Enumeration of Escherichia coli and determination of Salmonella spp. and verotoxigenic Escherichia coli in shellfish (Mytilus galloprovincialis and Ruditapes decussatus) harvested in Sardinia, Italy

  • Sonia Lamon Department of Veterinary Medicine, University of Sassari, Italy.
  • Francesca Piras Department of Veterinary Medicine, University of Sassari, Italy.
  • Domenico Meloni | dmeloni@uniss.it Dipartmento di Medicina Veterinaria, Università di Sassari, Italy.
  • Vanessa Agus Department of Veterinary Medicine, University of Sassari, Italy.
  • Gabriella Porcheddu Department of Veterinary Medicine, University of Sassari, Italy.
  • Margherita Pes Department of Veterinary Medicine, University of Sassari, Italy.
  • Maria Giovanna Cambula Department of Veterinary Medicine, University of Sassari, Italy.
  • Giuseppe Esposito Department of Veterinary Medicine, University of Sassari, Italy.
  • Federica Fois Department of Veterinary Medicine, University of Sassari, Italy.
  • Simonetta Gianna Consolati Department of Veterinary Medicine, University of Sassari, Italy.
  • Anna Mureddu Department of Veterinary Medicine, University of Sassari, Italy.


The aim of the present study was to evaluate the occurrence of Salmonella spp., Verotoxigenic E. Coli (VTEC) and enumerate E. coli in shellfish (Mytilus galloprovincialis and Ruditapes decussatus) collected before and after depuration from two class B harvesting areas located in Sardinia (Italy). All the samples were analyzed for Salmonella spp. detection according to European Commission Regulation (EC) 2073/2005 and examined using the five tube Most-Probable-Number (MPN) method for enumeration of E. coli in accordance with the European Union reference method ISO 16649-3:2015. E. coli VTEC was investigated following a direct multiplex Polymerase Chain Reaction (PCR) screening test. The enumeration of E. coli met the European law limit for Class A areas of 230 MPN/100g. The averaged enumeration of E. coli in samples of M. galloprovincialis and R. decussatus collected at the harvesting time was 39 and 37 MPN/100 g respectively. The average contamination levels in samples collected after purification were 58 MPN/100g (M. galloprovincialis) and 32 MPN/100 g (R. decussatus). E. coli VTEC was not detected, on the contrary, Salmonella ser. Typhimurium was detected in one sample of M. galloprovincialis and in one sample of R. decussatus collected at the harvesting time. No significant associations were observed between E. coli levels in shellfish and environmental parameters of water or with the detection of Salmonella ser. Typhimurium in M. galloprovincialis and R. decussatus samples. Nevertheless, the occurrence of Salmonella ser. Typhimurium, involved in human infection outbreaks, should be considered a potential risk for consumers.



PlumX Metrics


Download data is not yet available.


Avery LM, Williams AP, Killham K, Jones DL, 2008. Survival of Escherichia coli O157:H7 in waters from lakes, rivers, puddles and animal-drinking troughs. Sci Total Environ 389:378-385. DOI: https://doi.org/10.1016/j.scitotenv.2007.08.049

Balière C, Rincé A, Thevenot D, Gourmelon M, 2015. Successful detection of pathogenic Shiga-toxin-producing Escherichia coli in shellfish, environmental waters and sediment using the ISO/TS-13136 method. Lett Appl Microbiol 60:315–320. DOI: https://doi.org/10.1111/lam.12386

Bazzardi R, Fattaccio MC, Salza S, Canu A, Marongiu E, Pisanu M, 2014. Preliminary study on norovirus, hepatitis A virus, Escherichia coli and their potential seasonality in shellfish from different growing and harvesting areas in Sardinia region. Ital J Food Safety 3:125–130. DOI: https://doi.org/10.4081/ijfs.2014.1601

Bazzoni AM, Mudadu AG, Esposito G, Urru R, Ortu S, Mara L, Uda MT, Arras I, Lorenzoni G, Sanna G, Bazzardi R, Marongiu A, Virgilio S, Meloni D, 2019. Bacteriological and viral investigation combined with determination of phytoplankton and algal biotoxins in mussels and water from a Mediterranean coastal lagoon. J Food Prot 82, 9: 1501-1511. DOI: https://doi.org/10.4315/0362-028X.JFP-18-569

Caprioli A, Morabito S, Brugere H, Oswald E, 2005. Enterohaemorrhagic Escherichia coli: Emerging issues on virulence and modes of transmission. Vet Res 36: 289–311. DOI: https://doi.org/10.1051/vetres:2005002

Carraro V, Sanna C, Brandas V, Sanna A, Pinna A, Coroneo V, 2015. Hygiene and health risks associated with the consumption of edible lamellibranch molluscs. Int J Food Microbiol 201: 52–57. DOI: https://doi.org/10.1016/j.ijfoodmicro.2015.02.014

Catalao Dionisio LP, Ferreiro JP, Fidalgo ML, Garcia Rosado ME, Borrego JJ, 2000. Occurrence of Salmonella spp. in estuarine and coastal waters of Portugal. Antonie Van Leeuwenhoek 78:99-106. DOI: https://doi.org/10.1023/A:1002733516539

Donovan TJ, Gallacher S, Andrews N, Greenwood M, Graham J, Russell J, Roberts S, Lee R, 1998. Modification of the standard method used in the United Kingdom for counting Escherichia coli in live bivalve molluscs. Commun Dis Public Health 1:188-196.

European Food Safety Authority and European Centre for Disease Prevention and Control (EFSA and ECDC), 2016. The European Union summary report on trends and sources of zoonoses, zoonotic agents and food-borne outbreaks in 2015. EFSA J 14, 12, 4634. DOI: https://doi.org/10.2903/j.efsa.2016.4634

European Commission, 2015. Regulation No 2015/2285 of 8 December 2015 amending annex II to Regulation (EC) No 854/ 2004 of the European Parliament and of the Council laying down specific rules for the organisation of official controls on products of animal origin intended for human consumption as regards certain requirements for live bivalve molluscs, echinoderms, tunicates and marine gastropods and annex I to Regulation (EC) No 2073/2005 on microbiological criteria for foodstuffs. In: Official Journal, L 323:2–4.

Karch H, Meyer T, 1989. Single primer pair for amplifying segments of distinct Shiga-like-toxin genes by Polymerase Chain-Reaction. J Clin Microbiol 27:2751–2757. DOI: https://doi.org/10.1128/JCM.27.12.2751-2757.1989

ISO, 2002. ISO 6579:2002: Microbiology of food and animal feeding stuffs -- Horizontal method for the detection of Salmonella spp. Geneva, Switzerland: International Organization for Standardization (ISO).

ISO, 2003. ISO 6887-3:2003: Microbiology of food and animal feeding stuffs – Preparation of test samples, initial suspension and decimal dilutions for microbiological examination - Part 3: Specific rules for the preparation of fish and fishery products. Geneva, Switzerland: International Organization for Standardization (ISO).

ISO, 2005. ISO/FDIS 16649-3 Microbiology of the food chain - Horizontal method for the enumeration of beta-glucuronidase-positive Escherichia coli -Part 3: Detection and most probable number technique using 5-bromo-4-chloro-3-indolyl-ß-Dglucuronide. Geneva, Switzerland: International Organization for Standardization (ISO).

Iwamoto M, Ayers T, Mahon BE, Swerdlow DL, 2010. Epidemiology of seafood-associated infections in the United States. Clin Microbiol Rev 23: 399-410. DOI: https://doi.org/10.1128/CMR.00059-09

Lamon S, Bastardo A, Meloni D, Consolati SG, Fois F, Porcheddu G, Agus V, Pes M, Cambula MG, Mureddu A, Romalde J, 2019a. Clonal relationship among Vibrio parahaemolyticus isolated from Mediterranean mussels (Mytilus galloprovincialis) and Grooved carpet shells (Ruditapes decussatus) harvested in Sardinia (Italy). Food Microbiol 84:103258. DOI: https://doi.org/10.1016/j.fm.2019.103258

Lamon S, Consolati SG, Fois F, Cambula MG, Pes M, Porcheddu G, Agus V, Esposito G, Mureddu A, Meloni D, 2019b. Occurrence, seasonal distribution and molecular characterization of V. vulnificus, V. cholerae and V. parahaemolyticus in shellfish (Mytilus galloprovincialis and Ruditapes decussatus) collected in Sardinia (Italy). J Food Prot 82,11: 1851-1856. DOI: https://doi.org/10.4315/0362-028X.JFP-19-021

Leoni F, Talevi G, Masini L, Ottaviani D, Rocchegiani E 2016. Trh (tdh−/trh+) gene analysis of clinical, environmental and food isolates of Vibrio parahaemolyticus as a tool for investigating pathogenicity. Int J Food Microbiol 225:43–53. DOI: https://doi.org/10.1016/j.ijfoodmicro.2016.02.016

Leoni F, Chierichetti S, Santarelli S, Talevi G, Masini L, Bartolini C, Rocchegiani E, Naceur Haouet M, Ottaviani D, 2017. Occurrence of Arcobacter spp. and correlation with the bacterial indicator of faecal contamination Escherichia coli in bivalve molluscs from the Central Adriatic, Italy. Int J Food Microbiol 245: 6–12. DOI: https://doi.org/10.1016/j.ijfoodmicro.2017.01.006

Malham SK, Rajko-Nenow P, Howlett E, Tuson KE, Perkins TL, Pallett DW, Wang H, Jago JF, Jones DL, McDonald JE, 2014. The interaction of human microbial pathogens, particulate material and nutrients in estuarine environments and their impacts on recreational and shellfish waters. Environ Sci Process Impact 16 (9): 2145-2155. DOI: https://doi.org/10.1039/C4EM00031E

Marceddu M, Lamon S, Consolati SG, Ciulli S, Mazza R, Mureddu A, Meloni D, 2017. Determination of Salmonella spp., E. coli VTEC, Vibrio spp., and Norovirus GI-GII in Bivalve Molluscs Collected from Growing Natural Beds in Sardinia (Italy). Foods, Special Issue Seafood Products: Safety and Quality, 6, 88: 1-8. DOI: https://doi.org/10.3390/foods6100088

Obiri-Danso K, Jones K, 2000. Intertidal sediments as reservoirs for Hippurate negative campylobacters, salmonellae and faecal indicators in three EU recognised bathing waters in North West England. Water Res 34: 519-527. DOI: https://doi.org/10.1016/S0043-1354(99)00146-3

Paton JC, Paton AW, 1998. Pathogenesis and diagnosis of Shiga toxin-producing Escherichia coli infections. J Clin Microbiol 11: 450–479. DOI: https://doi.org/10.1128/CMR.11.3.450

Prato R, Martinelli D, Tafuri S, Barbuti G, Quarto M, Germinario CA, Chironna M, 2013. Safety of shellfish and epidemiological pattern of enterically transmitted diseases in Italy. Int J Food Microbiol 162: 125–128. DOI: https://doi.org/10.1016/j.ijfoodmicro.2012.12.025

Rubini S, Galletti G, D’Incau M, Govoni G, Boschetti L, Berardelli C, Barbieri S, Merialdi G, Formaglio A, Guidi E, Bergamini M, Piva S, Serraino A, Giacometti S, 2018. Occurrence of Salmonella enterica subsp. enterica in bivalve molluscs and associations with Escherichia coli in molluscs and faecal coliforms in seawater. Food Control 84: 429-435. DOI: https://doi.org/10.1016/j.foodcont.2017.08.035

Sferlazzo G, Meloni D, Lamon S, Marceddu M, Mureddu M, Consolati SG, Pisanu M, Virgilio S, 2018. Evaluation of short purification cycles in naturally contaminated Mediterranean mussels (Mytilus galloprovincialis) harvested in Sardinia (Italy). Food Microbiol 74: 86-91. DOI: https://doi.org/10.1016/j.fm.2018.03.007

Touchon M, Hoede C, Tenaillon O, Barbe V, Baeriswyl S, Bidet P, Bingen E, Bonacorsi S, Bouchier C, Bouvet O, Calteau A, Chiapello H, Clermont O, Cruveiller S, Danchin A, Diard M, Dossat C, Karoui ME, Frapy E, Garry L, Ghigo JM, Gilles AM, Johnson J, Le Bouguenec C, Lescat M, Mangenot S, Martinez-Jehanne V, Matic I, Nassif X, Oztas S, Petit MA, Pichon C, Rouy Z, Ruf CS, Schneider D, Tourret J, Vacherie B, Vallenet D, Medigue C, Rocha EP, Denamur E, 2009. Organised genome dynamics in the Escherichia coli species results in highly diverse adaptive paths. PLoS Genetics 5(1): e1000344. DOI: https://doi.org/10.1371/journal.pgen.1000344

Williams AP, Gordon H, Jones DL, Strachan NJC, Avery LM, Killham K, 2008. Leaching of bioluminescent Escherichia coli O157:H7 from sheep and cattle faeces during simulated rainstorm events. J Appl Microbiol 105:1452–1460. DOI: https://doi.org/10.1111/j.1365-2672.2008.03898.x

Original Articles
Mytilus galloprovincialis; Ruditapes decussatus; Purification; E. coli; Salmonella ser. Typhimurium
  • Abstract views: 309

  • PDF: 152
  • HTML: 0
How to Cite
Lamon S, Piras F, Meloni D, Agus V, Porcheddu G, Pes M, Cambula MG, Esposito G, Fois F, Consolati SG, Mureddu A. Enumeration of <em>Escherichia coli</em> and determination of <em>Salmonella</em> spp. and verotoxigenic <em>Escherichia coli</em> in shellfish (<em>Mytilus galloprovincialis</em> and <em>Ruditapes decussatus</em&gt;) harvested in Sardinia, Italy. Ital J Food Safety [Internet]. 2020Dec.3 [cited 2021May13];9(4). Available from: https://pagepressjournals.org/index.php/ijfs/article/view/8625

Most read articles by the same author(s)