ESTABLISHMENT OF MULTIPLEX PCR ASSAY FOR SIMULTANEOUS DETECTION OF SALMONELLA SPP., SHIGELLA SPP. AND ENTAMOEBA HISTOLYTICA
Main Article Content
Abstract
Objective: To establish a novel multiplex PCR assay for simultaneous detection
of foodborne pathogens, including Salmonella spp., Shigella spp., and Entamoeba
histolytica, with high sensitivity, specificity, cost-effectiveness and time-saving.
Method: Multiplex PCR, Semi-Nested PCR and melting analysis for specifically
amplifying and distinguishing three highly conserved target gene regions (invA,
ipaH, 18S rRNA), in order to simultaneously detect Salmonella spp., Shigella spp.,
Entamoeba histolytica in a single reaction tube. Specific pathogens are characterized
based on the melting profiles of the PCR products collected on a realtime PCR
instrument following the second round of amplification.
Results: The established multiplex PCR assay is capable of simultaneously detecting
Salmonella spp., Shigella spp., and Entamoeba histolytica at a limit of detection of
5.86 copies/µl, 24.4 copies/µl và 53.6 copies/µl, respectively (confidence interval of
95%), the specificity reached 100% when evaluated with multiple microbial species
provided by the Department of Microbiology, 103 Military Hospital.
Conclusion: The novel multiplex PCR assay for simultaneous detection of Salmonella
spp., Shigella spp., Entamoeba histolytica has been successfully established.
Article Details
Keywords
Multiplex PCR, Semi-Nested PCR, Salmonella spp., Shigella spp., Entamoeba histolytica.
References
Rec, 72(14): p. 97-9.
[2] Havelaar, A. H., M. D. Kirk, P. R.
Torgerson, H. J. Gibb, T. Hald, R. J.
Lake, N. Praet, D. C. Bellinger, N. R. de
Silva, N. Gargouri, N. Speybroeck, A.
Cawthorne, C. Mathers, C. Stein, F. J.
Angulo, B. Devleesschauwer, and Group
World Health Organization Foodborne
Disease Burden Epidemiology
Reference, (2015),”World Health
Organization Global Estimates and
Regional Comparisons of the Burden of
Foodborne Disease in 2010”. PLoS Med,
12(12): p. e1001923.
[3] Alena Klochko, Francisco Talavera,
John L Brusch et al., (2022) “Salmonella
Infection (Salmonellosis) Treatment &
Management”. Medscape.
[4] Mohle-Boetani, J. C., M. Stapleton,
R. Finger, N. H. Bean, J. Poundstone,
P. A. Blake, and P. M. Griffin,
(1995),”Communitywide shigellosis:
control of an outbreak and risk factors
in child day-care centers”. Am J Public
Health, 85(6): p. 812-6.
[5] Nyenje, M. E., C. E. Odjadjare, N.
F. Tanih, E. Green, and R. N. Ndip,
(2012),”Foodborne pathogens recovered
from ready-to-eat foods from roadside
cafeterias and retail outlets in Alice,
Eastern Cape Province, South Africa:
public health implications”. Int J Environ
Res Public Health, 9(8): p. 2608-19.
[6] Nygren, B. L., K. A. Schilling, E. M.
Blanton, B. J. Silk, D. J. Cole, and E.
D. Mintz, (2013),”Foodborne outbreaks
of shigellosis in the USA, 1998-2008”.
Epidemiol Infect, 141(2): p. 233-41.
[7] Rabsch, W., H. Tschape, and A. J.
Baumler, (2001),”Non-typhoidal
salmonellosis: emerging problems”.
Microbes Infect, 3(3): p. 237-47.
[8] Tanyuksel, M. and W. A. Petri, Jr.,
(2003),”Laboratory diagnosis of amebiasis”.
Clin Microbiol Rev, 16(4): p. 713-29.
[9] Waites, W. M. and J. P. Arbuthnott,
(1990),”Foodborne illness: an overview”.
Lancet, 336(8717): p. 722-5.