Detección de Pneumocystis jirovecii mediante PCR anidada en pacientes VIH negativos con enfermedad pulmonar

Nested PCR can be used to determine the status of Pneumocystis jirovecii infection in other lung diseases. This study sought to detect a target DNA fragment (mitochondrial large subunit rRNA or mtL SUrRNA) of P. jirovecii in patients with lung disease who underwent bronchoscopy with collection of bronchoalveolar lavage (BAL). The results from toluidine blue staining were compared with those obtained using molecular methods that included an “in house” DNA extraction procedure, PCR and nested PCR. Fifty-five BAL samples from patients with atypical chest X-rays were screened for P. jirovecii. None of the samples was positive for P. jirovecii using toluidine blue staining. In contrast, P. jirovecii DNA was detected by nested PCR in BAL samples from 36 of 55 patients (65.5%). The lung diseases in the patients included cancer, pneumonia, tuberculosis, and chronic obstructive pulmonary disease (COPD). Other chronic problems in the patients included hypertension, diabetes, smoking, and alcoholism. Nested PCR showed high sensitivity for detecting P. jirovecii, especially when compared with toluidine blue staining. Using this method, P. jirovecii infection was detected in HIV-negative patients with lung disease.3428388El diagnóstico de laboratorio mediante la técnica de PCR anidada permite determinar estados de infección por Pneumocystis jirovecii en otras enfermedades pulmonares. El objetivo de este estudio fue detectar fragmentos de ADN mitocondrial (mtLSU rRNA) de P. jirovecii en muestras de lavado broncoalveolar (LBA) de pacientes con enfermedades pulmonares, sometidos a broncoscopia. Se compara la técnica de coloración con azul de toluidina para la microscopia, con los métodos moleculares PCR y PCR anidada; se realizó una extracción in house de ADN para las reacciones moleculares. La presencia de P. jirovecii fue estudiada en 55 muestras de LBA de pacientes que presentaron patrones radiográficos de tórax atípicos. Ninguna de las muestras fue positiva para P. jirovecii con la técnica de coloración con azul de toluidina. Por la técnica de PCR anidada se detectó el ADN de P. jirovecii en 36 de los 55 pacientes (65,5%). Las enfermedades pulmonares de los pacientes fueron cáncer, neumonía, tuberculosis y enfermedad pulmonar obstructiva crónica (EPOC). Las otras enfermedades crónicas presentadas por los pacientes fueron hipertensión, diabetes, alcoholismo y tabaquismo. La PCR anidada mostró ser altamente sensible en la detección de P. jirovecii en comparación con la coloración por azul de toluidina. Este método permite detectar infecciones por P. jirovecii en pacientes VIH negativos con enfermedades pulmonares

Detección De Pneumocystis Jirovecii Mediante Pcr Anidada En Pacientes Vih Negativos Con Enfermedad Pulmonar

Nested PCR can be used to determine the status of Pneumocystis jirovecii infection in other lung diseases. Aims This study sought to detect a target DNA fragment (mitochondrial large subunit rRNA or mtL SUrRNA) of P. jirovecii in patients with lung disease who underwent bronchoscopy with collection of bronchoalveolar lavage (BAL). Methods The results from toluidine blue staining were compared with those obtained using molecular methods that included an “in house” DNA extraction procedure, PCR and nested PCR. Results Fifty-five BAL samples from patients with atypical chest X-rays were screened for P. jirovecii. None of the samples was positive for P. jirovecii using toluidine blue staining. In contrast, P. jirovecii DNA was detected by nested PCR in BAL samples from 36 of 55 patients (65.5%). The lung diseases in the patients included cancer, pneumonia, tuberculosis, and chronic obstructive pulmonary disease (COPD). Other chronic problems in the patients included hypertension, diabetes, smoking, and alcoholism. Conclusions Nested PCR showed high sensitivity for detecting P. jirovecii, especially when compared with toluidine blue staining. Using this method, P. jirovecii infection was detected in HIV-negative patients with lung disease. © 2016 Asociación Española de Micología342838