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- Abdeldaim, Guma, 1969-, et al.
(författare)
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Multiplex quantitative PCR for detection of lower respiratory tract infection and meningitis caused by Streptococcus pneumoniae, Haemophilus influenzae and Neisseria meningitidis
- 2010
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Ingår i: BMC Microbiology. - : Springer Science and Business Media LLC. - 1471-2180. ; 10, s. 310-
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Tidskriftsartikel (refereegranskat)abstract
- Background. Streptococcus pneumoniae and Haemophilus influenzae cause pneumonia and as Neisseria meningitidis they are important agents of meningitis. Although several PCR methods have been described for these bacteria the specificity is an underestimated problem. Here we present a quantitative multiplex real-time PCR (qmPCR) for detection of S. pneumoniae (9802 gene fragment), H. influenzae (omp P6 gene) and N. meningitidis (ctrA gene). The method was evaluated on bronchoalveolar lavage (BAL) samples from 156 adults with lower respiratory tract infection (LRTI) and 31 controls, and on 87 cerebrospinal fluid (CSF) samples from meningitis patients. Results. The analytical sensitivity was not affected by using a combined mixture of reagents and a combined DNA standard (S. pneumoniae/H. influenzae/N. meningitidis) in single tubes. By blood- and BAL-culture and S. pneumoniae urinary antigen test, S. pneumoniae and H. influenzae were aetiological agents in 21 and 31 of the LTRI patients, respectively. These pathogens were identified by qmPCR in 52 and 72 of the cases, respectively, yielding sensitivities and specificities of 95% and 75% for S. pneumoniae, and 90% and 65% for H. influenzae, respectively. When using a cut-off of 105 genome copies/mL for clinical positivity the sensitivities and specificities were 90% and 80% for S. pneumoniae, and 81% and 85% for H. influenzae, respectively. Of 44 culture negative but qmPCR positive for H. influenzae, 41 were confirmed by fucK PCR as H. influenzae. Of the 103 patients who had taken antibiotics prior to sampling, S. pneumoniae and H. influenzae were identified by culture in 6% and 20% of the cases, respectively, and by the qmPCR in 36% and 53% of the cases, respectively. In 87 CSF samples S. pneumoniae and N. meningitidis were identified by culture and/or 16 S rRNA in 14 and 10 samples and by qmPCR in 14 and 10 samples, respectively, giving a sensitivity of 100% and a specificity of 100% for both bacteria. Conclusions. The PCR provides increased sensitivity and the multiplex format facilitates diagnosis of S. pneumoniae, H. influenzae and N. meningitidis and the assay enable detection after antibiotic treatment has been installed. Quantification increases the specificity of the etiology for pneumonia.
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| 2. |
- Strålin, Kristoffer, et al.
(författare)
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Multiplex PCR fir bacteruak etiology using branchoalveolar lavage in adults with lower respiratory tract infection
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Study objective: To study the usefulness of a diagnostic multiplex PCR (mPCR) for Streptococcus pneumoniae, Haemophilus influenzae, Mycoplasma pneumoniae, and Chlamydophila pneumoniae applied to bronchoalveolar lavage (BAL) in lower respiratmy tract infection (LRTI).Design: Prospective diagnostic study.Setting: Silkeborg County Hospital, Silkeborg, Denmark.Patients and participants: Hospitalized adult LRTI patients (n=I56) and adult controls investigated on suspicion of malignancy (n=36).Interventions: After fiberoptic bronchoscopy (FOB) BAL fluid was analysed with bacterial culture and mPCR. S. pneumoniae and H. influenzae etiologies were established by cultures from blood, BAL and sputum, and urinary antigen test for S. pneumoniae. M. pneumoniae etiology was established by singleplex PCR on BAL and throat swab, and C. pneumoniae etiology by singleplex PCR and culture on BAL and throat swab.Measurements and Results: S. pneumoniae, H. influenzae, M. pneumoniae, and C. pneumoniae were etiologies in 14%, 21%, 3.2%, and 0, of the LRTI patients, respectively. These pathogens were identified by BAL mPCR in 28%, 47%, 3.2%, and 0.6%, respectively. The sensitivities of BAL mPCR were 0.86 for S. pneumoniae, 0.88 for H. influenzae, 1.0 for M. pneumoniae, and 0/0 for C. pneumoniae. The specificities were 0.81 for S. pneumoniae, 0.64 for H. influenzae, 1.0 for M. pneumoniae, and 0.99 for C. pneumoniae. In 103 patients with antibiotics taken prior to FOB, BAL culture and BAL mPCR identified S. pneumoniae in 2.9% and 31%, respectively, and H. influenzae in 20% and 50%, respectively. Among the controls, BAL culture and mPCR identified S. pneumoniae in 8.3% and 11%, respectively, and H. influenzae in 11% and 39%, respectively. No M. pneumoniae or C. pneumoniae was identified among the controls.Conclusions: In LRTI patients, BAL mPCR can be useful for identification of S. pneumoniae, M. pneumoniae, and C. pneumoniae. The method appears particularly useful in patients treated with antibiotics.
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