Liotta DJ, Cabanne G, Campos R, Andrés TS

Language: English
References: 23
Page: 82-87
PDF size: 184.88 Kb.

ABSTRACT

Most molecular protocols for Dengue virus detection described so far are time consuming and cumbersome with mosquito samples. In order to count with a sensitive and specific molecular detection system for monitoring possible Dengue outbreaks and circulating viral serotypes in field-caught Aedes aegypti populations from Northeastern Argentina, a RT-PCR and RFLP assay was developed. The original RT-PCR assay proposed by Sudiro et al. for human serum was optimized for mosquito samples. Modifications were done at the RNA extraction-purification and at the thermal profile steps. The generic 230 bp amplicon was validated by RFLP assay and cycle sequencing. Results showed that, due to the generic characteristic of the primers used, certain mosquito genome regions could be co-amplified, making confirmation of the Dengue specific amplicon by RFLP assay a required step. Under these conditions, the proposed method can be employed as a Dengue viral generic screening procedure in Aedes aegypti mosquito samples, giving in our hands an estimated 99.52% of confirmed negatives (207/208 tested samples).

REFERENCES

1. Acha, P. and Szyfres, B., 1997. In: Zoonosis y enfermedades transmisibles comunes al hombre y a los animales. Pan American Health Organization (Editor), Washington D.C.; Scientific publication No. 503, pp 302-305.

2. Avilés, G., Rangeón, G., Vorndam, V., Briones, A., Baroni, P., Enria, D., Sabattini, M.S., 1999. Dengue Reemergence in Argentina. Emerg Infect Dis 5(4): 575-578.

3. Avilés, G., Paz, V., Rangeón, G., Ranaivoarisoa, M. Verzeri, N., Roginski, S., Baroni, P., Enria, 2003. Laboratory Surveillance of Dengue in Argentina, 1995-2001. Emerg Infect Dis 9(6): 738-742.

4. Boffi, R. 1998. Programa de prevención del Dengue y control del Aedes aegypti. In: Temas de Zoonosis y enfermedades emergentes. II Congreso Argentino de Zoonosis, I Congreso Argentino y Latinoamericano de Enfermedades Emergentes y Asociación Argentina de Zoonosis (Editors), pp 413-419.

5. Boom, R., Sol, C.J., Salimans, M.M., Jansen, C.L., Wertheim-van Dillen, P.M., van der Noorda, J., 1990. Rapid and simple method for purification of nucleic acids. J Clin Microbiol 28: 495-503.

6. Chow, V., Chan, Y., Yong, R., Lee, M., Lim, L., Chung, Y., Lam-Phua, S., Tan, B., 1998. Monitoring of Dengue Viruses in Field-Caught Aedes aegypti and Aedes albopictus Mosquitoes by a Type-specific Polimerase Chain Reaction and Cycle Sequencing. Am J Trop Med Hyg 58(5): 578-586.

7. Consoli, R. and de Oliveira, R., 1994. Principais Mosquitos de Importancia Sanitaria no Brasil. Fiocruz Editor, Brazil, pp 104-117.

8. Deubel, V., Laille, V., Hugnot, J., Chunge, E., Guesdon, J., Drouet, M., Bassot, S., Chevrier, D., 1990. Identification of Dengue sequences by genomic amplification: rapid diagnosis of Dengue Virus serotypes in peripheral blood. J Virol Methods 30: 41-54.

9. Harris, E., Roberts, G., Smith, L., Selle, J., Kramer, L., Valle, S., Sandoval, E., Balmaseda, A., 1998. Typing of Dengue viruses in clinical specimens and mosquitoes by single-tube multiplex Reverse Transcriptase PCR. J Clin Microbiol 36: 2634-2639.

10. Jensen, T., 1994. Comparison of Bi-directional Fay, Omni-directional, CDC, and duplex cone traps for sampling adult Aedes albopictus and Aedes aegypti in north Florida. J Am Mosq Control Assoc 10(1): 74-78.

11. Kourí, G., Guzmán, M.G., Valdés, L., Carbonel, I., del Rosario, D., Vazquez, S., Laferté, J., Delgado, J., Cabrera, M., 1998. Reemergence of Dengue in Cuba: A 1997 Epidemic in Santiago de Cuba. Emerg Infect Dis 4(1): 89-92.

12. Kuno, G., 1998. Universal diagnostic RT-PCR protocol for Arboviruses. J Virol Methods 72: 27-41.

13. Lanciotti, R.S., Calisher, C., Gubler, D., Chang, G., Vorndam, V., 1992. Rapid detection and typing of Dengue Viruses from clinical samples by using Reverse Transcriptase-Polymerase Chain Reaction. J Clin Virol 30: 545-551.

14. León, T., 2001. Epidemiology Chief, National Ministery of Health, Republic of Paraguay. Personal communication.

15. Martínez Torres, E., 1998. Dengue y Dengue Hemorrágico. Universidad Nacional de Quilmes & Elea Laboratorio (Editors), pp 43-56.

16. Miagostovich, M.P., Nogueira, R.M., Schatzmayr, H., Lanciotti, R., 1998. Molecular Epidemiology of Den-2 Virus in Brazil. Mem Inst Oswaldo Cruz 93(5): 625-626.

17. Pan American Health Organization, 1994. Dengue and Dengue Haemorrhagic Fever: guidelines for prevention and control. Washington D.C. Scientific publication No. 548.

18. Peters, C.J., 1997. Viral Hemorrhagic Fevers. In: Viral Pathogenesis. Nathanson N. (Editor). Lippincott-Raven Publishers, Philadelphia, pp 779-799.

19. Sabattini, M.S., Avilés, G., Monath, T.P., 1998. Historical, epidemiological, and ecological aspects of arboviruses in Argentina: Flaviviridae, Bunyaviridae and Rhabdoviridae. In: An overview of arbovirology in Brazil and neighboring countries. Travassos da Rosa, Vasconcelos, Travassos da Rosa (Editors). Belem, Brazil. Instituto Evandro Chagas, pp 113-134.

20. Sudiro, T., Ishiko, H., Green, S., Vaughn, D., Nisalak, A., Kalayanarooj, S., Rothman, A., 1997. Rapid diagnosis of Dengue viremia by Reverse Transcriptase-Polymerase Chain Reaction using 3´-noncoding region universal primers. Am J Trop Med Hyg 56: 424-429.

21. Tanaka, M., 1993. Rapid identification of Flavivirus using the Polymerase Chain Reaction. J Virol Methods 41: 311-322.

22. Wallace, D.M., 1987. In: Methods in Enzymology. Berger and Kimmar (Editors), Academic Press. Molecular Cloning Techniques Series 152, pp 33-41.

23. World Health Organization, 1997. Dengue Haemorrhagic Fever: Diagnosis, treatment, prevention and control. 2nd Edition, Geneva, pp 60-66.