Measuring Conjugated Linoleic Acid (CLA) Production by Bifidobacteria.

The biological significance of conjugated fatty acids (CFAs) has been linked to positive health effects based on biomedical, in vitro, and clinical studies. Of note, conjugated linoleic acids (CLAs) are the most widely characterized fatty acids as geometric isomers cis-9,trans-11 and trans-10,cis-12 CLA occur naturally in ruminant fats, dairy products, and hydrogenated oils. Concerning CLAs, it is known that bacterial biohydrogenation, a process whereby ruminal bacteria or starter cultures of lactic acid bacteria have the ability to synthesize CLA by altering the chemical structure of essential fatty acids via enzymatic mechanisms, produces a multitude of isomers with desirable properties. Bifidobacterium species are classed as food grade microorganisms and some of these strains harness molecular determinants that are responsible for the bioconversion of free fatty acids to CLAs. However, molecular mechanisms have yet to be fully elucidated. Reports pertaining to CLAs have been attributed to suppressing tumor growth, delaying the onset of diabetes mellitus and reducing body fat in obese individuals. Given the increased attention for their bioactive properties, we describe in this chapter the qualitative and quantitative methods used to identify and quantify CLA isomers produced by bifidobacterial strains in supplemented broth media. These approaches enable rapid detection of potential CLA producing strains and accurate measurement of fatty acids in biological matrices.

The impact of probiotic supplementation on metabolic health in healthy women of reproductive age: a systematic review.

Products containing probiotics are targeted at healthy or at-risk individuals as a preventative measure to minimise disease risk. Most studies assessing the efficacy of probiotics in humans include a mixture of healthy and unhealthy populations, while studies that focus solely on female populations are largely limited to pregnancy or those with health conditions. Pre-conception is a significant time-point during the life-course, and improving female health status during this period may positively influence future offspring. The objective of this review is to assess the effect of probiotics administered in oral capsule formulation, on metabolic and immune markers in healthy, non-pregnant women of reproductive age. This review followed the PRISMA guidelines. Pubmed, EMBASE, CINAHL, and Web of Science were searched for relevant studies. English language articles relating to randomised-controlled trials were included. The search returned 3250 publications after duplicates were removed. Title (2516), abstract (642), and full text (87) screening excluded 3993 studies from consideration. Five papers were identified with outcomes of interest, and analysis of these showed no conclusive evidence that probiotic capsule supplementation elicited positive effects in this healthy population. This study highlights the need for further research to investigate the role that probiotics play during the pre-conception period, on female metabolic and immune health.

Bifidobacterium breve UCC2003 Exopolysaccharide Modulates the Early Life Microbiota by Acting as a Potential Dietary Substrate.

BACKGROUND: Bifidobacterium represents an important early life microbiota member. Specific bifidobacterial components, exopolysaccharides (EPS), positively modulate host responses, with purified EPS also suggested to impact microbe-microbe interactions by acting as a nutrient substrate. Thus, we determined the longitudinal effects of bifidobacterial EPS on microbial communities and metabolite profiles using an infant model colon system. METHODS: Differential gene expression and growth characteristics were determined for each strain; Bifidobacterium breve UCC2003 and corresponding isogenic EPS-deletion mutant (B. breve UCC2003del). Model colon vessels were inoculated with B. breve and microbiome dynamics monitored using 16S rRNA sequencing and metabolomics (NMR). RESULTS: Transcriptomics of EPS mutant vs. B. breve UCC2003 highlighted discrete differential gene expression (e.g., eps biosynthetic cluster), though overall growth dynamics between strains were unaffected. The EPS-positive vessel had significant shifts in microbiome and metabolite profiles until study end (405 h); with increases of Tyzzerella and Faecalibacterium, and short-chain fatty acids, with further correlations between taxa and metabolites which were not observed within the EPS-negative vessel. CONCLUSIONS: These data indicate that B. breve UCC2003 EPS is potentially metabolized by infant microbiota members, leading to differential microbial metabolism and altered metabolite by-products. Overall, these findings may allow development of EPS-specific strategies to promote infant health.

Compositional assessment of bacterial communities in probiotic supplements by means of metagenomic techniques.

Health promoting or probiotic bacteria are commonly incorporated into a variety of functional foods and drug formulations, due to their purported ability to confer benefit to host health. Despite the extensive commercial exploitation of probiotic formulations there are still major knowledge gaps regarding the precise molecular mechanism of action and corresponding genetic/genomic properties of probiotic bacteria. In the current study, we describe a metagenomic approach which allows determination of the composition of probiotic supplements through next-generation sequencing analyses based on rRNA-associated sequences to assess bacterial composition of the product combined with a shotgun metagenomics approach directed to decode the genome sequences of the probiotic strains for each product assayed. The here developed approach has been tested for 10 probiotic supplements, revealing inconsistencies between the identified probiotic strains and the declared strains as indicated by the producers. Furthermore, the decoded bacterial genome sequence of Bifidobacterium animalis subsp. lactis BB-12 from a 1995 frozen dried stock revealed genetic evidence for genome evolution and stability of this microorganism when compared with the re-constructed genome of the identical strain from a probiotic supplement of 2017.

In Vitro Characteristics of Phages to Guide 'Real Life' Phage Therapy Suitability.

The increasing problem of antibiotic-resistant pathogens has put enormous pressure on healthcare providers to reduce the application of antibiotics and to identify alternative therapies. Phages represent such an alternative with significant application potential, either on their own or in combination with antibiotics to enhance the effectiveness of traditional therapies. However, while phage therapy may offer exciting therapeutic opportunities, its evaluation for safe and appropriate use in humans needs to be guided initially by reliable and appropriate assessment techniques at the laboratory level. Here, we review the process of phage isolation and the application of individual pathogens or reference collections for the development of specific or "off-the-shelf" preparations. Furthermore, we evaluate current characterization approaches to assess the in vitro therapeutic potential of a phage including its spectrum of activity, genome characteristics, storage and administration requirements and effectiveness against biofilms. Lytic characteristics and the ability to overcome anti-phage systems are also covered. These attributes direct phage selection for their ultimate application as antimicrobial agents. We also discuss current pitfalls in this research area and propose that priority should be given to unify current phage characterization approaches.

Metabolism of a plant derived galactose-containing polysaccharide by Bifidobacterium breve UCC2003.

In this study, we describe the functional characterization of the Bifidobacterium breve UCC2003 gal locus, which is dedicated to the utilization of galactan, a plant-derived polysaccharide. Using a combination of molecular approaches we conclude that the galA gene of B. breve UCC2003 encodes a ß-1,4-endogalactanase producing galacto-oligosaccharides, which are specifically internalized by an ABC transport system, encoded by galBCDE, and which are then hydrolysed to galactose moieties by a dedicated intracellular ß-galactosidase, specified by galG. The generated galactose molecules are presumed to be fed into the fructose-6-phosphate phosphoketolase pathway via the Leloir pathway, thereby allowing B. breve UCC2003 to use galactan as its sole carbon and energy source. In addition to these findings we demonstrate that GalR is a LacI-type DNA-binding protein, which not only appears to control transcription of the galCDEGR operon, but also that of the galA gene.

Bacterial vitamin B2, B11 and B12 overproduction: An overview.

Consumers are becoming increasingly health conscious and therefore more discerning in their food choices. The production of fermented food products with elevated levels of B-vitamins increase both their commercial and nutritional value, and eliminate the need for subsequent fortification with these essential vitamins. Such novel products could reduce the incidence of inadequate vitamin intake which is common in many parts of the world, not only in developing countries, but also in many industrialised countries. Moreover, the concept of in situ fortification by bacterial fermentation opens the way for development of food products targeted at specific groups in society such as the elderly and adolescents. This review looks at how vitamin overproduction strategies have been developed, some of which have successfully been tested in animal models. Such innovative strategies could be relatively easily adapted by the food industry to develop novel vitamin-enhanced functional foods with enhanced consumer appeal.

Riboflavin production in Lactococcus lactis: potential for in situ production of vitamin-enriched foods.

This study describes the genetic analysis of the riboflavin (vitamin B(2)) biosynthetic (rib) operon in the lactic acid bacterium Lactococcus lactis subsp. cremoris strain NZ9000. Functional analysis of the genes of the L. lactis rib operon was performed by using complementation studies, as well as by deletion analysis. In addition, gene-specific genetic engineering was used to examine which genes of the rib operon need to be overexpressed in order to effect riboflavin overproduction. Transcriptional regulation of the L. lactis riboflavin biosynthetic process was investigated by using Northern hybridization and primer extension, as well as the analysis of roseoflavin-induced riboflavin-overproducing L. lactis isolates. The latter analysis revealed the presence of both nucleotide replacements and deletions in the regulatory region of the rib operon. The results presented here are an important step toward the development of fermented foods containing increased levels of riboflavin, produced in situ, thus negating the need for vitamin fortification.