Etiologic Diagnosis of Lower Respiratory Tract Bacterial Infections Using Sputum Samples and Quantitative Loop-Mediated Isothermal Amplification

Abstract

Etiologic diagnoses of lower respiratory tract infections (LRTI) have been relying primarily on bacterial cultures that often fail to return useful results in time. Although DNA-based assays are more sensitive than bacterial cultures in detecting pathogens, the molecular results are often inconsistent and challenged by doubts on false positives, such as those due to system- and environment-derived contaminations. Here we report a nationwide cohort study on 2986 suspected LRTI patients across P. R. China. We compared the performance of a DNA-based assay qLAMP (quantitative Loop-mediated isothermal AMPlification) with that of standard bacterial cultures in detecting a panel of eight common respiratory bacterial pathogens from sputum samples. Our qLAMP assay detects the panel of pathogens in 1047(69.28%) patients from 1533 qualified patients at the end. We found that the bacterial titer quantified based on qLAMP is a predictor of probability that the bacterium in the sample can be detected in culture assay. The relatedness of the two assays fits a logistic regression curve. We used a piecewise linear function to define breakpoints where latent pathogen abruptly change its competitive relationship with others in the panel. These breakpoints, where pathogens start to propagate abnormally, are used as cutoffs to eliminate the influence of contaminations from normal flora. With help of the cutoffs derived from statistical analysis, we are able to identify causative pathogens in 750 (48.92%) patients from qualified patients. In conclusion, qLAMP is a reliable method in quantifying bacterial titer. Despite the fact that there are always latent bacteria contaminated in sputum samples, we can identify causative pathogens based on cutoffs derived from statistical analysis of competitive relationship.Trial RegistrationClinicalTrials.gov NCT00567827

Bacterial pathogens were detected from human exhaled breath using a novel protocol

“…Here, we have found that H. influenza was the leading bacteria responsible for the respiratory infections of the patients we have recruited in this study. Similar to our work, Kang et al (2012) reported that the most common respiratory bacterial infections are caused by H. influenzae, S. pneumoniae, respiratory syncytial virus and so on. Similar studies also demonstrated that the S. pneumoniae, Fig.…”

A preliminary study on the potential of Mycoplasma pneumoniae to induce dyskaryotic change in respiratory epithelium in adult community-acquired pneumonia

“…China) as described elsewhere. [56] A presumptive diagnosis of acute Mpinfection was made by a positive IgM result. Clinical characteristics of patients were also recorded.…”

“…[6] Nine respiratory viruses in BALF were detected using FTD respiratory pathogens 21 plus multiplex real-time polymerase chain reaction kit (Fast-track Diagnostics, Junglinster, Luxembourg). Meanwhile, the assay using conventional sputum culture for each patient was carried out.…”

Development of a bead-based suspension array for the detection of pathogens in acute respiratory tract infections

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