Purpose: Pilot test an optimized detection method to identify both matrix inhibition and norovirus contamination on produce, hand-rinse, and agricultural water samples collected from northern Mexico.
Methods: From farms and packing facilities, composite rinses of equivalent surface area from cantaloupes (n=6 samples; 6 units/rinse), jalapeños (n=6 samples; 42 units/rinse), tomatoes (n=6 samples; 54 units/rinse), and hands (n=18 samples; 3 workers’ hands/rinse) were collected in 0.1% peptone solution. Source and irrigation water samples (n=14 samples; triplicate 1.5L) were collected. A two-step virus concentration/elution procedure (1% BSA elution and 12% PEG precipitation) was followed by RNA extraction and RT-qPCRs (GI and GII) with an internal amplification control (IAC) to detect inhibition.
Results: An algorithm based on target and IAC cycle threshold (Ct) values was used to classify samples as presumptively positive, negative, undetermined, or inhibited. Thirty-nine of 50 samples had matrix-associated inhibition; repeating the RT-qPCR diluting template 1:4 removed inhibition in 69% of these. Using a Ct value cutoff <40, 12% (6/50) of the samples were presumptively positive, six undetermined, and 12 inhibited even after dilution. While at least one sample of each type (hand-rinse, water, produce) appeared positive, water had the highest prevalence (50%; 3/6). One cantaloupe rinsate was presumptively positive. Presumptively positive samples were mostly norovirus GII with varying Ct values (32.1 - 38.6). Sequence confirmation is pending.
Significance: This study provides evidence of norovirus RNA in the agricultural environment demonstrating hands and water as potential vehicles of contamination. The amount of RNA due to infectious virus remains unclear. This approach is useful for estimating norovirus RNA presence on various agricultural matrices.