Multicenter evaluation of the FilmArray Meningitis/Encephalitis assay in a routine setting.

Introduction. The FilmArray Meningitis/Encephalitis (FA-ME) Panel (Biofire, Salt Lake City, Utah, US) enables fast and automated detection of 14 pathogens in cerebrospinal fluid (CSF).Gap statement. The performance of the FA-ME panel in a real routine setting has not yet been described and could lead to better patient management in cases of good performance.Aim. This multicenter study verified the FA-ME panel analytical performance in a routine hospital setting.Methodology. Between April 2016 and April 2018, 454 CSF samples were analysed with the FA-ME panel and compared with routine diagnostics. In cases of discrepancy or lack of a comparator result, a profound analysis based on patient records and other laboratory results was performed.Results. A first analysis of 65 frozen samples, suspicious for meningitis had a 89 % concordance with routine diagnostics. The limit of detection (LOD) was confirmed for all pathogens except for Streptococcus agalactiae and a strain of Haemophilus influenzae (Escherichia coli K1 and Cryptococcus gattii LOD experiments were not performed). The routine evaluation showed a positive result in 114 (25 %) clinical samples for at least one target. In three samples co-infections were found. After discrepancy analysis, overall sensitivity was 98 % (false negative FA-ME results for one HSV2, two HSV1 and two parechovirus). Four FA-ME results were considered false positive (two HHV6, one VZV and one E. coli K1), resulting in an overall specificity of >99 %. A clinical added value of the assay was seen in the diagnosis of eight cases of bacterial meningitis.Conclusion. Because of its rapidity and ease of use, the FA-ME panel has great potential in the diagnosis of central nervous infections. Implementation can improve clinical management, but costs and analytical limitations need to be addressed to convince clinicians and laboratories of its value.

Follow-up testing of borderline SARS-CoV-2 patients by rRT-PCR allows early diagnosis of COVID-19.

Detection of SARS-CoV-2 RNA in nasopharyngeal samples using the real-time reverse transcription polymerase chain reaction (rRT-PCR) is the gold standard for diagnosing COVID-19. Determination of SARS-CoV-2 RNA by rRT-PCR sometimes results in an inconclusive test result due to a high cycle threshold-value. We retrospectively analyzed 30,851 SARS-CoV-2 rRT-PCR test results. Borderline positivity was considered as the presence of ≤25 viral copies per milliliter, while no amplification was considered as a negative test result. Of all test results, 204 were answered as borderline, of which 107 were accompanied by a follow-up test within 96 hours. Of the 107 follow-up samples, 10 (9.35%) were found positive for SARS-CoV-2. COVID-19 symptoms were not predictive for testing positive in the follow-up test. The positive SARS-CoV-2 samples in the follow-up group represented 0.92% of all positive test results, highlighting the need for retesting and increased hygienic measures for borderline SARS-CoV-2 patients [NCT04636294].

Vertical transmission of SARS-CoV-2 infection and preterm birth.

Viral infections are common complications of pregnancy, with a wide range of obstetric and neonatal sequelae. Currently, there are limited data on whether SARS-CoV-2 is vertically transmitted in pregnant women tested positive for the virus. Here we describe a case of a known SARS-CoV-2-positive woman giving preterm birth to two fetuses with SARS-CoV-2 positive testing in placental tissue and amniotic fluid. The placental histological examinations showed chronic intervillositis and extensive intervillous fibrin depositions with ischemic necrosis of the surrounding villi.

Checklist for optimization and validation of real-time PCR assays.

Real-time polymerase chain reaction (PCR) is a frequently used technique in molecular diagnostics. To date, practical guidelines for the complete process of optimization and validation of commercial and in-house developed molecular diagnostic methods are scare. Therefore, we propose a practical guiding principle for the optimization and validation of real-time PCR assays. Based on literature, existing guidelines, and personal experience, we created a checklist that can be used in different steps of the development and validation process of commercial and in-house developed real-time PCR assays. Furthermore, determination of target values and reproducibility of internal quality controls are included, which allows a statistical follow-up of the performance of the assay. Recently, we used this checklist for the development of various qualitative and quantitative assays for microbiological and hematological applications, for which accreditation according to ISO 15189:2007 was obtained. In our experience, the use of the proposed guidelines leads to a more efficient and standardized optimization and validation. Ultimately, this results in reliable and robust molecular diagnostics. The proposed checklist is independent of environment, equipment, and specific applications and can be used in other laboratories. A worldwide consensus on this kind of checklist should be aimed at.