Optimization of an Adsorption-Elution Method with a Negatively Charged Membrane to Recover Norovirus from Lettuce.

Abstract

Viral pathogens, such as norovirus (NoV), are frequently associated with foodborne gastroenteritis worldwide, and the detection of NoV in food requires appropriate methods and the use of process controls. In this study, an adsorption-elution concentration method using negatively charged membranes was optimized to recover NoV from lettuce, using murine norovirus 1 (MNV-1) as a human NoV (HuNoV) surrogate. Initially, three elution buffers were evaluated by direct elution using a Stomacher® apparatus with a filter bag and different concentrations of MNV-1 genomic copies. The eluates were filtered in a Stericup® and concentrated by a Centriprep Concentrator®, and the viral RNA was quantified by real-time PCR that was preceded by reverse transcription. The MNV-1 recovery efficiency varied based on the buffers used, ranging from 5.2 to 9.8% for PBS pH 7.2, 0.2-18% for glycine NaCl pH 9.5 and 10.8-33.3% for glycine Tris-HCl pH 9.5. Further analysis of the glycine Tris-HCl pH 9.5 buffer revealed that gentle-shaking, direct elution could replace the use of a Stomacher®, with recovery rates reaching 66 and 32% for MNV-1 and HuNoV, respectively, all of which suggested that this procedure is a quick and efficient method for recovering NoV from lettuce.