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Recent advances in genetic tools for engineering probiotic lactic acid bacteria.

ABSTRACT: Synthetic biology has grown exponentially in the last few years, with a variety of biological applications. One of the emerging applications of synthetic biology is to exploit the link between microorganisms, biologics, and human health. To exploit this link, it is critical to select effective synthetic biology tools for use in appropriate microorganisms that would address unmet needs in human health through the development of new game-changing applications and by complementing existing technological capabilities. Lactic acid bacteria (LAB) are considered appropriate chassis organisms that can be genetically engineered for therapeutic and industrial applications. Here, we have reviewed comprehensively various synthetic biology techniques for engineering probiotic LAB strains, such as clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 mediated genome editing, homologous recombination, and recombineering. In addition, we also discussed heterologous protein expression systems used in engineering probiotic LAB. By combining computational biology with genetic engineering, there is a lot of potential to develop next-generation synthetic LAB with capabilities to address bottlenecks in industrial scale-up and complex biologics production. Recently, we started working on Lactochassis project where we aim to develop next generation synthetic LAB for biomedical application.

SUBMITTER: Mugwanda K

PROVIDER: S-EPMC9842951 | biostudies-literature | 2023 Jan

REPOSITORIES: biostudies-literature

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Recent advances in genetic tools for engineering probiotic lactic acid bacteria.

Mugwanda Kanganwiro K Hamese Saltiel S Van Zyl Winschau F WF Prinsloo Earl E Du Plessis Morne M Dicks Leon M T LMT Thimiri Govinda Raj Deepak B DB

Bioscience reports 20230101 1

Synthetic biology has grown exponentially in the last few years, with a variety of biological applications. One of the emerging applications of synthetic biology is to exploit the link between microorganisms, biologics, and human health. To exploit this link, it is critical to select effective synthetic biology tools for use in appropriate microorganisms that would address unmet needs in human health through the development of new game-changing applications and by complementing existing technolo ...[more]

PMID: 36597861

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Project description:The antibiotic resistances of 45 lactic acid bacteria strains belonging to the genera Lactobacillus, Streptococcus, Lactococcus, Pediococcus, and Leuconostoc were investigated. The objective was to determine antibiotic resistances and to verify these at the genetic level, as is currently suggested by the European "qualified presumption of safety" safety evaluation system for industrial starter strains. In addition, we sought to pinpoint possible problems in resistance determinations. Primers were used to PCR amplify genes involved in beta-lactam antibiotic, chloramphenicol, tetracycline, and erythromycin resistance. The presence of ribosomal protection protein genes and the ermB gene was also determined by using a gene probe. Generally, the incidences of erythromycin, chloramphenicol, tetracycline, or beta-lactam resistances in this study were low (<7%). In contrast, aminoglycoside (gentamicin and streptomycin) and ciprofloxacin resistances were higher than 70%, indicating that these may constitute intrinsic resistances. The genetic basis for ciprofloxacin resistance could not be verified, since no mutations typical of quinolone resistances were detected in the quinolone determining regions of the parC and gyrA genes. Some starter strains showed low-level ampicillin, penicillin, chloramphenicol, and tetracycline resistances, but no known resistance genes could be detected. Although some strains possessed the cat gene, none of these were phenotypically resistant to chloramphenicol. Using reverse transcription-PCR, these cat genes were shown to be silent under both inducing and noninducing conditions. Only Lactobacillus salivarius BFE 7441 possessed an ermB gene, which was encoded on the chromosome and which could not be transferred in filter-mating experiments. This study clearly demonstrates problems encountered with resistance testing, in that the breakpoint values are often inadequately identified, resistance genes may be present but silent, and the genetic basis and associated resistance mechanisms toward some antibiotics are still unknown.

Project description:Lactic acid bacteria (LAB) are of major concern due to their health benefits. Fermented food products comprise variable LAB demonstrating probiotic properties. Discovering and evaluating new probiotics in fermented food products poses a global economic and health importance. Therefore, the present work aimed to investigate and evaluate the probiotic potentials of LAB strains isolated from Egyptian fermented food. In this study, we isolated and functionally characterized 100 bacterial strains isolated from different Egyptian fermented food sources as probiotics. Only four LAB strains amongst the isolated LAB showed probiotic attributes and are considered to be safe for their implementation as feed or dietary supplements. Additionally, they were shown to exert antimicrobial activities against pathogenic bacteria and anticancer effects against the colon cancer cell line Caco-2. The Enterococcus massiliensis IS06 strain was exclusively reported in this study as a probiotic strain with high antimicrobial, antioxidant, and anti-colon cancer activity. Hitherto, few studies have focused on elucidating the impact of probiotic supplementation in vivo. Therefore, in the current study, the safety of the four strains was tested in vivo through the supplementation of rats with potential probiotic strains for 21 days. The results revealed that probiotic bacterial supplementation in rats did not adversely affect the general health of rats. The Lactiplantibacillus plantarum IS07 strain significantly increased the growth performance of rats. Furthermore, the four strains exhibited increased levels of antioxidants such as superoxide dismutase and glutathione in vivo. Consistently, all strains also showed high antioxidant activity of the superoxide dismutase enzyme in vitro. Overall, these findings demonstrated that these isolated potential probiotics harbor desirable characteristics and can be applied widely as feed additives for animals or as dietary supplements for humans to exert their health benefits and combat serious diseases.

Dataset Information

Recent advances in genetic tools for engineering probiotic lactic acid bacteria.

Publications

Recent advances in genetic tools for engineering probiotic lactic acid bacteria.

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