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Beef products have had a major role in C. jejuni outbreaks in the U.S. over the past five years. The Foodborne Outbreak Response and Surveillance Unit of the Centers for Disease Control (CDC) reported five confirmed outbreaks of campylobacteriosis due to ground beef or ground beef products since 1999, which have affected 199 individuals. A C. jejuni prevalence of 57 to 73% in feedlot cattle has been estimated, based on fecal samples (Lee et al., 2004). However, the prevalence of C. jejuni on carcass meat is unclear. In our studies, C. jejuni was detected in 94% of the carcasses sampled along the ventral midline cut (300 cm2). This region is frequently trimmed and sent to the grinders for the production of ground beef. Moreover, we have found that the majority of C. jejuni isolates recovered from feedlot cattle invade epithelial cells and/or survive in macrophages. Schouls et al. (2003) typed Campylobacter strains from humans (n=83), cattle (n=31), and poultry (n=59) and unexpectedly found that cattle strains more closely resembled human strains than poultry strains. Collectively, these in vitro findings suggest that C. jejuni isolates recovered from cattle are pathogenic to humans. While these five outbreaks demonstrate that ground beef is a risk factor for C. jejuni infection, most cases of campylobacteriosis are sporadic in nature. We believe that the number of humans infected with C. jejuni from the consumption of ground beef is grossly underestimated. The underestimation of beef products as a risk factor for C. jejuni infection may be, in part, due to the fact that the contaminated food product is rarely traced back to the source for a sporadic infection. Based on our preliminary data, we hypothesize that virulent C. jejuni are introduced into ground beef from trimmings, and that the C. jejuni-contaminated trimmings pose a serious risk factor for consumers. <P>The objectives of this study are to:<OL> <LI>isolate and quantify C. jejuni from various stages of beef cattle (farm-to-fork), including their environment for source-tracking and HACCP evaluation<LI>determine the genetic diversity of the C. jejuni isolates to allow grouping into epidemiologically relevant clusters<LI>assess the antibiotic susceptibilities and virulence properties of the C. jejuni isolates. </OL>The hypothesis will be accomplished by sampling feedlot cattle from range to processing, isolating C. jejuni from cattle beginning at the range, at the feedlot, and right through to the production of fine ground beef. In addition, environmental samples will be obtained from birds residing at the feedlot, feed, feed bunks, watering units, and the floor of pens designated for the holding of the calves. At the processing stage, we will sample carcasses using both the USDA-APHIS and ventral midline methods, in order to evaluate differences in screening. Isolates obtained from these samples will be examined for genotypic differences, antibiotic resistance, and virulence traits. Data generated from this research will be critical in the development of intervention and control methods to reduce the number of cases of campylobacteriosis.
NON-TECHNICAL SUMMARY: Recent studies have shown a decrease in Campylobacter jejuni prevalence in poultry, however, campylobacteriosis continues to be a leading cause of gastroenteritis in the U.S. This indicates that other sources for C. jejuni contamination are present in the food chain. Preliminary data from our laboratory demonstrated a prevalence of 73.5% of C. jejuni in feedlot cattle and a prevalence of 94.7% in samples obtained from the ventral midline of carcasses. Since trimmings for grinding into ground beef are often taken from this portion of the carcass, we believe that the consumption of ground beef poses a significant health risk to humans. Therefore, we hypothesize that C. jejuni are introduced into ground beef from trimmings due to carcass contamination and pose a significant risk to consumers. We will test this hypothesis by sampling and obtaining quantitative C. jejuni load data from feedlot cattle from range through processing. In addition, C. jejuni presence will be determined from environmental samples, including from birds at the feedlot, from the feed, watering units, bunks, and pens. The lineage of these isolates will be compared to C. jejuni isolated from feedlot cattle to determine the source of contamination. Isolates obtained from cattle and environmental samples will also be examined for antibiotic resistance, and virulence traits. Data generated from this research will be critical in the development of intervention and control methods to reduce the number of cases of campylobacteriosis. <P>APPROACH: The University of Arizona maintains a 86,785 acre ranch, with approximately 500 head of cattle, with calving beginning in February. Calves are branded, vaccinated and castrated during June. At about six months of age (400-450 lbs), the calves are moved to the College of Agriculture feedlot near Tucson, fed a high grain diet for 180 days and slaughtered at the Meat Science Laboratory. This closed system provides us with a unique opportunity and a controlled environment to conduct a longitudinal study to estimate the incidence of C. jejuni in a cohort of beef cattle from range to slaughter, along with an opportunity to evaluate currently accepted sampling methods for the presence of pathogens on carcasses. Doves and pigeons are present in high numbers around the feedlot, feeding on the grain. Fecal droppings left behind can contaminate the feed. If the birds are colonized with C. jejuni, it is probable that the calves will also become colonized with this pathogen. <P>The specific aims are:<BR> Specific Aim 1. Isolation of C. jejuni from the feedlot environment and beef cattle from range to slaughter. Doves and pigeons residing at the feedlot will be live-trapped and fecal samples examined for C. jejuni. Swipes from feed bunks, feed, watering units, and floors will also be examined for viable C. jejuni. The C. jejuni load will then be determined in feedlot cattle at weaning, at intervals while residing in the feedlot, and at four control points during processing through to the grinding of the meat into fine ground beef. In our studies, C. jejuni was detected in 94% of the carcasses sampled along the ventral midline cut. This region is frequently trimmed and sent to the grinders for the production of ground beef. However, this region is not tested by USDA-APHIS. Carcasses during processing will be sampled at regions recommended by USDA-APHIS and at the ventral midline cut, to determine differences in detection. <BR>Specific Aim 2. Determine the genetic diversity of C. jejuni isolates. Isolates will be genotyped by PFGE and MLST, to search for associations between genotypes and hosts. <BR>Specific Aim 3. Assess the antibiotic susceptibilities and virulence properties of bovine C. jejuni isolates. Half of the calves will receive a natural diet and the other half a similar diet with rumensin, tylosin and growth promoting implants of estradiol and progesterone. These diets will allow us to examine the transfer of macrolide resistance to C. jejuni during a feedlot setting. Antibiotic sensitivities will be determined for all isolates to determine if there is a correlation between antibiotic resistance and carcass survival. Selected isolates will be examined for virulence in cell culture and in the piglet model established by our laboratory to determine virulence levels. Standard logistic regression modeling techniques will be used, such as assessment of model goodness-of-fit, %correct classification, the Hosmer-Lemeshow test, and examination of standardized residuals for violations of model assumptions, along with repeated-measures analyses-of-variance models and pairwise comparisons to determine when significant changes of organism load occur.