This thesis meanly deals with the selection of starter culture, with oleuropenolytic activity, tailored to produce Sicilian table olives at low salt content. In detail, the aim of this study project was to: (i) develop a selected starter cultures for table olives fermentation at low salt content; (ii) investigate gene expression of β-glucosidase enzyme of selected strains, under stress condition such as brine; and (iii) elucidate on the effect of the selected starter cultures on microbial composition and function during table olives fermentation through an omics approach. In detail, this thesis is composed by: An Introduction and three Chapters describes as following. Introduction: provides an overview of the omics techniques currently applied to the study of the table olive microbiota. It also highlights the possibility of using an integrated approach capable of understanding in detail the dynamics and variability of this complex microbial consortium. Chapter 1 showed how the in vitro study of selection criteria revealed that the ability to grow at different pH, salt concentration and temperature is a strain-dependent condition, and that low temperature is the parameter that most negatively affected the survival ability of strains under simulated process conditions. The comprehension of these capabilities allows the discrimination at strain level and the appropriate selection of strains tailored for fermentation processes. Although subsequent transcriptomic and proteomic studies are required to validate the use of selected b-glucosidase-positive strains, the results of Chapter 2 showed that, through transcriptomic profiling, the designed primers were able to detect the gene encoding for the b-glucosidase enzyme directly involved in oleuropein hydrolysis. In addition, the use of mass spectrometry techniques made it possible to detect intermediate molecules (Hy-EDA) and thus to propose an alternative metabolic pathway for the formation of the final hydrolysis compounds. The application of β-glucosidase-positive starter strains was further validated in the study reported in Chapter 3. By applying a dual approach based on culture-dependent, metagenetic and volatilomics, it was possible to detect differences in microbial variability and VOC profiles between table olive samples with and without the addition of starter cultures. Besides sharply reducing the pH values in the medium, making it safer, the use of starter cultures contributes to the development of pleasant compounds in the final product, as well as preserving it from being spoilage microorganisms responsible for the appearance of unpleasant substances.
Il presente elaborato finale riguarda la selezione di una coltura starter, con attività oleuropenolitica, adattata alla produzione di olive da tavola siciliane a basso contenuto di sale. In dettaglio, lo scopo di questo progetto di studio è stato quello di: (i) sviluppare una coltura starter selezionata per la fermentazione delle olive da tavola a basso contenuto di sale; (ii) studiare l'espressione genica dell'enzima β-glucosidasi dei ceppi selezionati, in condizioni di stress come la salamoia; e (iii) chiarire l'effetto delle colture starter selezionate sulla composizione e la funzione microbica durante la fermentazione delle olive da tavola attraverso un approccio omico. In dettaglio, questa tesi è composta da: un'introduzione e tre capitoli descritti come segue. L'introduzione fornisce una panoramica delle tecniche omiche attualmente applicate allo studio del microbiota delle olive da tavola. Evidenzia inoltre, la possibilità di utilizzare un approccio integrato in grado di comprendere nel dettaglio le dinamiche e la variabilità di questo complesso consorzio microbico. Il capitolo 1 ha mostrato come lo studio in vitro dei criteri di selezione abbia rivelato che la capacità di crescere a diversi pH, concentrazione salina e temperatura è una condizione ceppo-dipendente, e che la bassa temperatura è il parametro che più ha influenzato negativamente la capacità di sopravvivenza dei ceppi, in condizioni simulate di processo. La comprensione di queste capacità ha permesso la discriminazione a livello di ceppo e la selezione appropriata di ceppi su misura per i processi fermentativi. Sebbene siano necessari successivi studi trascrittomici e proteomici per convalidare l'uso di ceppi b-glucosidasi-positivi selezionati, i risultati del Capitolo 2 hanno mostrato che, attraverso il profilo trascrittomico, i primer progettati sono stati in grado di rilevare il gene che codifica per l'enzima b-glucosidasi direttamente coinvolto nell'idrolisi dell'oleuropeina. Inoltre, l'uso di tecniche di spettrometria di massa ha permesso di rilevare molecole intermedie (Hy-EDA) e quindi di proporre una via metabolica alternativa per la formazione dei composti finali dell'idrolisi. L'applicazione di ceppi starter β-glucosidasi-positivi è stata ulteriormente convalidata nello studio riportato nel capitolo 3. Applicando un doppio approccio basato sulla coltura-dipendente, metagenetica e volatilomica, è stato possibile rilevare differenze nella variabilità microbica e nei profili di VOCs tra campioni di olive da tavola con e senza l'aggiunta di colture starter. Oltre a ridurre drasticamente i valori di pH nel mezzo, rendendolo più sicuro, l'uso di colture starter contribuisce allo sviluppo di composti gradevoli nel prodotto finale, oltre a preservarlo da microrganismi alteranti, responsabili della comparsa di sostanze sgradevoli.
Study of stress adaptation, versatility and functionality of selected Lactiplantibacillus plantarum strains used as starter cultures for table olives fermentation
VACCALLUZZO, AMANDA
2021
Abstract
This thesis meanly deals with the selection of starter culture, with oleuropenolytic activity, tailored to produce Sicilian table olives at low salt content. In detail, the aim of this study project was to: (i) develop a selected starter cultures for table olives fermentation at low salt content; (ii) investigate gene expression of β-glucosidase enzyme of selected strains, under stress condition such as brine; and (iii) elucidate on the effect of the selected starter cultures on microbial composition and function during table olives fermentation through an omics approach. In detail, this thesis is composed by: An Introduction and three Chapters describes as following. Introduction: provides an overview of the omics techniques currently applied to the study of the table olive microbiota. It also highlights the possibility of using an integrated approach capable of understanding in detail the dynamics and variability of this complex microbial consortium. Chapter 1 showed how the in vitro study of selection criteria revealed that the ability to grow at different pH, salt concentration and temperature is a strain-dependent condition, and that low temperature is the parameter that most negatively affected the survival ability of strains under simulated process conditions. The comprehension of these capabilities allows the discrimination at strain level and the appropriate selection of strains tailored for fermentation processes. Although subsequent transcriptomic and proteomic studies are required to validate the use of selected b-glucosidase-positive strains, the results of Chapter 2 showed that, through transcriptomic profiling, the designed primers were able to detect the gene encoding for the b-glucosidase enzyme directly involved in oleuropein hydrolysis. In addition, the use of mass spectrometry techniques made it possible to detect intermediate molecules (Hy-EDA) and thus to propose an alternative metabolic pathway for the formation of the final hydrolysis compounds. The application of β-glucosidase-positive starter strains was further validated in the study reported in Chapter 3. By applying a dual approach based on culture-dependent, metagenetic and volatilomics, it was possible to detect differences in microbial variability and VOC profiles between table olive samples with and without the addition of starter cultures. Besides sharply reducing the pH values in the medium, making it safer, the use of starter cultures contributes to the development of pleasant compounds in the final product, as well as preserving it from being spoilage microorganisms responsible for the appearance of unpleasant substances.File | Dimensione | Formato | |
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https://hdl.handle.net/20.500.14242/73771
URN:NBN:IT:UNICT-73771