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There is an increasing demand for dairy products, but the presence of food-spoilage bacteria seriously affects the development of the dairy industry. Bacteriocins are considered to be a potential antibacterial or antibiofilm agent that can be applied as a preservative. In this study, bacteriocin BM173 was successfully expressed in the Escherichia coli expression system and purified by a 2-step method. Furthermore, it exhibited a broad-spectrum antibacterial activity, high thermal stability (121°C, 20 min), and broad pH stability (pH 3-11). Moreover, the minimum inhibitory concentration values of BM173 against E. coli ATCC 25922 and Staphylococcus aureus ATCC 25923 were 14.8 μg/mL and 29.6 μg/mL, respectively. Growth and time-kill curves showed that BM173 exhibited antibacterial and bactericidal activity. The results of scanning electron microscopy and transmission electron microscopy demonstrated that BM173 increased membrane permeability, facilitated pore formation, and even promoted cell lysis. The disruption of cell membrane integrity was further verified by propidium iodide uptake and lactic dehydrogenase release. In addition, BM173 exhibited high efficiency in inhibiting biofilm formation. Therefore, BM173 has promising potential as a preservative used in the dairy industry., (Copyright © 2021 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Some lactobacilli strains had beneficial effects on human beings due to their antioxidant activities. In this study lactobacilli strains stored in our laboratory were screened for potential antioxidant activities by investigating their 1,1-diphenyl-2-picrylhydrazyl free radical scavenging activity, oxygen radical absorbance capacity, resistance to H 2 O 2 , and hydroxyl free radical scavenging activity; then the antioxidant activities of the screened strains were evaluated by cellular antioxidant assay and protection for HT-29 cells against H 2 O 2 injury assay. The results showed that Lactobacillus plantarum Y44 could scavenge oxygen free radicals, inhibit the production of intracellular reactive oxygen species without creating obvious cytotoxic effects, and protect HT-29 cells against H 2 O 2 injury evidenced by the significant decrease of the Bcl-2-associated X protein (Bax)/B-cell lymphoma 2 (Bcl-2) ratio and heat shock protein 70 expression, increase of superoxide dismutase and glutathione peroxidase activities, and decrease of malondialdehyde level of HT-29 cells damaged by H 2 O 2 . It was speculated that L. plantarum Y44 protect HT-29 cells against oxygen radical injury through scavenging reactive oxygen species and activating intracellular antioxidant enzymes. A significant correlation was observed among the results of the hydroxyl radical scavenging assay, protection assay for HT-29 cells against H 2 O 2 injury, and the cellular antioxidant assay. The findings indicated that L. plantarum Y44 could be a probiotic candidate with antioxidant properties and combining several chemical antioxidant methods and antioxidant cellular models could be an effective procedure to screen lactobacilli strains with antioxidant activity., (Copyright © 2018 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

This study was conducted to examine if adding microbial inoculants or propionic acid to alfalfa silages contaminated with Escherichia coli O157:H7 would inhibit the growth of the pathogen during or after ensiling. Alfalfa forage was harvested at the early bloom stage, wilted to a dry matter concentration of 54%, chopped to 19-mm lengths, and ensiled after treatment with one of the following: (1) distilled water (control); (2) 1×10(5) cfu/g of E. coli O157:H7 (EC); (3) EC and 1×10(6) cfu/g of Lactobacillus plantarum (EC+LP); (4) EC and 1×10(6) cfu/g of Lactobacillus buchneri (EC+LB); and (5) EC and 2.2g/kg of propionic acid (EC+PA). Each treatment was ensiled in quadruplicate in laboratory silos for 0, 3, 7, 16, and 100d and analyzed for EC counts, pH, and organic acids. In addition, samples from d 100 were analyzed for chemical composition, ammonia-N, counts of yeasts and molds, and aerobic stability. Escherichia coli O157:H7 was detected in all silages until d 7, but by d 16 it was not detected in those treated with EC+LB and EC+LP, though it was still detected in EC and EC+PA silages. However, by d 100, the pathogen was not detected in any silage. The rate of pH decrease to 5.0 was fastest for the EC+LP silage (7d), followed by the EC+LB silage (16d). Nevertheless, all silages had attained a pH of or less than 5.0 by d 100. The rapid decrease in pH in EC+LP and EC+LB silages was observed due to higher lactate and acetate concentrations, respectively, relative to the other silages during the early fermentation phase (d 3-16). Propionic acid was only detected in the EC+PA silage. Yeast counts were lowest in EC+LB and EC+PA silages. Subsamples of all d-100 silages were reinoculated with 1×10(5) cfu/g of EC immediately after silo opening. When the pathogen was subsequently enumerated after 168h of aerobic exposure, it was not detected in silages treated with EC+PA, EC+LB, or EC+LP, which all had pH values less than 5.0. Whereas the EC silage had a pH value of 5.4 and 2.3 log cfu/g of the pathogen. Certain bacterial inoculants can hasten the inhibition of E. coli O157:H7 during ensiling, such as propionic acid, and they can also prevent its growth on silage contaminated with the pathogen after ensiling., (Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Lactobacillus spp. from Mexican Cocido cheese were shown to produce bacteriocin-like substances (BLS) active against Staphylococcus aureus,Listeria innocua,Escherichia coli, andSalmonella typhimurium by using the disk diffusion method. Crude extracts of Lactobacillus fermentum showed strong inhibitory activity against Staph. aureus, L. innocua, E. coli, and Salmonella cholerae. Complete inactivation of antimicrobial activity was observed after treatment of crude extracts with proteinase K, pronase, papain, trypsin, and lysozyme, confirming their proteinaceous nature. However, antimicrobial activity was partly lost for some of the crude extracts when treated with α-amylase, indicating that carbohydrate moieties were involved. The antimicrobial activity of the crude extracts was stable at 65°C for 30min over a wide pH range (2-8), and addition of potassium chloride, sodium citrate, ethanol, and butanol did not affect antibacterial activity. However, antimicrobial activity was lost after heating at 121°C for 15min, addition of methanol or Tween 80. Fourteen out of 18 Lactobacillus spp. showed antimicrobial activity against different test microorganisms, and 12 presented bacteriocin-like substances. Generation time and growth rate parameters indicated that the antimicrobial activity of crude extracts from 3 different strains was effective against the 4 indicator microorganisms. One of the crude extracts showed inhibition not only against gram-positive but also against gram-negative bacteria. Bacteriocin-like substances produced by this specific Lactobacillus strain showed potential for application as a food biopreservative., (Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Very limited information exists on the exclusion of pathogens by probiotics in the gut of the host challenged with pathogens. In this study, we tested probiotic characteristics in vitro and anticolonization ability of Lactobacillus paracasei FJ861111.1 in mice infected with selected pathogenic microorganisms. The in vitro results indicated that L. paracasei FJ861111.1 had a high survival in acidic conditions at pH 2.5 and bile salt concentration at 0.3%, and strong inhibition ability against common pathogens including Shigella dysenteriae, Staphylococcus aureus, Cronobacter sakazakii, Escherichia coli, and Candida albicans. The cell adhesion assays showed that L. paracasei FJ861111.1 exhibited strong adherence to HT-29 cells and excluded the adhesion of selected food-borne pathogens to HT-29 cells. The in vivo results showed that fermented milk with L. paracasei and viili (a Nordic yogurt product) significantly improved the population of total bacteria and of Lactobacillus in the feces of mice, and significantly inhibited the colonization of C. albicans to the intestines of mice post-C. albicans infection. Thus, it appears that this strain could be used as a probiotic organism for manufacturing functional fermented milk., (Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Exogenous protease addition may be an option to increase proteolysis of zein proteins and thus starch digestibility in rehydrated and high-moisture corn (HMC) ensiled for short periods. In addition, microbial inoculation may accelerate fermentation and increase acid production and thus increase solubilization of zein proteins. Four experiments were performed to evaluate the effect on fermentation profile, N fractions, and ruminal in vitro starch digestibility (ivSD) of the following: (1) rehydration and ensiling of dry ground corn; (2) exogenous protease addition to rehydrated un-ensiled and ensiled corn; (3) exogenous protease addition or inoculation in rehydrated ensiled corn; and (4) exogenous protease addition or inoculation in HMC. Experiments 1, 2, and 3 were performed with 7 treatments: dry ground corn (DGC); DGC rehydrated to a targeted dry matter content of 70% (REH); REH treated with exogenous protease (REH+); REH ensiled for 30 d (ENS); ENS treated with exogenous protease (ENS+); ENS treated with a microbial inoculant containing Lactobacillus plantarum, Lactobacillus casei, Enterococcus faecium, and Pediococcus sp. (ENSI); and ENS treated with exogenous protease and microbial inoculant (ENSI+). Experiment 1 compared DGC, REH, and ENS with ivSD being greater for ENS (64.9%) than DGC and REH (51.7% on average). Experiment 2 compared REH and ENS without or with exogenous protease addition (REH+ and ENS+, respectively). Ensiling and exogenous protease addition increased ivSD, but exogenous protease addition was more effective in ENS than REH (6.4 vs. 2.6 percentage unit increase). Experiment 3 compared the effects of exogenous protease addition and inoculation in ENS corn (ENS, ENS+, ENSI, and ENSI+). The addition of protease, but not inoculant, increased ivSD. Inoculation reduced pH and acetate, propionate, and ethanol concentrations, and increased lactate and total acid concentrations. In experiment 4, 8 treatments were a combination of HMC noninoculated or inoculated with 1 of 3 microbial inoculants and with or without exogenous protease addition. The inoculant treatments contained (1) Lactobacillus buchneri 40788 and Pediococcus pentosaceus, (2) L. buchneri 40788, and (3) a mixture of P. pentosaceus and Propionibacterium freudenreichii. Protease, but not inoculation, increased ivSD by 7.5 percentage units (44.4 vs. 51.9%). Protease addition increased ivSD in rehydrated corn and HMC. Microbial inoculation improved fermentation profiles but did not affect ivSD., (Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Some strains of Lactobacillus genus have antiproliferative activities against cancer cells. However, until now, the exact effector molecules of Lactobacillus strains with anticancer activity have not been identified. The aim of the present study was to explore which fraction of the Lactobacillus cells exerts the highest antiproliferative effect. For this purpose, the heat-killed bacterial cells, bacterial cell wall extract, and genomic DNA of 8 Lactobacillus strains were prepared to assess their antiproliferative activities against human myeloid leukemia cell lines K562. The heat-killed bacterial cells of the 8 lactobacilli strains exerted antiproliferative effect on K562 cells, and the inhibition rates exerted by the heat-killed bacterial cells of the strains G15AL, M5AL, SB31AL, SB5AL, and T3AL were significantly higher than those exerted by the cell walls and genomic DNA of the strains. The bacterial DNA of G15AL exerted higher antiproliferative effect on K562 cells. The exact effector molecules and the effect mechanism of the strains should be further explored for the application of these strains as probiotic strains or bioactive probiotic molecules., (Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

The objective of this work was to evaluate the effects of biological and chemical silage additives on the production of volatile organic compounds (VOC; methanol, ethanol, 1-propanol, methyl acetate, and ethyl acetate) within corn silage. Recent work has shown that silage VOC can contribute to poor air quality and reduce feed intake. Silage additives may reduce VOC production in silage by inhibiting the activity of bacteria or yeasts that produce them. We produced corn silage in 18.9-L bucket silos using the following treatments: (1) control (distilled water); (2) Lactobacillus buchneri 40788, with 400,000 cfu/g of wet forage; (3) Lactobacillus plantarum MTD1, with 100,000 cfu/g; (4) a commercial buffered propionic acid-based preservative (68% propionic acid, containing ammonium and sodium propionate and acetic, benzoic, and sorbic acids) at a concentration of 1 g/kg of wet forage (0.1%); (5) a low dose of potassium sorbate at a concentration of 91 mg/kg of wet forage (0.0091%); (6) a high dose of potassium sorbate at a concentration of 1g/kg of wet forage (0.1%); and (7) a mixture of L. plantarum MTD1 (100,000 cfu/g) and a low dose of potassium sorbate (91 mg/kg). Volatile organic compound concentrations within silage were measured after ensiling and sample storage using a headspace gas chromatography method. The high dose of potassium sorbate was the only treatment that inhibited the production of multiple VOC. Compared with the control response, it reduced ethanol by 58%, ethyl acetate by 46%, and methyl acetate by 24%, but did not clearly affect production of methanol or 1-propanol. The effect of this additive on ethanol production was consistent with results from a small number of earlier studies. A low dose of this additive does not appear to be effective. Although it did reduce methanol production by 24%, it increased ethanol production by more than 2-fold and did not reduce the ethyl acetate concentration. All other treatments increased ethanol production at least 2-fold relative to the control, and L. buchneri addition also increased the 1-propanol concentration to approximately 1% of dry matter. No effects of any treatments on fiber fractions or protein were observed. However, L. buchneri addition resulted in slightly more ammonia compared with the control. If these results hold under different conditions, a high dose of potassium sorbate will be an effective treatment for reducing VOC production in and emission from silage. Regulations aimed at reducing VOC emission could be ineffective or even increase emission if they promote silage additives without recognition of different types of additives., (Copyright © 2014 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.)

Lactobacillus gasseri LA39 and LA158 isolated from human-infant feces produce bacteriocins named gassericins A and T, respectively. Both gassericins have high heat stability (121 degrees C, 10 min), good pH tolerance (pH 2-11), and strong bactericidality against many gram-positive bacteria, especially lactic acid bacteria, and thus are expected to be effective food preservatives. A microwell plate assay against 12 strains of custard cream spoilage bacteria showed that the gassericins had broader antibacterial spectra than nisin A. Although the gassericins allowed gram-negative isolates to grow, they successfully inhibited the growth of all tested bacterial strains in microwells with the addition of glycine. Glycine was bacteriostatic against many strains except lactic acid bacteria. For practical use, gassericin A was efficiently produced by cultivation in a food-grade medium improved using cheese whey, nourishing proteose peptone, and surfactant yolk lecithin. The practical preservative effect of gassericin A and glycine was verified from the viability of 4 isolated strains, Bacillus cereus, Lactococcus lactis ssp. lactis, Achromobacter denitrificans, and Pseudomonas fluorescens, in custard creams. Custard cream containing 123 arbitrary units of gassericin A per milliliter entirely growth-inhibited the 2 gram-positive strains. In custard cream containing an insufficient amount of gassericin A (49 arbitrary units/mL), the gram-positive strains gradually grew but were completely inhibited by the addition of 0.5% (wt/wt) glycine. The 2 gram-negative strains did not multiply even in the additive-free custard cream, probably because of the unsuitable growth environment. This is the first report showing the combined effect of bacteriocin and glycine and their application for food preservation, which may be helpful for future use in the food industry.

Allergic diseases are reported to be caused by a skew in the balance between T helper type 1 and 2 cells. Because some lactic acid bacteria have been shown to stimulate IL-12 (p70) production, which in turn shifts the balance between the T helper type 1 and 2 cell response from the latter to the former, they have the potential to either prevent or ameliorate disease conditions or both. They have therefore been extensively studied in the recent past for their probiotic activities. Nevertheless, much less information is available concerning the microbial factors that determine the strain-dependent ability to affect the production of cytokines. The objectives of our study were first to select potentially probiotic lactobacilli that strongly stimulate cytokine production in vitro, and then to determine whether the selected Lactobacillus strains could suppress antigen-specific IgE production in vivo by using allergic model animals. Finally, our investigation was extended to analyze which bacterial components were responsible for the observed biological activity. Twenty strains of heat-killed lactobacilli isolated from humans were screened for their stimulatory activity for the production of IL-12 (p70) by murine splenocytes in vitro. The results showed that some strains of Lactobacillus plantarum and Lactobacillus gasseri had a higher stimulatory activity for IL-12 (p70) production than the other lactobacilli tested; however, this effect was strain dependent rather than species dependent. Oral administration of the heat-killed strains that showed higher stimulatory activity for IL-12 (p70) production tended to reduce the serum antigen-specific IgE levels in ovalbumin-sensitized BALB/c mice compared with the controls. Among the lactobacilli tested, L. gasseri OLL2809 showed the highest activity in reducing the level of antigen-specific IgE. Furthermore, the stimulatory activity for IL-12 (p70) production was found to be reduced after treating the lactobacilli with N-acetyl-muramidase and to be positively correlated with the amount of peptidoglycan in the cells. The present data suggest that L. gasseri OLL2809 is a good candidate for potential probiotics in terms of either the prevention or amelioration of allergic diseases or both. In addition, the strain-dependent stimulatory activity for IL-12 (p70) production was found to be due, at least in part, to the amount of peptidoglycan present in the cells.

Eleven strains of lactobacilli were studied for their acid and bile tolerance. Possible mechanisms of cholesterol removal by strains of lactobacilli were examined. Cholesterol assimilation as determined by the difference in cholesterol content in the medium before and after the incubation period showed that all lactobacilli strains were able to assimilate cholesterol at varying levels ranging from 12.03 to 32.25 microg/mL. Cholesterol removal was associated with growth of cultures. Binding of cholesterol to lactobacilli cells was determined using growing, heat-killed, and resting cells in phosphate buffer. Cholesterol removed by dead and resting cells ranged from 0.79 to 3.82 mg/g of dry weight compared with growing cells, which ranged from 4.53 to 16.03 mg/g of dry weight. Fatty acid methyl esters, as quantified using gas chromatography, showed changes in lipid profiles in cells grown in the presence of cholesterol compared with those grown without cholesterol. Fatty acid profiles, especially of hexadecanoic, octadecanoic, total saturated, and unsaturated acids suggested that cholesterol from the medium was incorporated into the cellular membrane. These findings suggest that strains of lactobacilli could remove cholesterol via various mechanisms and may be promising candidates for use as a dietary adjunct to lower serum cholesterol in vivo.