Publicação
Evaluation of the delivery of an anti-Listeria endolysin via CRISPR-Cas9 engineered probiotic Saccharomyces boulardii
| Resumo: | Listeriosis is a foodborne infection caused by Listeria monocytogenes that causes febrile gastroenteritis and central nervous system infections and that can often lead to fatality. Upon consumption of contaminated food, Listeria is able to survive a number of gastrointestinal stressors, including competition with the host microbiota. The emergence of antibiotic-resistant clones of L. monocytogenes, together with the side effects of antibiotic treatment, highlights the need for alternatives or additives for its treatment and prevention. Saccharomyces boulardii is a probiotic yeast that is often used alongside antibiotics to minimize side effects since it is not affected by them as a result of its eukaryotic nature. Furthermore, it can be engineered to produce a wide range of molecules. We previously engineered Saccharomyces cerevisiae through CRISPR-Cas9 integration to produce Ply511, a bacteriophage endolysin active against L. monocytogenes, showing the potential of engineered yeast to produce endolysins for biocontrol. In this study, we extended this approach to the probiotic yeast S. boulardii and directly compared the two yeasts as secretion hosts for Ply511. Using a simulated human gastrointestinal environment, we evaluated their ability to retain endolysin activity and reduce L. monocytogenes levels. We then tested the cell extracts from both yeasts in a bacterial consortium termed SImplified HUman intestinal MIcrobiota (SIHUMI), confirming a specificity for Listeria. Finally, we evaluated their activity in a simulated intestinal fermentation using fecal samples from human donors. Overall, this study demonstrates the potential of delivering endolysins to the gut via engineered probiotic S. boulardii. |
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| Autores principais: | Sáez, David |
| Outros Autores: | Carvalho, João Paulo; Murray, Ellen; Colombo, Natalia Soledad Rios; Lamas, Alexandre; Cobas, Alejandra Cardelle; Hill, Colin; Azeredo, Joana; Domingues, Lucília |
| Assunto: | CRISPR-Cas systems CRISPR-Cas9 Genome Editing Listeria Metabolic Engineering Saccharomyces cerevisiae Bacillus subtilis |
| Ano: | 2026 |
| País: | Portugal |
| Tipo de documento: | artigo |
| Tipo de acesso: | acesso aberto |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | Listeriosis is a foodborne infection caused by Listeria monocytogenes that causes febrile gastroenteritis and central nervous system infections and that can often lead to fatality. Upon consumption of contaminated food, Listeria is able to survive a number of gastrointestinal stressors, including competition with the host microbiota. The emergence of antibiotic-resistant clones of L. monocytogenes, together with the side effects of antibiotic treatment, highlights the need for alternatives or additives for its treatment and prevention. Saccharomyces boulardii is a probiotic yeast that is often used alongside antibiotics to minimize side effects since it is not affected by them as a result of its eukaryotic nature. Furthermore, it can be engineered to produce a wide range of molecules. We previously engineered Saccharomyces cerevisiae through CRISPR-Cas9 integration to produce Ply511, a bacteriophage endolysin active against L. monocytogenes, showing the potential of engineered yeast to produce endolysins for biocontrol. In this study, we extended this approach to the probiotic yeast S. boulardii and directly compared the two yeasts as secretion hosts for Ply511. Using a simulated human gastrointestinal environment, we evaluated their ability to retain endolysin activity and reduce L. monocytogenes levels. We then tested the cell extracts from both yeasts in a bacterial consortium termed SImplified HUman intestinal MIcrobiota (SIHUMI), confirming a specificity for Listeria. Finally, we evaluated their activity in a simulated intestinal fermentation using fecal samples from human donors. Overall, this study demonstrates the potential of delivering endolysins to the gut via engineered probiotic S. boulardii. |
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