Sara Cleto

Bio: Sara Cleto is an academic researcher from Massachusetts Institute of Technology. The author has contributed to research in topics: Siderophore & Synthetic biology. The author has an hindex of 9, co-authored 21 publications receiving 857 citations. Previous affiliations of Sara Cleto include University of Minho & Harvard University.

Genetically Engineered Phages: a Review of Advances over the Last Decade

Abstract: Soon after their discovery in the early 20th century, bacteriophages were recognized to have great potential as antimicrobial agents, a potential that has yet to be fully realized. The nascent field of phage therapy was adversely affected by inadequately controlled trials and the discovery of antibiotics. Although the study of phages as anti-infective agents slowed, phages played an important role in the development of molecular biology. In recent years, the increase in multidrug-resistant bacteria has renewed interest in the use of phages as antimicrobial agents. With the wide array of possibilities offered by genetic engineering, these bacterial viruses are being modified to precisely control and detect bacteria and to serve as new sources of antibacterials. In applications that go beyond their antimicrobial activity, phages are also being developed as vehicles for drug delivery and vaccines, as well as for the assembly of new materials. This review highlights advances in techniques used to engineer phages for all of these purposes and discusses existing challenges and opportunities for future work.

Corynebacterium glutamicum Metabolic Engineering with CRISPR Interference (CRISPRi)

Abstract: Corynebacterium glutamicum is an important organism for the industrial production of amino acids. Metabolic pathways in this organism are usually engineered by conventional methods such as homologous recombination, which depends on rare double-crossover events. To facilitate the mapping of gene expression levels to metabolic outputs, we applied CRISPR interference (CRISPRi) technology using deactivated Cas9 (dCas9) to repress genes in C. glutamicum. We then determined the effects of target repression on amino acid titers. Single-guide RNAs directing dCas9 to specific targets reduced expression of pgi and pck up to 98%, and of pyk up to 97%, resulting in titer enhancement ratios of l-lysine and l-glutamate production comparable to levels achieved by gene deletion. This approach for C. glutamicum metabolic engineering, which only requires 3 days, indicates that CRISPRi can be used for quick and efficient metabolic pathway remodeling without the need for gene deletions or mutations and subsequent selection.

Synthetic biology and microbioreactor platforms for programmable production of biologics at the point-of-care

Abstract: Current biopharmaceutical manufacturing systems are not compatible with portable or distributed production of biologics, as they typically require the development of single biologic-producing cell lines followed by their cultivation at very large scales. Therefore, it remains challenging to treat patients in short time frames, especially in remote locations with limited infrastructure. To overcome these barriers, we developed a platform using genetically engineered Pichia pastoris strains designed to secrete multiple proteins on programmable cues in an integrated, benchtop, millilitre-scale microfluidic device. We use this platform for rapid and switchable production of two biologics from a single yeast strain as specified by the operator. Our results demonstrate selectable and near-single-dose production of these biologics in <24 h with limited infrastructure requirements. We envision that combining this system with analytical, purification and polishing technologies could lead to a small-scale, portable and fully integrated personal biomanufacturing platform that could advance disease treatment at point-of-care. Current biopharmaceutical manufacturing platforms use single biologic-producing cell lines cultured at large scales. Here the authors develop a small-scale, portable biomanufacturing platform to produce single dose IFNa2b and rHGH from a single engineered strain of P. pastoris.

A review of current and emergent biofilm control strategies

Abstract: Microbial adhesion to surfaces and the consequent biofilm formation has been documented in many different environments. Biofilms constitute a protected mode of growth that allows microorganisms to survival in hostile environments, being their physiology and behavior significantly different from their planktonic counterparts. In dairy industry, biofilms may be a source of recalcitrant contaminations, causing food spoilage and are possible sources of public health problems such as outbreaks of foodborne pathogens. Biofilms are difficult to eradicate due to their resistant phenotype. However, conventional cleaning and disinfection regimens may also contribute to inefficient biofilm control and to the dissemination of resistance. Consequently, new control strategies are constantly emerging with main incidence in the use of biosolutions (enzymes, phages, interspecies interactions and antimicrobial molecules from microbial origin). The present review will focus on describing the mechanisms involved in biofilm formation and behavior, deleterious effects associated with their presence, and some of the current and emergent control strategies, providing new insight of concern for food industry.

Antibacterial activity and mode of action of ferulic and gallic acids against pathogenic bacteria

Abstract: The increased resistance of pathogenic microorganisms is frequently attributed to the extreme and inadequate use of antibiotics and transmission of resistance within and between individuals. To counter the emergence of resistant microorganisms, considerable resources have been invested in the search for new antimicrobials. Plants synthesize a diverse array of secondary metabolites (phytochemicals) known to be involved in defense mechanisms, and in the last few years it is recognized that some of these molecules have health beneficial effects, including antimicrobial properties. In this study, the mechanism of action of gallic (GA) and ferulic (FA) acids, a hydroxybenzoic acid and a hydroxycinnamic acid, was assessed on Escherichia coli, Pseudomonas aeruginosa, Staphylococcus aureus, and Listeria monocytogenes. The targets of antimicrobial action were studied using different bacterial physiological indices: minimum inhibitory concentration (MIC), minimum bactericidal concentration (MBC), membrane ...