Production of plant-based fermented beverages possessing functional ingredients antioxidant, γ-aminobutyric acid and antimicrobials using a probiotic Lactiplantibacillus plantarum strain L42g as an efficient starter culture.

Plant-based probiotic beverages have gained increasing interest due to demand from health-conscious consumers. In this study, we aimed to isolate and screen lactic acid bacteria possessing functional properties for use as a starter culture of fermented almond and coix beverages. Lactiplantibacillus plantarum L42g isolated from fermented beef was selected. Both intact cells and cell free supernatant of this strain exhibited high antioxidant activity based on 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging at 38.2% and 44.9%, respectively. L. plantarum L42g grown in MRS broth supplemented with 1% (w v-1) monosodium glutamate (MSG) produced a large amount of γ-aminobutyric acid (GABA) at 496.7 µg mL-1. Moreover, strain L42g displayed remarkable antibacterial activity against several potential foodborne bacterial pathogens, including Bacillus cereus, Listeria monocytogenes, Listeria inocua, Staphylococcus aureus, Streptococcus agalactiae, Escherichia coli, Salmonella enterica subsp. enterica serovar Typhimurium, Shigella sp., Vibrio cholerae and Vibrio parahaemolyticus. Strain L42g also possessed additional probiotic properties including abilities to tolerate gastrointestinal conditions, adhere to gut mucosa, co-aggregate with pathogens, be susceptible to antibiotics, and produce protease. Probiotic strain L42g was subsequently employed in fermenting almond and coix juices containing MSG (1%) supplementation. Levels of antioxidant, GABA and antibacterial formation along with cell growth were clearly higher in fermented almond juice than in fermented coix juice. Nonetheless, both fermented almond and coix juices meet the standards required for the consumption of fermented beverages. Therefore, L. plantarum strain L42g represents a promising starter culture for producing functional plant-based probiotic beverages.

Mixed culture of Bacillus aerius B81e and Lactiplantibacillus paraplantarum L34b-2 derived from in vivo screening using hybrid catfish exhibits high probiotic effects on Pangasius bocourti.

A mixed culture of probiotics, one from the genus Bacillus and one lactic acid bacterium (LAB), was developed to be used as a feed additive for enhancing growth, innate immunity and disease resistance in Pangasius bocourti. From our earlier work, three probiotic Bacillus species, Bacillus siamensis B44v, Bacillus sp. B51f and Bacillus aerius B81e, and three probiotic LABs, Streptococcus lutetiensis L7c, Lactiplantibacillus paraplantarum (synonym. Lactobacillus paraplantarum) L34b-2 and Lactiplantibacillus plantarum (synonym. Lactobacillus plantarum) L42g, were selected for comparison. These bacteria, which express probiotic properties including bacteriocin-like activity against Aeromonas hydrophila, were subjected to in vivo screening in hybrid catfish (Clarias macrocephalus × Clarias gariepinus). A 30-day feed-trial followed by a challenge test in screening experiments resulted in the prominent B. aerius B81e and L. paraplantarum L34b-2 being selected. A mixture of these bacteria was added to a diet for P. bocourti. After 60-day feeding, the fish fed with mixed probiotics had weight gain, specific growth rate and feed conversion ratio improved significantly (p < 0.01) when compared to the control. Both humoral and cellular immunity were significantly higher in probiotic-fed fish. Following the 60-day feeding experiment, P. bocourti fed with the diet containing mixed probiotics had a higher survival rate than the control fish after injection with a virulent A. hydrophila. It can be concluded that a combination of B. aerius strain B81e and L. paraplantarum strain L34b-2 markedly improved growth performance, innate immunity and disease resistance of P. bocourti.

Bioactive homogentisic acid derivatives from fruits and flowers of Miliusa velutina.

Chromatographic separation of fruits and flowers of the Thai medicinal plant, Miliusa velutina, resulted in the isolation of five new rare homogentisic acid derivatives, miliusanal (1) and miliusanones A-D (2-5), together with fifteen known secondary metabolites (6-20). Their structures were determined through the use of extensive spectroscopic data. The absolute configurations of homogentisic acid derivatives 2-7 were identified using NOESY data and a comparison of experimental and calculated ECD spectral data. Compounds 2, 3, 6, and 7 showed antimalarial activity with IC50 values in the range of 3.3-5.2 µg/mL. Compound 6 also showed activity against Mycobacterium tuberculosis with an MIC value of 50 µg/mL. Compounds 1-3, 6 and 7 exhibited cytotoxicity againt KB, MCF-7, NCI-H187 and Vero cell lines with IC50 values in the range of 5.8-40.4 µg/mL. In addition, compounds 1, 2 and 6 showed moderate antibacterial activities against three Gram-positive bacteria (Bacillus cereus, Staphylococcus aureus, and Methicillin resistant S. aureus) with MICs in the range of 32-64 µg/mL.

Amycolatopsis silviterrae sp. nov., isolated from forest soil.

A novel actinobacterial strain, designated C12CA1T, was isolated from forest soil in the conservation area of Chulabhorn dam, Thailand, and its taxonomic position was determined by using a polyphasic approach. Strain C12CA1T contained meso-2,6 diaminopimelic acid in the cell-wall peptidoglycan, and arabinose and galactose as diagnostic sugars of the whole-cell hydrolysate. On the basis of morphological and chemotaxonomic characteristics, strain C12CA1T was classified in the genus Amycolatopsis. It contained MK-9(H4) as the predominant menaquinone, C16 : 0, iso-C15 : 0 and iso-C16 : 0 as the major cellular fatty acids, and several phospholipids consisting of diphosphotidylglycerol, phosphatidylglycerol, phosphatidylmethylethanolamine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, hydroxyphosphatidylethanolamine, phosphatidylinositol mannosides and an unidentified glucosamine-containing phospholipid. Based on 16S rRNA gene sequence and phylogenetic analyses, strain C12CA1T was closely related to Amycolatopsis vancoresmycina DSM 44592T (98.96 %) and Amycolatopsis pretoriensis JCM 12673T (98.82 %). The strain exhibited low DNA-DNA relatedness values with A. vancoresmycina DSM 44592T (6.9±0.2-11.6±1.9 %) and A. pretoriensis JCM 12673T (8.8±0.3-9.2±1.8 %). The DNA G+C content of strain C12CA1T was 69.8 mol%. Based on the results of polyphasic characterization, strain C12CA1T represents a novel species of the genus Amycolatopsis, for which the name Amycolatopsis silviterrae sp. nov. is proposed. The type strain is C12CA1T (=TBRC 1456T=NBRC 111116T).

Evaluation of probiotic Bacillus aerius B81e isolated from healthy hybrid catfish on growth, disease resistance and innate immunity of Pla-mong Pangasius bocourti.

Infectious diseases have been found to be a major cause of mortality in fish hatcheries. Probiotics have been introduced to replace antibiotics commonly used for treatment of bacterial infection in aquaculture. This study was conducted to isolate, screen, and evaluate the probiotic Bacillus spp. for potential use as a feed supplement to enhance fish growth, disease resistance and innate immunity of Pla-mong Pangasius bocourti. Bacillus aerius strain B81e was selectively isolated from the intestine of healthy catfish and chosen based on its probiotic properties both in vitro and in vivo. This bacterium produced a bacteriocin-like substance and exhibited a broad-spectrum antibacterial activity inhibiting both Gram-positive and Gram-negative bacteria especially the fish pathogens Aeromonas hydrophila and Streptococcus agalactiae. The susceptibility to all 8 antibiotics tested implies that it is unlikely to be an antibiotic-resistant bacterium. B. aerius strain B81e possessed interesting adhesion properties as shown by its high percentages of hydrophobicity, auto-aggregation, co-aggregation with fish pathogens A. hydrophila FW52 and S. agalactiae F3S and mucin binding. The strain B81e survived simulated gastrointestinal conditions, producing protease and lipase but not ß-haemolysin. The study also evaluated the effects of dietary supplementation with strain B81e on growth performance, innate immunity, and the disease resistance of P. bocourti against A. hydrophila infection. Fish with a mean body weight of 69 g were fed strain B81e at 0 (control) and 107 CFU g-1 feed (test) for 60 days. Various growth and immune parameters were examined at 30 and 60 days post-feeding. Fish were challenged with A. hydrophila 60 days post-feeding and mortalities were recorded over 14 days post-infection. Results showed that the administration of strain B81e for 60 days had significant effects (p < 0.05) on weight gain, specific growth rate and feed utilization efficiency of P. bocourti. Dietary administration of strain B81e increased the serum lysozyme and bactericidal activities of P. bocourti significantly throughout the experimental period whereas the alternative complement, phagocytic and respiratory burst activities were significantly (p < 0.05) higher in the test fish compared to the control fish after 60 days of feeding. In addition, the fish fed a strain B81e supplemented diet had a significantly higher (p < 0.05) post-challenge survival rate than the control fish. The results in this study indicate that B. aerius B81e has beneficial effects on growth performance, innate immunity and disease resistance of P. bocourti. This is the first report on the probiotic roles of B. aerius in aquaculture.

A novel probiotic Bacillus siamensis B44v isolated from Thai pickled vegetables (Phak-dong) for potential use as a feed supplement in aquaculture.

The use of probiotic bacteria to control bacterial infection in farmed fish is of clear practical interest. The aims of this study were to isolate and select a probiotic Bacillus sp. and to evaluate the effects of its supplementation on the growth and disease resistance of hybrid catfish. Bacillus siamensis strain B44v, selectively isolated from Thai pickled vegetables (Phak-dong), displayed a high potential as a probiotic in catfish culture. This bacterium produced a bacteriocin-like substance and exhibited a broad-spectrum antibacterial activity inhibiting both Gram-positive and Gram-negative bacteria, especially the fish pathogens Aeromonas hydrophila and Streptococcus agalactiae. The susceptibility to all 14 antibiotics tested implies its less possibility to be the antibiotic-resistant bacterium. Bacillus siamensis strain B44v possessed interesting adhesion properties, as shown by its high percentages of hydrophobicity (64.8%), auto-agglutination (73.8%), co-aggregation (67.2% with A. hydrophila FW52 and 63.5% with S. agalactiae F3S), and mucin binding (88.7%). The strain B44v survived simulated gastrointestinal conditions and produced protease and cellulase enzymes. Hybrid catfish (C. macrocephalus × C. gariepinus) were employed in the feed-trial experiments. Fish fed diet containing strain B44v (107 CFU/g feed) displayed not only no mortality but also growth improvement. At the end of the feed trial, fish were challenged by an intraperitoneal injection of Aeromonas hydrophila FW52. The Bacillus siamensis strain B44v fed fish survived (75.0%; p < 0.05) better than the controls (36.7%; p < 0.05) after a two week challenge. These collective results present for the first time the potential of Bacillus siamensis strain B44v for use as a bacterial probiotic in aquaculture.

Isolation of thermophilic L-lactic acid producing bacteria showing homo-fermentative manner under high aeration condition.

By applying non-sterile open fermentation of food waste, various thermotolerant l-lactic acid-producing bacteria were isolated and identified. The predominant bacterial isolates showing higher accumulation of l-lactic acid belong to 3 groups of Bacillus coagulans, according to their 16S rRNA gene sequence similarities. B. coagulans strains M21 and M36 produced high amounts of l-lactic acid of high optical purity and lactic acid selectivity in model kitchen refuse medium and glucose-yeast extract-peptone medium. Other thermotolerant isolates resembling to Bacillus humi, B. ruris, B. subtilis, B. niacini and B. soli were also identified. These bacteria produced low amounts of l-lactic acid of more than 99% optical purity. All isolated strains showed the highest growth rate at temperatures around 55-60°C. They showed unique responses to various oxygen supply conditions. The majority of isolates produced l-lactic acid at a low overall oxygen transfer coefficient (KLa); however, acetic acid was produced instead of l-lactic acid at a high KLa. B. coagulans M21 was the only strain that produced high, consistent, and reproducible amounts of optically pure l-lactic acid (>99% optical purity) under high and low KLa conditions in a homo-fermentative manner.

Differently environment stable bio-silver nanoparticles: study on their optical enhancing and antibacterial properties.

Generally, limited research is extended in studying stability and applicational properties of silver nanoparticles (Ag NPs) synthesized by adopting 'green chemistry' protocol. In this work, we report on the synthesis of stable Ag NPs using plant-derived materials such as leaf extract of Neem (Azadirachta indica) and biopolymer pectin from apple peel. In addition, the applicational properties of Ag NPs such as surface-enhanced Raman scattering (SERS) and antibacterial efficiencies were also investigated. As-synthesized nanoparticles (NPs) were characterized using various instrumentation techniques. Both the plant materials (leaf extract and biopolymer) favored the synthesis of well-defined NPs capped with biomaterials. The NPs were spherical in shape with an average particle size between 14-27 nm. These bio-NPs exhibited colloidal stability in most of the suspended solutions such as water, electrolyte solutions (NaCl; NaNO3), biological solution (bovine serum albumin), and in different pH solutions (pH 7; 9) for a reasonable time period of 120 hrs. Both the bio-NPs were observed to be SERS active through displaying intrinsic SERS signals of the Raman probe molecule (Nile blue A). The NPs were effective against the Escherichia coli bacterium when tested in nutrient broth and agar medium. Scanning and high-resolution transmission electron microscopy (SEM and HRTEM) images confirmed cellular membrane damage of nanoparticle treated E. coli cells. These environmental friendly template Ag NPs can be used as an antimicrobial agent and also for SERS based analytical applications.