Sangnam Oh

Short communication: Development of a direct in vivo screening model to identify potential probiotic bacteria using Caenorhabditis elegans

Caenorhabditis elegans is an accepted model host to study host-bacteria interactions in the gut, in addition to being a simple model with which to study conserved aspects of biological signaling pathways in intestinal environments, because these nematode worms have similar intestinal cells to those of humans. Here, we used C. elegans to develop a new in vivo screening system for potential probiotic lactic acid bacteria (LAB). Initially, critical colonization ability of LAB strains isolated from Korean infant feces was screened in the worm intestinal tract over a period of 5 d. Furthermore, we investigated host health-promoting activities, including longevity-extending effects and immune-enhancing activities against foodborne pathogen infection. We identified 4 LAB strains that were highly persistent in the nematode gut and that significantly prolonged the longevity of C. elegans and improved the survival of C. elegans in response to infection by Staphylococcus aureus. The 4 LAB strains we identified showed resistance to acid and bile conditions, assimilated cholesterol, and were able to attach to a mucus layer. The 4 LAB isolates were identified as Lactobacillus plantarum using 16S rRNA sequencing analysis. Taken together, we developed a direct in vivo screening system using C. elegans to study host health-promoting LAB. Our system is simple, rapid, cost-effective, and reliable, and we anticipate that this system will result in the discovery of many more potential probiotic bacteria for dairy foods.

Bacillus licheniformis Isolated from Traditional Korean Food Resources Enhances the Longevity of Caenorhabditis elegans through Serotonin Signaling

In this study, we investigated potentially probiotic Bacillus licheniformis strains isolated from traditional Korean food sources for ability to enhance longevity using the nematode Caenorhabditis elegans as a simple in vivo animal model. We first investigated whether B. licheniformis strains were capable of modulating the lifespan of C. elegans. Among the tested strains, preconditioning with four B. licheniformis strains significantly enhanced the longevity of C. elegans. Unexpectedly, plate counting and transmission electron microscopy (TEM) results indicated that B. licheniformis strains were not more highly attached to the C. elegans intestine compared with Escherichia coli OP50 or Lactobacillus rhamnosus GG controls. In addition, qRT-PCR and an aging assay with mutant worms showed that the conditioning of B. licheniformis strain 141 directly influenced genes associated with serotonin signaling in nematodes, including tph-1 (tryptophan hydroxylase), bas-1 (serotonin- and dopamine-synthetic aromatic amino acid decarboxylase), mod-1 (serotonin-gated chloride channel), ser-1, and ser-7 (serotonin receptors) during C. elegans aging. Our findings suggest that B. licheniformis strain 141, which is isolated from traditional Korean foods, is a probiotic generally recognized as safe (GRAS) strain that enhances the lifespan of C. elegans via host serotonin signaling.

Improved functionality of fermented milk is mediated by the synbiotic interaction between Cudrania tricuspidata leaf extract and Lactobacillus gasseri strains

This study was designed to investigate the cooperative effect of selected Lactobacillus gasseri strains and Cudrania tricuspidata (CT) leaf extract in enhancing the health-promoting activities of fermented milk. Addition of CT increased total bacterial counts and proteolysis during fermentation of milk with L. gasseri strains. Antioxidant capacities were determined by measuring the ABTS, DPPH, and peroxyl radical scavenging activities and ferric reducing power. The antioxidant capacity of CT-supplemented milk was greater than that of milk without supplementation; moreover, the antioxidant activity of CT-supplemented milk was synergistically improved by fermentation with L. gasseri strains. In particular, CT-supplemented milk fermented by L. gasseri 505 showed the highest antioxidant activity. The phenolic compounds in CT, such as neo-chlorogenic, chlorogenic, and caffeic acid, were metabolized during fermentation with L. gasseri strains, and 3,4-dihydroxy-hydrocinnamic acid was produced as a fermentation metabolite. Moreover, the liberation of bioactive peptides of fermented milk was increased by the proteolytic activity of L. gasseri strains. In particular, six peptides, which were mainly derived from β-casein, were newly identified in this study. These findings suggest that L. gasseri strains metabolize the phenolic acids in the CT and the bioactive peptides released through this interaction improve the antioxidant activity of the fermented milk.

Short communication: In vivo screening platform for bacteriocins using Caenorhabditis elegans to control mastitis-causing pathogens

This study aimed to develop an in vivo screening platform using Caenorhabditis elegans to identify a novel bacteriocin for controlling the mastitis-causing pathogen Staphylococcus aureus strain RF122 in dairy cows. Using Bacillus spp. isolated from traditional Korean foods, we developed a direct in vivo screening platform that uses 96-well plates and fluorescence image analysis. We identified a novel bacteriocin produced by Bacillus licheniformis strain 146 (lichenicin 146) with a high in vivo antimicrobial activity using our liquid C. elegans-Staph. aureus assay. We also determined the characteristics of lichenicin 146 using liquid chromatography-mass spectrometry and confirmed that it shared homologous sequences with bacteriocin family proteins. In addition, RNA-sequencing analysis revealed genes encoding cell surface or membrane proteins (SAB0993c, SAB0150, SAB0994c, and SAB2375c) that are involved in the bactericidal activity of lichenicin 146 against Staph. aureus strain RF122 infection as well as those encoding transcriptional regulators (SAB0844c and SAB0133). Thus, our direct in vivo screening platform facilitates simple, convenient, cost-effective, and reliable screening of potential antimicrobial compounds with applications in the dairy field.

Microbiological characterization and functionality of set-type yogurt fermented with potential prebiotic substrates Cudrania tricuspidata and Morus alba L. leaf extracts.

The objective of this study was to investigate the effect of 2 plant leaf extracts on fermentation mechanisms and health-promoting activities and their potential as a nutraceutical prebiotics ingredient for application in dairy products. The individual active phenolic compounds in the plant extract-supplemented milk and yogurts were also identified. Compared with control fermentation, the plant extracts significantly increased the growth and acidification rate of Streptococcus thermophilus and Lactobacillus delbrueckii ssp. bulgaricus. In particular, plant extract components, including monosaccharides, formic acid, and hydroxycinnamic acid, such as neo-chlorogenic, chlorogenic, and caffeic acid, together play a stimulatory role and cause this beneficial effect on the growth of yogurt culture bacteria through fermentation. In addition, supplementation with the plant extracts enhanced antioxidant activities with increased total phenolic contents, especially the highest antioxidant activity was observed in yogurt supplemented with Cudrania tricuspidata leaf extract.

Enhanced protection of pathogenic Escherichia coli ingested by a soil nematode Caenorhabditis elegans against sanitizer treatments

We employed Caenorhabditis elegans as a model to study the effectiveness of sanitizers in killing pathogenic Escherichia coli strains ingested by free-living nematodes. Adult worms that had fed on six pathogenic E. coli strains (highly persistent in the nematode intestine) were treated with three chemical solutions. In planktonic cells, none of the H2O2 and acetic acid treatments influenced the survival of the pathogenic E. coli strains, whereas sodium hypochlorite critically decreased the viability of the strains. Importantly, the survival of the E. coli strains was dramatically increased by persistence in the C. elegans gut under 0.1% sodium hypochlorite, and several strains could survive at a concentration of 0.5%. In addition, all pathogenic E. coli strains in the C. elegans gut survived on the lettuce for 5 days even though they were washed with 0.1% sodium hypochlorite. Taken together, our results indicate that pathogenic E. coli ingested by C. elegans may be protected against washing treatment with commercial sanitizers on raw food materials. Graphical Abstract Pathogenic E. coli are highly resistant to digestion by C. elegans serve as carriers or vectors of pathogens from soil resources and they can colonize in the nematode intestine.