Evaluation of yield of currently available diagnostics by sample type to optimize detection of respiratory pathogens in patients with a community-acquired pneumonia

Background: For the detection of respiratory pathogens, the sampling strategy may influence the diagnostic yield. Ideally, samples from the lower respiratory tract are collected, but they are difficult to obtain. Objectives: In this study, we compared the diagnostic yield in sputum and oropharyngeal samples (OPS) for the detection of respiratory pathogens in patients with community-acquired pneumonia (CAP), with the objective to optimize our diagnostic testing algorithm. Methods: Matched sputum samples, OPS, blood cultures, serum, and urine samples were taken from patients (>18 years) with CAP and tested for the presence of possible respiratory pathogens using bacterial cultures, PCR for 17 viruses and five bacteria and urinary antigen testing. Results: When using only conventional methods, that is, blood cultures, sputum culture, urinary antigen tests, a pathogen was detected in 49·6% of patients (n = 57). Adding molecular detection assays increased the yield to 80%. A pathogen was detected in 77 of the 115 patients in OPS or sputum samples by PCR. The sensitivity of the OPS was lower than that of the sputum samples (57% versus 74%). In particular, bacterial pathogens were more often detected in sputum samples. The sensitivity of OPS for the detection of most viruses was higher than in sputum samples (72% versus 66%), except for human rhinovirus and respiratory syncytial virus. Conclusion: Addition of PCR on both OPS and sputum samples significantly increased the diagnostic yield. For molecular detection of bacterial pathogens, a sputum sample is imperative, but for detection of most viral pathogens, an OPS is sufficient

Multi-centre evaluation of real-time multiplex PCR for detection of carbapenemase genes OXA-48, VIM, IMP, NDM and KPC

Background: Resistance to carbapenem antibiotics is emerging worldwide among Enterobacteriaceae. To prevent hospital transmission due to unnoticed carriage of carbapenemase producing micro-organisms in newly admitted patients, or follow-up of patients in an outbreak setting, a molecular screening method was developed for detection of the most prevalent carbapenemase genes; blaOXA-48, blaVIM, blaIMP, blaNDM and blaKPC.Methods: A real-time multiplex PCR assay was evaluated using a collection of 86 Gram negative isolates, including 62 carbapenemase producers. Seven different laboratories carried out this method and used the assay for detection of the carbapenemase genes on a selection of 20 isolates.Results: Both sensitivity and specificity of the multiplex PCR assay was 100%, as established by results on the strain collection and the inter-laboratory comparisons.Conclusions: In this study, we present a multiplex real-time PCR that is a robust, reliable and rapid method for the detection of the most prevalent carbapenemases blaOXA-48, blaVIM, blaIMP, blaNDM and blaKPC, and is suitable for screening of broth cultured rectal swabs and for identification of carbapenemase genes in cultures