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Master's Dissertation
DOI
10.11606/D.9.1998.tde-11082006-115017
Document
Author
Full name
Jane Harumi Atobe
E-mail
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 1998
Supervisor
Committee
Mamizuka, Elsa Masae (President)
Schmal, Manoel Reinaldo
Takei, Kioko
Title in Portuguese
Amplificação de DNA de Neisseria meningitidis em amostras de líquido cefalorraquidiano pela reação em cadeia da polimerase-multiplex
Keywords in Portuguese
Diagnóstico molecular
Meningite bacteriana
Neisseria meningitidis
Abstract in Portuguese
Padronizou-se a PCR-Multiplex para a detecção do DNA de Neisseria meningitidis. Para tanto os primers escolhidos foram: RW01, DG74 e COR28 baseados na subunidade menor do ribossomo (16S rRNA) que apresenta regiões de seqüências conservadas encontradas em todas as bactérias conhecidas. Os primers RW01 e DG74 amplificaram o fragmento universal bacteriano de 370 bp e, os primers RW01 e COR28, o fragmento específico de N. meningitidis de 279 bp em uma única etapa. Os resultados obtidos nas amostras de LCR de 168 pacientes pelos métodos de cultura e PCR-Multiplex quando comparados à bacterioscopia mostraram que tal técnica apresentou alta sensibilidade (91,3%) no estudo de amostras de LCR de bacterioscopia positiva, enquanto que a cultura apresentou resultados menores (19,7%). Nas amostras de LCR com bacterioscopia negativa a sensibilidade da PCR-Multiplex (57,8%) também foi mais elevada do que da cultura (10%). Estes dados sugerem que a técnica aqui padronizada é altamente promissora para ser utilizada como método diagnóstico da meningite meningocócica, especialmente nos casos de pacientes submetidos à terapia antibiótica prévia.
Title in English
Multiplex-PCR for the diagnosis of meningococcal meningitis in cerebral spinal fluid
Keywords in English
Bacterial meningitis
Molecular diagnosis
Neisseria meningitidis
Abstract in English
The PCR-multiplex technique was standardized to detect N.meningitidis DNA. It was used universal primer for all bacteria showing sequence of minor subunit of 16S ribossome regions (RW01, DG74) by amplification of 370bp fragment and another (COR28) for specific sequence of N. meningitidis, amplifying 279bp fragment in one step. The results obtained in CSF samples of 168 patients by culture and PCR-Multiplex technique when compared with microscopy showed high sensitivity (91,3%) in samples with positive microscopy (81) to Gram negative diplococcus, however the culture presented only 19, 7% of positivity in the same samples. In other hand the CSF samples with negative bacterioscopy (67) the PCR-Multiplex sensitivity (57,8%) was higher to culture (10,0%) too. These data indicate a high sensitivity and specificity of PCR as a tool for a rapid diagnosis of meningococcal meningitis, mainly in that patient submitted to previous antibiotic therapy as in case of this work (90% of patients) besides the possibility of a rational practice of specific treatment.
 
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DISSERTACAO.pdf (1.01 Mbytes)
Publishing Date
2007-01-26
 
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