Binding Dynamics of Human Norovirus to Berries and Berry-associated Microflora

Introduction: Fresh berries, including raspberries and strawberries, have been associated with human norovirus (HuNoV) outbreaks.  It is likely that HuNoV bind to berries, but the dynamics of such binding are unknown.  Candidate mechanisms of binding include histo-blood group-like antigens (HBGAs) or unknown ligands present in foods; native berry microflora; biofilms; and/or electrostatic interactions.

Purpose: To elucidate the binding dynamics of HuNoV to berries. 

Methods: Three varietals of strawberries and five of raspberries were collected and concentrates prepared.  Natural berry microflora was isolated, whole colony sequencing performed, and biofilm production of selected strains was quantified using a crystal violet biofilm assay.   Affinity of HuNoV GI.6 and GII.4 virus-like particles (VLPs) binding to berries and biofilm-producing bacteria was determined by ELISA, as was the presence of HBGA-like moieties (A, B, H, Le^a, Le^y). Virus capture efficiency of GI.6 and GII.4 HuNoV to berries and isolates was determined by RT-qPCR. 

Results: A total of 172 (n = 98 from raspberries; n = 74 from strawberries) bacterial isolates were sequenced, with microbial profiles differing by berry type. Biofilm production also varied greatly, but strains of Pantoea, Pseudomonas, Klebsiella, and Enterobacteriaceae were the strongest biofilm producers (A₄₀₅ > 2.2).  ELISA results indicated minimal binding of GI.6 and GII.4 VLPs to berry concentrates (+/- absorbance ratio of 0.2 - 0.7).  Only low levels of Le^a HBGA-like moieties were present in strawberries (P < 0.05), and a minimal level of A, B, H and Le^y moieties were detected in biofilm-producing bacteria isolates. Capture efficiency of GI.6 and GII.4 HuNoV to berries and selected isolates was ~1 log (91.7%) or less.   

Significance: HuNoV bind weakly to raspberries and strawberries, berry microflora, and associated biofilms. Berries and the bacterial isolates do not contain significant levels of HBGA-like moieties. We hypothesize that fecal material may facilitate HuNoV binding to berries and further studies to confirm this are underway.