Attachment, Internalization and Dissemination of Human Norovirus Surrogates in Romaine Lettuce

Introduction: Fresh produce is a major vehicle for transmission of human norovirus because they are easily contaminated both pre- and post-harvest. In fact, norovirus accounts for more than 40% of produce-associated foodborne illness. However, the attachment, internalization and dissemination of human norovirus in fresh produce are poorly understood. Internalized viruses would pose a significant risk to consumers because the viruses would be protected from all sanitation measures. 

Purpose: The purpose of this research is to determine whether human norovirus surrogates can be internalized and disseminated in growing plant tissues. 

Methods: Romaine lettuce was grown hydroponically in feed water containing two human norovirus surrogates (murine norovirus, MNV-1; and Tulane virus, TV).  The hydroponic feed water was inoculated with either MNV-1 or TV to have a starting titer of 1×10⁶ pfu/ml and plants were allowed to grow for 2 weeks. Leaves, shoots, and roots were homogenized and viral titers were determined by plaque assays at multiple time points throughout the study.

Results:  We demonstrated that both MNV-1 and TV can be efficiently internalized and disseminated in Romaine lettuce grown hydroponically. Both viruses can attach tightly to roots and became internalized and disseminated to shoots and leaves at day 1 following virus inoculation. At day 3 post inoculation, the titer of MNV-1 in the shoots and leaves reached 3.3×10⁴ pfu/g and 8.9×10³ pfu/g, respectively. Similarly, the titer of TV in the shoots and leaves reached 2.8x10⁵ pfu/g and 9.1×10⁴ pfu/g, respectively, at day 3 post inoculation.  The MNV-1 and TV viral titers detectable in the roots, shoots, and leaves increased gradually over 14 days to a final titer of 10⁶ pfu/g.

Significance:  These results indicate that human norovirus surrogates can be internalized in the roots and disseminated to shoot and leaf portions of the plant. A better understanding of the virus-fresh produce interaction would lead to the development of novel interventions to minimize virus contamination in fresh produce.