Publicação
Vir-ID-L: Viral proteins to develop an improved diagnosis of Listeria monocytogenes
| Resumo: | Listeria monocytogenes is a gram-positive bacterium with ubiquitous distribution in nature. It is responsible for listeriosis, a foodborne disease with particular impact on immunocompromised individuals, pregnant women and newborns. The main route of transmission is through consumption of contaminated foods. Therefore, fast and accurate bacterial detection is crucial for detecting L. monocytogenes in food processing environments, as well as in clinical context to correctly diagnose, prevent and treat the disease. Traditional detection methods have various limitations and bacteriophages, as well as their proteins, have emerged as a solution for fast and specific bacterial detection. In this project, two functional proteins were identified, with ability to bind to L. monocytogenes: one receptor binding protein (RBP) and one cell-wall binding domain (CBD). Proteins were identified through bioinformatic analysis and their fusion with green fluorescent protein (GFP) allowed for functional analysis. GFP-tagged versions of both proteins were successfully expressed, purified, and shown to bind L. monocytogenes cells, with signal detection by epifluorescence microscopy. Binding specificity was evaluated by epifluorescence microscpy against strains belonging to different Listeria serovars and other bacterial genera, to which other Listeria CBDs reportedly bind to. Results were promising, particularly for the RBP, shown to be specific for L. monocytogenes, binding to 1/2 serovars, known to be among the most relevant in strains isolated from foods and clinical environments. The CBD also decorated cells of strains belonging to 1/2 serovars, but showed weaker binding to other Listeria species and other genera. This behaviour goes along with other Listeria CBDs, that are also not specific to this genera. This research contributes to the development of a novel detection technique for Listeria, particularly L. monocytogenes, using phage proteins, offering a promising alternative to traditional techniques. |
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| Autores principais: | Silva, Isabel Maria Ferreira da |
| Assunto: | Bacterial detection Bacterial diagnosis Bacteriophage proteins epifluorescence microscopy Listeria monocytogenes Deteção de bactérias Diagnóstico bacteriano Proteínas de bacteriófagos Microscopia de epifluorescência |
| Ano: | 2024 |
| País: | Portugal |
| Tipo de documento: | dissertação de mestrado |
| Tipo de acesso: | acesso embargado |
| Instituição associada: | Universidade do Minho |
| Idioma: | inglês |
| Origem: | RepositóriUM - Universidade do Minho |
| Resumo: | Listeria monocytogenes is a gram-positive bacterium with ubiquitous distribution in nature. It is responsible for listeriosis, a foodborne disease with particular impact on immunocompromised individuals, pregnant women and newborns. The main route of transmission is through consumption of contaminated foods. Therefore, fast and accurate bacterial detection is crucial for detecting L. monocytogenes in food processing environments, as well as in clinical context to correctly diagnose, prevent and treat the disease. Traditional detection methods have various limitations and bacteriophages, as well as their proteins, have emerged as a solution for fast and specific bacterial detection. In this project, two functional proteins were identified, with ability to bind to L. monocytogenes: one receptor binding protein (RBP) and one cell-wall binding domain (CBD). Proteins were identified through bioinformatic analysis and their fusion with green fluorescent protein (GFP) allowed for functional analysis. GFP-tagged versions of both proteins were successfully expressed, purified, and shown to bind L. monocytogenes cells, with signal detection by epifluorescence microscopy. Binding specificity was evaluated by epifluorescence microscpy against strains belonging to different Listeria serovars and other bacterial genera, to which other Listeria CBDs reportedly bind to. Results were promising, particularly for the RBP, shown to be specific for L. monocytogenes, binding to 1/2 serovars, known to be among the most relevant in strains isolated from foods and clinical environments. The CBD also decorated cells of strains belonging to 1/2 serovars, but showed weaker binding to other Listeria species and other genera. This behaviour goes along with other Listeria CBDs, that are also not specific to this genera. This research contributes to the development of a novel detection technique for Listeria, particularly L. monocytogenes, using phage proteins, offering a promising alternative to traditional techniques. |
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