterica. At least one Salmonella fecal-shedding cow was found in 9 of the 16 herds (56%) and the within-herd prevalence varied in our study from 0% in 7 herds to a maximum of 37% in 2 herds, with a mean prevalence among Salmonella-positive herds of 17%. Seventeen different serotypes were isolated, representing 7 different Salmonella serogroups. There were 2 or more different serogroups and serotypes present on 7 of the 9 Salmonella-positive farms. Serotypes Montevideo and Muenster were the most frequent and widespread. From our data, it appears that subclinical colonization with Salmonella enterica is relatively common on dairy farms and is represented by diverse serotypes on US dairy farms.

Seasonal shedding of Escherichia coli O157:H7 in ruminants is well documented; however, viable explanations for this phenomenon are lacking. The fecal prevalence of E. coli O157:H7 in ruminants is the highest in the summer months, decreasing to low or undetectable levels in the winter. To determine if day length is correlated to seasonal shedding, nine reports of E. coli O157:H7 prevalence in cattle were selected for similarity of sampling technique, culture methodology, cattle type, and ability to determine approximate sampling month and location, representing three regions in North America (southern Canada, midwestern United States, and the southern United States/Mexico). Correlation and regression analysis revealed a positive correlation existed between day length (r = 0.67; R2 = 0.45; P = 0.0009) and, to a lesser extent, ambient temperature (r = 0.43; R2 = 0.19; P = 0.05) and E. coli O157:H7 prevalence. Based on these results, a study was conducted in a commercial feedlot to examine the effect of artificial lighting on E. coli O157 prevalence. Four pens of cattle received approximately 5 hours of artificial lighting daily (in addition to the natural light) for 60 days and four pens served as controls. Freshly voided fecal samples (representing approximately 30% of the steers/pen) were collected from the pen floor and cultured for E. coli O157 after 25 and 53 days of artificial lighting and 28 and 43 days following cessation of the lighting treatment. Fecal prevalence of E. coli O157 remained constant in the lighted pens whereas prevalence was lower (P< 0.01) in the control treatment after 53 days of lighting. Forty-three days following cessation of the lighting treatment, prevalence decreased in the lighted treatment group to levels comparable (P > 0.20) to control levels. Understanding the mechanisms underlying the seasonality of this important foodborne pathogen will enable the implementation of effective control strategies when it is most prevalent.