Max cdiff

From June 2018 to February 2020 a total of 2545 fecal samples from outpatients with acute gastroenteritis, both children and adults, were collected for bacterial and viral enteropathogens prospective detection by real-time multiplex PCR. The BD Max enteric viral panel combined with the enteric bacterial panel, and the C. difficile toxin B panel (BD Max Cdiff) (Becton Dickinson, Sparks, MD, USA) when requested, were used at the Clinical Microbiology Laboratory of the Hospital Clínico Universitario of Valencia to analyze the stools. Positive samples for enteropathogenic bacteria were further cultivated in the appropriate bacteriological media, and those positive for enteric viruses were kept at −20 °C for viral strain characterization. The BD Max enteric viral panel detects norovirus genogroup I (GI) and GII, rotavirus type A, adenovirus type F 40/41, human astrovirus and sapovirus (genogroups I, II, IV, and V). The testing procedure was performed according to the manufacturer’s instructions and all samples were anonymized once the diagnosis was done. The present study was carried out in accordance with the Declaration of Helsinki and was approved by the Ethics Committee of the Hospital Clínico Universitario of Valencia (Approval No. F-CE-GEva-15, 26 May 2015).

For determination of the limit of detection, 400 μl of a homogenous stool solution, tested negative for the respective pathogens, was spiked with 2 × 50μl of a 0.5 McF suspension of two pathogens each: C. jejuni (DSM4688), S. cholerasuis (ATCC554), Y. enterocolitica (ATCC9610), S. flexneri (ATCC29903), C. difficile (DSM27544), Aeromonas hydrophila (DSM30187), Vibrio cholerae (DSM100200). Thereafter, a 1:10 dilution series was produced (1 E7/ml to 1 E0/ml) and tested by the multiplex PCR as well as colony counting (for exact CFU determination) on the respective agar media. The test was repeated once with a 1:10 dilution series and then five times with a 1:3 dilution around the limit of detection. Moreover, 159 clinical stool samples (samples from the routine diagnostics in Heidelberg and samples from Lucerne) that had been analyzed before and that had been stored at − 80 °C for up 2 years were tested. For these samples, a culture result and/or a molecular result (BD MAX Enteric Panel, Biofire Filmarray, BD MAX C.diff) were available. In case of discrepant results, the samples were reanalyzed once by the same method. A composite of culture and/or molecular result was used as reference for the evaluation of the Seegene multiplex assay.