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个人简介
My PhD research focused on optimizing plasmid DNA production fermentation processes using Escherichia coli. This research was a part of an R&D project whose purpose was the creation of a pilot plant for the production of DNA vaccines. My approach to this topic of research is unique because I’ve developed and implemented new tools for bioprocess monitoring. I’ve also performed important studies on the impact of several culture conditions on plasmid stability and cell physiology which gave researchers a new perspective for optimizing their processes. All the information gathered during this project was applied to optimize cell growth and plasmid DNA titres using benchtop bioreactors, yielding a very reproducible process that could be implemented at a pilot scale. The central component to my research has been the study of the influence of plasmid DNA fermentation strategies in the host cell, while using that knowledge to further optimize the process. However, the best yield isn’t necessarily the best process. My research tackled this issue in the field of cell physiology and plasmid DNA segregational stability analysis where new techniques and new methodologies such as flow cytometry and real-time PCR can provide new insights about the performance of the fermentation process. When I began my PhD project, I noticed that my production system and culture conditions were imposing a high metabolic burden to the host strain, E. coli, which needed to be overcome. In order to pursue this goal, we have been implementing new flow cytometric protocols using newly developed far-red dyes thanks to the collaborations established with Biostatus Limited. The fluorescent stain used, DRAQ5, allowed the development of a new flow cytometric protocol for the analysis of cell cycle progression during bacterial cell growth in Gram-negative bacteria, which is an important tool for evaluating bacterial population heterogeneity and the impact of metabolic burden on bacterial cell replication. While performing my PhD, I got involved in other ongoing projects related with Microbiology. One of such projects was developed in collaboration with Molecular Probes and involved the development of a flow cytometric protocol using a far-red fluorescent dye for cell cycle analysis. After a preliminary optimization step, this flow cytometric protocol was successfully applied to ciprofloxacin antibacterial susceptibility testing using flow cytometry, which enable us to distinguish between ciprofloxacin bacteriostatic and bactericidal activities against Escherichia coli, providing a faster and easier method for the evaluation of ciprofloxacin antibacterial activity. The latest project I got involved with was related with the study of the antimicrobial properties of essential oils used as food additives. This research was focused on coriander (Coriandrum sativum L.) essential oil antibacterial and antifungal actions on several pathogens. My work in this project was related with the implementation of flow cytometric protocols for the evaluation of cellular functions both in bacterial and yeast cells, such as efflux activity, membrane permeability and polarization, cell cycle and metabolic activity. The assessment of these cellular functions is of great relevance in order to assess the possible mechanism of action of the essential oil.
In my post-doc, I'm expanding my research to the field of Food Science, pursuing an investigation more related to antimicrobial food packaging and functional foods with antimicrobial properties. During my PhD, I came across other researchers working with food-related pathogens, such as Helicobacter spp. and Arcobacter spp and the pathogeneicity of these microorganisms and other foodborne pathogens has captured my attention. I see potential in the development of new strategies for food preservation since one of the major challenges for current food industry is to prevent the transmission of microbial foodborne pathogens throughout the food chain while satisfying the growing consumers’ demands for better foods, which are nutrient and vitamin-rich and minimally processed. The WHO, in their 2002 food strategic planning meeting, has already stated that the extent of microbiological problems in food safety is of major importance and new methods for food preservation and packaging need to be developed in order to circumvent today’s limitations and hazards. Therefore, the development of new antimicrobial food packaging or new functional foods which could circumvent today’s limitations is of great relevance and could have a tremendous impact in world health. These new food preservation strategies should include the use of natural products due to their proven benefits and also due to the increased resistance of foodborne pathogens to commonly used food preservatives and antibiotics.
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FOOD PACKAGING AND SHELF LIFE (2023): 101177-101177
Marta Quatorze,Filomena S. G. Silva,Ana Duarte, Júlio Roquete Cardoso,Carolina Caetano,João Ramalho‐Santos, Paulo J. Oliveira,Sara Varela Amaral
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