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

Autor: Wim Boersma, Ruud Jansen, Ruud Duijkers, M. L. van Schaik, W. A. van der Reijden, W. Rozemeijer, Nienke Paternotte
Jazyk: angličtina
Rok vydání: 2019
Předmět:
0301 basic medicine
Male
Pilot Projects
medicine.disease_cause
Quantitative PCR
0302 clinical medicine
Community-acquired pneumonia
80 and over
Pathogen
Pneumococcal/diagnosis
Aged
80 and over
General Medicine
Middle Aged
Streptococcus pneumoniae
Real-time polymerase chain reaction
LytA
030220 oncology & carcinogenesis
Pharynx/microbiology
Pneumococcal pneumonia
Original Article
Female
Mouth/microbiology
Pneumonia
Pneumococcal/diagnosis
Adult
Real-Time Polymerase Chain Reaction/methods
Real-Time Polymerase Chain Reaction
Microbiology
03 medical and health sciences
Young Adult
In vivo
Genetics
medicine
Journal Article
Humans
Molecular Biology
Aged
Mouth
business.industry
Autolysin
Reproducibility of Results
Pneumonia
Pneumonia
Pneumococcal
medicine.disease
030104 developmental biology
ROC Curve
Pharynx
Feasibility Studies
business
Zdroj: Molecular Biology Reports, 46(1), 1013. Springer Netherlands
Molecular Biology Reports
ISSN: 0301-4851
Popis: 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. Electronic supplementary material The online version of this article (10.1007/s11033-018-4558-0) contains supplementary material, which is available to authorized users.
Databáze: OpenAIRE
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