Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Salmonella enterica in Low-Resource Settings

Authors

DOI:

https://doi.org/10.47430/ujmr.25103.011

Keywords:

Typhoid fever, Salmonella enterica, loop-mediated isothermal amplification (LAMP), rapid detection

Abstract

Study’s Excerpt:

  • LAMP assay developed for rapid detection of Salmonella enterica.
  • Works at 63°C for 90 mins using a simple water bath.
  • Visual detection enabled by SYBR Green fluorescence.
  • Higher sensitivity (90%) compared to PCR (85%).
  • Cost-effective and ideal for low-resource settings.

Full Abstract:

Typhoid fever remains a significant global health burden, particularly in regions with inadequate sanitation and limited access to safe water supplies.  Traditional diagnostic methods, such as blood culture and biochemical assays, although considered the best standards, are labor-intensive and time-consuming, thereby limiting their practical utility in resource-constrained settings.  This study aimed to develop a loop-mediated isothermal amplification (LAMP) assay for the rapid detection of S. enterica using conserved genomic regions (367 as targets.  Unlike conventional PCR, which requires expensive thermal cyclers, our LAMP protocol utilizes a water bath to maintain a constant reaction temperature of 63°C for 90 minutes, significantly reducing equipment costs.  Amplified products were rapidly detected by the addition of SYBR Green dye, which produced a distinct green fluorescence under blue light, enabling easy visual interpretation.  Comparative analysis revealed that the LAMP assay demonstrated higher sensitivity—detecting Salmonella in 90% of isolates—compared to 85% for PCR, with both methods showing high specificity and no cross-reactivity with non-Salmonella species.  These findings suggest that the LAMP assay provides a rapid, cost-effective, and sensitive alternative for typhoid diagnosis, offering promise for enhanced disease surveillance and management in low-resource settings.

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References

Akinyemi, K. O., Oyefolu, A. O. B., Mutiu, W. B., Iwalokun, B. A., Ayeni, E. S., Ajose, S. O., & Obaro, S. K. (2018). Typhoid Fever: Tracking the Trend in Nigeria. The American Journal of Tropical Medicine and Hygiene, 99(3_Suppl), 41–47. https://doi.org/10.4269/ajtmh.18-0045

Anejo-Okopi, J., Okojokwu, J., Ebonyi, A. O., Ejeliogu, E., Isa, S. E., Audu, O., Akpakpan, E. E., Nwachukwu, E. E., Ifokwe, C. K., Ali, M., Lar, P., & Oguche, S. (2016). Molecular characterization of cryptosporidium in children aged 0- 5 years with diarrhea in Jos, Nigeria. Pan African Medical Journal, 25. https://doi.org/10.11604/pamj.2016.25.253.10018

Bell, R. L., Jarvis, K. G., Ottesen, A. R., McFarland, M. A., & Brown, E. W. (2016). Recent and emerging innovations in Salmonella detection: a food and environmental perspective. Microbial Biotechnology, 9(3), 279–292. https://doi.org/10.1111/1751-7915.12359

Berenger, B. M., Chui, L., Ferrato, C., Lloyd, T., Li, V., & Pillai, D. R. (2022). Performance of four commercial real-time PCR assays for the detection of bacterial enteric pathogens in clinical samples. International Journal of Infectious Diseases, 114, 195–201. https://doi.org/10.1016/j.ijid.2021.10.035

Cheesbrough, M. (2006). District laboratory practice in tropical countries: Part 2. (2nd ed.). Cambridge University Press. https://www.cabidigitallibrary.org/doi/full/10.5555/20063178567

Debellut, F., Friedrich, A., Baral, R., Pecenka, C., Mugisha, E., & Neuzil, K. M. (2024). The cost of typhoid illness in low-and middle-income countries, a scoping review of the literature. PLoS One, 16(6), e0305692.

Dhayananth, B. (2024). Phenotypic and Molecular Characterization of Salmonellaserovars Isolated from Pigs, Pork and their Associated Environment. Indian Veterinary Research Institute.

Edel, B., Glöckner, S., Stoll, S., Lindig, N., Boden, K., Wassill, L., Simon, S., Löffler, B., & Rödel, J. (2023). Development of a rapid diagnostic test based on loop-mediated isothermal amplification to identify the most frequent non-typhoidal Salmonella serovars from culture. European Journal of Clinical Microbiology and Infectious Diseases, 42(4), 461–470. https://doi.org/10.1007/S10096-023-04571-3/TABLES/4

Fan, F., Du, P., Kan, B., & Yan, M. (2015). The Development and Evaluation of a Loop-Mediated Isothermal Amplification Method for the Rapid Detection of Salmonella enterica serovar Typhi. PloS One, 10(4), e0124507. https://doi.org/10.1371/journal.pone.0124507

Garg, N., Ahmad, F. J., & Kar, S. (2022). Recent advances in loop-mediated isothermal amplification (LAMP) for rapid and efficient detection of pathogens. Current Research in Microbial Sciences, 3, 100120. https://doi.org/10.1016/j.crmicr.2022.100120

Goay, Y. X., Chin, K. L., Tan, C. L. L., Yeoh, C. Y., Ja’afar, J. N., Zaidah, A. R., Chinni, S. V., & Phua, K. K. (2016). Identification of Five Novel Salmonella Typhi-Specific Genes as Markers for Diagnosis of Typhoid Fever Using Single-Gene Target PCR Assays. BioMed Research International, 2016, 8905675. https://doi.org/10.1155/2016/8905675

Hara-Kudo, Y., Yoshino, M., Kojima, T., & Ikedo, M. (2005). Loop-mediated isothermal amplification for the rapid detection of Salmonella. FEMS Microbiology Letters, 253(1), 155–161. https://doi.org/10.1016/J.FEMSLE.2005.09.032

Kuhn, K. G., Emborg, H. D., Krogfelt, K. A., & Mølbak, K. (2015). Detecting non-typhoid Salmonella in humans by enzyme-linked immunosorbent assays (ELISAs): Practical and epidemiological aspects. Salmonella: Methods and Protocols, 117–126. https://doi.org/10.1007/978-1-4939-1625-2_8

Lee, J. E., Mun, H., Kim, S. R., Kim, M. G., Chang, J. Y., & Shim, W. B. (2020). A colorimetric Loop-mediated isothermal amplification (LAMP) assay based on HRP-mimicking molecular beacon for the rapid detection of Vibrio parahaemolyticus. Biosensors and Bioelectronics, 151, 111968. https://doi.org/10.1016/j.bios.2019.111968

Mahmoud, A., Oluyemisi, A., Uwishema, O., Sun, J., Jobran, A. W., David, S., Wireko, A. A., Adanur, I., Dost, B., & Onyeaka, H. (2023). Recent advances in the diagnosis and management of typhoid fever in Africa: A review. International Journal of Health Planning and Management, 38(2), 317–329. https://doi.org/10.1002/hpm.3599

Manesh, A., Meltzer, E., Jin, C., Britto, C., Deodhar, D., Radha, S., & Rupali, P. (2021). Typhoid and paratyphoid fever: a clinical seminar. . Journal of Travel Medicine, 28(2), 12. https://doi.org/10.1093/jtm/taab012

Mina, S. A., Debnath, P., Hossain, A. K. M. Z., Hasan, M. Z., & Azad Chowdhury, A. M. M. (2024). Screening and identification of multiple antibiotic- resistant genes containing Salmonella Typhi from drinking water: A severe public health concern in Bangladesh. Heliyon, 10(22), e40523. https://doi.org/10.1016/j.heliyon.2024.e40523

Murthy, S., Hagedoorn, N. N., Faigan, S., Rathan, M. D., Marchello, C. S., & Crump, J. A. (2025, February 12). Global typhoid fever incidence: an updated systematic review with meta-analysis. https://doi.org/10.12688/verixiv.403.1

Nagamine, K., Hase, T., & Notomi, T. (2002). Accelerated reaction by loop-mediated isothermal amplification using loop primers. Molecular and Cellular Probes, 16(3), 223–229. https://doi.org/10.1006/MCPR.2002.0415

Neupane, D. P., Dulal, H. P., & Song, J. (2021). Enteric Fever Diagnosis: Current Challenges and Future Directions. Pathogens, 10(4), 410. https://doi.org/10.3390/pathogens10040410

Notomi, T., Mori, Y., Tomita, N., & Kanda, H. (2015). Loop-mediated isothermal amplification (LAMP): principle, features, and future prospects. Journal of Microbiology, 53(1), 1–5. https://doi.org/10.1007/s12275-015-4656-9

Oliveira, B. B., Veigas, B., & Baptista, P. V. (2021). Isothermal Amplification of Nucleic Acids: The Race for the Next “Gold Standard.” Frontiers in Sensors, 2. https://doi.org/10.3389/fsens.2021.752600

Ou, H., Wang, Y., Gao, J., Bai, J., Zhang, Q., Shi, L., Wang, X., & Wang, C. (2021). Rapid detection of Salmonella based on loop-mediated isothermal amplification. Annals of Palliative Medicine, 10(6), 6850–6858. https://doi.org/10.21037/apm-21-1387

Rao, X., & Sun, J. (2015). Development of SYBR Green I based real-time RT-PCR assay for specific detection of watermelon silver mottle Virus. Iranian Journal of Biotechnology, 13(3), 20. https://doi.org/10.15171/ijb.1124

Sayad, A. A., Ibrahim, F., Uddin, S. M., Pei, K. X., Mohktar, M. S., Madou, M., & Thong, K. L. (2016). A microfluidic lab-on-a-disc integrated loop mediated isothermal amplification for foodborne pathogen detection. Sensors and Actuators B. Chemical, 227, 600–609. https://doi.org/10.1016/j.snb.2015.10.116

Shi, C., Singh, P., Ranieri, M. L., Wiedmann, M., & Moreno Switt, A. I. (2015). Molecular methods for serovar determination of Salmonella. Critical Reviews in Microbiology, 41(3), 309–325. https://doi.org/10.3109/1040841X.2013.837862

Sohrab, M., Majidzadeh, K., Morovvati, A., & Soleimani, M. (2022). Real-Time Loop-Mediated Isothermal Amplification (LAMP) Method for Quantitative Salmonella Typhi Detection Based on ViaB Gene. Journal of Advanced Biomedical Sciences. https://doi.org/10.18502/jabs.v11i2.8769

Soroka, M., Wasowicz, B., & Rymaszewska, A. (2021). Loop-Mediated Isothermal Amplification (LAMP): The Better Sibling of PCR? Cells, 10(8), 1931. https://doi.org/10.3390/cells10081931

Sukri, L., Banza, A., Shafer, K., Sanoussi, Y., Neuzil, K. M., & Sani, R. (2024). Typhoid intestinal perforation in Francophone Africa, a scoping review. PLOS Global Public Health, 4(3), e0003056. https://doi.org/10.1371/journal.pgph.0003056

Sunar, N. M., Stewart, D. I., Fletcher, L. A., Stentiford, E. I., & Ashraf, M. A. (2016). Rapid propidium monoazide cPCR assay for exclusive quantification of viable Salmonella spp. cells. Journal of Environmental Biology, 37(5), 1057.

Tegene, B. A., & Eshetie, M. T. (2025). Evaluating alternatives to the Widal test for typhoid fever diagnosis in developing countries: A targeted literature review. Journal of Public Health in Africa, 16(1), 700. https://doi.org/10.4102/jphia.v16i1.700

Teh, C. S. J., Chua, K. H., & Thong, K. L. (2014). Paratyphoid Fever: Splicing the Global Analyses. International Journal of Medical Sciences, 11(7), 732–741. https://doi.org/10.7150/ijms.7768

Wang, Y., Zhang, L., Chen, Y., & Zhang, X. (2018). Comparative evaluation of loop-mediated isothermal amplification and PCR for rapid detection of Salmonella in food samples. Food Control, 90, 119–125.

WHO. (2024.). Measures for the control of non-typhoidal Salmonella spp. in poultry meat ... - Google Books. 2024. Retrieved January 14, 2025, from https://books.google.com.ng/books?hl=en&lr=&id=3aMOEQAAQBAJ&oi=fnd&pg=PA9&dq=who+2024+wash+services+salmonella&ots=4w20j9rfw-&sig=CH2QAO6UKG3k8W6e78D_ZG9MLHM&redir_esc=y#v=onepage&q&f=false

Yamasaki, E., Matsuzawa, S., Takeuchi, K., Morimoto, Y., Ikeda, T., Okumura, K., & Kurazono, H. (2021). Rapid Serotyping of Salmonella Isolates Based on Single Nucleotide Polymorphism-Like Sequence Profiles of a Salmonella- Specific Gene. Foodborne Pathogens and Disease, 18(1), 31–40. https://doi.org/10.1089/fpd.2020.2823

Yang, Q., Domesle, K. J., & Ge, B. (2018). Loop-Mediated Isothermal Amplification for Salmonella Detection in Food and Feed: Current Applications and Future Directions. Foodborne Pathogens and Disease, 15(6), 309–331. https://doi.org/10.1089/fpd.2018.2445

Zou, Y., Mason, M. G., & Botella, J. R. (2020). Evaluation and improvement of isothermal amplification methods for point-of-need plant disease diagnostics. PLOS ONE, 15(6), e0235216. https://doi.org/10.1371/journal.pone.0235216

Published

30-06-2025

How to Cite

Nasir, U. M., Musa, Y. I., Dantata, A., Rabiu, T. N., & Mohammed, A. G. (2025). Loop-Mediated Isothermal Amplification (LAMP) Assay for Rapid Detection of Salmonella enterica in Low-Resource Settings. UMYU Journal of Microbiology Research (UJMR), 10(3), 97–106. https://doi.org/10.47430/ujmr.25103.011