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A two-step strategy combining assisted benchmark testing (entry controls) and External Quality Assessments (EQAs) with blinded simulated clinical specimens to enhance and maintain the quality of nucleic acid amplification testing was developed. This strategy was successfully applied to 71 diagnostic laboratories in The Netherlands when upscaling the national diagnostic capacity during the SARS-CoV-2 pandemic. The availability of benchmark testing in combination with advice for improvement substantially enhanced the quality of the laboratory testing procedures for SARS-CoV-2 detection. The three subsequent EQA rounds demonstrated high quality testing with regard to specificity (99.6% correctly identified) and sensitivity (93.3% correctly identified). Even with the implementation of novel assays, changing workflows using diverse equipment and a high degree of assay heterogeneity, the overall high quality was maintained using this two-step strategy. We show that in contrast to the limited value of Cq value for absolute proxies of viral load, these Cq values can, in combination with metadata on strategies and techniques, provide valuable information for laboratories to improve their procedures. In conclusion, our two-step strategy (preparation phase followed by a series of EQAs) is a rapid and flexible system capable of scaling, improving, and maintaining high quality diagnostics even in a rapidly evolving (e.g. pandemic) situation.

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.