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Dott.ssa Selene Rubiola

Phd thesis

Next generation sequencing approaches for the resistome characterization of milk and milk production environments.

The extensive use of antibiotics in the livestock production systems can lead to the presence of antimicrobial resistance (AMR) genes in bacteria naturally occurring in milk and dairy products or that may contaminate such products. Microorganisms harboring antibiotic resistance genes are thereby introduced into the food chain. In this context, high-throughput next-generation sequencing (NGS) technologies can be used as valuable tools to improve fast AMR genes identification and characterization. Among them, the whole metagenome sequencing (WMS) has not yet been widely applied to explore the resistome of milk, dairy products and environments thereof, notwithstanding its huge potential; this is mainly due to the challenges related to its use, including the complexity and dimension of generated data and the vast amount of background host DNA. Given the current paucity of literature concerning the application of WMS on milk, the first aim of this thesis was to evaluate the effect of different sequencing depths, host DNA depletion methods and matrices to characterize the resistome and the microbiome of bulk tank milk. Secondly, the present thesis aimed to investigate the resistome and the microbiome of different dairy farms taking advantage of bulk tank milk filters as promising though yet unexplored tools. Herein, the WMS strategy applied has allowed the identification of numerous AMR genes, overcoming the host DNA issue; notably most of the species involved were predicted to be Gram-negative bacteria belonging to the Moraxellaceae and to the Enterobacteriaceae families, pointing out the role of these bacterial families as reservoirs of AMR determinants. Besides, the full length 16S metabarcoding was integrated with the WMS technique to evaluate the microbial resolution of this third-generation sequencing method, whose application in food and food-related matrices is yet in its infancy. In accordance with our results, bulk tank milk can be considered a source of bacteria harboring AMR genes; this points out the importance of properly informing food operators about the risk associated with poor hygiene practices in the dairy production environment and the potential risk for public health, highlighting the importance of broadening a One Health perspective. Translating these findings as risk assessment outputs heralds the next generation of food safety controls.

Research activities


Panebianco, F, Rubiola, S, Di Ciccio, PA (2022). The Use of Ozone as an Eco-Friendly Strategy against Microbial Biofilm in Dairy Manufacturing Plants: A Review. Microorganisms. 10(1):162.

Rubiola, S, Moroni, B, Carisio, L, Rossi, L, Chiesa, F, Martano, G, Cavallo, E, Rambozzi, L (2021). Risk Factors for Bovine Cysticercosis in North-West Italy: A Multi-Year Case-Control Study. Animals. 11(11):3049.

Panebianco, F, Rubiola, S, Chiesa, F, Civera, T, Di Ciccio, PA (2021). Effect of Gaseous Ozone on Listeria monocytogenes Planktonic Cells and Biofilm: An In Vitro Study. Foods. 10(7):1484.

Rubiola, S, Civera, T, Panebianco, F, Vercellino, D, Chiesa, F (2021). Molecular detection of cattle Sarcocystis spp. in North-West Italy highlights their association with bovine eosinophilic myositis. Parasites Vectors 14, 223.

Cuccato, M, Rubiola, S, Giannuzzi, D, Grego, E, Pregel, P, Divari, S, Cannizzo, F T (2021). 16S rRNA Sequencing Analysis of the Gut Microbiota in Broiler Chickens Prophylactically Administered with Antimicrobial Agents. Antibiotics 10, no. 2: 146.

Rubiola, S, Chiesa, F, Dalmasso, A, Di Ciccio, P, Civera, T (2020). Detection of Antimicrobial Resistance Genes in the Milk Production Environment: Impact of Host DNA and Sequencing Depth. Front. Microbiol. 11.

Rubiola, S, Civera, T, Ferroglio, E, Zanet, S, Zaccaria, T, Brossa, S, Cipriani, R, Chiesa, F (2020). Molecular differentiation of cattle Sarcocystis spp. by multiplex PCR targeting 18S and COI genes following identification of Sarcocystis hominis in human stool samples.

Di Ciccio, P, Rubiola, S, Grassi, MA, Civera, T, Abbate, F, Chiesa, F (2020). Fate of Listeria monocytogenes in the presence of resident cheese microbiota on common packaging materials. Front. Microbiol. 11.

Rubiola, S, Chiesa, F, Zanet, S, Civera, T (2018). Molecular identification of Sarcocystis spp. in cattle: partial sequencing of Cytochrome C Oxidase subunit 1(COI). Italian Journal of Food Safety, 7(4): 7725. doi: 10.4081/ijfs.2018.7725

Tramuta, C, Gallina, S, Bellio, A, Bianchi, DM, Chiesa, F, Rubiola, S, Romano, A, Decastelli, L (2018). A Set of Multiplex Polymerase Chain Reactions for Genomic Detection of Nine Edible Insect Species in Foods. Journal of Insect Science, 18(5): 3; 1-6.

Last update: 08/05/2022 12:43

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