A study of the bacterial ecology of a chocolate ingredient production environment and characterisation of Salmonella species a zoonotic food-borne bacteria of importance to this food industry

Abstract

The safe production of chocolate is a critical issue to both the consumer and the brand, requiring the application of Hazard Analysis Critical Control Points (HACCP) system across the production process. These production environments are challenged on occasion with different hazards of importance to food safety. Control of bacterial hazards as exemplified by pathogenic bacteria like Salmonella is an important part of the process management. Salmonella is a Gram-negative, facultatively anaerobic bacterium that causes food-borne illness with significant health effects. The application of next-generation sequencing methods were used to investigate these bacterial isolates. MinION-based rapid sequencing was used to identify pathogenic bacteria of importance to chocolate production contained in these environmental samples. Full length 16S rRNA amplicon-mediated sequencing was used to describe and analyse the core microbiome of the swabbed samples taken from a variety of locations within this chocolate production environment. Results from this study provided a species-specific description of the core microbiome of these samples.
56 Salmonella isolates were chosen and analysed phenotypically and by whole-genome sequencing. Phenotypic experiments relevant to assessing their colonising potential, were conducted to characterise this selection of Salmonella. A bioinformatics pipeline was applied to confirm species identification, identify key virulence factors, evaluate antimicrobial resistance genotypes and calculate average nucleotide identity. An analysis of the AMR genotypes detected in silico identified 19 different antimicrobial classes and which largely confirmed the AST tests carried out. Salmonella infections using two distinct host models -Galleria mellonella and mice was investigated. Data obtained provides valuable insights into categorising different Salmonella at crucial points during the infection process, such as the onset of mortality and potential variations in susceptibility among individual larvae, arising from these infections. Subsequently, the immune responses were evaluated using ELISA for five key cytokines namely-KC, IL-6, IL-1β, IL-17, and MCP-1.

Thesis embargoed until 31 December 2029.

Date of Award	Dec 2024
Original language	English
Awarding Institution	Queen's University Belfast
Sponsors	Biotechnology & Biological Sciences Research Council & Mondelez UK R&D Ltd
Supervisor	Seamus Fanning (Supervisor), Helina Marshall (Supervisor), Beckie Ingram (Supervisor) & Caroline Meharg (Supervisor)