Screening of Fruit Products for Norovirus and the Difficulty of Interpreting Positive PCR Results

Abstract

Despite recent norovirus (NoV) outbreaks related to consumption of fruit products, little is known regarding the NoV load on these foods. Therefore, 75 fruit products were screened for NoV presence by using an evaluated in-house NoV detection methodology consisting of a NoV extraction method and a reverse transcription quantitative PCR assay. Additionally, the fruit samples were screened for bacterial pathogens and bacterial hygiene indicators. Results of the NoV screening showed that 18 of 75 samples tested positive for GI and/or GII NoV despite a good bacteriological quality. The recovery of murine norovirus 1 virus particles acting as process control was successful in 31 of 75 samples with a mean recovery efficiency of 11.32% ± 6.08%. The level of detected NoV genomic copies ranged between 2.5 and 5.0 log per 10 g. NoV GI and/or GII were found in 4 of 10, 7 of 30, 6 of 20, and 1 of 15 of the tested raspberries, cherry tomatoes, strawberries, and fruit salad samples, respectively. However, confirmation of the positive quantitative PCR results by sequencing genotyping regions in the NoV genome was not possible. Due to the nature of the method used (reverse transcription quantitative PCR) for detection of genomic material, no differentiation was possible between infectious and noninfectious viral particles. No NoV outbreaks related to the tested fruit product types were reported during the screening period, which hampers a conclusion as to whether these unexpected high numbers of NoV-positive results should be perceived as a public health threat. These results, however, may indicate a prior NoV contamination of the tested food samples throughout the fresh produce chain.

Persistence of human norovirus GII.4 and GI.4, murine norovirus, and human adenovirus on soft berries as compared with PBS at commonly applied storage conditions

“…The attribution of hNoV illness to food is, however, challenging, due to e.g. difficulties in detecting hNoV (Le Guyader et al, 2004;Rutjes et al, 2006;Stals et al, 2011), the absence of a systematic surveillance for foodborne viral disease (Newell et al, 2010), and the fact that secondary person to person transmission may mask foodborne transmission of hNoV (Carter, 2005;Verhoef et al, 2010). As a consequence, the contribution of food matrices in causing foodborne hNoV outbreaks is not clear and the significance of foodborne hNoV disease to public health may be estimated incorrectly.…”

Scientific Opinion on the risk posed by pathogens in food of non-animal origin. Part 2 (Salmonellaand Norovirus in tomatoes)

“…The analyses used methods similar to the standardised methods described in ISO/TS 15216-1 and ISO/TS 15216-2, in general or specific aspects. Stals et al (2011a) detected Norovirus in 7 out of 30 samples of cherry tomatoes whereas in two samples of cherry tomatoes both Norovirus genogroups I and genogroup II signals were present. It should be noted that usually one and maximally two out of four replicate RT-qPCRs per sample gave a positive signal in samples where NoV genomic presence was detected, which can be explained by the fact that most detected NoV signals were close to the presumed detection limit of the NoV RT-qPCR methodology (also noticeable from associated Ct values of the positive samples ranging between 37 and 42).…”

Evaluation of the swab sampling method to recover viruses from fomites

“…Its use as ICP for detecting viruses in foods such as shellfish, soft red fruits and ready-to-eat foods has also been described (Kingsley et al, 2007;Stals et al, 2009Stals et al, , 2011aStals et al, , 2011b. However, some studies focusing on PP7 as ICP for food and environmental samples have been described (Rajal et al, 2007;Fumian et al, 2010;Brandão et al, 2014), mainly because PP7 propagation is easier than MNV-1, not requiring the set-up of cell cultures to produce stocks of those viruses (Rajal et al, 2007).…”