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United States Department of Agriculture

Agricultural Research Service

Research Project: MICROBIAL ECOLOGY OF HUMAN PATHOGENS RELATIVE TO POULTRY PROCESSING
2011 Annual Report


1a.Objectives (from AD-416)
1)Determine the role that outside environmental sources of Listeria monocytogenes play in the presence of this pathogen in poultry further processing facilities. 2)Develop and test intervention strategies to eliminate L. monocytogenes and Campylobacter from meat products or processing plant surfaces. 3)Evaluate gene expression profiles of L. monocytogenes and C. jejuni in conditions relevant to poultry processing environments. 4)Evaluate the influence of animal agriculture on Campylobacter in the environment.


1b.Approach (from AD-416)
Poultry products are commonly associated with the human pathogens Campylobacter and Listeria. The processing plant is a post-harvest site in which numerous manipulations are made to the poultry product, many of which may impact the microbial quality of the product. This research project is designed to study the distribution and dispersion of bacterial pathogens in poultry processing plants and poultry products. The goal is to develop information so that knowledge-based methods can be developed to improve the microbiological quality of poultry products. Sub-types of organisms will be studied to determine if they are specific for the poultry processing ecosystems from which they are derived. Gene expression will be examined in order to find genes that are regulated in response to environmental conditions or are growth phase specific. As work progresses, the results of these efforts may uncover possibilities for definition of critical control points or potential intervention strategies.


3.Progress Report
Detecting and isolating Campylobacter from highly contaminated poultry associated samples can be difficult due to the high number of competing bacteria. In collaboration with scientists, hatchery debris, specifically downy fluff, was collected from a commercial broiler house, spiked with low levels of Campylobacter and used for development of a novel polymerase chain reaction and pyrosequencing method to detect Campylobacter and determine other organisms present. This sample type was found to have a very heavy load of a wide range of bacteria and further used for a comparison of several types of plating media, both commercially available and under development, for detection, isolation and identification of Campylobacter.

The affect of broiler processing chemicals on the antimicrobial resistance and subtype of Campylobacter is not entirely clear. A culture collection of Campylobacter from a previous study includes individual isolates exposed to pH extremes during broiler processing. The antimicrobial resistance profile off all isolates was determined in order to determine if the pH treatments selected for organisms more resistant to traditional antimicrobial treatments. No difference was noted; at this point we have no plans to further characterize isolates in this culture collection.

In earlier work we found that Listeria monocytogenes can be harbored in the floor drains of commercial chicken processing plants. One concern is that such contamination may be inadvertently spread to food contact surfaces or ready-to-eat meat products during plant wash down procedures. A project was designed and preliminary studies have been completed to examine the dangers associated with L. monocytogenes as a biofilm inside floor drains due to a 2 second hose spray. The complete transmission cycle of Campylobacter jejuni remains unknown. Transmission in water is a potential mechanism. C. jejuni has been collected for five years in an ecological study on microbial populations in water ways of the Oconee river basin. One large yearly (85 site) and three quarterly collections were conducted. Forty-four independent isolates of C. jejuni that were been recovered that have been characterized by multi-locus sequence typing (MLST). The types for majority of the isolates that were recovered were unique to environmental water. Most of the types that were not unique are common in every environment. Evidence was seen for gene transfers between types, but these were only in the water types. It appears that agricultural practices did not substantially impact the Campylobacter population in the Oconee river in the course of the study.


4.Accomplishments
1. Contamination of poultry carcasses during processing. Leakage of gut contents during automated feather picking causes an increase in Campylobacter numbers. A study was completed to examine the affect of several novel variations on standard broiler processing on the numbers of Campylobacter on broiler carcasses. Therefore, carcass orientation, order of processing events and plugging the cloaca were tested as possible means to prevent gut contents from coming in contact with the edible surfaces of carcasses. We found that neither hanging carcasses in a vent side down orientation or plugging the vent with spray foam were reliable means to prevent gut content contamination during feather picking. However, changing processing order to eviscerate prior to scalding and feather picking was effective to significantly moderate the increase in Campylobacter that is co-incident with feather removal.

2. Cross-contamination of poultry during carriage to the processing plant. Broiler live haul transport coops can be a platform for Campylobacter cross contamination from a positive flock to the next flock transported. Most U.S processors do not wash cages between uses and when cages are washed, the conventional spray method is only partially effective. Our earlier work showed that allowing feces deposited on coop flooring during transport to dry out was more effective to eliminate detectable Campylobacter but also more time consuming. Two potential solutions to the time problem were tested. Studies were completed to examine the utility of cornstarch as a desiccant and hot air as a drying aid to hasten the desiccation caused death of Campylobacter in broiler feces on transport cage flooring. Both methods show promise as means to significantly lower Campylobacter contamination in transport cages in times of 15 minutes or less.

3. Species identification of Campylobacter based on the ribosomal genes. The ribosomal genes for Campylobacter jejuni and C. coli are almost indistinguishable. But there is a section of genetic sequence between these genes called the Intergenic Sequence Region (ISR) that is usually very different between the two species. However, we discovered a strain that had copies of the region characteristic of both species, an indication of exchange of genetic material between the two. We devised a procedure to monitor the same kind of cross-over in more than 100 isolates. We found that changes in the region could be found in about 10% of the isolates but the exchange between the two species was not found again. Therefore, the cross-over appears to be rare and diagnostic procedures based on identifying the ISR will not commonly be erroneous.


Review Publications
Musgrove, M.T., Cox Jr, N.A., Berrang, M.E., Buhr, R.J., Richardson, L.J., Mauldin, J.M. 2010. Effect of inoculation and application methods on the performance of chemicals used to disinfect salmonella contaminated broiler hatching eggs. Journal of Applied Poultry Research. 19(4):387-392.

Meinersmann, R.J., Ladely, S.R., Lindsey, R.L. 2010. Ribosomal operon intergenic sequence region (ISR) heterogeneity in Campylobacter coli and Campylobacter jejuni. Letters in Applied Microbiology. 51(5):539-545.

Last Modified: 10/24/2014
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