CONTROL OF FOOD-BORNE PATHOGENS IN PRE- AND POST-HARVEST ENVIRONMENTS

• Sponsoring Institution: National Institute of Food and Agriculture
• Project Status: TERMINATED
• Funding Source: HATCH
• Reporting Frequency: Annual
• Accession No.: 0225213
• Project No.: MIS-501140
• Multistate No.: S-1033
• Project Start Date: Oct 1, 2007
• Project End Date: Sep 30, 2012

Project Director
Kim, T.

Recipient Organization
MISSISSIPPI STATE UNIV
(N/A)
MISSISSIPPI STATE,MS 39762

Performing Department
Food Science & Technology

Non Technical Summary
Salmonella is responsible for the largest number of outbreaks in the USA. However, source of illness remains a puzzle in many of the outbreaks. In many cases the source of illness is only speculated or arrived at by probability and not by complete identification. Another problem is that many food products that are imported or otherwise processed (frozen) that may contain Salmonella and other pathogens are not detected. The reason for this is that the pathogen may be in a nonculturable state, i.e., cannot be isolated utilizing normal culturing conditions. This affects our economy, including our state. The ability to have better, more reliable and sensitive methods to identify contaminated food products will allow us to compete more fairly in the global market and insure a safer food supply to the consumers. In addition, the ability to better identify sources of pathogens can help us pinpoint critical steps needed to produce safer food products. In specific, environmental (water, soil, manure, plant, and other samples) and food (fresh, frozen, etc) samples will be obtained (in some cases from collaborators/colleagues, in other cases from importers and distributors, in other cases by our group) and codified to be able to traceback the origins. These samples will then be split (keeping some in storage) and assayed for Salmonella and E. coli 0157:H7 by conventional (BAM, 2000; USDA, 2010) and unconventional methods (incubation for days without selective broth, incubation for days with selective broth, incubation with other nutrients, etc.) to determine the viability and recovery of pathogens. Once a pathogen is recovered as VNBC, then identification and typing will be done, in addition to confirmation by PCR. We will also check for patterns that are not common in that species (enzyme deficiency or other marker) to further study its survivability. Depending on the number of VNBC isolates and their source, we will then proceed to identify their genetic makeup (LSBI) to further identify characteristics of survivability. These isolates will then be exposed to different environmental conditions (pH, temperature, nutrients, etc) to discern their optimum growth characteristics to then design a proper recovery method. The method will then be validated with further samples. The method can then be used for epidemiological and related studies.

Animal Health Component

(N/A)

Research Effort Categories

Basic

(N/A)

Applied

(N/A)

Developmental

(N/A)

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
712	5010	1000	30%
712	5010	1040	30%
712	5010	1100	40%

Knowledge Area
712 - Protect Food from Contamination by Pathogenic Microorganisms, Parasites, and Naturally Occurring Toxins;

Subject Of Investigation
5010 - Food;

Field Of Science
1100 - Bacteriology; 1000 - Biochemistry and biophysics; 1040 - Molecular biology;

Keywords

salmonella

viable-but-non-culturable

Goals / Objectives
New methods to prevent, reduce or eliminate foodborne disease agents at all points of the food chain, from farm to fork, are needed to improve the safety of the food supply to prevent illnesses and deaths and to prevent economic losses to the food industry. Objectives Develop or improve methods for control or elimination of pathogens in pre-and post harvest environments including meat, poultry, seafood, fruits and vegetables and nutmeats. Develop and validate mathematical modeling to gain understanding of pathogen behavior in macro- and micro-environments. Investigate factors leading to the emergence, persistence and elimination of antimicrobial resistance in food processing and animal production environments. Outputs: Validated decontamination methods that can be used by the fruit, vegetable, seafood, meat and poultry industry to enhance the safety of their finished product Outreach/extension education and training materials for regulatory personnel, producers, processors, consumers, extension agents Overall enhanced food safety for consumers Outcomes or projected Impacts: Enhanced safety of fruit, vegetable, seafood, meat, and poultry products Increased understanding of food safety measures by regulatory personnel, producers, processors, consumers, extension agents Overall enhanced food safety for consumers Milestones: (2007): Pre-harvest food safety: Initiate work on antimicrobial films, high pressure processing of viruses. Modeling: Develop and validate wind tunnel to validate heat transfer models. Antimicrobial drug resistance: Tetracycline resistance genes in the environment. Total bacterial population genomic DNA extracted from fecal samples and analyzed for presence of all tetracycline resistance (Tc-R) genes. (2008): Pre-harvest food safety: Initiate efforts on sanitizers, high pressure processing of vibrios. Modeling: Collect growth data of Salmonella in chicken and beef at isothermal conditions. Develop neural network model and compare its performance to statistical models. Antimicrobial drug resistance: Environmental sample collection from antibiotic free and antibiotic receiving farms and molecular analysis. Analyze tetracycline resistant isolates for (expected) tetracycline resistance genes with PCR. (2009): Pre-harvest food safety: Investigate optimization of high pressure processing in RTE seafoods. Modeling: Collect growth data of E. coli in ground beef for different fat content at isothermal conditions. Antimicrobial drug resistance: Data analysis and manuscript preparation (2010): Pre-harvest food safety: Initiate outreach activities. Modeling: Current FSIS risk assessment model for E. coli in ground beef is based on models developed using broth. (2011): Pre-harvest food safety: continue outreach activities and publish research results. Modeling: Develop heat transfer to obtain temperature profile in shell eggs during cooling. Expected Outputs we hope to discover viable-but-non-culturable (VBNC)/stressed pathogens in food and environmental samples through the development of rapid and easy-to-use procedures.

Project Methods
Since we recently found that nontyphoidal Salmonella spp. was resistant to culture in isolation on conventional nonselective/selective media including buffered peptone water, RV and tetrathionate broth, Brilliant green and XLD agar and PCR test, it is important to develop techniques to culture "unculturable Salmonella spp. on a novel recovery techniqu(s). Optimization for the growth of unculturable Salmonella spp. includes simulated natural environmental conditions including the need for specific nutrients, pH conditions, incubation temperatures or levels of oxygen in the atmosphere and a reliance on beneficial bacterial interactions within the source environment. Combinatorial approaches will also enhance optimization to improve isolation of unculturable pathogenic bacteria that can be cultured in the laboratory.

Progress 01/01/12 to 12/31/12

Outputs
OUTPUTS: Project research goal was to development of single tube assays for independent detection of Vibrio vulnificus from seafood utilizing unique biochemical features of this organism. V. vulnificus is the leading cause of seafood-associated illnesses and deaths in the U.S. Reliable, rapid, and efficient assays not requiring expensive equipment or reagents would greatly facilitate analysis of seafood samples. Assays under development would reduce analytical cost by up to 90%. This would expand researchers' capacity for outbreak investigations, evaluation of proposed mitigations and market or environmental surveys needed to support current risk assessments. This test method would expedite validation of post-harvest processes (PHP) and procedures to reduce or eliminate this pathogen from seafood. Increased analytical capacity fit for the above purposes was urgently needed to evaluate and improve the effectiveness of current U.S. shellfish safety policy. FDA initiated work to validate a real-time PCR method using naturally incurred vibrios in oyster and clam samples from the Atlantic and Gulf coasts. Dr. Kim was invited to visit the Gulf Coast Seafood Laboratory to share validation samples and reference methods results. Suggestion was made to expand this collaboration to include development of a V. vulnificus assay using a similar single tube. Researchers at MSU and FDA hypothesized that a test kit incorporating a two-phase medium provides adequate sensitivity, specificity, and differentiation to reliably determine V. vulnificus levels in post-harvest processed oysters. Upper liquid phase was formulated to have optimal pH and NaCl concentrations for V. vulnificus recovery. Compounds that enhance the recovery of stressed/injured cells were included to optimize the sensitivity and recovery of this organism. Growth of background microflora was suppressed in the solid phase by incorporation of antimicrobial compounds and by a shift to elevated temperatures after a period of incubation under optimal conditions for V. vulnificus that allowed stressed cells recovery. Differential aspects were achieved in the solid phase based on unique biochemical utilization patterns of V. vulnificus and indicator compounds. Initial testing indicated that V. vulnificus single tube assay had potential utility for detection of this pathogen in naturally contaminated samples. Based on the results of some pure culture testing as well as the real world samples, different formulations were evaluated based on the expertise and discussions between the investigators. In order to reduce media preparation and analysis time, assay was adapted to a 96-well plate format. Format was applied to pure isolate analysis to evaluate performance with regard to specificity and for most probable number analysis (MPN) of seafood samples. When compared to the single tube format, 96-well plate format gave equivalent results with a reduction in sample handling time. After satisfactory results were obtained with pure cultures, inoculated seafood samples were used to validate the assay. Experiments were initiated in December 2012. PARTICIPANTS: Taejo Kim, Angelo DePaola, Jessica Jones, Juan L. Silva, and Demarcus Carter. Participants include industry members in Mississippi, faculty members and graduate research assistants at Mississippi State University. TARGET AUDIENCES: Farmers, food/seafood industry, biotechnology industry, and the microbiology class at Mississippi State University. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
Vibrio analytical capability and capacity in the U.S. is extremely limited due to the high cost of equipment, supplies and highly skilled analysts needed to test oysters or other seafood using currently available and accepted methods. Adequate capability exists at a few state and federal shellfish control authorities and a handful of academic laboratories but not at any commercial laboratories in the Gulf region. In general this capability is directed at research and PHP validation/verification mainly for large firms handling high product volumes. Laboratories with vibrio analytical capability generally lack resources for monitoring at needed spatial and temporal granularity to identify high risk products or for surge capacity during outbreaks. Validation of the proposed two-phase testing approach for V. vulnificus would provide an alternative method that would eliminate the need for expensive equipment and highly trained analysts and reduce the cost of expendable supplies by a factor of 10-fold relative to currently used methods such as real-time PCR. The above reductions in analytical resources and technical complexity would allow virtually all state shellfish control authorities to quickly achieve sufficient vibrio analytical capabilities and capacity to meet all monitoring needs and the surge capacity to manage outbreak response. Affordable testing could be provided by commercial laboratories to assist with validation and verification of PHP oysters by industry. Field testing capability could potentially expand testing to processing plants located in remote oyster growing areas. Taken together this expansion of capability and capacity would be transformational for risk management based on vibrio analysis as MASGC is already funding a similar project for V. vulnificus.

Publications

• Kim, T. 2012. Confidential Disclosure Agreement between Mississippi State University, FDA and Neogen on Developing Vibrio Assay Kit (2012-2015).
• Gawborisut, S., Kim.T.J., and Silva J.L. 2012. Microbial quality and safety of x-ray irradiated fresh catfish (Ictalurus punctatus) fillets stored under CO2 atmosphere. Journal of Food Science. 77:M533-538.
• Samala, A., Srinivasan, R., Yadav, M., Kim, T.J., and Prewitt, L. 2012. Production of xylo-oligosaccharides from corn fiber by autohydrolysis. Bioresources Journal. 7:3030-3050.
• Kim, T.J., and Silva, J.L. 2012. Development of innovative one-tube Salmonella detection kit and its application for detecting Salmonella in food and environmental samples. Institute of Food Technologists Annual Meeting, Las Vegas, NV.
• Arroyo-Llantin N., Silva, J.L., and Kim T.J. 2012. Incidence of Salmonella in fish during 2 seasons in a seafood processing plant. Institute of Food Technologists Annual Meeting, Las Vegas, NV.
• Gu, Qian, Silva, J.L, and Kim, T. 2012. Development of detection method for E. coli O157H7 and non O157:H7 in catfish. Journal of the Mississippi Academy of Sciences. 57:39.
• Arroyo-Llantin, N., Kim, T., and Silva, J.L. 2012. Relationship of indicator and pathogenic microorganisms on catfish products. Journal of the Mississippi Academy of Sciences. 57:40.
• Liao, Y.C, Kim, T., and Silva, J.L. 2012. Enhanced production of glucose, cellobiose and xylose during germination of mold spores on corn cob. Southeast Biofuels and Renewable Energy Conference, Jackson, MS. Aug. 8-9 (Poster, student competition)
• Silva, J.L., Kim, T., Arroyo-Llantin, N., Armbrust, K., and Sampedro-Parra, F. 2012. Risk assessment of the presence of antibiotic residues and chemical contaminants in aquaculture products. AQUA 2012 Meeting, Prague, Czech Republic. Sept 1-5, 2012. (Invited)

Progress 10/01/07 to 09/30/12

Outputs
Target Audience: Sincewe developed the novel pathogen test kits Salmonella, E.coli O157:H7 and Listeria spp, wetransferred thetechnologies forpractical implementation to demonstrate their impacts for fresh producer and farmers at Mississippi. We assistedfarms andfood processorsfor controlling on-farm and in-plant foodborne pathogens and expected seasonal prevalence of the pathogens and currently have a power to solve emergency pathogen incidence cases by early reporting. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Nothing Reported How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? Genetic tools (PFGE and RT-PCR)have been developed for in-house pathogenscreeningon pre and post-harvest end products and their processing environmental samples. We anticipate to usethescreened dataas pathogen monitoring tools for enhancing HACCPprogram of the food processing plant that will help Mississippi farmers and processors to prevent Salmonella, Listeria and E. coli O157:H7 contamination on their food products

Publications

• Type: Journal Articles Status: Published Year Published: 2012 Citation: Aditya Samala, Radhakrishnan Srinivasan, Madhav Yadav, T.J. Kim, and Lynn Prewitt. 2012. Production of xylo-oligosaccharides from corn fiber by autohydrolysis. Bioresources Journal. 7:3038-3050
• Type: Journal Articles Status: Published Year Published: 2012 Citation: S. Gawborisut, T.J. Kim, and J.L. Silva. 2012. Microbial Quality and Safety of X-Ray irradiated Fresh Catfish (Ictalurus punctatus) Fillets Stored under CO2 atmosphere. Journal of Food Science. 77(9):M533-8
• Type: Journal Articles Status: Published Year Published: 2012 Citation: P. Parakulsuksatid P., Dja Shin Wang, J.L. Silva, T.J. Kim and T.Y. Koo. 2012. Optimization of Enzymatic Skinning on Catfish Nuggets Using Response Surface Methodology. Advanced Materials Research. 472:3094-3103.
• Type: Journal Articles Status: Published Year Published: 2012 Citation: S. Kin, M.W. Schilling, B.S. Smith, J.L. Silva, T.J. Kim. 2012. Effects of liquid and wood smoking combined with potassium lactate and acetate on the quality and inhibition of Listeria monocytogenes growth in ready-to-eat smoked catfish fillets. Journal of Aquatic Food Product Technology. 21:338-350.
• Type: Journal Articles Status: Published Year Published: 2011 Citation: T.J. Kim, J.L. Silva, and Y.S. Jung. 2011. Enhanced functional properties of tannic acid after thermal hydrolysis. Food Chemistry. 126:116-120.
• Type: Journal Articles Status: Published Year Published: 2011 Citation: T.J. Kim, M. Corbitt, and J.L. Silva. 2011. Optimization of hot water processing for removing microbial biofilm on fresh blueberry surface. Journal of Food Science. 76:M353-M360
• Type: Journal Articles Status: Published Year Published: 2011 Citation: S. Kin, M.W. Schilling, B.S. Smith, J.L. Silva, T.J. Kim, S. Campano, A.J. Pham. 2011. Potassium acetate and potassium lactate enhance the microbiological and physical properties of marinated catfish fillets. Journal of Food Science. 76:S242-S250.
• Type: Journal Articles Status: Published Year Published: 2010 Citation: R. Pyla, T.J. Kim, J.L. Silva, and Y.S. Jung. 2010. Proteome analysis of Azotobacter vinelandii ?arrF mutant that overproduces poly-?-hydroxybutyrate polymer. Applied Microbiology and Biotechnology. 88:1343-54.
• Type: Journal Articles Status: Published Year Published: 2010 Citation: T.J. Kim, J.L. Silva, M.G. Kim and Y.S. Jung. 2010. Enhanced antioxidant capacity and antimicrobial activity of tannic acid by thermal processing. Food Chemistry. 118:740-746.
• Type: Journal Articles Status: Published Year Published: 2010 Citation: T.J. Kim, W. L. Weng, J. L. Silva, and D. Marshall. 2010. Identification of Natural Antimicrobial Substances in Red Muscadine Juice against Enterobacter sakazakii. Journal of Food Science. 75:M150-M154.
• Type: Journal Articles Status: Accepted Year Published: 2010 Citation: B.Y. Chen, R. Pyla, T.J. Kim, J. L. Silva, and Y.S. Jung. 2010. Prevalence and contamination patterns of Listeria monocytogenes in fresh catfish fillets and their processing plants. Food Microbiology. 27:645-52.
• Type: Journal Articles Status: Published Year Published: 2010 Citation: B.Y. Chen, R. Pyla, T.J. Kim, J. L. Silva, and Y.S. Jung. 2010. Incidence and persistence of Listeria monocytogenes in catfish processing environment and fresh fillets. Journal of Food Protection 73:1641-50
• Type: Journal Articles Status: Published Year Published: 2010 Citation: R. Pyla, T.J. Kim, J.L. Silva, and Y.S. Jung. 2010. Enhanced antimicrobial activity of starch-based film impregnated with thermally processed-tannic acid, a strong antioxidant. International Journal of Food Microbiology. 137: 154-160
• Type: Journal Articles Status: Published Year Published: 2010 Citation: B.Y. Chen, R. Pyla, T.J. Kim, J. L. Silva, and Y.S. Jung. 2010. Antibiotic resistance in Listeria species isolated from catfish fillets and processing environment. Letters in Applied Microbiology. 50:662-632.
• Type: Journal Articles Status: Published Year Published: 2010 Citation: S. Kin, M.W. Schilling, J.L. Silva, B.S. Smith, V. Jackson, T.J. Kim. Phosphate Type Affects the Quality of Injected Catfish Fillets. 2010. Journal of Food Science. 75: S74-S80.
• Type: Journal Articles Status: Published Year Published: 2010 Citation: T.J. Kim, J.L. Silva, A. Tokitkla, and F.B. Matta. Modified Atmosphere Storage Influences Quality Parameters and Shelf Life of Tifblue Blueberries. 2010. Journal of the Mississippi Academy of Sciences, 55(2-3):143-148
• Type: Journal Articles Status: Published Year Published: 2009 Citation: B.Y. Chen, T.J. Kim, J.L. Silva, and Y.S. Jung. 2009. Positive correlation between the expression of inlA and inlB genes on Listeria monocytogenes and its attachment strength on glass. Food Biophysics. 4: 304-311
• Type: Journal Articles Status: Published Year Published: 2009 Citation: T.J. Kim, J.L. Silva, W.L. Weng, W.W. Chen, M. Corbitt, Y.S. Jung and Y.S. Chen. 2009. Inactivation of Enterobacter sakazakii by water-soluble muscadine seed extracts. International Journal of Food Microbiology. 129:3295-3299.
• Type: Journal Articles Status: Published Year Published: 2009 Citation: T.J. Kim, J.L. Silva and Y.S. Jung. 2009. Antimicrobial activity of fresh and processed red muscadine juice and the role of their polar compounds on Escherichia coli O157:H7. Journal of Applied Microbiology. 107(2):533-539.
• Type: Journal Articles Status: Published Year Published: 2009 Citation: S. Kin, M.W. Schilling, J.L. Silva, B.S. Smith, V. Jackson, T.J. Kim. 2009. Effects of phosphate type on the quality of vacuum tumbled catfish fillets. Journal of Aquatic and Food Product Technology. 18:400-415.
• Type: Journal Articles Status: Published Year Published: 2009 Citation: R. Pyla, T.J. Kim, J.L. Silva, and Y.S. Jung. 2009. Overproduction of poly-?-hydroxybutyrate in the Azotobacter vinelandii mutant that did not produce small RNA ArrF. Applied Microbiology and Biotechnology. 84:717-724.
• Type: Other Status: Published Year Published: 2009 Citation: Kin, S., Schilling, M.W., Smith, B.S., Silva, J.L., Jackson, V., Kim. T., Williams, J.B. 2009. Ingredient Optimization in Fresh, Marinated Catfish Fillets. Mississippi Agricultural and Forestry Experiment Station. Research Report. December 2009. 24(7), pp:1-6
• Type: Book Chapters Status: Published Year Published: 2009 Citation: DAbramo, L., Silva, J.L. and T. Kim. 2009. Optimization of crustacean quality trough husbandry and adherence to post-harvest standards for processing. In Shellfish Safety and Quality, Part 3- Improving Crustacean Safety and Quality, S. Shumway (Ed.), Woodhead Publishing Limited (ISBN 1-84569-152-0)(Book Chapter).
• Type: Journal Articles Status: Published Year Published: 2008 Citation: T.J. Kim, W.L. Weng, J. Stojanovic, Y. Lu, Y. S. Jung and J. L. Silva. 2008. Antimicrobial effect of water-soluble muscadine seed extracts on Escherichia coli O157:H7. Journal of Food Protection. 71:1465-1468.
• Type: Journal Articles Status: Published Year Published: 2008 Citation: B.Y. Chen, T.J. Kim, Y.S. Jung and J.L. Silva. 2008. Attachment strength of Listeria monocytogenes and its Internalin negative mutants. Food Biophysics. 3:329-332.
• Type: Journal Articles Status: Published Year Published: 2007 Citation: T.J. Kim, Y.S. Jung, J.L. Silva and S. Danviriyakul. 2007. Detection and rapid purification of internalin B as a protein marker in Listeria monocytogenes. Food Biotechnology. 21:161-168.
• Type: Journal Articles Status: Published Year Published: 2008 Citation: V.D. Zheljazkov, J.L. Silva, M. Patel, J. Stojanovic, Y. Lu, and T.J. Kim. 2008. Uncomposted human hair as nutrient sources for horticultural crops. HortTechnology. 18 (4): 592-596.
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Arroyo-Llantin N., Silva, J.L., Kim T.J. 2012. Incidence of Salmonella in fish Druing 2 Seasons in a Seafood Processing Plant. IFT mtg., Las Vegas, NV.
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: G. Qian, T.J. Kim, J.L. Silva. 2012. Innovative Application of classic Microbiology for Detecting Escherichia coli O157:H7 in food Samples. IFT mtg., Las Vegas, NV.
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: T.J. Kim, J.L. Silva. 2012. Development of Innovative One-Tube Salmonella Detection Kit and Its Application for Detecting Salmonella in Food and Environmental Samples. IFT mtg., Las Vegas, NV.
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Sheibani, E., Silva J.L., Kim T.J. 2012. Applicatioin of Response Surface Methodology (RSM) in Optimization of Hot Water Treatemnt of Extending Postharves Shelf Life of Sweet Potatoes (lpomoea batatas L. Lam). IFT mtg., Las Vegas, NV.
• Type: Conference Papers and Presentations Status: Published Year Published: 2012 Citation: Qian Gu, Juan L Silva, Tae Jo Kim. 2012. Development of detection method for E.coli O157H7 and non O157:H7 in catfish. Volume 57:39. Mississippi Academy of Sciences Lake Terrace Convention Center, Hattiesburg, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2011 Citation: Norman N. Arroyo-Llantin, Taejo Kim, Juan L. Silva. Relationship of indicator and pathogenic microorganisms on catfish products. Volume 57:40. Mississippi Academy of Sciences Lake Terrace Convention Center, Hattiesburg, MS.
• Type: Journal Articles Status: Published Year Published: 2011 Citation: Abdallah A., Kim T.J., Silva J.L., Arroyo N., Durango A. 2011. Incidence of Listeria and indicator microorganisms of sanitation in catfish ponds and processing plants. IFT mtg., New Orleans, LA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2011 Citation: Kim T.J. and Silva J.L. 2011. Enhanced functional properties of tannic acid after thermal hydrolysis. IFT mtg., New Orleans, LA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2011 Citation: Gu Q., Kim T.J., and Silva J.L. 2011. Incidence and development of methodology of generic and pathogenic Escherichia coli in catfish farms and processing plants. IFT mtg., New Orleans, LA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2011 Citation: Sheibani E., Silva J.L., Kim T.J., and Arancibia R. 2011. Effect of curing and ethylene treatment on sensory and other quality traits of sweet potatoes. IFT mgt., New Orleans, LA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2010 Citation: T.J. Kim, Juan Silva and Yean-Sung Jung. 2010. Processing for the product of gallic acid, pyrogallol and purpurogallin form tannic acid. J. Miss. Academy Sci. 55:36.
• Type: Conference Papers and Presentations Status: Published Year Published: 2010 Citation: Bang-Yuan Chen, T.J. Kim, Yean-Sung Jung, Juan Silva. 2010. Prevalence of Listeria monocytogenes and Listeria sp. in catfish and process environment and equipment by molecular typing methods. J. Miss. Academy Sci. 55:32.
• Type: Conference Papers and Presentations Status: Published Year Published: 2010 Citation: Bang-Yuan Chen, Rajkumar Pyla, T.J. Kim, Yean-Sung Jung, Juan Silva. 2010. Antibiotic resistance in Listeria species isolated form catfish fillets and processing surfaces. J. Miss. Academy Sci. 55:33.
• Type: Conference Papers and Presentations Status: Published Year Published: 2010 Citation: Weiwei Chen, T.J. Kim, Donna Marshall Stephen Stringer, Frank Matta, and Juan Silva. 2010. Evaluation of the antimicrobial capacity and active compounds of extracts form different sections of various muscadine cultivars. J. Miss. Academy Sci. 55:38.
• Type: Conference Papers and Presentations Status: Published Year Published: 2010 Citation: Norman Arroyo, T.J. Kim and Juan Silva. 2010. Enhancing the efficacy of isolation procedure for Salmonella spp. in fresh and frozen aquaculture foods. J. Miss. Academy Sci. 55:39
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: Sovann Kin, M. Wes Schilling, Juan L. Silva, Brian Smith, Viodelda Jackson, T.J. Kim. 2009. Effect of phosphate type on the quality of vacuum tumbled catfish fillets. IFT mtg., Anaheim, CA.
• Type: Journal Articles Status: Published Year Published: 2009 Citation: Juan L. Silva, T.J. Kim, Yean-Sung Jung, Youkai Lu, Bang-Yuan Chen, Parvaneh Kosravi. 2009. Effect of preharvest and postharvest factors on quality and safety of channel catfish (Ictalurus punctatus) fillets. IFT mtg., Anaheim, CA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: Reginald C Johnson, Diane K. Tidwell, M. Wes Schilling, Juan L. Silva, T.J. Kim. 2009. Formulation of a fruit slush using evaporated sweet whey. IFT mtg., Anaheim, CA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: T.J. Kim , Juan Silva, Yean-Sung Jung, Moon G. Kim. 2009. Enhanced antimicrobial and antioxidant activities of tannic acid by thermal processing. IFT mtg., Anaheim, CA
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: Bang-Yuan Chen, T.J. Kim, Yean-Sung Jung, Juan Silva. 2009. Occurrence and identification of sources of Listeria monocytogenes and Listeria spp. in catfish processing. IFT mtg., Anaheim, CA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: Melody Corbitt, T.J. Kim, Juan L. Silva. 2009. Recovery methods influence microbial counts on fresh and frozen blueberries. IFT mtg., Anaheim, CA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: T.J. Kim, J.L. Silva, Y.S. Jung. 2009. Processed plant polyphenol (tannic acid) enhances antimicrobial activity on foodborne pathogens. Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Journal Articles Status: Published Year Published: 2009 Citation: M Corbitt, T.J. Kim and J.L. Silva. 2009. Comparison of microbial recovery methods on yeast, mold, and aerobic counts of frozen blueberries (Vaccinium ashei), Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: W.W. Chen, T.J. Kim, F. Matta, and J.L. Silva. 2009. Effect of grape variety and solvent extraction methods on the antipathogenic activity of muscadine (Vitis Rotundifolia michx.) seed extracts. Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: T.J. Kim, J.L. Silva, and Y.S. Jung. 2009. Inhibtion of E.coli O157:H7 by fresh and processed red muscadine muscadine juice and antimicrobial role of their polar compounds. Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: W.L. Chen, T.J. Kim, and J.L. Silva. 2009. Inhibitory effect of Enterobacter sakazakii by muscadine juices and antimicrobial activity of their identified compounds at pH 3.2. Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: J. L. Silva, T.J. Kim, Y.S. Jung, Y. Lu, D. Minchew, and C. Hoque. 2009. Preharvest and postharvest factors that affect color and quality of catfish (Ictalurtus punctatus) fillets. Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2009 Citation: B.Y. Chen, T.J. Kim, Y.S. Jung and J.L. Silva. 2009. Occurrence of Listeria monocytogenes and Listeria spp. catfish processing plants and identification of contamination sources. Mississippi Academy of Sciences. Olive Branch, MS.
• Type: Conference Papers and Presentations Status: Published Year Published: 2008 Citation: T.J. Kim , W.L. Chen, Y. Lu, Y.J. Jung and J.L. Silva. 2008. Antimicrobial effects of water-soluble muscadine seed extract on Escherichia coli O157:H7. IFT mtg.,New Orleans, LA.
• Type: Conference Papers and Presentations Status: Published Year Published: 2008 Citation: T.J. Kim, Y.S. Jung and J. Silva. 2008. Antimicrobial effects of fresh and processed bronze muscadine (Vitis Rotundifolia Michx) juice on selected human pathogens. J. Miss. Acad. Sci. 53(1): 34.
• Type: Conference Papers and Presentations Status: Published Year Published: 2008 Citation: T.J. Kim, Y.S. Jung and J. Silva. 2008. Muscadine juices and seed extracts inhibit the growth of Enterobacter sakazakii. Mississippi Acad. Sci. mtg, Feb. 20-22, Oliver Branch, MS. (2nd place, oral competition). 2008. J. Miss. Acad. Sci. 53(1): 32.
• Type: Conference Papers and Presentations Status: Published Year Published: 2008 Citation: T.J. Kim, Y.S. Jung and J. Silva. 2008. Comparison of attachment strength of Listeria monocytogenes and its internalin negative mutants. Mississippi Acad. Sci. mtg, Feb. 20-22, Oliver Branch, MS. (3nd place, oral competition). 2008. J. Miss. Acad. Sci. 53(1): 35.

Progress 01/01/11 to 12/31/11

Outputs
OUTPUTS: An invention was developed. This is a novel single tube Salmonella detection kit useful for detecting and quantifying the presence of Salmonella in food and environment samples in laboratory, field or industrial settings with a high sensitivity. The invention provides a simple, inexpensive mechanism to detect and quantify Salmonella in food and environment samples with a very high sensitivity while also preventing the occurrence of false-positives, false-negatives or other statistical errors. A novel combination and gradient technology of selective ingredients and temperature is utilized to allow the growth of Salmonella in the detection kit and enhance the physiological activities of cell motility/infusion and enzyme production of Salmonella which causes pH change under facultative anaerobic conditions. The invention, through the detection mechanism accomplished by the novel combination of selective ingredients and the gradient technology, allows for real-time detection of a single cell of Salmonella within 24 hours or less of introducing a pre-enriched sample to the one tube Salmonella detection kit. An additional benefit of this invention is that it does not require the use of additional machinery or equipment to detect the presence of Salmonella in the test sample. The invention does not require additional machinery or equipment to read or interpret the results of the detection mechanism; unlike the current state of technology. It is the object of this invention to provide a method and kit to detect Salmonella in food and environment samples, in laboratory and industrial settings, through the use of a single test tube Salmonella detection kit. It is an additional object of this invention to describe a methodology for detecting Salmonella in food and environment samples in a single test tube Salmonella detection kit that reduces the opportunity for Type-I or Type-II errors while also limiting the time and additional resources traditionally required for detecting the presence of Salmonella. The invention relies on a unique combination of selective media, chemicals, temperature and decarboxylase activity of Salmonella that promote the facultative anaerobic growth and migration of Salmonella while also inhibiting the growth of other microorganisms. For example, malachite green oxalate in Vassiliadis R10 broth and novobiocin in brilliant green agar are examples of chemical that are known in the industry to increase selectivity and migration of Salmonella at the exclusion of other microorganisms. Utilizing the combination of these selective media and chemicals and gradient technology, the single test tube kit can quickly increase selectivity of Salmonella in a test sample; this enhanced selectivity can then be evaluated and evidenced through the single test tube kit by altering the characteristics of a signal indicating medium, such as phenol red as a pH indicator which, in turn, will provide visible evidence of the presence of Salmonella. PARTICIPANTS: Taejo Kim, Assistant Research Professor Juan Silva, Professor Qian Gui, Graduate Research Assistant TARGET AUDIENCES: Farmers, Food Industry, Biotechnology Industry, Microbiology Class PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Impacts
A commercial patent was developed from this. As a result of the patent, a Non-Disclosure Agreement between Neogen Corporation and Mississippi State University on behalf of its Office of Technology Commercialization was developed on July 17, 2011. The commercial value of the invention could be more than US $10 million. More than 200 food processors, suppliers and allied industries have been contacted for the use of this technology.

Publications

• Kim, T. and Silva, J.L. Development of the Rapid Salmonella Test Kit. Invention Disclosure Entitled, "One-tube Salmonella Test Kit." MSU Tech ID No.: 2010.0792.
• Gu, Q., Kim T.J. and Silva J.L. 2011. Incidence and development of methodology of generic and pathogenic Escherichia coli in catfish farms and processing plants. IFT mtg., New Orleans, LA. Abstract No. 31-04, p. 14. Available at: http://www.ift.org/Meetings-and-Events/Past-Meeting-Resources/Technic al%20Abstract%20Search%20Details.aspxid=53052