Purpose: To assess viability of vegetable wash and brackish tidal surface waters in survival and detection of enteric viruses. Here, methods for the evaluation of murine norovirus (MNV) persistence were assessed for future evaluation of water samples.
Methods: Samples (200-L) of vegetable wash and brackish tidal surface waters were collected from two sites in the Mid-Atlantic and subsamples, in duplicates, were inoculated with MNV (5.0-log PFU/ml) as a surrogate for human norovirus, and stored at 12°C. RNA was extracted from each sample on the day of inoculation and days 1, 3, 6, and 9 and stored at -20°C. RNA was evaluated by qPCR and compared to MNV standard curves. Aerobic plate counts were evaluated on uninoculated wash and surface waters using TSA incubated at 37°C overnight.
Results: Over nine days, MNV persisted in each sample, each day, at concentrations consistent with the original inoculum concentration (4.0 log PFU/ml) as compared to standard curves created with each qPCR run. Ct values were as follows: 25.22 for days 0 and 1, 21.72 for days 3 and 6, and 23.25 for day 9. Ct values for all samples were within acceptable range of their corresponding standard curve, where R2 values were 0.99. Aerobic plate counts revealed bacterial concentrations ranging from 5.96×102 to 5.63×102CFU/ml for the two vegetable wash water samples and 18.3 to 41.7 CFU/ml for the two brackish tidal surface water samples.
Significance: Neither bacterial populations nor quality/chemical characteristics interfered with qPCR. MNV persisted in surface and vegetable wash waters collected in Fall, 2016, allowed survival of MNV, warranting further studies on human enteric virus detection and survival.