Source: University of Maryland Eastern Shore submitted to NRP
INVESTIGATING THE PREVALENCE OF FOODBORNE PATHOGENS AND PROFILING THE ANTIMICROBIAL RESISTANCE OF SALMONELLA IN COMMERCIAL PET FOODS
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
ACTIVE
Funding Source
OTHER GRANTS
Reporting Frequency
Annual
Accession No.
1031926
Grant No.
2024-38821-42105
Cumulative Award Amt.
$600,000.00
Proposal No.
2023-09291
Multistate No.
(N/A)
Project Start Date
Apr 1, 2024
Project End Date
Mar 31, 2027
Grant Year
2024
Program Code
[EQ]- Research Project
Recipient Organization
University of Maryland Eastern Shore
11868 College Backborne Road
Princess Anne,MD 21853
Performing Department
(N/A)
Non Technical Summary
The increasing trend of humanization and premiumization of pets, along with the use of raw and commercial pet foods, poses a heightened risk of transmitting foodborne pathogens to both pets and their owners. Salmonella, a common foodborne pathogen frequently linked to pet foods, can lead to severe illnesses in humans through handling or cross-contamination. Moreover, the prevalence of antimicrobial-resistant (AMR) pathogens like Salmonella in commercial pet foods exacerbates this concern. Our proposed investigation aims to assess the prevalence of foodborne pathogens, particularly Salmonella, in U.S. commercial pet foods, and to characterize Salmonella's antibiotic resistance profile. Through isolating and identifying major pathogens like Salmonella across all pet food categories, we will genotype Salmonella to determine serotypes and detect resistance genes. By assessing these serotypes' sensitivity to medically relevant antibiotics, we will gain crucial insights into AMR in pet foods and potential human health risks linked to pet food handling. This research will aid in pinpointing high-risk products and contaminants, facilitating targeted interventions, and improving mitigation strategies to curb the spread of foodborne pathogens in the pet food supply chain. Additionally, our findings will inform regulatory agencies, pet food manufacturers, and consumers, fostering enhanced industry practices, consumer awareness, and responsible pet ownership.
Animal Health Component
80%
Research Effort Categories
Basic
10%
Applied
80%
Developmental
10%
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
72352301103100%
Knowledge Area
723 - Hazards to Human Health and Safety;

Subject Of Investigation
5230 - Feed and feed additives;

Field Of Science
1103 - Other microbiology;
Goals / Objectives
Pets are becoming an integral part of human life. The increasing trend of humanization and premiumization of pets is also leading to increased risk to pet owners from handling pet foods, especially raw pet foods. This project aims to evaluate commercially available pet foods and their microbiological quality focusing on Salmonella spp. Along with their antibiotic resistance profiling.The overall goal of this project is to improve human public health and safety by gaining knowledge on the prevalence, identification, and characterization of Salmonella associated with commercial pet foods. The pet food HACCP course, offered to the stakeholders, will provide the pet food manufacturers with the knowledge and understanding of the importance of developing food safety plans and protocols in the facility.Objective 1 A-C: ResearchObjective 1A: Isolate foodborne Salmonella from commercial pet foods.Objective 1B: Phenotypic characterization of the Salmonella, and screen for antibiotic sensitivity profile.Objective 1C: (Evaluate an acidulant antimicrobial compound against the isolated MDR Salmonella isolates in-vitro and in food matrices.Objective 2: ExtensionObjective 2: (Byron Chaves) Develop and deliver a virtual Pet Food HACCP composed of three modules:1) Introduction to Pet Food Safety,2) Regulatory Framework for the Safety of Pet Foods, and3) Applications of the Hazard Analysis and Critical Control Points (HACCP)System to the Pet Food Industry.
Project Methods
Objective 1Procurement and sampling of pet foods: Different categories of pet foods will be procured from local grocery stores and/or online suppliers. To ensure the study is representative of the commercial pet food market, factors such as type of pet foods, brand diversity, ingredient variation, and product popularity will be considered. A total of 360 dry pet foods, 360 raw pet foods, 180 semi-moist, 180 pet treats, and 180 canned pet foods will be collected. Starting from year 0.5 until the end of year 2 (18 months) approximately 20 samples each of dry and raw, and 10 each of semi-moist, treat, and canned foods will be collected each month. Proper sampling protocols will be followed to maintain randomness, and sample uniformity and prevent contamination during collection and transportation to the laboratory for analysis. Additionally, samples will be properly labeled and stored to facilitate accurate identification and tracking throughout the research process.Pathogen isolation, identification, and serotype characterization of Salmonella:Pet foods (dry kibbles, semi-moist, canned food, raw food, treats, etc.) will be collected from different retail stores in Maryland. The samples will be transported to the laboratory and stored in proper storage condition as described in the package until processed. Each small-sized bag of dry food or each bag/pouch of wet food or individual canned food will be considered one sample. On each collection visit, products from different lots and dates will be procured to make the sampling as representative and diverse as possible. Similarly, to maintain randomness, samples will be procured from different stores in different cities in the state. The moisture percentage of dry and semi-moist feed will be evaluated, and feed samples will be processed for microbiological evaluation. Samples will be processed using the standard BAM protocol of Salmonella isolation and identification procedure. The positive samples from TSI and LIA slants will be proceeded for biochemical and serological tests. All Salmonella-positive cultures will be serotyped following the World Health Organization guidelines at the USDA National Veterinary Service Laboratories (NVSL) in Ames, IA.The antibiotic resistance profile of the isolates will be tested using agar Minimal Inhibitory Concentration methods. Following the Sensititre® micro-broth dilution method according to the guidelines of the Clinical and Laboratory Standards Institute (CLSI), MICs of all the isolates will be determined using 96-well microtiter plates, against 15 most common medically important antibiotics used in human and veterinary health. Using a recently established protocol.Determination of virulent genes in Salmonella: Salmonella isolates will be tested for the presence of virulent genes known to cause infection in humans. The Salmonella isolates will be analyzed for the following virulent genes: invA, spvC, and pagC by PCR using the methods previously established.Effect of acidulant against MDR Salmonella isolates in-vitro and in food matrices:An in-vitro model of the experiment will be conducted where an acidulant, sodium bisulfate (SBS) at various concentrations will be subjected to MDR Salmonella. Minimum inhibitory concentrations of SBS will be determined against different serotypes of MDR Salmonella using a protocol described above. Similarly, the efficacy of SBS will be evaluated in a dry pet food kibble.Objective 2 (Extension)A "Pet Food HACCP" training course will be developed, piloted, and implemented. The curriculum will be based on the guidelines of the International HACCP Alliance and will be modified to include examples and case studies relevant to the pet food industry. Additionally, the FDA FSMA Preventive Controls for Animal Food will be consulted to reconcile language and any aspects of regulatory compliance pertinent to the pet food industry. Once developed, the training course will be turned into several modules: Technical and Regulatory Overview, Food Safety Hazards, Good Manufacturing Practices, Seven Principles of HACCP, and Food Safety Recalls. Each module's content will be peer-reviewed for accuracy by professionals in academia and industry. After module development, the course will be piloted in a hybrid format: The participants will be responsible for watching the recordings and completing a small test after each module to move on to the next one. Then, a one-day virtual training will be conducted for the practical aspects of the course. Participants who complete the virtual modules and the practical session within three months will be provided with a Certificate of Course Completion with the seal of the International HACCP Alliance.EvaluationThe overall progress of the project will be assessed based on the milestones outlined in the timetable. The team will meet quarterly to evaluate the project's progress, identify challenges, and recommend research or extension modifications (if necessary). Additionally, the team will regularly seek advisory committee guidance, recommendations, and feedback from the advisory committee during research and extension activities throughout the project's duration. The success of the project will be evaluated in a comprehensive and multifaceted manner, focusing on several key aspects. These include increased student and faculty interest in pet food safety and AMR research programs at UMES, FAMU, and UNL. Increased involvement, increase in underrepresented communities, and student participation in the research activity are rise in the visibility of UMES, FAMU, and UNL in newsletters, media, scientific meetings, and community through research publications, workshops, training, and demonstrations. The efficacy of research facilities and research conduct will be assessed based on the quality and quantity of research projects conducted and scientific publications published during the grant period.

Progress 04/01/24 to 03/31/25

Outputs
Target Audience:During this reporting period, the primary target audience included pet food manufacturers, graduate and undergraduate students, and pet owners concerned with food safety. These groups were strategically selected due to their direct roles in the production or consumption of commercial pet food and their vested interest in preventing foodborne illness and antimicrobial resistance. Pet food manufacturers and industry stakeholders were engaged through informal outreach and information exchanges. This audience is critical, as they are directly responsible for sourcing, formulating, and processing commercial pet food, and thus are in a position to adopt improved safety and quality control measures based on the research findings. Graduate and undergraduate students, especially those in animal science, food safety, and microbiology programs, were involved through laboratory instruction, research internships, and hands-on data collection experiences. These efforts provided valuable experiential learning opportunities, enhanced scientific literacy, and helped train the next generation of researchers in pet food safety and antimicrobial resistance. Pet owners were indirectly targeted through extension-related discussions and educational material drafts aimed at increasing public awareness about the importance of proper pet food handling, the risks of foodborne pathogens, and the implications of antimicrobial resistance. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided?The project has supported multiple opportunities for both training and professional development: Two graduates (one at UMES and another at UNL) and one undergraduate student are actively working on both objectives 1 and 2. Undergraduate involvement included presentation opportunities, resulting in a first-place award at the 2024 Undergraduate Symposium at the University of Maryland Eastern Shore. The presentation focused on the prevalence and antimicrobial resistance of Salmonella in household pets. Participation in major scientific events included: A presentation at the 2025 Graduate Research Symposium at UMES. Attend the Pet Food Forum (April 28 - 30, 2025) in Kansas City, Missouri. Involvement in an invited symposium titled Food Safety and Regulatory Considerations for Raw Pet Foods at IAFP-2024. The project was also featured on the Pet Food Science Podcast Show, providing outreach and visibility. https://www.youtube.com/watch?v=nidHkEFVO08 How have the results been disseminated to communities of interest?Findings have been shared with both scientific and general audiences through various outreach and dissemination channels such as publications, symposiums, conference presentations, and media outreach. Publications: Dhakal, J.*, Cancio, L. P., Deliephan, A., Tubene, S., Chaves, B. D. 2024. Salmonella Presence and Risk Mitigation in Pet Foods, a Growing Challenge with Implications for Human Health. Comprehensive Reviews in Food Science and Food Safety. DOI:10.1111/1541-4337.70060 Scientific abstracts Presentations and oral presentation. Otwey, R.Y., Chapagain, S., Dhakal, J. 2025. Investigating the Prevalence and Antimicrobial Resistance Profile of Salmonella in Commercial Pet Foods in Delmarva. Graduate Research Symposium. University of Maryland Eastern Shore, Princess Anne, MD 21853. Invited Symposium: Steven Moore, Jenny Murphy, Jasmine Kataria, and Janak Dhakal. Food Safety and Regulatory Considerations for Raw Pet Foods: Challenges and Opportunities. Symposium, IAFP-2024, Long Beach, CA Media: Featured in the Pet Food Science Podcast Show- Invited guest. https://www.youtube.com/watch?v=nidHkEFVO08 What do you plan to do during the next reporting period to accomplish the goals?During the next reporting period, the project will focus on the Completing the sampling of pet food samples to provide a holistic view of the prevalence of Salmonella in commercial pet food. Continuing with serotyping and antimicrobial resistance profiling. This will help identify the specific characteristics of Salmonella contaminating commercial pet foods and provide deeper insights into effective mitigation strategies. Continuing work on developing and delivering a virtual Pet Food HACCP. This will enhance outreach and support to pet food manufacturers.

Impacts
What was accomplished under these goals? Objective 1A-C Research: Between January and March 31st, 2025, 242 pet food samples were collected from retail outlets across the Delmarva region. These samples were categorized into dry, raw-wet, semi-moist, canned, and treats. Standard microbiological techniques, including enrichment and selective isolation, followed by PCR confirmation targeting the invA gene, were used to detect Salmonella. Isolated strains were tested for resistance to 16 antibiotics, covering all the groups of antibiotics. Out of 242 pet food samples, 11 (4.5 %) tested positive for Salmonella, with prevalence rates of 8.5 % in raw/mildly cooked (7/82), 5.7 % in dry food (2/35), and 3.4 % in treats (2/59). Canned and semi-moist foods did not show any positives for Salmonella; (0/13) and (0/53) respectively. All eleven isolates were screened for antimicrobial resistance. All isolates were multidrug resistant (MDR), with 5 categorized as extensively drug resistant (XDR). Isolates have been sent for stereotyping, results are pending. Extension Objective 2: The development of a virtual Pet Food Hazard Analysis and Critical Control Points(HACCP) training composed of three modules is underway. A graduate student has been assigned to plan and execute the course content related to pet food safety.

Publications

  • Type: Peer Reviewed Journal Articles Status: Published Year Published: 2024 Citation: 1. Dhakal, J.*, Cancio, L. P., Deliephan, A., Tubene, S., Chaves, B. D. 2024. Salmonella Presence and Risk Mitigation in Pet Foods, a Growing Challenge with Implications for Human Health. Comprehensive Reviews in Food Science and Food Safety. DOI:10.1111/1541-4337.70060
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: Steven Moore, Jenny Murphy, Jasmine Kataria, and Janak Dhakal. Food Safety and Regulatory Considerations for Raw Pet Foods: Challenges and Opportunities. Symposium, IAFP-2024, Long Beach, CA
  • Type: Conference Papers and Presentations Status: Accepted Year Published: 2025 Citation: Otwey, R.Y., Chapagain, S., Dhakal, J. 2025. Investigating the Prevalence and Antimicrobial Resistance Profile of Salmonella in Commercial Pet Foods in Delmarva. Graduate Research Symposium. University of Maryland Eastern Shore, Princess Anne, MD 21853.
  • Type: Other Status: Published Year Published: 2025 Citation: Featured in the Pet Food Science Podcast Show- Invited guest. https://www.youtube.com/watch?v=nidHkEFVO08
  • Type: Conference Papers and Presentations Status: Published Year Published: 2024 Citation: The Prevalence and Antimicrobial Resistance of Salmonella in Household Pets (Dogs and Cats) in Lower Eastern Shore, Maryland