Validation of a DIVA qPCR Duplex Assay to Differentiate Primun Salmonella T Vaccine from Salmonella enterica subsp. enterica Serovar Typhimurium Wild Strains
Source abstract
Abstract
Salmonella enterica subsp. enterica serovar Typhimurium is an important foodborne pathogen, and poultry products are a major source of human infection. Live attenuated vaccines for poultry are an effective tool for reducing the prevalence of infection, but vaccine strains must be differentiated from wild strains to ensure effective disease surveillance and control. This study reports the validation of the SalTypm&PriSal-T qPCR Duplex kit, a DIVA qPCR assay for the differentiation of the Primun Salmonella T vaccine from wild strains using DNA extracted from isolated colonies. Analytical specificity and sensitivity, as well as diagnostic specificity and sensitivity, were evaluated with optimal results. This qPCR assay significantly reduces the time required to obtain a diagnostic result compared to reference methods based on antibiogram differentiation. Notably, this is the first qPCR test available worldwide for distinguishing this vaccine from wild strains, providing a valuable tool for improving the efficiency and accuracy of Salmonella surveillance programs in poultry production systems.
Source: crossref · Quality: baseline
Research context
OpenAlex topics: Salmonella and Campylobacter epidemiology · Bacteriophages and microbial interactions · Viral gastroenteritis research and epidemiology
SciVX discovery tags are terminology-based. OpenAlex topics are source-provided classifications.
Affiliations
Department of Microbiology, University Miguel Hernández, 03312 Orihuela, Alicante, Spain
Genetic PCR Solutions™, 03300 Orihuela, Alicante, Spain
Laboratorios Calier S.A., 24231 Onzonilla, León, Spain
References (24)
Show source-provided references
1. Maskell, D., and Mastroeni, P. (2006). Host-Specificity of Salmonella Infections in Animal Species. Salmonella Infections: Clinical, Immunological and Molecular Aspects, Cambridge University Press. Advances in Molecular and Cellular Microbiology.10.1017/CBO9780511525360 ↗
2. EFSA, and ECDC (2024). The European Union One Health 2023 Zoonoses Report. EFSA J., 22, e9106.10.2903/j.efsa.2024.9106 ↗
3. Nazir, J., Manzoor, T., Saleem, A., Gani, U., Bhat, S.S., Khan, S., Haq, Z., Jha, P., and Ahmad, S.M. (2025). Combatting Salmonella: A Focus on Antimicrobial Resistance and the Need for Effective Vaccination. BMC Infect. Dis., 25.10.1186/s12879-025-10478-5 ↗
4. Molecular-Based Identification and Detection of Salmonella in Food Production Systems: Current Perspectives10.1111/jam.13888 ↗
5. Salmonellosis: The Role of Poultry Meat10.1016/j.cmi.2015.12.004 ↗
6. Shaji, S., Selvaraj, R.K., and Shanmugasundaram, R. (2023). Salmonella Infection in Poultry: A Review on the Pathogen and Control Strategies. Microorganisms, 11.10.3390/microorganisms11112814 ↗
7. Rakov, A.V., Mastriani, E., Liu, S.-L., and Schifferli, D.M. (2019). Association of Salmonella Virulence Factor Alleles with Intestinal and Invasive Serovars. BMC Genom., 20.10.1186/s12864-019-5809-8 ↗
8. Associations Between the Level of Biosecurity and Occurrence of Dermanyssus gallinae and Salmonella spp. in Layer Farms10.1637/11327-111415-Reg ↗
9. Ruvalcaba-Gómez, J.M., Villagrán, Z., Valdez-Alarcón, J.J., Martínez-Núñez, M., Gomez-Godínez, L.J., Ruesga-Gutiérrez, E., Anaya-Esparza, L.M., Arteaga-Garibay, R.I., and Villarruel-López, A. (2022). Non-Antibiotics Strategies to Control Salmonella Infection in Poultry. Animals, 12.10.3390/ani12010102 ↗
10. Salmonella Infections: Immune and Non-Immune Protection with Vaccines10.1080/03079450601113167 ↗
11. Population Dynamics and Antimicrobial Resistance of the Most Prevalent Poultry-Associated Salmonella Serotypes10.3382/ps/pew342 ↗
12. Antimicrobial Resistance among Invasive Nontyphoidal Salmonella enterica Isolates in the United States: National Antimicrobial Resistance Monitoring System, 1996 to 200710.1128/AAC.01333-10 ↗
13. (2013). Microbiology of Food and Animal Feed—Preparation of Test Samples, Initial Suspension and Decimal Dilutions for Microbiological Examination (Standard No. ISO 6887-6:2013). Available online: https://www.iso.org/standard/53664.html.
14. (2017). Microbiology of the Food Chain—Horizontal Method for the Detection, Enumeration and Serotyping of Salmonella (Standard No. ISO 6579-1:2017). Available online: https://www.iso.org/standard/56712.html.
15. World Organization for Animal Health (2025, February 14). Terrestrial Manual, Chapter 2.2.3. Development and Optimisation of Nucleic Acid Detection Assays. Online Access. Available online: https://www.woah.org/en/what-we-do/standards/codes-and-manuals/terrestrial-manual-online-access/.
16. (2017). General Requirements for the Competence of Testing and Calibration Laboratories (Standard No. ISO/IEC 17025:2017). Available online: https://www.iso.org/standard/66912.html.
17. Martínez-Murcia, A., Navarro, A., Garcia-Sirera, A., Pérez, L., and Bru, G. (2023). Internal Validation of a Real-Time qPCR Kit Following the UNE/EN ISO/IEC 17025:2005 for Detection of the Re-Emerging Monkeypox Virus. Diagnostics, 13.10.3390/diagnostics13091560 ↗
18. Bru, G., Martínez-Candela, M., Romero, P., Navarro, A., and Martínez-Murcia, A. (2023). Internal Validation of the ASFV MONODOSE Dtec-qPCR Kit for African Swine Fever Virus Detection under the UNE-EN ISO/IEC 17025:2005 Criteria. Vet. Sci., 10.10.3390/vetsci10090564 ↗
19. Internal Validation of GPSTM MONODOSE CanAur Dtec-qPCR Kit Following the UNE/EN ISO/IEC 17025:2005 for Detection of the Emerging Yeast Candida Auris10.1111/myc.12834 ↗
20. Research Note: Validation of a New Differentiation Approach Using the Commercial ASAPTM Media to Detect the Salmonella 441/014 Vaccine Strain10.1016/j.psj.2024.103679 ↗
21. Aribam, S.D., Nakayama, M., Ichimura, S., Tokuyama, K., Hara, Y., Ogawa, Y., Shimoji, Y., and Eguchi, M. (2023). Differentiation of Salmonella Vaccinated and Infected Animals by Serological Detection of Antibody to T3SS Effector SsaK in an Indirect ELISA. J. Microbiol. Methods, 209.10.1016/j.mimet.2023.106729 ↗
22. Gebauer, J., Kudlackova, H., Kosina, M., Kovarcik, K., Tesarik, R., Osvaldova, A., Faldyna, M., and Matiasovic, J. (2016). A Proteomic Approach to the Development of DIVA ELISA Distinguishing Pigs Infected with Salmonella Typhimurium and Pigs Vaccinated with a Salmonella Typhimurium-Based Inactivated Vaccine. BMC Vet. Res., 12.10.1186/s12917-016-0879-1 ↗
23. Development, Validation and Field Evaluation of a Quantitative Real-Time PCR Able to Differentiate Between Field Mycoplasma synoviae and the MS-H-Live Vaccine Strain10.1080/03079457.2017.1296105 ↗
24. Vidanović, D., Tešović, B., Šekler, M., Debeljak, Z., Vasković, N., Matović, K., Koltsov, A., Krstevski, K., Petrović, T., and De Leeuw, I. (2021). Validation of TaqMan-Based Assays for Specific Detection and Differentiation of Wild-Type and Neethling Vaccine Strains of LSDV. Microorganisms, 9.10.3390/microorganisms9061234 ↗