Aptamer-nanobody based ELASA for detection of Vibrio cholerae O1
Background and Objectives: In recent years, the prevalence of diseases caused by Vibrio spp. is increasing in the world, and among them species, Vibrio cholerae is the most important Vibrio associated with pandemic and epidemic cholera outbreaks. Therefore, the development of a reliable method for early and accurate detection of V. cholerae for management of diseases is a real need. Aptamers with the ability to detect targets with high specificity and accuracy can be one of the candidates used for the whole cell and thereby V. cholerae detection.
Materials and Methods: In this research high-affinity DNA aptamers against with two major serotypes of Inaba (ATCC 39315) and Ogawa (clinical sample) were selected from DNA aptamer library through 12 rounds of Systematic Evolution of Ligands by Exponential (SELEX) enrichment procedure using live cells as a target which monitored with flow cytometry.
Results: The binding efficiency and dissociation constant of the isolated aptamers V.ch47 and V.ch27 were 56.4%, 53.3% and 15.404 ± 4.776 pM, 20.186 ± 3.655 pM, respectively. A sandwich Enzyme-linked aptamer sorbent assay (ELASA) was developed with the biotinylated V.ch47 aptamer and our previously developed nanobody anti-Lipopolysaccharides (LPS). We optimized this system with V. cholerae O1 and analyzed their cross reactivity with close physiological bacteria. The threshold of detection was obtained 104 CFU/ml in the sandwich ELASA process.
Conclusion: Our results showed that the sandwich ELASA is sensitive enough for the rapid detection of V. cholerae from other bacteria.
2. Sánchez J, Holmgren J. Cholera toxin—a foe & a friend. Indian J Med Res 2011;133: 153-163.
3. Chowdhury FR, Nur Z, Hassan N, von Seidlein L, Dunachie S. Pandemics, pathogenicity and changing molecular epidemiology of cholera in the era of global warming. Ann Clin Microbiol Antimicrob 2017;16:10.
4. Lipp EK, Rivera IN, Gil AI, Espeland EM, Choopun N, Louis VR, et al. Direct detection of Vibrio cholerae and ctxA in Peruvian coastal water and plankton by PCR. Appl Environ Microbiol 2003;69:3676-3680.
5. Gubala AJ. Multiplex real-time PCR detection of Vibrio cholerae. J Microbiol Methods 2006;65:278-293.
6. Blackstone GM, Nordstrom JL, Bowen MD, Meyer RF, Imbro P, DePaola A. Use of a real time PCR assay for detection of the ctxA gene of Vibrio cholerae in an environmental survey of Mobile Bay. J Microbiol Methods 2007;68:254-259.
7. Jyoung J-Y, Hong S, Lee W, Choi J-W. Immunosensor for the detection of Vibrio cholerae O1 using surface plasmon resonance. Biosens Bioelectron 2006;21:2315-2319.
8. Chakraborty S, Alam M, Scobie HM, Sack DA. Adaptation of a simple dipstick test for detection of Vibrio cholerae O1 and O139 in environmental water. Front Microbiol 2013;4: 320.
9. Yi-Xian W, Zun-Zhong Y, Cheng-Yan S, Yi-Bin Y. Application of aptamer based biosensors for detection of pathogenic microorganisms. Chinese J Anal Chem 2012;40:634-642.
10. Van Dorst B, Mehta J, Bekaert K, Rouah-Martin E, De Coen W, Dubruel P, et al. Recent advances in recognition elements of food and environmental biosensors: a review. Biosens Bioelectron 2010;26:1178-1194.
11. Bitaraf FS, Rasooli I, Mousavi Gargari SL. DNA aptamers for the detection of Haemophilus influenzae type b by cell SELEX. Eur J Clin Microbiol Infect Dis 2016;35:503-510.
12. Moon J, Kim G, Park S, Lim J, Mo C. Comparison of whole-cell SELEX methods for the identification of Staphylococcus aureus-specific DNA aptamers. Sensors (Basel) 2015;15:8884-8897.
13. Mirzakhani K, Gargari SLM, Rasooli I, Rasoulinejad S. Development of a DNA aptamer for screening Neisseria meningitidis Serogroup B by cell SELEX. Iran Biomed J 2017;22:193-201.
14. Ebrahimizadeh W, Mousavi Gargari S, Rajabibazl M, Safaee Ardekani L, Zare H, Bakherad H. Isolation and characterization of protective anti-LPS nanobody against V. cholerae O1 recognizing Inaba and Ogawa serotypes. Appl Microbiol Biotechnol 2013;97:4457-4466.
15. Mondal B, Ramlal S, Lavu PS, Murali HS, Batra HV. A combinatorial systematic evolution of ligands by exponential enrichment method for selection of aptamer against protein targets. Appl Microbiol Biotechnol 2015;99:9791-9803.
16. Moon J, Kim G, Lee S, Park S. Identification of Salmonella Typhimurium-specific DNA aptamers developed using whole-cell SELEX and FACS analysis. J Microbiol Methods 2013;95:162-166.
17. Baranova DE, Levinson KJ, Mantis NJ. Vibrio cholerae O1 secretes an extracellular matrix in response to antibody-mediated agglutination. PLoS One 2018;13(1):e0190026.
18. Rasoulinejad S, Gargari SLM. Aptamer-nanobody based ELASA for specific detection of Acinetobacter baumannii isolates. J Biotechnol 2016;231:46-54.
19. Wandtke T, Woźniak J, Kopiński P. Aptamers in diagnostics and treatment of viral infections. Viruses 2015;7:751-780.
20. Dassie JP, Hernandez LI, Thomas GS, Long ME, Rockey WM, Howell CA, et al. Targeted inhibition of prostate cancer metastases with an RNA aptamer to prostate-specific membrane antigen. Mol Ther 2014;22:1910-1922.
21. Kimoto M, Yamashige R, Matsunaga K-i, Yokoyama S, Hirao I. Generation of high-affinity DNA aptamers using an expanded genetic alphabet. Nat Biotechnol 2013;31:453-457.
22. Alfavian H, Mousavi Gargari SL, Rasoulinejad S, Medhat A. Development of a DNA aptamer that binds specifically to group A Streptococcus serotype M3. Can J Microbiol 2017;63:160-168.
23. Almasi F, Mousavi Gargari SL, Bitaraf F, Rasoulinejad S. Development of a single stranded DNA aptamer as a molecular probe for LNCap cells using cell-SELEX. avicenna. Avicenna J Med Biotechnol 2016;8:104-111.
24. Lee YJ, Han S, Maeng J-S, Cho Y-J, Lee S-W. In vitro selection of Escherichia coli O157:H7-specific RNA aptamer. Biochem Biophys Res Commun 2011;417:414-420.
25. Yu X, Chen F, Wang R, Li Y. Whole-bacterium SELEX of DNA aptamers for rapid detection of E. coli O157:H7 using a QCM sensor. J Biotechnol 2018;266:39-49.
26. Toh SY, Citartan M, Gopinath SC, Tang T-H. Aptamers as a replacement for antibodies in enzyme-linked immunosorbent assay. Biosens Bioelectron 2015;64:392-403.
27. Groff K, Brown J, Clippinger AJ. Modern affinity reagents: Recombinant antibodies and aptamers. Biotechnol Adv 2015;33:1787-1798.
28. Wang P, Hatcher KL, Bartz JC, Chen SG, Skinner P, Richt J, et al. Selection and characterization of DNA aptamers against PrP(Sc). Exp Biol Med (Maywood) 2011;236:466-476.
29. Chen F, Hu Y, Li D, Chen H, Zhang X-L. CS-SELEX generates high-affinity ssDNA aptamers as molecular probes for hepatitis C virus envelope glycoprotein E2. PLoS One 2009;4(12):e8142.
30. Payandeh Z, Rasooli I, Mousavi Gargari SL, Rajabi Bazl M, Ebrahimizadeh W. Immunoreaction of a recombinant nanobody from camelid single domain antibody fragment with Acinetobacter baumannii. Trans R Soc Trop Med Hyg 2014;108:92-98.
31. Tuteja U, Kumar S, Shukla J, Kingston J, Batra HV. Simultaneous direct detection of toxigenic and non-toxigenic Vibrio cholerae from rectal swabs and environmental samples by sandwich ELISA. J Med Microbiol 2007;56:1340-1345.
32. Nato F, Boutonnier A, Rajerison M, Grosjean P, Dartevelle S, Guénolé A, et al. One-step immunochromatographic dipstick tests for rapid detection of Vibrio cholerae O1 and O139 in stool samples. Clin Diagn Lab Immunol 2003;10:476-478.
33. Yamasaki E, Sakamoto R, Matsumoto T, Morimatsu F, Kurazono T, Hiroi T, et al. Development of an immunochromatographic test strip for detection of cholera toxin. Biomed Res Int 2013;2013:679038.
34. Bayat M, Khabiri A, Hemati B. Development of IgY-based sandwich ELISA as a robust tool for rapid detection and discrimination of toxigenic Vibrio cholerae. Can J Infect Dis Med Microbiol 2018;2018:4032531.
35. Schofield CL, Field RA, Russell DA. Glyconanoparticles for the colorimetric detection of Cholera toxin. Anal Chem 2007;79:1356-1361.
This work is licensed under a Creative Commons Attribution-NonCommercial 4.0 International License.