Diferenciación de especies de Rhodococcus mediante una prueba de PCR-RFLP basada en los genes codificantes para la subunidad 16S ribosomal

Diferenciación de especies de Rhodococcus mediante una prueba de PCR-RFLP basada en los genes codificantes para la subunidad 16S ribosomal

Considering the difficulty to differentiate the species ofRhodococcusby biochemical tests, a PolymeraseChain Reaction-Restriction Fragment Length Polymorphism (PCR-RFLP) has been developed to distinguishthe species of the genusRhodococcus.R. equi, R. rhodniiand other bacterial samples were cultured on Blood agar and BHI at 26 and 37∞C. The obtained DNA was amplified with the primers described by Hypsaand Dale. Products of amplification were submitted to digestion with multiple restriction enzymes and therestriction patterns were confirmed by usingin silicoanalysis. The expected amplification fragment of 1300pb was obtained in all tested samples.R. equiwas distinguished fromR. rhodniiby using endonucleases such as PstI and HindIII, and from other bacteria with endonucleasesPstI, HindIII, SstI, BamHIandEcoRI. Theobtained restriction patterns were confirmed by usingin silicoanalysis. This PCR-RFLP establishes analternative for the differentiation among species of the genusRhodococcussuch asR. equiandR. rhodnii.

1. Murria P, Rosenthal K, Kobayashi G. Nocardia, Rhodococcus and related Actinomycetes. En: Medical Microbiology, Tercera edición, Editorial Mosby, USA 1998.
2. Kedlaya I, Ing MB, Wong SS. Rhodococcus equi infections in immunocompetent hosts: case report and review. Clin Infect Dis 2001; 32: 39-46.
3. Weinstock DM, Brown AE. Rhodococcus equi: an emerging pathogen. Clin Infect Dis 2002; 34:1379-1385.
4. Prescott JF. Rhodococcus equi: an animal and human pathogen. Clin Microbiol Rev 1991; 4: 20-34.
5. Hill P, Campbell JA, Petrie LA. Rhodnius prolixus and its symbiotic actinomycete: a microbiological, physiological and behavioural study. Proc R Soc Lond B, Biol Sci Sp 1976; 194: 501-525.
6. Yassin AF. Rhodococcus triatomae sp. nov., isolated from a bloodsucking bug. Int J Syst Evol Microbiol 2005; 55: 1575-1579.
7. Eichler S, Schaub, GA. Development of symbionts in triatomine bugs and the effects of infections with
8. trypanosomatids. Exp Parasitol 2002; 100: 17-27.
9. Beard C, Cordon-Rosales C, Durvasula R. Bacterial symbionts of the triatominae and their potential use in control of Chagas disease transmission. Annu Rev Entomol 2002; 47: 123-141.
10. Beard C, Mason P, Aksoy S, Tesh R, Richards F. Transformation of an insect symbiont and expression of a
11. foreign gene in the Chagas´ disease vector Rhodnius prolixus. Am J Trop Med Hyg 1992; 46: 195-200.
12. Dotson EM, Plikaytis B, Shinnick TM, Durvasula RV, Beard CB. Transformation of Rhodococcus rhodnii, a symbiont of the Chagas disease vector Rhodnius prolixus, with integrative elements of the L1 mycobacteriophage. Infect Genet Evol 2003; 3: 103-109.
13. Shoemaker SA, Fisher JH, Jones WD Jr. Scoggin CH. Restriction fragment analysis of chromosomal DNA defines
14. different strains of Mycobacterium tuberculosis. Am Rev Respir Dis 1986; 134: 210-213.
15. Sambrook JF, Russell DW. Molecular Cloning, A Laboratory Manual. Tercera edición, Cold Spring Harbor Laboratory Press, New York 2001.
16. Hypsa V, Dale C. In vitro culture and phylogenetic analysis of .Candidatus Arsenophonus triatominarum. an intracellular bacterium from the triatomine bug, Triatoma infestans. Int J
17. Syst Bacteriol 1997; 47: 1140-1144.
18. Bell KS, Philp JC, Aw DW. The genus Rhodococcus. J Appl Microbiol 1998; 85: 195-210.
19. Yamada H, Kobayashi M. Nitrile hydratase and its application to industrial production of acrylamide. Biosci Biotechnol Biochem 1996; 60: 1391-1400.
20. ----------------------------------------------------------------------------
21. DOI: http://dx.doi.org/10.22490/24629448.332