Czech J. Food Sci., X:X | DOI: 10.17221/101/2025-CJFS

Occurrence of Norovirus genogroup II in leafy greens in the region of Marrakech using a molecular methodOriginal Paper

Salma Berrouch ORCID...1,2, Nouhaila El Fellaki1, Abdelkader Biary1, Hibatallah Lachkar1, Houda Rafi1, Halima Rouane1, Simeon Goïta1, Jamal Eddine Hafid ORCID...1
1 Laboratory of Bioresources and Food Safety, Faculty of Sciences and Technology, Cadi Ayyad University, Marrakech, Morocco
2 Higher School of Technology of El Kelaa des Sraghna, Cadi Ayyad University, El Kelaa des Sraghna, Morocco

Leafy greens are important vectors for enteric viruses, including human noroviruses (HuNoV), which are a leading cause of foodborne disease. These viruses can contaminate the agricultural environment through untreated wastewater or direct contamination. While studies on HuNoV in vegetables have been conducted, few have focused on Morocco. This study aimed to detect HuNoV in leafy greens collected in the region of Marrakech region over a fifteen-week period (March–June 2023). For this purpose, 112 samples (coriander, lettuce and parsley) were collected and analysed using the ISO 15216-2:2019 method with minor modifications, after validation. The method involved eluting viruses using an alkaline buffer, concentrating them through polyethylene glycol precipitation, and detecting viral RNA via real-time RT-PCR. The applied method yielded variable recovery rates among the tested matrices, with coriander showing the highest recovery (1.5%), followed by lettuce (1.2%) and parsley (0.6%), confirming a significant matrix-dependent variation in HuNoV recovery (ANOVA, P < 0.001). This method enabled the assessment of leafy green contamination, which was found to be 0.89% (1/112). This study underscores the need to enhance detection methods to better assess the risks associated to noroviruses in leafy greens, with implications for human health.

Keywords: nteric viruses, fresh vegetables; real-time RT-PCR detection; survey; market

Received: June 28, 2025; Revised: January 27, 2026; Accepted: January 29, 2026; Prepublished online: April 13, 2026 

Share...
Download citation
  • BibTeX (.bib)
  • Bookends (.ris)
  • EasyBib (.ris)
  • EndNote (.enw)
  • EndNote 8 (.xml)
  • ISI WoS (.isi)
  • Medlars (.medlars)
  • Mendeley (.ris)
  • MODS (.xml)
  • Papers (.ris)
  • RefWorks (.txt)
  • RefManager (.ris)
  • RIS (.ris)
  • MS Word (.xml)
  • Zotero (.ris)
    • Open full article

    References

    1. Aiyedun S.O., Onarinde B.A., Swainson M., Dixon R.A. (2021): Foodborne outbreaks of microbial infection from fresh produce in Europe and North America: A systematic review of data from this millennium. International Journal of Food Science and Technology, 56: 2215-2223. Go to original source...
    2. Barril P.A., Oteiza J.M., Pardo J., Leotta G.A., Signorini M.L. (2022): Meta-analysis of the prevalence of the main human pathogens in vegetables, with emphasis on lettuce. Food Research International, 160: 111727. Go to original source... Go to PubMed...
    3. Bounagua R., Rhaffouli H.E., Bouderra A., Boukbir L., Laaouissi M., Yassin Z., El Kasraoui H., Sekhsoukh Y., Fassi-Fihri O. (2021): Molecular evaluation of enteric viruses' contamination of raw vegetables grown in the Errachidia Region. Saudi Journal of Pathology and Microbiology, 6: 323-328. Go to original source...
    4. Chatziprodromidou I.P., Bellou M., Vantarakis G., Vantarakis A. (2018): Viral outbreaks linked to fresh produce consumption: A systematic review. Journal of Applied Microbiology, 124: 932-942. Go to original source... Go to PubMed...
    5. Cook N., Williams L., D'Agostino M. (2019): Prevalence of Norovirus in produce sold at retail in the United Kingdom. Food Microbiology, 79: 85-89. Go to original source... Go to PubMed...
    6. Da Silva A.K., Le Saux J.C., Parnaudeau S., Pommepuy M., Elimelech M., Le Guyader F.S. (2007): Evaluation of removal of noroviruses during wastewater treatment, using real-time reverse transcription-PCR: Different behaviors of genogroups I and II. Applied and Environmental Microbiology, 73: 7891-7897. Go to original source... Go to PubMed...
    7. Dubois E., Agier C., Traoré O., Hennechart C., Merle G., Crucière C., Laveran H. (2002): Modified concentration method for the detection of enteric viruses on fruits and vegetables by reverse transcriptase-polymerase chain reaction or cell culture. Journal of Food Protection, 65: 1962-1969. Go to original source... Go to PubMed...
    8. Ekundayo T.C., Ijabadeniyi O.A. (2023): Human norovirus contamination challenge in fresh produce: A global prevalence and meta-analytic assessment. Journal of Applied Microbiology, 134: lxac009. Go to original source... Go to PubMed...
    9. Elfellaki N., Berrouch S., Biary A., Goïta S., Rafi H., Lachkar H., Dehhani O., de Rougemont A., Bourlet T., Hafid J.E. (2024): Comparison of four concentration methods of adenovirus, norovirus and rotavirus in tap water. Journal of Virological Methods, 330: 115013. Go to original source... Go to PubMed...
    10. Gaskin J.W., Risse L.M., Segars W.I., Harris G.H. (2009): Beneficial Reuse of Municipal Biosolids in Agriculture. Agriculture Pollution Prevention Factsheet Series, University of Georgia.
    11. Kageyama T., Kojima S., Shinohara M., Uchida K., Fukushi S., Hoshino F.B., Takeda N., Katayama K. (2003): Broadly reactive and highly sensitive assay for Norwalk-like viruses based on real-time quantitative reverse transcription-PCR. Journal of Clinical Microbiology, 41: 1548-1557. Go to original source... Go to PubMed...
    12. Keraita B., Jimenez B., Drechsel P. (2008): Extent and implications of agricultural reuse of untreated, partly treated and diluted wastewater in developing countries. CABI Reviews, 2008. Go to original source...
    13. Kokkinos P., Kozyra I., Lazic S., Bouwknegt M., Rutjes S., Willems K., Moloney R., de Roda Husman A.M., Kaupke A., Legaki E., D'Agostino M., Cook N., Rzezutka A., Petrovic T., Vantarakis A. (2012): Harmonised investigation of the occurrence of human enteric viruses in the leafy green vegetable supply chain in three European countries. Food and Environmental Virology, 4: 179-191. Go to original source... Go to PubMed...
    14. Loisy F., Atmar R.L., Guillon P., Le Cann P., Pommepuy M., Le Guyader F.S. (2005): Real-time RT-PCR for norovirus screening in shellfish. Journal of Virological Methods, 123: 1-7. Go to original source... Go to PubMed...
    15. Prez V. E., Martínez L.C., Victoria M., Giordano M.O., Masachessi G., Ré V.E., Pavan J.V., Colina R., Barril P.A., Nates S.V. (2018): Tracking enteric viruses in green vegetables from central Argentina: potential association with viral contamination of irrigation waters. Science of the Total Environment, 637: 665-671. Go to original source... Go to PubMed...
    16. Randazzo W., Vásquez-García A., Bracho M.A., Alcaraz M.J., Aznar R., Sánchez G. (2018): Hepatitis E virus in lettuce and water samples: A method-comparison study. International Journal of Food Microbiology, 277: 34-40. Go to original source... Go to PubMed...
    17. Raymond P., Paul S., Perron A., Deschênes L., Hara K. (2021): Extraction of human noroviruses from leafy greens and fresh herbs using magnetic silica beads. Food Microbiology, 99: 103827. Go to original source... Go to PubMed...
    18. Shaheen M.N.F., Elmahdy E.M., Chawla-Sarkar M. (2019): Quantitative PCR-based identification of enteric viruses contaminating fresh produce and surface water used for irrigation in Egypt. Environmental Science and Pollution Research, 26: 21619-21628. Go to original source... Go to PubMed...
    19. Sorensen J.P.R., Aldous P., Bunting S.Y., McNally S., Townsend B.R., Barnett M.J., Harding T., La Ragione R.M., Stuart M.E., Tipper H.J., Pedley S. (2021): Seasonality of enteric viruses in groundwater-derived public water sources. Water Research, 207: 117813. Go to original source... Go to PubMed...
    20. Stoufer S., Soorneedi A.R., Kim M., Moore M.D. (2024): Sample processing and concentration methods for viruses from foods and the environment prior to detection. Annual Review of Food Science and Technology 15: 455- 472. Go to original source... Go to PubMed...
    21. Suther C., Moore M.D. (2019): Quantification and discovery of PCR inhibitors found in food matrices commonly associated with foodborne viruses. Food Science and Human Wellness, 8: 351-355. Go to original source...
    22. Tang M., Liao N., Tian P., Shen K., Liu C., Ruan L., Wu G. (2023): Use of bentonite-coated activated carbon for improving the sensitivity of RT-qPCR detection of norovirus from vegetables and fruits: The ISO 15216-1:2017 standard method extension. Food Microbiology, 110: 104165. Go to original source... Go to PubMed...
    23. Teunis P.F.M., Moe C.L., Liu P., Miller S.E., Lindesmith L., Baric R.S., Le Pendu J., Calderon R.L. (2008): Norwalk virus: How infectious is it? Journal of Medical Virology, 80: 1468-1476. Go to original source... Go to PubMed...
    24. Teunis P.F.M., Xu M., Fleming K.K., Yang J., Moe C.L., LeChevallier M.W. (2010): Enteric Virus infection risk from intrusion of sewage into a drinking water distribution network. Environmental Science & Technology, 44: 8561-8566. Go to original source... Go to PubMed...
    25. Torok V.A., Hodgson K.R., Jolley J., Turnbull A., McLeod C. (2019): Estimating risk associated with human norovirus and hepatitis A virus in fresh Australian leafy greens and berries at retail. International Journal of Food Microbiology, 309: 108327. Go to original source... Go to PubMed...
    26. WHO & FAO (2023): FAO/WHO Expert Committee Conducts Review of Foodborne Viruses to Support Codex Guidance Updates. Brimingham, Food Safety Magazine. Available at https://www.food-safety.com/articles/8974-fao-who-expert-committee-conducts-review-of-foodborne-viruses-to-support-codex-guidance-updates.
    27. Wilson I.G. (1997): Minireview: Inhibition and facilitation of nucleic acid amplification. Applied and Environmental Microbiology, 63: 3741-3751. Go to original source... Go to PubMed...
    28. Xie Y., Du X., Li D., Wang X., Xu C., Zhang C., Sun A., Schmidt S., Liu X. (2021): Seasonal occurrence and abundance of norovirus in pre- and postharvest lettuce samples in Nanjing, China. LWT - Food Science and Technology, 152: 112226. Go to original source...

    This is an open access article distributed under the terms of the Attribution-NonCommercial 4.0 International (CC BY-NC 4.0.), which permits non-comercial use, distribution, and reproduction in any medium, provided the original publication is properly cited. No use, distribution or reproduction is permitted which does not comply with these terms.


    Return