Volume 6, Issue 4 (12-2019)                   nbr 2019, 6(4): 374-381 | Back to browse issues page

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Kabiri K, Majidzadeh K. The molecular detection of the causative agent of plague on the basis of the pla gene. nbr. 2019; 6 (4) :374-381
URL: http://nbr.khu.ac.ir/article-1-3084-en.html
Department of Microbiology, North Tehran Branch, Islamic Azad University, Tehran, Iran , Kabiri.kh93@ gmail.com
Abstract:   (48 Views)
Yersinia pestis, a gram-negative rod belonging to the Enterobacteriaceae family, is the causative agent of plague. Classical methods of detecting the organisms are time-consuming, expensive and dangerous. The aim of the study was to design a Real-time PCR assay on the basis of the pla gene of Yersinia pestis. In this research the Real- time PCR test was optimized by using special primers for targeting pla gene. After preparing 10-fold serial dilutions of the pla and their analysis by the assay, the last dilution showing a fluorescent signal was confirmed as the limit of detection (LOD). A standard curve based on the Ct values was depicted, so the assay was developed to quantify the target gene. The analytical specificity was determined by subjecting the genome of some control negative bacteria to the assay. In this experiment, negative control genomes did not show detectable signals in the assay. The last dilution of pla plasmid which showed a fluorescent signal was 4.5 fg. So, the lower detectable copy numbers of the gene in a 20 μl PCR reaction was calculated as 1×103.
Full-Text [PDF 872 kb]   (23 Downloads)    
Type of Study: Original Article | Subject: Microbiology
Received: 2018/02/11 | Revised: 2020/01/20 | Accepted: 2019/05/12 | Published: 2020/01/8 | ePublished: 2020/01/8

1. Arya, M., Shergill, I.S., Williamson, M., Gommersall, L., Arya, N. and Patel, H.R. 2005. Basic principles of real-time quantitative PCR. - Expert Rev. Mol. Diagn. 5: 209-219. [DOI:10.1586/14737159.5.2.209]
2. Ayyadurai, S., Flaudrops, C., Raoult, D. and Drancourt, M. 2010. Rapid identification and typing of Yersinia pestis and other Yersinia species by matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) mass spectrometry. - BMC Microbiol. 10: 1-7. [DOI:10.1186/1471-2180-10-285]
3. Azizi, M.H. and Azizi, F. 2010. A history of the human plague in Iran. - Arch. Iran Med. 13: 563-569.
4. Chase, C.J., Ulrich, M.P., Wasieloski, L.P., Kondig, J.P., Garrison, J. and Lindler, L.E. 2005. Real-time PCR assays targeting a unique chromosomal sequence of Yersinia pestis. - Adv. Clin. Chem. 51: 1778-1785. [DOI:10.1373/clinchem.2005.051839]
5. Drancourt, M. and Raoult, D. 2002. Molecular insights into the history of plague . - Microbes Infect. 4: 105-109. [DOI:10.1016/S1286-4579(01)01515-5]
6. Engelthaler, D.M., Gage, K.L., Montenieri, J.A., Chu, M. and Carter, L.G. 1999. PCR detection of Yersinia pestis in fleas: comparison with mouse inoculation. - J. Clin. Microbiol. 37: 1980-1984. [DOI:10.1128/JCM.37.6.1980-1984.1999]
7. Esamaeili, S., Azadmanesh, K., Naddaf, S.R. Rajerison, M., Carniel, E. and Mostafavi, E. 2013. Serologic survey of plague in animals, western Iran. - J. Emerg. Infect. Dis. 19: 1549-1551.. [DOI:10.3201/eid1909.121829]
8. Espy, M.J., Uhl, J.R., Sloan, L.M., Buckwalter, S.P., Jones, M.F., Vetter, E.A., Yao, J.D., Wengenack, N.L., Rosenblatt, J.E., Cockerill, F.R. and Smith, T.F. 2006. Real-time PCR in clinical microbiology: applications for routine laboratory testing. - Clin. Microbiol. Rev. 19: 165-256. [DOI:10.1128/CMR.19.1.165-256.2006]
9. Filippov .A.A., Sergueev, K.V., He, Y., Huang, X.Z., Gnade, B.T. and Mueller, A.J. 2011. Bacteriophage-resistant mutants in Yersinia pestis: identification of phage receptors and attenuation for mice. - PLOS ONE 6: 1-11. [DOI:10.1371/journal.pone.0025486]
10. Kenny, J.H., Zhou, Y., Schriefer, M.E. and Bearden, S.W. 2008. Detection of viable Yersinia pestis by fluorescence in situ hybridization using peptide nucleic acid probes. - J. Microbiol. Methods. 75: 293-301. [DOI:10.1016/j.mimet.2008.06.021]
11. Leal, N.C and Almeida, A.M. 1999. Diagnosis of plague and identification of virulence markers in Yersinia pestis by multiplex PCR. - Rev. Inst. Med. Trop. Sao Paulo. 41: 339-342. [DOI:10.1590/S0036-46651999000600002]
12. Lindler, L.E., Fan, W. and Jahan, N. 2001. Detection of ciprofloxacin-resistantYersinia pestis by fluorogenic PCR using the light Cycler. - Eur. J. Clin. Microbiol. Infect. Dis. 39: 3649-3655. [DOI:10.1128/JCM.39.10.3649-3655.2001]
13. Loiez, C., Herwegh, S., Wallet, F., Armand, S., Guinet, F. and Courcol, R.J. 2003. Detection of Yersinia pestis in sputum by real-time PCR. - Eur. J. Clin. Microbiol. 10: 190-212. [DOI:10.1128/JCM.41.10.4873-4875.2003]
14. Mackay, I.M. 2004. Real‐time PCR in the microbiology laboratory. - Clin. Microbiol. Infect. 10: 190-212. [DOI:10.1111/j.1198-743X.2004.00722.x]
15. Matero, P., Pasanen, T., Laukkanen, R., Tissari, P., Tarkka, E. and Vaara M. 2009. Real‐time multiplex PCR assay for detection of Yersinia pestis and Yersinia pseudotuberculosis. - Apmis. 117:34-44. [DOI:10.1111/j.1600-0463.2008.00013.x]
16. O'Hara, C.M. 2005. Manual and automated instrumentation for identification of Enterobacteriaceas and other aerobic gram-negative bacilli. - Clin. Microbiol. Rev. 18: 147-162. [DOI:10.1128/CMR.18.1.147-162.2005]
17. Perry, R.D. and Fetherston, J.D. 1999. Yersinia pestis etiologic agent of plague. - Clin. Microbiol. Rev. 10: 35-66. [DOI:10.1128/CMR.10.1.35]
18. Qu, S., Shi, Q,. Zhou, L., Guo, Z., Zhou, D. and Zhai, J. 2010. Ambient stable quantitative PCR reagents for the detection of Yersinia pestis. - PLOS ONE 4: 1-7. [DOI:10.1371/journal.pntd.0000629]
19. Rachwal, P.A., Rose, H.L., Cox ,V., Lukaszewski, R.A., Murch, A.L. and Weller, S.A. 2012. The potential of TaqMan Array Cards for detection of multiple biological agents by real-time PCR. - PLOS ONE 7: 1-7. [DOI:10.1371/journal.pone.0035971]
20. Riehm, J.M., Rahalison, L., Scholz, H.C., Thoma, B., Pfeffer, M. and Razanakoto, L.M. 2011. Detection of Yersinia pestis using real-time PCR in patients with suspected bubonic plague. - Mol. Cell Probes. 25: 8-12. [DOI:10.1016/j.mcp.2010.09.002]
21. Rollins, S.E., Rollins, S.M. and Ryan, E.T. 2003. Yersinia pestis and the plague. - Am. J. Clin. Pathol. 119: 78-85. [DOI:10.1309/DQM93R8QNQWBFYU8]
22. Schofield, D.A., Molineux, I.J. and Westwate, C. 2009. Diagnostic bioluminescent phage for detection of Yersinia pestis. - J. Clin. Microbiol. 47: 3887-3894. [DOI:10.1128/JCM.01533-09]
23. Sergueev, K.V., Borschel, R.H., Nikolich, M.P. and Filippov, A.A. 2010. Rapid and sensitive detection of Yersinia pestis using amplification of plague diagnostic bacteriophages monitored by real-time PCR. - PLOS ONE 5: 1-10. [DOI:10.1371/journal.pone.0011337]
24. Stewart, A., Satterfield. B., Cohen, M., O'Neill, K. and Robison, R. 2008. A quadruplex real-time PCR assay for the detection of Yersinia pestis and its plasmids. - J. Med. Microbiol. 57: 324-331. [DOI:10.1099/jmm.0.47485-0]
25. Suzuki, S., Sakakibara, H. and Hotta, S. 1997. Latex agglutination tests for measurement of antiplague antibodies. - Eur. J. Clin. Microbiol. Infect. Dis. 6: 332-336.
26. Tomaso, H., Reisinger, E.C., Dahouk, S., Frangoulidis, D., Rakin., A. and Landt, O. 2006. Rapid detection of Yersinia pestis with multiplex real‐time PCR assays using fluorescent hybridisation probes. - FEMS Immunol. Med. Microbiol. 38: 117-26. [DOI:10.1016/S0928-8244(03)00184-6]
27. Tomaso, H., Thullier, P., Seibold, E., Guglielmo, V., Buckendahl, A. and Rahalison, L. 2007. Comparison of hand-held test kits, immunofluorescence microscopy, enzyme-linked immunosorbent assay, and flow cytometric analysis for rapid presumptive identification of Yersinia pestis. - Eur. J. Clin. Microbiol. Infect. Dis. 45: 3404-3407. [DOI:10.1128/JCM.00458-07]
28. Tomaso, H., Reisinger, E.C., Dahouk, S., Frangoulidis, D., Rakin, A. and Landt, O. 2003. Rapid detection of Yersinia pestis with multiplex real‐time PCR assays using fluorescent hybridisation probes. - FEMS Microbiol. Immunol. 38: 117-126. [DOI:10.1016/S0928-8244(03)00184-6]
29. Wang, J.Z., Duan, R., Liang, J.R., Huang, Y., Xiao, Y.C, Qiu, H. and Wang, X. 2014. Real-time TaqMan PCR for Yersinia enterocolitica detection based on the ail and fox A Genes - Eur. J. Clin. Microbiol. Infect. Dis. 52: 4443-4444. [DOI:10.1128/JCM.02528-14]
30. Wilmoth, B.A., Chu, M.C. and Quan, T.J. 1996. Identification of Yersinia pestis by BBL crystal enteric / nonfermentor identification system. - Eur. J. Clin. Microbiol. Infect. Dis. 34: 2829-2830. [DOI:10.1128/JCM.34.11.2829-2830.1996]
31. Woron, A.M., Nazarian, E.J., Egan, C., McDonough, K.A., Cirino, N.M. and Limberger, R.J. 2006. Development and evaluation of a 4-target multiplex real-time polymerase chain reaction assay for the detection and characterization of Yersinia pestis. - Diagn. Microbiol. Infect. Dis. 56: 261-268 [DOI:10.1016/j.diagmicrobio.2006.06.009]
32. Zhang. Z., Liang. Y., Yu. D., Xia. L. and Hai. R. 2013. Development of a multiplex polymerase chain reaction (PCR) with an internal control method to detect Yersinia pestis in the plague foci surveillance. - Afr. J. Microbiol. Res.7: 698-700.

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