Feasibility of a quantitative polymerase chain reaction assay for diagnosing pneumococcal pneumonia using oropharyngeal swabs.

Autor: van Schaik ML; Department Pulmonology, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands.; Department Molecular Biology, Regional Laboratory for Medical Microbiology and Public Health, Haarlem, The Netherlands.; Department of Pulmonology, Isala clinics Zwolle, Dr. van Heesweg 2, 8025AB, Zwolle, The Netherlands., Duijkers R; Department Pulmonology, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands., Paternotte N; Department Pulmonology, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands., Jansen R; Department Molecular Biology, Regional Laboratory for Medical Microbiology and Public Health, Haarlem, The Netherlands., Rozemeijer W; Department Medical Microbiology, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands., van der Reijden WA; Department Molecular Biology, Regional Laboratory for Medical Microbiology and Public Health, Haarlem, The Netherlands., Boersma WG; Department Pulmonology, Noordwest Ziekenhuisgroep, Alkmaar, The Netherlands. w.boersma@NWZ.nl.; Department of Pulmonary Diseases, Northwest Hospital Group, P.O. Box 501, 1800AM, Alkmaar, The Netherlands. w.boersma@NWZ.nl.
Jazyk: angličtina
Zdroj: Molecular biology reports [Mol Biol Rep] 2019 Feb; Vol. 46 (1), pp. 1013-1021. Date of Electronic Publication: 2018 Dec 19.
DOI: 10.1007/s11033-018-4558-0
Abstrakt: Streptococcus pneumoniae is the most important pathogen causing community-acquired pneumonia (CAP). The current diagnostic microbial standard detects S. pneumoniae in less than 30% of CAP cases. A quantitative polymerase chain reaction (PCR) targeting autolysin (lytA) is able to increase the rate of detection. The aim of this study is validation of this quantitative PCR in vitro using different available strains and in vivo using clinical samples (oropharyngeal swabs). The PCR autolysin (lytA) was validated by testing the intra- and inter-run variability. Also, the in vitro specificity and sensitivity, including the lower limit of detection was determined. In addition, a pilot-study was performed using samples from patients (n = 28) with pneumococcal pneumonia and patients (n = 28) with a pneumonia without detection of S. pneumoniae with the current diagnostic microbial standard, but with detection of either a viral and or another bacterial pathogen to validate this test further. The intra- and inter-run variability were relatively low (SD's ranging from 0.08 to 0.96 cycle thresholds). The lower limit of detection turned out to be 1-10 DNA copies/reaction. In-vitro sensitivity and specificity of the tested specimens (8 strains carrying lytA and 6 strains negative for lytA) were both 100%. In patients with pneumococcal and non-pneumococcal pneumonia a cut-off value of 6.000 copies/mL would lead to a sensitivity of 57.1% and a specificity of 85.7%. We were able to develop a quantitative PCR targeting lytA with good in-vitro test characteristics.
Databáze: MEDLINE
Externí odkaz: