Abstract
Pneumonia and meningitis continue to present an enormous public health burden and pose a major threat to young children. Among the causative organisms of pneumonia and meningitis, bacteria are the most common causes of serious disease and deaths. It is challenging to accurately and rapidly identify these agents. To solve this problem, we developed and validated a 12-plex PCR coupled with matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) method (bacterial pathogen-mass spectrometry, BP-MS) that can be used to simultaneously screen for 11 key bacterial pathogens related to pneumonia and meningitis. Forty-six nasopharyngeal swabs and 12 isolates were used to determine the specificity of the method. The results showed that, using the BP-MS method, we could accurately identify the expected bacteria without cross-reactivity with other pathogens. For the 11 target bacterial pathogens, the analytical sensitivity of the BP-MS method was as low as 10 copies/reaction. To further evaluate the clinical effectiveness of this method, 204 nasopharyngeal swabs from hospitalized children with suspected pneumonia were tested using this method. In total, 81.9% (167/204) of the samples were positive for at least one of the 11 target pathogens. Among the 167 bacteria-positive samples, the rate of multiple infections was 55.7% (93/167), and the most frequent combination was Streptococcus pneumoniae with Haemophilus influenzae, representing 46.2% (43/93) two-pathogen mixed infections. We used real-time PCR and nested PCR to confirm positive results, with identical results obtained for 81.4% (136/167) of the samples. The BP-MS method is a sensitive and specific molecular detection technique in a multiplex format and with high sample throughput. Therefore, it will be a powerful tool for pathogen screening and antibiotic selection at an early stage of disease.
Highlights
Pneumonia and meningitis continue to be an enormous public health burden
Taking advantage of the system’s multiplex capability and high sample throughput, we developed and validated a bacterial pathogen-MS panel (BP-MS) to simultaneously screen 11 key bacterial pathogens associated with pneumonia and meningitis, including Streptococcus pneumoniae, Haemophilus influenzae, Neisseria meningitidis, Klebsiella pneumoniae, Acinetobacter baumannii, Pseudomonas aeruginosa, Staphylococcus aureus, Moraxella catarrhalis, L. pneumophila, M. pneumoniae, and Bordetella pertussis
We developed a 12-plex method to simultaneously detect 11 key bacterial pathogens associated with pneumonia and meningitis, using HBB as a nucleic acid extraction control
Summary
Pneumonia and meningitis continue to be an enormous public health burden. According to the Global Burden of Disease 2015 study, more than 3.1 million deaths due to pneumonia and meningitis were estimated to have occurred in 2015. The two diseases together accounted for more than 15% of deaths in children under 5 years of age. Viruses, and fungi can all cause pneumonia and meningitis, but bacteria are the leading cause of serious disease and deaths (GBD 2015 Mortality and Causes of Death Collaborators, 2016). The selection of appropriate antibiotics forms the basis of sound clinical management. This complex etiology and overlapping clinical presentations make it challenging to identify the causative organism
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