ARTICULOS ORIGINALES

Detección e identificación por PCR de microorganismos causantes de meningoencefalitis bacteriana.

Detection and identification by PCR of microorganisms causing bacterial meningocephalitis.

Yanet Climent1, Isabel Martínez1, Niurys Nuñez1, Mónica Ginebra1, Leydis Zamora1, Mercedes Gutiérrez1, Francklin Sotolongo1, Armando Acosta1, Luis W Climent2.

RESUMEN

Reportamos el empleo de cebadores para la detección e identificación de microorganismos causantes de meningoencefalitis bacteriana (MEB) en 24 muestras de líquido cefalorraquídeo (LCR) y a partir de ADN genómico de cepas de referencia previamente purificado. Se emplearon cebadores generales y específicos derivados del gen 16S ARNr (ARN ribosómico). Los cebadores generales amplificaron en todas las especies bacterianas analizadas: Neisseria meningitidis, Haemophilus influenzae tipo b, Streptococcus sp y Staphylococcus aureus, en una misma banda de 825 pb. Los cebadores específicos solamente amplificaron en el microorganismo del que se derivan, obteniendo bandas de 667, 478 y 253 pb para N. meningitidis, H. influenzae y Streptococcus sp, respectivamente. También se diferenció Streptococcus agalactiae mediante reacción en cadena de la polimerasa (PCR), donde se obtuvo una banda de 478 pb. Al evaluar estos cebadores en muestras clínicas de LCR se constató que hubo una sensibilidad del 100% de las MEB por PCR; la especificidad fue del 100%. Los productos de PCR se analizaron por digestión Hae III, encontrando correspondencia en los patrones de restricción de las muestras clínicas y las cepas de referencia.

Palabras claves: ARN ribosómico, Bacterias, gen, LCR, MEB, PCR.

ABSTRACT

Our work reports the use of genomic DNA primers previously purified from reference strains to detect and identify the microorganisms that cause bacterial meningocephalitis (BME) in 24 samples of cerebrospinal fluid (CSF). General and specific primers derived from the 16S rRNA (ribosomal RNA) gene were used. The general primers were amplified in all the bacterial species analyzed:Neisseria meningitidis, type b Haemophylus influenzae and Streptococcus sp. in the same 825 pb band, but the specific ones only amplified in the microorganism from which they were derived. Bands of 667, 478 and 253 pb were respectively obtained for N. meningitidis,H. influenzae and Streptococcus sp. Streptococcus agalactiae was also detected by PCR in which a 478 pb band was obtained. Upon evaluating the primers in clinical CSF samples, 100% detection of the BME causing microorganisms was observed. The specificity was 100%. The PCR products were analyzed by Hae III digestion, and correspondence found for the restriction patterns of the clinical samples and the reference strains.

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