In-house validation of real-time PCR methods for detecting the INV A and TTR genes of Salmonella spp. in food
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2018
Authors
Dmitrić, MarkoVidanović, Dejan
Matović, Kazimir
Šekler, Milanko
Sarić, Ljubiša
Arsić, Milos
Karabasil, Nedjeljko
Article (Published version)
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The microbiological standard method for the detection of Salmonella spp. is time-consuming, and consequently there is a need for an alternative rapid methodology for its detection. In this study, two open-formula real-time PCR methods for detecting the inv A and ttr gene Salmonella spp. have been successfully optimized and in-house validated. Different DNA extraction procedures were used (boiling, Chelex resin and standard I - Biorad). The relative sensitivity and false positive ratio for the alternative methods were 100% and 0.0%, respectively. The relative trueness was in range from 96.8% to 99.2%. No false negative results were detected. The lowest C-t values were obtained using the protocol for detecting the ttr gene after DNA extraction by the Chelex. The results were compared in a large number of food and environmental samples to those of the SRPS EN ISO 6579:2008 standard method and commercial kit (IQ check((R))Salmonella II kit, Bio-Rad, USA). Practical applicationsThe whole pr...ocedure of real-time PCR methods in this study was approximately 20 hr, in contrast to 4-5 days of analysis time for the standard method (SRPS EN ISO 6579:2008). The real-time PCR method also provides a high level of sensitivity and specificity with a low risk of cross-contamination. The implementation of the method for routine analysis will help improve safety in the food production chain, by providing results compatible with the ISO standard, but more rapidly.
Source:
Journal of Food Processing and Preservation, 2018, 42, 2, e13455-Publisher:
- Wiley, Hoboken
DOI: 10.1111/jfpp.13455
ISSN: 0145-8892
WoS: 000424261300040
Scopus: 2-s2.0-85027527561
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Fakultet veterinarske medicineTY - JOUR AU - Dmitrić, Marko AU - Vidanović, Dejan AU - Matović, Kazimir AU - Šekler, Milanko AU - Sarić, Ljubiša AU - Arsić, Milos AU - Karabasil, Nedjeljko PY - 2018 UR - https://vet-erinar.vet.bg.ac.rs/handle/123456789/1641 AB - The microbiological standard method for the detection of Salmonella spp. is time-consuming, and consequently there is a need for an alternative rapid methodology for its detection. In this study, two open-formula real-time PCR methods for detecting the inv A and ttr gene Salmonella spp. have been successfully optimized and in-house validated. Different DNA extraction procedures were used (boiling, Chelex resin and standard I - Biorad). The relative sensitivity and false positive ratio for the alternative methods were 100% and 0.0%, respectively. The relative trueness was in range from 96.8% to 99.2%. No false negative results were detected. The lowest C-t values were obtained using the protocol for detecting the ttr gene after DNA extraction by the Chelex. The results were compared in a large number of food and environmental samples to those of the SRPS EN ISO 6579:2008 standard method and commercial kit (IQ check((R))Salmonella II kit, Bio-Rad, USA). Practical applicationsThe whole procedure of real-time PCR methods in this study was approximately 20 hr, in contrast to 4-5 days of analysis time for the standard method (SRPS EN ISO 6579:2008). The real-time PCR method also provides a high level of sensitivity and specificity with a low risk of cross-contamination. The implementation of the method for routine analysis will help improve safety in the food production chain, by providing results compatible with the ISO standard, but more rapidly. PB - Wiley, Hoboken T2 - Journal of Food Processing and Preservation T1 - In-house validation of real-time PCR methods for detecting the INV A and TTR genes of Salmonella spp. in food VL - 42 IS - 2 SP - e13455 DO - 10.1111/jfpp.13455 ER -
@article{ author = "Dmitrić, Marko and Vidanović, Dejan and Matović, Kazimir and Šekler, Milanko and Sarić, Ljubiša and Arsić, Milos and Karabasil, Nedjeljko", year = "2018", abstract = "The microbiological standard method for the detection of Salmonella spp. is time-consuming, and consequently there is a need for an alternative rapid methodology for its detection. In this study, two open-formula real-time PCR methods for detecting the inv A and ttr gene Salmonella spp. have been successfully optimized and in-house validated. Different DNA extraction procedures were used (boiling, Chelex resin and standard I - Biorad). The relative sensitivity and false positive ratio for the alternative methods were 100% and 0.0%, respectively. The relative trueness was in range from 96.8% to 99.2%. No false negative results were detected. The lowest C-t values were obtained using the protocol for detecting the ttr gene after DNA extraction by the Chelex. The results were compared in a large number of food and environmental samples to those of the SRPS EN ISO 6579:2008 standard method and commercial kit (IQ check((R))Salmonella II kit, Bio-Rad, USA). Practical applicationsThe whole procedure of real-time PCR methods in this study was approximately 20 hr, in contrast to 4-5 days of analysis time for the standard method (SRPS EN ISO 6579:2008). The real-time PCR method also provides a high level of sensitivity and specificity with a low risk of cross-contamination. The implementation of the method for routine analysis will help improve safety in the food production chain, by providing results compatible with the ISO standard, but more rapidly.", publisher = "Wiley, Hoboken", journal = "Journal of Food Processing and Preservation", title = "In-house validation of real-time PCR methods for detecting the INV A and TTR genes of Salmonella spp. in food", volume = "42", number = "2", pages = "e13455", doi = "10.1111/jfpp.13455" }
Dmitrić, M., Vidanović, D., Matović, K., Šekler, M., Sarić, L., Arsić, M.,& Karabasil, N.. (2018). In-house validation of real-time PCR methods for detecting the INV A and TTR genes of Salmonella spp. in food. in Journal of Food Processing and Preservation Wiley, Hoboken., 42(2), e13455. https://doi.org/10.1111/jfpp.13455
Dmitrić M, Vidanović D, Matović K, Šekler M, Sarić L, Arsić M, Karabasil N. In-house validation of real-time PCR methods for detecting the INV A and TTR genes of Salmonella spp. in food. in Journal of Food Processing and Preservation. 2018;42(2):e13455. doi:10.1111/jfpp.13455 .
Dmitrić, Marko, Vidanović, Dejan, Matović, Kazimir, Šekler, Milanko, Sarić, Ljubiša, Arsić, Milos, Karabasil, Nedjeljko, "In-house validation of real-time PCR methods for detecting the INV A and TTR genes of Salmonella spp. in food" in Journal of Food Processing and Preservation, 42, no. 2 (2018):e13455, https://doi.org/10.1111/jfpp.13455 . .