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      Genome editing of lactic acid bacteria: opportunities for food, feed, pharma and biotech

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          Abstract

          This mini-review provides a perspective of traditional, emerging and future applications of lactic acid bacteria (LAB) and how genome editing tools can be used to overcome current challenges in all these applications. It also describes available tools and how these can be further developed, and takes current legislation into account. Genome editing tools are necessary for the construction of strains for new applications and products, but can also play a crucial role in traditional ones, such as food and probiotics, as a research tool for gaining mechanistic insights and discovering new properties. Traditionally, recombinant DNA techniques for LAB have strongly focused on being food-grade, but they lack speed and the number of genetically tractable strains is still rather limited. Further tool development will enable rapid construction of multiple mutants or mutant libraries on a genomic level in a wide variety of LAB strains. We also propose an iterative Design–Build–Test–Learn workflow cycle for LAB cell factory development based on systems biology, with ‘cell factory’ expanding beyond its traditional meaning of production strains and making use of genome editing tools to advance LAB understanding, applications and strain development.

          Abstract

          Traditional, emerging and future applications of lactic acid bacteria can all benefit from genome editing and a proposed Design–Build–Test–Learn workflow cycle for advancement of strain development.

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          Most cited references107

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          Increasing the genome-targeting scope and precision of base editing with engineered Cas9-cytidine deaminase fusions

          Base editing is a recently developed approach to genome editing that uses a fusion protein containing a catalytically defective Streptococcus pyogenes Cas9, a cytidine deaminase, and an inhibitor of base excision repair to induce programmable, single-nucleotide changes in the DNA of living cells without generating double-strand DNA breaks, without requiring a donor DNA template, and without inducing an excess of stochastic insertions and deletions 1 . Here we report the development of five new C→T (or G→A) base editors that use natural and engineered Cas9 variants with different protospacer-adjacent motif (PAM) specificities to expand the number of sites that can be targeted by base editing by 2.5-fold. Additionally, we engineered new base editors containing mutated cytidine deaminase domains that narrow the width of the apparent editing window from approximately 5 nucleotides to as little as 1 to 2 nucleotides, enabling the discrimination of neighboring C nucleotides that would previously be edited with comparable efficiency, thereby doubling the number of disease-associated target Cs that can be corrected preferentially over nearby non-target Cs. Collectively, these developments substantially increase the targeting scope of base editing and establish the modular nature of base editors.
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            Expanding the biotechnology potential of lactobacilli through comparative genomics of 213 strains and associated genera

            Lactobacilli are a diverse group of species that occupy diverse nutrient-rich niches associated with humans, animals, plants and food. They are used widely in biotechnology and food preservation, and are being explored as therapeutics. Exploiting lactobacilli has been complicated by metabolic diversity, unclear species identity and uncertain relationships between them and other commercially important lactic acid bacteria. The capacity for biotransformations catalysed by lactobacilli is an untapped biotechnology resource. Here we report the genome sequences of 213 Lactobacillus strains and associated genera, and their encoded genetic catalogue for modifying carbohydrates and proteins. In addition, we describe broad and diverse presence of novel CRISPR-Cas immune systems in lactobacilli that may be exploited for genome editing. We rationalize the phylogenomic distribution of host interaction factors and bacteriocins that affect their natural and industrial environments, and mechanisms to withstand stress during technological processes. We present a robust phylogenomic framework of existing species and for classifying new species.
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              Antibacterial activities of bacteriocins: application in foods and pharmaceuticals

              Bacteriocins are a kind of ribosomal synthesized antimicrobial peptides produced by bacteria, which can kill or inhibit bacterial strains closely-related or non-related to produced bacteria, but will not harm the bacteria themselves by specific immunity proteins. Bacteriocins become one of the weapons against microorganisms due to the specific characteristics of large diversity of structure and function, natural resource, and being stable to heat. Many recent studies have purified and identified bacteriocins for application in food technology, which aims to extend food preservation time, treat pathogen disease and cancer therapy, and maintain human health. Therefore, bacteriocins may become a potential drug candidate for replacing antibiotics in order to treat multiple drugs resistance pathogens in the future. This review article summarizes different types of bacteriocins from bacteria. The latter half of this review focuses on the potential applications in food science and pharmaceutical industry.

                Author and article information

                Journal
                FEMS Microbiol Lett
                FEMS Microbiol. Lett
                femsle
                FEMS Microbiology Letters
                Oxford University Press
                0378-1097
                1574-6968
                18 December 2018
                January 2019
                18 December 2018
                : 366
                : 1
                : fny291
                Affiliations
                [1]The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kemitorvet B220, 2800 Kongens Lyngby, Denmark
                Author notes
                Corresponding author: The Novo Nordisk Foundation Center for Biosustainability, Technical University of Denmark, Kemitorvet B220, 2800 Kongens Lyngby, Denmark. Tel: +4593511710; Fax: not available; E-mail: elfebo@ 123456biosustain.dtu.dk
                Author information
                http://orcid.org/0000-0001-6189-7753
                Article
                fny291
                10.1093/femsle/fny291
                6322438
                30561594
                6a43deea-9576-4af4-9846-96493c825a84
                © FEMS 2018.

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                : 27 August 2018
                : 16 December 2018
                Page count
                Pages: 12
                Funding
                Funded by: Marine Biotechnology
                Award ID: 5178–00003B
                Categories
                Minireview
                Biotechnology and Synthetic Biology
                Mini Review

                Microbiology & Virology
                genetic tool development,food fermentation,biotherapeutics,phytotherapeutics,synthetic biology,gmo regulation

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