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<p>The overall objective is to investigate the potential of four plant-derived molecules, namely tran-cinnamaldehyde (TC), carvacrol (CR), thymol (TH) and eugenol (EG) to improve the microbiological safety of shell eggs. This will be accomplished through science-based, applied research at pre-harvest and post-harvest levels, and communication of research findings to stakeholders, using a variety of educational methods. The research objectives are </p>
<p>1. To determine the effect of TC, CR, TH and EG as dietary supplements to reduce the colonization of S. Enteritidis in the intestinal and reproductive tracts, and eggs of layer chickens. </p>
<p>2. To determine the efficacy of TC, CR, TH or EG as an antimicrobial wash for inactivating S. Enteritidis on shell eggs </p>
<p>3. To determine if consumers can detect a difference in eggs obtained from layer chickens treated with TC, CR, TH and EG as dietary supplements when compared to eggs from hens on non-treated diets. </p>
<p>4. To determine if consumers can detect a difference in eggs washed with TC, CR, TH or EG when compared to eggs washed in a standard commercial wash. </p>
<p>The Extension/Outreach Objectives include </p>
<p>1. To develop, implement and evaluate S. Enteritidis reduction programs through the use of plant derived antimicrobials by egg producers. </p>
<p>2. To provide extension programs, workshops, and educational materials designed to educate egg producers on the value of using plant-derived antimicrobials, as a part of a total egg quality assurance program (EQAP) to reduce Salmonella in/on eggs.</p>
<p>NON-TECHNICAL SUMMARY:<br/> S. Enteritidis is a major food-borne pathogen in the U.S., with eggs serving as the most common source of infection. Contamination of eggs with S. Enteritidis occurs by spread of the bacterium from layers' infected ovaries, or by penetration through eggshell from contaminated feces. Therefore effective methods for decreasing Salmonella in layers can reduce human infection from contaminated eggs. Our preliminary research revealed that several food-grade, plant ingredients, including trans-cinnamaldehyde, carvacrol, thymol, and eugenol killed S. Enteritidis in chicks, suggesting they may be effective in controlling Salmonella in layers and eggs. The plant ingredients were also effective in killing Salmonella on shell surface when used as a wash. This project will investigate the (1) effect of plant ingredients to reduce S. Enteritidis in layers
and eggs when given to birds through feed, (2) effect of plant ingredients for killing Salmonella on eggs as an antimicrobial wash, (3) consumer acceptance of eggs laid by layers fed with the plant ingredients or eggs washed with plant ingredients, and (4) safety of plant ingredients in chickens to make that they are not harmful to birds. This project will develop, implement and evaluate Salmonella control programs for poultry farmers, egg producers and processors through extension programs, workshops, and educational materials. This project will potentially lead to decreased salmonella infection from eggs, thus improving public health and economic opportunities for poultry farmers.<p>
APPROACH: <br/>The effect of TC, CR, TH and EG on the colonization of S. Enteritidis in layers will be studied in 18-20 week-old pullets. The bird will be randomly divided into groups, including 1. negative control (no S. Enteritidis challenge, no antimicrobial), 2. positive S. Enteritidis control (S. Enteritidis challenge, no compound antimicrobial), 3. positive compound control (no S. Enteritidis challenge, feed supplemented with compound at 1%), 4. S. Enteritidis challenged and feed supplemented with antimicrobial at 0.5%, 5. S. Enteritidis challenged and feed supplemented with antimicrobial at 0.75%, 6. S. Enteritidis challenged and feed supplemented with antimicrobial at 1%. On day 1 of the experiment, all the birds will be weighed and their fecal samples tested for Salmonella. The birds belonging to the different treatments will be provided feed containing
respective concentrations of plant compound and water ad libitum. On day 5 of the experiment, the birds will be infected orally with 1.0 ml of PBS containing ~ 8 log CFU of a 5-strain mixture of nalidixic acid-resistant S. Enteritidis by crop gavage. The birds not challenged with S. Enteritidis will be given 1.0 ml of sterile PBS. Following challenge, S. Enteritidis counts in the fecal samples and eggs (internal and external) from all the birds will be determined on every third day. Moreover, after challenge, ten birds from each treatment group will be sacrificed on weeks 1, 3, 5, 7, 9, 11, and 13 by carbon dioxide inhalation. The cloaca, cecum, liver, spleen, oviduct, ovaries, and formed eggs, if any, in the oviduct from each bird will be collected for bacteriological analysis. Further, the populations of S. Enteritidis in the cecal and clocal contents will also be determined. In
addition, the body weight and feed intake of treated and control birds will be determined. Finally, histopathological examination of internal organs will be done to rule out any toxicity of the compounds on birds. The efficacy of TC, CR, TH and EG as an antimicrobial egg wash on S. Enteritidis will be also be studied. Salmonella-inoculated eggs will be subjected to the washing treatment in water (negative control) or plant compound (0.25, 0.5 0r 0.75%) or chlorine (positive control) for 1 min, 2 min, 3 min or 5 min at 32 or 40C. The egg washing will be performed in a USDA-approved, KUHL KF-200 Egg Washer, which is commonly used by small/medium sized processors. After washing for each specified exposure time, S. Enteritidis counts on eggs will be determined. Finally the consumer acceptance of eggs laid by plant molecule-treated layers or eggs subjected to antimicrobial wash will be
determined using triangle tests. Our outreach activities will develop, implement and evaluate Salmonella control programs for egg producers and processors through extension programs, workshops, eXtension and educational materials. Our educational programs will emphasize the best practices for pathogen reduction, but would add new information about natural S. Enteritidis reduction methods that could also be used by the organic poultry industry.
</p><p>PROGRESS: <br/>2012/09 TO 2013/08 <br/>Target Audience: Poultry Scientists, Poultry Veterinarians and Poultry Extension Scientists Changes/Problems: Two plant compounds, namely carvacrol and eugenol had a negative effect on growth and body weight of chickens. Hence we included another plant compound, namely caprylic acid in the chicken trials. What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? A 3 hour 40 min symposium entitled ""Reducing Salmonella Enteritidis Contamination of Shell Eggs with an Integrated Research and Outreach Approach"" was presented at theannual meeting of the Poultry Science Associationheld in July 2013 at San Diego, CA.. The following people, including the project PDs presented the talks. Efficacy of Plant-derived Antimicrobials for
Reducing Egg-borne Transmission of Salmonella Enteritidis, (30min) Kumar Venkitanarayanan, University of Connecticut Practical Implications of Plant Molecules in Poultry Diets, (30 min) Michael Darre, University of Connecticut Pre-Harvest Egg Quality Assurance Extension and Outreach for the Poultry Industry, (20 min) Paul Patterson, Penn State University eXtension and Other Outreach for Egg Processors and Consumers, (20 min) Pat Curtis, Auburn University Molecular Characterization of Salmonella Enteritidis Isolated from Laying Hen Environments & Eggs (30 min) Eva Pendleton & Subhashinie Kariyawasam, Penn State University. On Farm Risk Factors for Salmonella Contamination, (20 min) Paul Patterson, Penn State University On Farm Opportunities for Intervention to Reduce the Risk of Salmonella Contamination of Eggs, (20 min) Darrell Trampel, Iowa State University What do you plan to
do during the next reporting period to accomplish the goals? Investigate the efficacy of another plant-derived compound (caprylic acid) as a feed additive for reducing egg-borne transmission of Salmonella Enteritidis.
<br/>PROGRESS: <br/>2011/09/01 TO 2012/08/31 <br/>OUTPUTS: In vivo efficacy of trans-cinnamaldehyde in reducing egg-borne transmission of S. Enteritidis. As a follow up to the previous year's experiment, this in vivo study investigated the efficacy of feed supplemented with TC for reducing S. Enteritidis colonization of oviduct and its egg-borne transmission in 40-week-old and 25 week-old white leghorn layers. In two separate experiments, birds were randomly assigned to six treatment groups (n=20, N=120): a negative control (no S. Enteritidis, no TC), a low dose compound control (no S. Enteritidis, 1% TC), high dose compound control (no S. Enteritidis, 1.5% TC), a positive control (inoculated with S. Enteritidis, no TC), a low dose group (inoculated with S. Enteritidis, 1% TC) and a high dose group (inoculated with S. Enteritidis, 1.5% TC). Water and feed were provided ad
libitum. On day 0, three birds from each trial were randomly selected and sacrificed to confirm that birds were devoid of any inherent Salmonella. On day 10, birds were inoculated with approximately 10.0 log CFU of a 5-strain mixture of nalidixic acid resistant S. Enteritidis by crop gavage. After 24 hours, three birds from each group were sacrificed and analyzed to ensure colonization of oviduct with the pathogen. Eggs were collected from day 4 after inoculation and the presence of S. Enteritidis on eggshell and in egg yolk was determined separately everyday for 10 weeks. At the end of 10 weeks, 10 birds were sacrificed per group and checked for S. Enteritidis colonization in the oviduct and cecum. PARTICIPANTS: K. Venkitanarayanan, M. Darre, M. I. Khan, I. Upadhyaya, and A. Kollanoor-Johny TARGET AUDIENCES: Poultry scientists, Veterinarians and Extension specialists. PROJECT
MODIFICATIONS: Not relevant to this project.
<br/>PROGRESS: <br/>2010/09/01 TO 2011/08/31 <br/>OUTPUTS: 1. Effect of trans-cinnamaldehyde on egg-borne transmission of Salmonella Enteritidis (SE) in chickens. This in vivo study investigated the efficacy of feed supplemented with TC for reducing S. Enteritidis colonization of oviduct and its egg borne transmission in 40-week-old white leghorn layers. Birds were randomly assigned to six treatment groups (n=20, N=120): a negative control (no S. Enteritidis, no TC), a low dose compound control (no S. Enteritidis, 1% TC), high dose compound control (no S. Enteritidis, 1.5% TC), a positive control (inoculated with S. Enteritidis, no TC), a low dose group (inoculated with S. Enteritidis, 1% TC) and a high dose group (inoculated with S. Enteritidis, 1.5% TC). Water and feed were provided ad libitum. On day 0, three birds from each trial were randomly selected and sacrificed to
confirm that birds were devoid of any inherent Salmonella. On day 10, birds were inoculated with approximately 10.0 log CFU of a 5-strain mixture of nalidixic acid resistant S. Enteritidis by crop gavage. After 24 hours, three birds from each group were sacrificed and analyzed to ensure colonization of oviduct with the pathogen. Eggs were collected from day 4 after inoculation and the presence of S. Enteritidis on eggshell and in egg yolk was determined separately everyday for 10 weeks. At the end of 10 weeks, 10 birds were sacrificed per group and checked for S. Enteritidis colonization in the oviduct and cecum. PARTICIPANTS: Venkitanarayanan, K, Darre, M., Khan, M. I., Upadhyaya, I, Upadhyay, A., Kollanoor Johny, A. TARGET AUDIENCES: Poultry industry, Poultry Extension agents, Poultry Scientists, Regulatory agencies. PROJECT MODIFICATIONS: Not relevant to this project.</p>