Pet and human food probiotics for health and safety

Abstract

Provided herein are probiotic compositions and methods for treating animals or humans, or human or animal feed, or a surface in contact therewith comprising: feeding or providing a feed, supplement, additive, or food to animals or humans with a composition comprising one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3AL3 A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 provided in an amount sufficient to be at least one of: bactericidal against bacteria selected from at least one of: spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation.

Description

PET AND HUMAN FOOD PROBIOTICS FOR HEALTH AND SAFETYCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims priority to U.S. Provisional Application Serial No. 63 / 682,097, filed August 12, 2024, the entire contents of which are incorporated herein by reference.TECHNICAL FIELD

[0002] The present invention relates in general to the field of animal and human probiotics, and more particularly, to the use of novel probiotic strains as pet food probiotics and feed to improve food safety and animal health or as supplements for human food, that are bactericidal.STATEMENT OF FEDERALLY FUNDED RESEARCH

[0003] Not applicable.INCORPORATION-BY-REFERENCE OF MATERIALS FILED ON COMPACT DISC

[0004] Not Applicable.BACKGROUND

[0005] Without limiting the scope of the invention, its background is described in connection with pet food and probiotics.

[0006] One such product is taught in U.S. Patent No. 7,842,329 filed by Saylock and Dixon entitled, “Pet Food Product and Method of Manufacture”. These inventors are said to teach a food that includes a sorbent food body and a fluid carrier comprising a functional ingredient absorbed into the body. The body is said to be relatively moist (a moisture content for example of from 35% to 60% by weight) and a firm structure that is resilient under initial biting by a pet animal. Protein content is preferably said to be at least 20% by weight, the absorptivity of the body is increased by depleting the body of a first liquid (e.g., by causing drying by exposure to a source of dry heat, such as in roasting, grilling, frying and baking).

[0007] Another such product is taught in U.S. Patent Publication No. 20240225048 filed by Frantz and Panasevich entitled, “Pet Food Composition”. These applicants are said to teach a pet food composition that combines a fermentate that contains a non-viable, non-pathogenic bacteria and a prebiotic that contains an oligosaccharide component to achieve a gut health benefit in a companion animal.

[0008] Another such product is taught in U.S. Patent Publication No. 20240000106 filed by Directo entitled, “Pet Food and Method Of Manufacture”. This applicant is said to teach a combination of upcycled ingredients and new ingredients and a careful combination of prebiotics and probiotics to provide pets with a 100% balanced flora. It is said that probiotics, referred to as the “beneficial bacteria,” and prebiotics, referred to as the “beneficial fiber”, work together to stabilize a pet's digestive system, promote good bacteria growth, and a healthy immune system.

[0009] Despite these advances, a need remains for pet food probiotics and feed that improve food safety and animal health, or as supplements for human food that are bactericidal.SUMMARY

[0010] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of treating animals or humans comprising, consisting essentially of, or consisting of: feeding or providing a feed, supplement, additive, or food to animals or humans with a composition comprising one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L.28, L3A, L20-B, J7, JI 4, J27, J43, JI 6, J34, J25, L5-A and J 19 provided in an amount sufficient to be at least one of: bactericidal against spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation. In one aspect, the probiotic bacteria are selected 2, 3, 4, 5, 6, 7, 8, 9,10, or 11 of the probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and JI 9. In another aspect, the spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, are selected from at least one of: Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enterobacter ssp.; Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enteropathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.; Plesiomonas shigelloides; Salmonella ssp.; Shigella ssp.; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enter ocolilica: and Listeria monocytogenes. In another aspect, the probiotic bacteria are provided in an amount selected from 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product. In another aspect, a combination of 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from 1.328, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19, comprise at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product. In another aspect, the composition is formulated into a liquid, a tablet, a paste, a powder, a fermentate, a supernatant, an extract, a pill, or capsule. In another aspect, the composition consist essentially of a feed for livestock, poultry, fish, birds, reptiles, or domestic animals that consists of at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30% w / w one or more probiotic bacteria selected from L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 as areplacement for in-feed antibiotics and treatment for gut infection and inflammation, wherein the composition is formulated for oral delivery. In another aspect, JI 9 is formulated for poultry feed.

[0011] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of treating an animal comprising: providing the animal a composition that comprises one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 in an amount sufficient to be at least one of: bactericidal against bacteria selected from at least one of: Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enterobacter ssp.; Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enteropathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.; Plesiomonas shigelloides; Salmonella ssp.; Shigella ssp.; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enter ocolilica: and Listeria monocytogenes in animal intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation, wherein the composition is formulated for oral delivery. In another aspect, JI 9 is formulated for poultry feed.

[0012] As embodied and broadly described herein, an aspect of the present disclosure relates to a composition comprising, consisting essentially of, or consisting of: one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 in an amount sufficient to be at least one of: bactericidal against bacteria selected from spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, at least one of: in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation. In one aspect, the probiotic bacteria are selected from 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L28, L3 A, L20-B, J7, J 14, J27, J43, J16, J34, J25, L5-A and J19. In another aspect, the probiotic bacteria are provided in an amount selected from 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product. In another aspect, a combination of 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19, comprise at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product. In another aspect, the composition is formulated into a liquid, a tablet, a paste, a powder, a fermentate, a supernatant, an extract, apill, or capsule. In another aspect, the one or more probiotic bacteria are effective against Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enter obacter ssp.; Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enter opathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.; Plesiomonas shigelloides; Salmonella ssp.; Shigella ssp.; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enlerocolilica: and Listeria monocytogenes . In another aspect, the composition consist essentially of a feed for livestock, poultry, fish, birds, reptiles, or domestic animals that consists of at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30% w / w one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 as a replacement for in-feed antibiotics and treatment for gut infection and inflammation, wherein the composition is formulated for oral delivery. In another aspect, J19 is formulated for poultry feed.

[0013] As embodied and broadly described herein, an aspect of the present disclosure relates to an antimicrobial composition comprising a human probiotic or a food comprising the human probiotic comprising, consisting essentially of, or consisting of: one or more probiotic lactic acid bacteria, supernatants, extracts, broth, or fermentate thereof, selected from L28, L3 A, L20-B, J7, J14, J27, J43, JI 6, J34, J25, L5-A and JI 9 in an amount sufficient to be at least one of: bactericidal against bacteria selected from at least one of: spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation. In one aspect, the probiotic bacteria are selected 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the bacterial strains selected from L28, L3A, L20-B, J7, J14, J27, J43, JI 6, J34, J25, L5-A and J19. In another aspect, the probiotic bacteria are provided in an amount selected from 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product. In another aspect, a combination of 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19, comprise at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product. In another aspect, the composition is formulated into a liquid, a tablet, a paste, a powder, a fermentate, a supernatant, an extract, a pill, or capsule. In another aspect, the one or more probiotic bacteria are effective against Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enterobacter ssp.;Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enter opathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.; Plesiomonas shigelloides; Salmonella ssp.; Shigella ssp.; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enlerocolilica: and Listeria monocytogenes . In another aspect, the composition consist essentially of a feed for livestock, poultry, fish, birds, reptiles, or domestic animals that consists of at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30% w / w one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 as a replacement for in-feed antibiotics and treatment for gut infection and inflammation, wherein the composition is formulated for oral delivery. In another aspect, J19 is formulated for poultry feed. In another aspect, the composition is formulated as a food additive. In another aspect, a food product is fermented with at least one of the L28, L3A, L20-B, J7, J14, J27, J43, JI 6, J34, J25, L5-A and JI 9 lactic acid bacteria. In another aspect, the probiotic lactic acid bacteria, extract, or supernatant thereof is bactericidal.

[0014] As embodied and broadly described herein, an aspect of the present disclosure relates to a method of treating a surface to eliminate a pathogenic bacteria comprising: contacting a surface with a composition comprising one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 provided in an amount sufficient to be at least one of: reduce or eliminate at least one of: sporeforming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof on the surface. In one aspect, the surface is steel, stainless steel, galvanized steel, carbon steel, alloy steel, chrome, steel chrome-plated, steel with nickel / silicon carbide composite coating, brass, brass- chrome plated, brass with nickel / silicon carbide composite, stainless chrome-plated, stainless with nickel / silicon carbide composite coating, carbonitrided steel, nickel carbide plated steel, tempered steel, copper, brass, bronze, aluminum, tinplate, manganese, nickel, zinc, titanium, tin, plastic, polymer, fiberglass, polypropylene, polytetrafluorethylene (PTFE), polyvinyl chloride (PVC), and high-density polyethylene, polycarbonate, carbon fiber, a composite material, ceramic, porcelain, coated with a non-stick surface, stone, rock, granite, glass, a coating, a plastic-coated metal, wood, a thermoplastic material, and polyether ether ketone (PEEK), or combinations thereof. In another aspect, the surface is a drain, a pipe, a pot, a dish, a bowl, a pan, tableware, a spoon, a fork, a knife, crockery, glasses, cutlery, colanders, mixers, food presses, food extruders, spreaders, countertops, cooking utensils, baking utensils, serving trays, a floor, or combinations thereof. In another aspect, the surface is on a kitchen utensil selected from group consisting of biscuit cutter, cake and pie server, bottle opener, tin opener, cork screw, nut cracker, crab cracker, pizza cutter, potato masher,rolling pin, corer, curler, pitter, peeler, sieve, scissors, scraper, shear, slicer, scaler, sifter, scoop, spatula, spider, chopper, squeezer, grater, grinder, blender, whisk, tong, ladle, bowl, dish, plate, platter, saucer, tray, sauce boat, gravy boat, creamer, salt shakers, pepper shaker, salt cellar, sugar bowl, jug, pitcher, cup, mug, glass, teapot, or coffee pot. In another aspect, the reduction in pathogenic bacteria is at least a Logw reduction of 0.5, 0.81, 0.87, 1.01, 1.06, 1.14, 1.21, 1.30, 1.33, 1.36, 1.54, 1.73, 1.81, 1.95, or 2.31.BRIEF DESCRIPTION OF THE DRAWINGS

[0015] For a more complete understanding of the features and advantages of the present disclosure, reference is now made to the detailed description of the disclosure along with the accompanying figures and in which:

[0016] FIG. 1 is a graph that compares the zone of inhibition in millimeters of Lactobacillus salivarious strain L28 broth compared to a broth from lactic acid bacteria strain NP51, against Salmonella, E. coli O157:H7, and Listeria monocytogenes. Zones greater than 0.5 mm are considered inhibitory.

[0017] FIG. 2 is a graph that compares the zone of inhibition in millimeters of Lactobacillus salivarious strain L28 broth compared to a broth from lactic acid bacteria strain NP51, against Clostridium perfringens at 37 °C. Zones greater than 0.5 mm are considered inhibitory.

[0018] FIG. 3 shows the Log reductions of Salmonella by varying doses of Lactobacillus salivarious L28 at 37 °C in cattle manure.

[0019] FIG. 4 shows the Log reductions of a cocktail of 7 STEC strains by varying doses of Lactobacillus salivarious L28 at 37 °C in cattle manure.

[0020] FIG. 5A is a graph that shows Log reduction of adherent L. monocytogenes after 24 h exposure to lactic acid bacteria strains. FIG. 5B shows the mass formation in culture and electron microscopy of biomass formation.

[0021] FIG. 6 is a graph that shows the Log reduction of Salmonella in inoculated chicken fat. After 72 h of Lactobacillus salivarious strain L28 treatment: Salmonella was not detectable (ND) by direct plating (based on three independent experiments).

[0022] FIG. 7 is a graph that shows lamb meal inhibition of Salmonella using Lactobacillus salivarious strain L28.

[0023] FIG. 8 is a graph that shows that after 72 h of Lactobacillus salivarious strain L28 treatment: Salmonella was not detectable (ND) by direct plating or enrichment (based on three independent experiments).

[0024] FIG. 9 is a direct comparison of the Lactobacillus salivarious strain L28 treatment (Probicon) compared to four different commercially available probiotics (A, B, C, and D).

[0025] FIG. 10 shows L. monocytogenes counts (Logw CFU / cm2) over time (Days 1, 3, 7, 14, and 21) for three treatment groups: Control (blue), Treatment 1 (pink), and Treatment 2 (green). Each boxplot illustrates the distribution of Logw CFU / cm2values for a treatment at each time point, with individual data points representing three biological replicates (colored by replicate: blue = Rep 1, red = Rep 2, green = Rep 3). The mean Logw CFU / mL values are shown above each group of boxplots. Different letters (a, b, c) above the boxes denote statistically significant differences among treatments at each time point based on post hoc analysis (p < 0.05), with overall significance evaluated by one-way ANOVA (p-values displayed at the bottom of each panel).

[0026] FIG. 11 is a graph that shows the reduction of Listeria monocytogenes on stainless steel over time using varying doses of J 19.

[0027] FIG.12 is a graph that shows the reduction of Listeria monocytogenes on drains over time using varying doses of J 19.

[0028] FIG. 13 is a graph that shows the reduction of Listeria monocytogenes on stainless steel over time using varying doses of LI 5.

[0029] FIG. 14 is a graph that shows the reduction of Listeria monocytogenes on drains over time using varying doses of LI 5.DETAILED DESCRIPTION

[0030] While the making and using of various aspects of the present disclosure are discussed in detail below, it should be appreciated that the present disclosure provides many applicable inventive concepts that can be embodied in a wide variety of specific contexts. The specific aspects discussed herein are merely illustrative of specific ways to make and use the disclosure and do not delimit the scope of the disclosure.

[0031] To facilitate the understanding of this disclosure, a number of terms are defined below. Terms defined herein have meanings as commonly understood by a person of ordinary skill in the areas relevant to the present disclosure. Terms such as “a”, “an” and “the” are not intended to refer to only a singular entity, but include the general class of which a specific example may be used for illustration. The terminology herein is used to describe specific aspects of the disclosure, but their usage does not delimit the disclosure, except as outlined in the claims.

[0032] Probiotics originate from the Greek term meaning “for life,” and represent living microorganisms that work to replenish gastrointestinal microflora. This gastrointestinal microflora aids in digestion, and leads to greater health. The present disclosure includes the isolation and formulation of probiotic bacteria into food or foodstuffs for humans and animals.

[0033] Previously, the present inventors have isolated lactic acid bacterial strains from a variety of sources, as reported by Ayala, et al., A Systematic Approach to Identify and Characterize the Effectiveness and Safety of Novel Probiotic Strains to Control Food Borne Pathogens, Frontiers in Microbiology, Vol. 10, 17 May 2019, Article 1108, pp. 1-16, relevant portions incorporated herein by reference.

[0034] The composition may provide a probiotic benefit to a host in a special dietary product. The special dietary product may be selected from the group consisting of an infant formula, a follow- on formula, processed cereal based food, canned baby food, an animal supplement or treatment, and / or a special food for a medical purpose, including an infant formula or meals that are formulated for special dietary needs.

[0035] The probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28 (ATCC Deposit No. PTA-127731), L3A, L20-B, .17, J 14, J27, J43, J 16, J34, J25, L5-A and JI 9, which have been found to be bactericidal against a number of human and animal pathogens and can be included as additives, supplements, or as standalone probiotics for animal or human consumption.

[0036] The probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and JI 9 and can be mixed with one or more proteins, e.g., hydrolyzed proteins. The one or more proteins may be selected from the group consisting of milk proteins, plant proteins, and combinations thereof. The one or more hydrolyzed proteins may be selected from the group consisting of hydrolyzed casein, hydrolyzed whey protein, hydrolyzed pea protein, hydrolyzed soy protein, and combinations thereof.

[0037] The probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J 19, and can be mixed with one or more disaccharides and / or oligosaccharides. The one or more disaccharides may be selected from the group consisting of sucrose, lactose, and combinations thereof. The one or more oligosaccharides may be selected from the group consisting of inulin, maltodextrins, dextrans, fructo-oligosaccharides (FOS), galactooligosaccharides (GOS), mannan-oligosaccharides (MOS), and combinations thereof. The one or more polysaccharides may be selected from the group consisting of carrageenan, guar gum, gum acacia, locust bean gum, starches, modified starches, and combinations thereof.

[0038] The probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L.28, L3 A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and JI 9, and can be mixed with one or more additional agents that may be selected from the group consisting of vitamins, minerals, micronutrients, carboxylic acid salts, tocopherols,and combinations thereof. The carboxylic acid salts may be selected from the group consisting of ascorbic acid salts and citric acid salts. The one or more additional agents may comprise one or more tocopherols and one or more carboxylic acid salts at a weight ratio from 1 :4 to 4: 1. Preferably, the one or more additional agents comprise vitamin E and sodium ascorbate at a weight ratio of 4: 1.

[0039] The probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and JI 9, and can be prepared into a solid or powder using one or more drying processes selected from the group consisting of air drying, vacuum-drying, fluid bed drying and spraydrying.

[0040] In one example, the present invention includes a method of making a formulation that can be used alone or in combination with a food or feed that includes: probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19, and optionally, one or more of the following: the one or more proteins, the one or more disaccharides, the one or more oligosaccharides, and the one or more polysaccharides in an alkali aqueous solvent to form a slurry; snap-freezing the slurry in liquid nitrogen to form solid frozen particles in the form of beads, droplets or strings; drying the solid frozen particles by evaporation, under vacuum, while maintaining the temperature of the particles above their freezing temperature, whereby a primarily dried formulation is formed; and optionally a second drying step at the same or a different level of vacuum and a heat source temperature of 20°C (or higher) for a time sufficient to reduce the water activity of the primarily dried formulation to 0.3 Aw or lower. The method may further include sterilizing the one or more hydrolyzed proteins, the one or more disaccharides, the one or more oligosaccharides, and the one or more polysaccharides before mixing with the one or more probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28, L3 A, L20-B, J7, J14, J27, J43, J 16, J34, J25, L5-A and J 19. The method may further comprise cutting, crushing, milling or pulverizing the composition into a free-flowing powder.

[0041] The composition that includes the one or more probiotic microorganism, extracts, fermentates, and / or supernatants may be selected from one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 will generally comprise an effective amount of the one or more viable probiotic microorganisms for providing a probiotic benefit to a host in a special dietary product. The special dietary product may be selected from the group consisting of an infant formula, a follow-on formula, processed cereal based food, canned baby food, an animal supplement or treatment, and / or a special food for a medical purpose.

[0042] In a particular embodiment, the amount of probiotics comprised in the food or feed, or as an additive for a feed or food, of the invention is of about UK)7, 5*107, U108, 2*108, 3*108, 4xl08, 5xl08, 6xl08, 7xl08, 8xl08, 9xl08, IxlO9, 2xl09, 3xl09, 4xl09, 5xl09, 6xl09, 7xl09, 8xl09, 9xl09, IxlO10, 2xlO10, 3xlO10, 4xlO10, 5xlO10, 5.5xlO10, 6xlO10, 6.2xlO10, 6.5xlO10, 6.7xlO10, 7xlO10, 7.2xlO10, 7.5xlO10, 7.7xio10, 8xlO10, 8. IxlO10, 8.2xlO10, 8.3xio10, 8.4xlO10, 8.5xlO10, 8.6xlO10, 8.7xlO10, 8.8xlO10, 8.9xlO10, 9xlO10, 9.1xlO10, 9.2xlO10, 9.3xio10, 9.4xlO10, 9.5xlO10, 9.6xlO10, 9.7xlO10, 9.8xlO10, 9.9xlO10, IxlO11, 1.U1011, 1.2xlOn, 1.3xl0n, 1.4xlOn, 1.5xlOn, 1.6xlOn, 1.7X1011, 1.8X1011, 1.9xlOn, 2xlOn, 2.2xlOn, 2.5xlOn, 2.7xlOn, 3xl0n, 3.5X1011, 4xlOn, 4.5X1011, 5xl0n, 6xlOn, 7xlOn, 8xl0n, 9xlOn, IxlO12, 2xl012, 3xl012, 4xl012, 5xl012, 6xl012, 7xl012, 8xl012, 9xl012, IxlO13, 5xl013, IxlO145xl014, IxlO15CFU of probiotic bacteria per dose or meal.

[0043] In another particular embodiment, the amount of probiotics comprised in the food or feed, or as an additive for a feed or food of the invention is of between IxlO7and IxlO15CFU of probiotic bacteria per dose or meal, between 1 x 108and 1 x 1013CFU of probiotic bacteria per dose or meal, between IxlO9and IxlO12CFU of probiotic bacteria per dose or meal, between IxlO10and UlO11CFU of probiotic bacteria per dose or meal, between 5xlO10and 5xl0nCFU of probiotic bacteria per dose or meal, between 7x IO10and 4x 1011CFU of probiotic bacteria per dose or meal, between 8xlOloand 2xlOnCFU of probiotic bacteria per dose or meal, between 8.5xlO10and 1.8xl0nCFU of probiotic bacteria per dose or meal, between 8.7xlO10and 1.7xlOnCFU of probiotic bacteria per dose or meal, between 8.7xlOloand 1.4xlOnCFU of probiotic bacteria per dose or meal, between 9xlO10and 1.7xlOnCFU of probiotic bacteria per dose or meal, between 9.2xlO10and 1.7x1011CFU of probiotic bacteria per dose or meal, between 9.2xlO10and 1.4xlOnCFU of probiotic bacteria per dose or meal, between U1011and 1.2xlOnCFU of probiotic bacteria per dose or meal, between 8.5xlO10and 2xlOnCFU of probiotic bacteria per dose or meal, or between 8.7xlOloand 1.7xlOnCFU of probiotic bacteria per dose ormeal. In one aspect, the number of probiotic bacteria in the biomaterial of the invention is of between 8.5 xlO10and 2xlOnCFU of probiotic bacteria per dose or meal. In another embodiment, the number of probiotic bacteria in the biomaterial of the invention is of between U1011and 1.2xlOnCFU of probiotic bacteria per dose or meal.

[0044] In another example, the present disclosure includes a method of treating a surface to eliminate a pathogenic bacteria comprising: contacting a surface with a composition comprising one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 provided in an amount sufficient to be at least one of: reduce or eliminate at least one of: spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof on the surface. In one aspect, the surfaceis steel, stainless steel, galvanized steel, carbon steel, alloy steel, chrome, steel chrome-plated, steel with nickel / silicon carbide composite coating, brass, brass-chrome plated, brass with nickel / silicon carbide composite, stainless chrome-plated, stainless with nickel / silicon carbide composite coating, carbonitrided steel, nickel carbide plated steel, tempered steel, copper, brass, bronze, aluminum, tinplate, manganese, nickel, zinc, titanium, tin, plastic, polymer, fiberglass, polypropylene, polytetrafluorethylene (PTFE), polyvinyl chloride (PVC), and high-density polyethylene, polycarbonate, carbon fiber, a composite material, ceramic, porcelain, coated with a non-stick surface, stone, rock, granite, glass, a coating, a plastic-coated metal, wood, a thermoplastic material, and polyether ether ketone (PEEK), or combinations thereof. In another aspect, the surface is a drain, a pipe, a pot, a dish, a bowl, a pan, tableware, a spoon, a fork, a knife, crockery, glasses, cutlery, colanders, mixers, food presses, food extruders, spreaders, countertops, cooking utensils, baking utensils, serving trays, a floor, or combinations thereof. In another aspect, the surface is on a kitchen utensil selected from group consisting of biscuit cutter, cake and pie server, bottle opener, tin opener, cork screw, nut cracker, crab cracker, pizza cutter, potato masher, rolling pin, corer, curler, pitter, peeler, sieve, scissors, scraper, shear, slicer, scaler, sifter, scoop, spatula, spider, chopper, squeezer, grater, grinder, blender, whisk, tong, ladle, bowl, dish, plate, platter, saucer, tray, sauce boat, gravy boat, creamer, salt shakers, pepper shaker, salt cellar, sugar bowl, jug, pitcher, cup, mug, glass, teapot, or coffee pot.

[0045] As it will be understood by a skilled person, the probiotics comprised in the biomaterial of the invention are almost all mixed into the biomaterial, such as a feed or food. Thus, in a particular aspect, the expression “the probiotics comprised in the feed or food”, as used all along the specification, refers to the probiotics combined into a dosage form that can be included with the feed or food, provided separately from the feed or food, mixed at least partially or fully into the feed or food, provided as a separate dose from the feed or food. As will be apparent the dosage form can be formulated into any form, such as solid, partially solid, gel, liquid, gel cap, chewable, or any other dosage form for oral or topical administration.

[0046] Animals such as livestock (cattle, sheep, horses, goats, and other domestic animals ordinarily raised or used on the farm), or pets such as dogs and cats must have a strong and healthy microflora because of the stress they induce on their intestines from their unfiltered eating habits. While probiotic products are available for livestock and / or pets, there are no distinctions for which probiotics to give the respective species.

[0047] As used herein, the term “effective amount” refers to the amount of the invention which gives rise to an inhibition of the bacterial growth or a reduction of the number of other bacteria from the food product.

[0048] As used herein, the term “bactericidal” refers to any type of treatment which effects the killing of bacteria (i.e. which reduce their numbers). This is in contrast to a “bacteriostatic effect” which refers to the situation where the treatment only inhibits the growth or reproduction of the bacteria. An agent is said to be bactericidal if the agent is able to kill one or more type of bacteria.

[0049] As used herein, the term “bacteriocins” refers to peptides, protein, or other molecules released extracellularly by the one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19, e.g., a supernatant, a fermentate, or extract, that is / are able to kill, or reduce the number of, or the symptoms, caused pathogenic bacteria by a mechanism by which the producer cell exhibits microbiocidal activity.

[0050] As used herein, the term “animal feed” refers to feed for an animal that provides basic nutrition and an improvement of the health of livestock, poultry, fish, birds, reptiles, and domestic animals. The animal feed may be in the form of powder, grain or liquid form and may be used in accordance with the feeding condition and installations of the farm and the target animal. Animal feedstuffs often include, e.g., green feed, silages, dried green feed, roots, tubers, fleshy fruits, grains and seeds, brewer's grains, pomace, brewer's yeast, distiller's spent grains, milling byproducts, byproducts of the production of sugar, starch and oil recovery and various food wastes. The animal feed may also include feed additives used alone or in conjunction with other well-known feed additives such as antioxidants or mixtures of various substances (mineral mixtures, vitamin mixtures) that can be added to such feeds for enhancement. Specific feeds may also be adapted for certain animal species depending on age and stages of development.

[0051] Base animal feeds suitable for use in conjunction with the present invention may be prepared as is well-known to the artisan skilled in the art of preparing feeds, e.g., they may use those as described in Kirk-Othmer, Encyclopedia of Chemical Technology, 4th Ed., vol. 10, pp. 288-300, Wiley, N.Y., 1993, relevant portions incorporated herein by reference. For example, the base feed may include one or more of the following ingredients: corn, sorghum, barley, wheat, soybean, peanut, canola, fish meal, milk products, fats and oils, vitamins and minerals.

[0052] As used herein, the terms “food product” and “food stuff’ refer to any food that is susceptible to spoilage as a result of bacterial growth and proliferation, e.g., but not limited to, meat, dairy products, vegetables, fruits and grains. Generally, food is used in the context of human consumption.

[0053] As used herein, the term “livestock” refers to, e.g., cattle, sheep, pigs, goats, horses, donkeys, mules, buffalo, oxen, or camels, namely, farm animals used in agriculture.

[0054] As used herein, the term “meat” refers to any meat product or meat by-product (including those processed) from an animal which is consumed by humans or animals, including, without limitation, meat from bovine, ovine, porcine, poultry, fish and crustaceous seafood. As used in thepresent application, the term “ready to eat meat product”, also referred to as RTE meat product, is intended to include any meat product which does not require cooking prior to consumption.

[0055] As used herein, the terms “refrigerated product” or “preserved in a refrigerated state” are equally used and refer to food products which are stored at temperatures ranging from to 2 to 10° C. The food product can be packaged, packaged under vacuum or packaged at modified atmosphere.

[0056] As used herein, the term “shelflife” refers to the period of time that a food product remains saleable to retail customers. In traditional meat processing, the shelf life of meat and meat byproducts is about 30 to 40 days after an animal has been slaughtered. Refrigeration of meat during this period of time is expected to largely arrest and / or retard the growth of pathogenic bacteria, and to a lesser extent, spoilage bacteria. After about 30 to 40 days, however, refrigeration is no longer able to effectively control the proliferation of spoilage bacteria below acceptable levels.

[0057] As used herein, the term “spoilage bacteria” refers to any type of bacteria that act to spoil food. Spoilage bacteria may grow and proliferate to such a degree that a food product is made unsuitable or undesirable for human or animal consumption. Bacteria are able to proliferate on food surfaces, such as meat surfaces, by assimilating sugars and proteins on such surfaces. By metabolizing these components, spoilage bacteria create by-products including carbon dioxide, methane, nitrogenous compounds, butyric acid, propionic acid, lactic acid, formic acid, sulfur compounds, and other undesired gases and acids. The production of such by-products alters the color of meat surfaces, often turning meat from a red color to a brown, grey or green color. Gaseous by-products generated by spoilage bacteria also give spoiled meat an undesirable odor. The color and odor alterations of meat due to the growth of spoilage bacteria on a surface of a meat product often make such food product unsaleable to consumers.

[0058] In addition to the control of spoilage bacteria, another significant concern in the food processing industry is controlling the growth of food-borne pathogenic bacteria. As used herein, the term “food-borne pathogenic bacteria” refers to any food poisoning organism that is capable of causing disease or illness in animals or humans. The term “pathogenic bacteria” will be understood to include bacteria that infect the food product (for instance meat) and thereby cause disease or illness, as well as bacteria that produce toxins that cause disease or illness. The pathogenic bacteria may be selected from the group that includes, but is not limited to: Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enterobacter ssp.; Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enter opathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.;Plesiomonas shigelloides; Salmonella ssp.; Shigella ssp.; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enter ocolitica; and Listeria monocytogenes.

[0059] Example 1. Reduction of Salmonella.

[0060] The L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 lactic acid bacteria (LAB) are as follows. Probicon can include one or more of these probiotic bacteria: L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19.

[0061] L28, Lactobacillus salivarius, (ATCC Deposit No. PTA-127731).

[0062] L3 A, Pediococcus acidilaclici. secretes a Pediocin, isolated from a bovine sample.

[0063] L20-B, Enterococcus faecium, secretes Enterocin B, Anterolysin A, isolated from a bovine sample.

[0064] J7, Enterococcus faecium, secretes Enterolysin A, Lactacin F, isolated from a grape sample.

[0065] J14, Enterococcus faecium, secretes Enterocin B, Enterolysin A, enterocin L50A, Lactacin F, isolated from a grape sample.

[0066] J27, Enterococcus faecium, secretes Enterolysin A, Lactacin F, Enterocin B, isolated from a grape sample.

[0067] J43, Enterococcus faecium, secretes Enterolysin A, Lactacin F, enterocin (2), isolated from a carrot sample.

[0068] JI 6, Enterococcus faecium, secretes Enterocin B, Enterolysin A, enterocin L50A, isolated from a grape sample.

[0069] J34, Enterococcus faecium, secretes Enterolysin A, Lactacin F, Enterocin B, enterocin L50A, isolated from a grape sample.

[0070] J25, Enterococcus faecium, secretes Enterolysin A, Enterocin B, isolated from a grape sample.

[0071] L5-A, Enterococcus faecium, secretes enterocin (2), Enterolysin A, Lactacin F, isolated from a bovine sample.

[0072] JI 9, Enterococcus faecium, secretes Colicin V, and enterocin producing cluster (4), isolated from a cabbage sample.

[0073] Experimental Design and Analysis: Fecal samples were collected from the Texas Tech University’s Burnett Center in New Deal, Texas from cattle fed base diets containing 3 variations: (1) no probiotics, tylosin or Moenensin (control); (2) Monensin with a probiotic, and (3) Monensin and Tylosin with no probiotic. Variations were studied because the presence of antimicrobial agents could potentially change the pathogen reduction. Samples were inoculated with a cocktail of three pathogenic E. coli strains (0157, Ol l i, and 026) to a final concentration of 105CFU / ml. Lactobacillus salivarius L28, was used as an environmental mitigation treatment at threeconcentrations (106, 107, and 108cfu / g). Samples were collected at 1 and 24 hrs after inoculation and incubation at 37°C. Pathogenic E. coli were enumerated on MacConkey agar with a thin-layer overlay of TSA to facilitate injured cell recovery.

[0074] Agar Spot Test of Lactic Acid Bacteria.

[0075] Method 1-1. Prepare cultures of E. coli O157:H7, Salmonella, Listeria monocytogenes and C. perfringens from frozen stock collections in TSA at 37° C for 24 hrs transferring at least two times to ensure that full recovery is obtained. (C. perfringens must be incubated in anaerobic jar). Use 3-4 strains of each pathogen. This will take 2-4 days in advance of actual experiment.2. Prepare TSA agar plates for the agar spot test by pre-pouring and allowing to dry overnight in the incubator. It is imperative that the surface is very dry so the cultures do not utilize the water on the surface to “move”.3. Obtain Lactobacillus salivarius L28 (Probicon), supernatant, broth, or fermentate thereof from freezer and rehydrate at the following concentrations: a. 1 g added to 10 ml sterile distilled water b. 1 g added to 100 ml of sterile distilled water c. 1 g added to 1000 ml of sterile distilled water4. Spot inoculate each culture onto the surface of the dried TSA plates with an inoculation needle. All three can be spot inoculated on a single plate in 3 separate regions. Prepare duplicate plates for each pathogen that will be evaluated (total of 8).5. Incubate the plates for 24 hrs at 37°C to allow cultures to grow. If there is no growth, allow another 24 hrs of growth.6. Prepare “soft” Tryptic Soy Agar (prepared from scratch with TSB and adding agar) in 10 ml tubes and temper at 37°C. A minimum of 8 of these tubes are generally used.7. Prepare a cocktail mixture of all strains for each pathogen individually (one set of cocktail for each pathogen) by adding 5 ml of each strain and mixing together in a sterile container.8. Add 1 ml of the prepared cocktail to the tempered soft TSA and mix well. There will be 8 tubes (two for each pathogen).9. Pour the inoculated soft agar onto the surface of the pre-grown Probicon TSA plates and allow to solidify.10. Incubate the plates face-up at 37°C. Measure zones of inhibition at 24 and 48 hrs using calipers and take pictures of each plate.

[0076] Method 2-1. Prepare cultures of E. coli O157:H7, Salmonella, Listeria monocytogenes and C. perfringens from frozen stock collections in TSA at 37°C for 24 hrs transferring at least two times to ensure that full recovery is obtained. C. perfringens must be incubated in anaerobic jar. Generally, use 1-4 strains of each pathogen. Prepare 2-4 days in advance of actual experiment.2. Prepare TSA agar plates for the agar spot test by pre-pouring and allowing to dry overnight in the incubator. It is imperative that the surface is very dry so the cultures do not utilize the water on the surface to “move”.3. Prepare a cocktail mixture of all strains for each pathogen individually (one set of cocktail for each pathogen) by adding 5 ml of each strain and mixing together in a sterile container.4. Using a cotton-tipped swab, fully cover the entire surface of a dried TSA plate with each individual pathogen cocktail and allow to dry / absorb into the agar for at least 1 hr at room temperature. Prepare two plates for each pathogen cocktail (total of 8).5. Obtain Probicon from freezer and rehydrate at the following concentrations: a. 1 g added to 10 ml sterile distilled water b. 1 g added to 100 ml of sterile distilled water c. 1 g added to 1000 ml of sterile distilled water6. Spot inoculate each concentration onto the pathogen-inoculated plates using an inoculation needle. Be sure to label each region on the plate.7. Incubate the plates face-up at 37°C. Measure zones of inhibition at 24 and 48 hrs using calipers and take pictures of each plate.

[0077] Competitive Exclusion Broth Culture Assay. Overnight cultures of each pathogen and LAB strains were prepared as described above for agar well diffusion assays and co-inoculated at 105 and 106 CFU / mL, respectively in Tryptic Soy Broth (TSB; Oxoid Ltd., Basingstoke, United Kingdom) supplemented with 1 g I’1Tween 80 (Acros, Organics, NJ, United States) and incubated at 37°C with slight agitation (130 rpm). Addition of tween to TSB allows growth of both LAB and gram-negative pathogens, including A. coli O157:H7 and Salmonella. Samples were diluted and plated onto Modified Oxford Agar (MOX; Becton, Dickinson and Company), Xylose Lysine Tergitol 4 Agar (XLT4; Becton, Dickinson and Company), MacConkey agar with sorbitol (SMAC; Criterion, Hardy Diagnostics, CA, United States), and MRS agar plates to enumerate L. monocytogenes, Salmonella, E. coli O157:H7, and lactic acid bacterial (LAB) strains above, respectively, at 0, 6, 12, and 24 h of coinoculation. MOX, XLT4, and SMAC plates were incubated at 37°C for 24 h, and MRS plates were incubated at 37°C for 48 h. Antagonistic activity of each LAB strain was determined by pathogenic reduction with respect to control samples (pathogencultures without LAB) at each time point. As for agar well diffusion assays, pathogenic reductions were summed up across all time points and across all three pathogens, and potential LAB probiotic strains were ranked based on their antagonistic effect.

[0078] FIG. 1 is a graph that compares the zone of inhibition in millimeters of Lactobacillus salivarious strain L28 broth compared to a broth from lactic acid bacteria strain NP51, against Salmonella, E. coli O157:H7, and Listeria monocytogenes. Zones greater than 0.5 mm are considered inhibitory.

[0079] FIG. 2 is a graph that compares the zone of inhibition in millimeters of Lactobacillus salivarious strain L28 broth compared to a broth from lactic acid bacteria strain NP51, against Clostridium perfringens at 37 °C. Zones greater than 0.5 mm are considered inhibitory.

[0080] FIG. 3 shows the Log reductions of Salmonella by varying doses of Lactobacillus salivarious L28 at 37 °C in cattle manure.

[0081] FIG. 4 shows the Log reductions of a cocktail of 7 STEC strains by varying doses of Lactobacillus salivarious L28 at 37 °C in cattle manure.

[0082] FIG. 5A is a graph that shows Log reduction of adherent L. monocytogenes after 24 h exposure to lactic acid bacteria strains. FIG. 5B shows the mass formation in culture and electron microscopy of biomass formation.

[0083] FIG. 6 is a graph that shows the Log reduction of Salmonella in inoculated chicken fat. After 72 h of Lactobacillus salivarious L28 treatment: Salmonella was not detectable (ND) by direct plating (based on three independent experiments).

[0084] FIG. 7 is a graph that shows lamb meal inhibition of Salmonella using Lactobacillus salivarious strain L28.

[0085] FIG. 8 is a graph that shows that after 72 h of Lactobacillus salivarious strain L28 treatment: Salmonella was not detectable (ND) by direct plating or enrichment (based on three independent experiments).

[0086] FIG. 9 is a direct comparison of the Lactobacillus salivarious strain L28 treatment (Probicon) in a blinded study compared to four different commercially available probiotics (A, B, C, and D).

[0087] Transcriptional profile of Salmonella and E. coli O157:h7 grown in presence of L. salivarius L28 metabolites.

[0088] Attachment and virulence-associated genes of Salmonella and E. coli O157:H7 were among the 25.2%, and 37.01% of the genes with significantly reduced expression, respectively, when grown with L29 metabolites. Down-regulation of virulence-associated genes. By way of explanation, but not a limitation of the present disclosure, the mechanism used in vivo by L. salivarius L28 to inhibit pathogen’s ability to: colonize the gastrointestinal tract, invade epithelialcells and replicate intracellularly, reduce carriage and fecal shedding of these foodborne pathogens in animals.

[0089] Table 1. Down regulation of Salmonella virulence-associated genes.Name log2 log2 total Fold Regulation P value Function total RPKM ChangeRPKM (Treatment) (control)Seq487 13.27 11.77 2.87 down 3.35E-03 Outer membraneOmpXSeq2088 8.78 7.71 2.09 down 8.30E-03 Motility and virulenceName log2 log2 total Fold Regulation P value Function total RPKM ChangeRPKM (Treatment) (control)Seq2097 13.79 12.59 2.829 down 3.18E-03 Outer membrane protein A precursor

[0090] Example 2. Reduction of Listeria monocytogenes in Deli Meat using Enterococcus faecium JI 9 and Lactobacillus sakei LI 5.

[0091] This example aimed to evaluate the antilisterial effectiveness of a characterized LAB strain, Enterococcus faecium (JI 9), and Lactobacillus sakei (LI 5) applied to surface-inoculated deli meat loaves (Ham) using sprays to control three strains of Listeria monocytogenes (L. monocytogenes) during storage in retail display cases at 4.0- 4.3 °C.

[0092] Product samples. Mainstream brand black forest uncured hams (10.7 g each, on average) were received in retail packaging shortly after in-store retail distribution. Samples were inoculated the next day prior to the experiment.

[0093] Listeria monocytogenes preparation of inoculum: A cocktail of L. monocytogenes was created using three strains: L. monocytogenes TTU N 1-002, L. monocytogenes TTU N 1-022, and L. monocytogenes TTU N 1 -014.

[0094] These strains were selected due to the presence of vital virulence-encoding genes. Fresh cultures of each strain were stored at -80 °C in 10% glycerol stocks and plated onto Brain Heart Infusion (BHI) agar to obtain isolated colonies before preparing the inoculum. Each strain was cultivated independently by introducing a well-isolated colony into 9.0 ml of BHI broth and incubating at 37 °C for 24 hours. Equal volumes (500 pl) of the 24-hour cultures from each strain were combined to create a three-strain cocktail. The mixed cultures were vortexed to ensure thorough blending, achieving a target concentration of approximately 109CFU / ml. For experimental treatments, the cocktail was diluted in sterile deionized-distilled water to reach a working concentration of around 105CFU / ml, which was then used to inoculate deli meat samples via immersion. Additionally, to ensure a robust inoculum, the strains were also cultured independently in 500 ml bottles of BHI supplemented with yeast extract (YE) and incubated for 24 hours at 37 °C before diluting and applying treatments.

[0095] Lactic- Acid-Bacteria (LAB) preparation of inoculum. E. faecium strain JI 9 and / .. sakei LI 5, both lactic acid bacteria (LAB), were prepared following protocols similar to those described for L. monocytogenes. Frozen stocks of J19 / L15 strains were stored at -80 °C and streaked on de Man, Rogosa, and Sharpe (MRS) agar to isolate single colonies. An isolated colony was transferred into MRS broth and incubated anaerobically at 37 °C for 48 hours. J19 / L15 were cultured in 1.0 L bottles of MRS broth for large-scale preparations and incubated for 24 hours. Following incubation, the cultures reached approximately 108CFU / ml. The pH of each broth culture was measured after incubation to confirm LAB metabolic activity. The cultures were then diluted in Buffered Peptone Water (BPW) to final working concentrations of approximately 107CFU / ml. These dilutions were applied to deli meat (Ham) treatments 1 and 2 via spraying.

[0096] Inoculation of samples. Three experimental replications were performed; for each experimental replication, the hams were purchased from a local grocery store using different lot numbers and stored at 4 °C. Prior to inoculation, hams were aseptically opened and surface-dried using sterilized absorbent paper. A three-strain L. monocytogenes cocktail was diluted in BHI broth to approximately 105CFU / ml, and ham squares (n=75 total) per treatment were submerged in 1.5 L (1.5 L*3 treatments) of BHI broth with the inoculum in sterilized 6.0-L stainless steel pans for 20 minutes to allow bacterial attachment. After exposure, the samples' excess broth was drained for an additional 20 minutes, resulting in a final attachment concentration of approximately 103— 104CFU / cm2Each treatment consisted of five ham squares per day over three treatments and five sampling days (DI, D3, D7, DI 4, and D21). A total of 75 samples were processed per biological repetition, with extra samples as backups.

[0097] Application of LAB by spraying onto the surface of Deli Meats. The spray intervention was conducted in vitro at the ICFIE Laboratory (BSL-2) at Texas Tech University (TTU), simulating industry conditions. Each ham square group was sprayed separately using a handheld sanitization sprayer, which was first flushed for one minute to remove residual sanitizer. Three treatments will be applied during the intervention, as follows: Control = Five ham squares with 105CFU / ml cocktail of / .. monocytogenes + sprayed with the blank of MRS broth; Treatment 1 = Five ham squares with 105CFU / ml cocktail of / .. monocytogenes + 107CFU / ml of JI 9 E. fciecium Treatment 2 = Five ham squares with 105CFU / ml cocktail of / .. monocytogenes + 107CFU / ml of LI 5 L. sakei.

[0098] Each treatment was prepared in a sterile 1.0 L spray bottle. Spraying occurred for 30 seconds per side, followed by draining and vacuum packaging in separate rooms to prevent crosscontamination. Ham samples were stored at 4°C and opened at designated time points for microbial enumeration.

[0099] Enumeration of Listeria monocytogenes. After vacuum packaging, ham samples were stored at approximately 4 °C in a retail display case in the ICFIE Food Microbiology Lab at Texas Tech University (TTU). At designated time points (DI, D3, D7, DI 4, and D21), all treatments — including control, treatment 1, and treatment 2 — were evaluated for reductions in / .. monocytogenes. One deli meat square was placed into a filtered Whirl-Pak stomacher bag containing 90.0 ml of BPW (1 : 10 dilution) for enumeration. Samples were hand-massaged for 30 seconds and then homogenized using a stomacher at 230 rpm for two minutes. Serial dilutions were prepared in 9 ml BPW tubes, and aliquots were plated onto Modified Oxford (MOX) Agar using the spread plate method. Plates were incubated for 24-48 hours, after which L. monocytogenes colonies were enumerated and reported in CFU / cm2

[0100] Statistical analysis. Each experiment included a control, treatment 1, and treatment 2, all inoculated with L. monocytogenes. Colonies on MOX agar plates were counted for total / .. monocytogenes on duplicate agar plates, converted to total CFU / cm2, averaged, and then transformed into Logic numbers. Analyses of Variance (ANOVA) identified differences among means using a one-way ANOVA conducted with R Studio software (Version: 2024.12.0+467). A post-hoc Tukey’s test was applied to compare mean values among treatments and identify significant differences (p < 0.05). Results were considered significant at a p < 0.05 level. The statistical analysis was performed as a two-way ANOVA, factoring in time and treatments. All reductions of L. monocytogenes were regarded as significant if (p < 0.05).

[0101] Figure 10 and Table 2 present L. monocytogenes counts (Logw CFU / cm2) on deli ham slices inoculated with 105CFU / mL of / .. monocytogenes and treated with probiotic cultures JI 9or L15 (107CFU / mL), stored at 4 °C for 21 days. On Day 1, the control showed 4.27 Logw CFU / cm2, while J19 and L15 achieved reductions of 0.69 and 0.48 Logw, respectively. By Day 3, J19 maintained a decrease of 0.65 logs, with LI 5 slightly lower at 0.37 logs. On Day 7, J19 reached a 0.96 Logw (-90%) reduction, while LI 5 achieved 0.52 logs. By Day 14, both treatments exhibited greater efficacy, with JI 9 reducing L. monocytogenes by 2.03 Logw (99.0%) and LI 5 by 1.64 Logw (97.71%). After 21 days, J19 achieved a 3.25 Logw reduction (-99.94%), outperforming LI 5 at 3.03 Logw (-99.9%).

[0102] From Day 7 onward, the control (sodium diacetate-treated ham) consistently showed higher L. monocytogenes levels than the JI 9 treatment, with a difference of nearly 1 log. Both probiotic strains significantly reduced pathogen levels over time, with JI 9 demonstrating superior antilisterial activity.

[0103] Throughout storage, temperatures fluctuated between 4 °C and 7 °C, occasionally rising to 10 °C during retail-style defrost cycles in open display cases, simulating worst-case storage conditions and accelerating shelf-life assessments.

[0104] It was found that under ideal conditions (4-5 °C), Listeria monocytogenes can increase by approximately 3 Logw within 14 days (Gandhi & Chikindas, 2007). To mitigate this risk, the USDA-FSIS Listeria Rule’s Alternative 2b mandates that ready-to-eat (RTE) products be reformulated with an antimicrobial growth inhibitor, such as sodium diacetate, to control L. monocytogenes proliferation (USDA-FSIS, 2014). This regulatory option requires that the product demonstrate no more than a 2 Logw increase in L. monocytogenes throughout its shelf life. Notably, the FSIS guidelines do not specify a fixed validation temperature or a defined shelf-life duration. However, most RTE deli meats are stored closer to 0°F during processing and retail distribution. Validation studies conducted at elevated temperatures (4-7 °C) are commonly used to simulate accelerated shelf-life conditions and worst-case scenarios during transport and retail storage.

[0105] Additionally, the National Advisory Committee on Microbiological Criteria for Foods (NACMCF) recommends limiting L. monocytogenes growth to no more than a 1 Logw increase over 1.25 times the product’s shelf life at both 4 °C and 7 °C (NACMCF, 2010). In the present study, both the sodium diacetate-treated and J 19 / L 15 -treated hams met the regulatory criteria established by FSIS and NACMCF. However, the J19 probiotic culture showed more significant inhibition of L. monocytogenes than the sodium diacetate formulation. Moreover, JI 9 and LI 5 provide the added benefit of supporting clean-label claims, which may enhance consumer appeal compared to synthetic additives like sodium diacetate.

[0106] Table 2. Listeria monocytogenes counts in this table show the mean Logw CFU / mL, standard deviation, and Logw reduction for Control, Treatment 1 (J 19), and Treatment 2 (LI 5) across Days 1 to 21.i Treatment i Mean i Standard deviation i Logw reductioni Treatment 1 Mean i Standard dev w reductionControl 7,346.39 N / AaI I (J 19) 4.09 0.33 3.25f4.31 0.61 3.03

[0107] - N / A= Not applicable

[0108] - Indicates a 1 Logw reduction, meaning a 90% decrease in bacteria.

[0109] - Indicates a 1.64 Logw reduction, meaning a 97.71% decrease in bacteria.

[0110] - Indicates a 2 Logw reduction, meaning a 99% decrease in bacteria.

[0111] eIndicates a 3.25 Logw reduction, meaning a 99.94% decrease in bacteria.

[0112] fIndicates a 3 Logw reduction, meaning a 99.9% decrease in bacteria.

[0113] Example 3. Listeria reduction on Surfaces utilizing Two Lactic Acid Bacteria strains (J 19 and LI 5).

[0114] One of the key factors that makes Listeria so hard to manage is its ability to adhere to surfaces. Even more so, if the surfaces are inadequately disinfected, Listeria strains can form biofilms, which are even harder to remove. In processing environments stainless steel and drains are commonly present and can be reservoirs for Listeria.

[0115] This example evaluated the effect of two different lactic acid bacteria (LAB) strains as interventions after Listeria has attached to the surface of stainless steel or drains.

[0116] The example was divided into two different phases, where the methodology remains the same, but two different strains of lactic acid bacteria were utilized.

[0117] Pathogen reactivation. Strains were removed from the freezer stock collections at Texas Tech University. Three strains of L. monocytogenes were used in this study: L. monocytogenes TTUN 1-002, L. monocytogenes TTUN 1-022, and / .. monocytogenes TTU N1-014. These strains were selected based on their association with foodbome outbreaks in the US and their virulence factors encoding. After a 24-hour incubation, the strains reached an approximately 109CFU / ml concentration. A cocktail for incubation was prepared by mixing equal volumes (lOmL) of each strain from the 24-hour cultures. The cocktail strains were diluted in sterile buffered peptone water (BPW) to obtain a concentration of approximately 106CFU / ml.

[0118] Lactic acid bacteria (LAB) reactivation. The lactic acid bacteria strains were prepared in a similar manner as the / .. monocytogenes strains. In this case, the strains utilized were Enterococcus faecium JI 9 and Lactobacillus sakei LI 5. After the 24-hour incubation, the LAB in MRS broth reached a concentration of approximately 108CFU / ml. The strains were diluted in BPW to obtain the following concentrations, 107CFU / ml), 106CFU / ml, and 105CFU / ml.

[0119] Inoculation of Surfaces. Two types of surfaces were utilized for this study: stainless steel and drains. The three-strain L. monocytogenes cocktail was diluted by performing decimal serial dilutions resulting in an approximate LOGio 5.00 or 105CFU / ml concentration. To inoculate, 1.2ml (to account for loss in spray bottle) was spray-inoculated onto the surface. Surfaces were dried for 45-75 minutes at room temperature using the biohazard hood for faster attachment time if needed. The final expected bacterial attachment concentration of L. monocytogenes in the surfaces was of approximately LOGio 4.00 or 3.00 CFU / cm2. Table 3. The different LAB treatments were inoculated.

[0120] Table 3. Treatments and description of each for both phases of the project.

[0121] *The concentration of the interventions applies to both strains of lactic acid bacteria.

[0122] Sampling and Plating. The tested surface area for stainless steel slate was of 100 cm2and for the drains it was 4-1 / 4 in. Using one sponge per sample, the surfaces were swabbed for 20 seconds on each side. The sponges were placed inside the sample bag and stomached for 30 seconds at 230 rpm. All treatment reductions and control of L. monocytogenes dipping methods were evaluated for each time point (Ohrs, 6hrs, and 24hrs) for each treatment. Serial dilutions were performed in 9 ml BPW tubes. The samples were spread-plated onto MOX agar, incubated at 37° for 24 hours. After the 24 hours, plate counts were reported in Log10CFU / cm2.

[0123] Statistical analysis. The statistical analysis consisted of a robust three-way ANOVA, which examined the effects of Treatment, Timepoint, and Surfaces (along with their interactions) on bacterial counts, measured as log CFU / cm2. An individual ANOVA was performed for each LAB strain.

[0124] JI 9 Strain Experiment. The reduction of L. monocytogenes counts based on the effect of each treatment can be observed in Table 4.

[0125] Table 4. Listeria monocytogenes counts with the mean Logw CFU / mL, standard deviation, and Logw reduction for Control, 107J 19, 106J 19, 105JI 9, from hour 0 to hour 24 in stainless steel.

[0126] *Negative reductions symbolize an increase in average logs, which may be affected by outliers.

[0127] Table 4 demonstrates the average reduction of Listeria per treatment on a stainless steel surface and Table 5 demonstrates the reduction on drains.

[0128] Table 5. Listeria monocytogenes counts with the mean Logw CFU / mL, standard deviation, and Logw reduction for Control, 107JI 9, 106J 19, 105JI 9, from hour 0 to hour 24 in drains.

[0129] FIG. 11 demonstrates the effect of the treatments through time on stainless steel while FIG. 12 focuses on the effect in drains. As can be observed in Table 2 and FIGS. 11 and 12, the control consistently had higher counts than the treatments with interventions. Both stainless steel samples and drain samples presented a similar pattern, with drains providing slightly higher counts. While the counts were higher at 0 hours, they significantly decreased by hour 6. By 24 hours, the counts slightly increased compared to hour 6 but still maintained lower values than hour 0. This indicates that the JI 9 strain might have an initial inhibitory effect on bacterial counts, reducing them within the first 6 hours, but this effect diminishes over time, allowing the counts to recover slightly by 24 hours. Compared to the negative control, all treatments were able to reduce Listeria, with the lower concentrations providing better results. As clearly stated in Table 2, most treatments were able to reduce up to 1 log in the different surfaces.

[0130] LI 5 Strain Experiment. The reduction of L. monocytogenes counts based on the effect of each treatment can be observed in Table 6.

[0131] Table 6. Listeria monocytogenes counts with the mean Logw CFU / mL, standard deviation, and Logw reduction for Control, Treatment 1 (1 : 10 L15), Treatment 2 (1 : 100 L15), and Treatment 3 (1 : 1000 L15) from hour 0 to hour 24 on stainless steel.

[0132] Table 6 demonstrates the average reduction of Listeria per treatment on stainless steel and Table 7 demonstrates average reduction on drains.

[0133] Table 7. Listeria monocytogenes counts with the mean Logw CFU / mL, standard deviation, and Logw reduction for Control, Treatment 1 (1 : 10 L15), Treatment 2 (1 : 100 L15), and Treatment 3 (1 : 1000 LI 5) from hour 0 to hour 24 on drains.

[0134] FIG. 13 demonstrates the effect of the treatments through time on stainless steel while FIG. 14 focuses on the effect in drains.

[0135] Similar to JI 9, the control consistently provided higher counts than the treatments with interventions, which can be observed in Table 3 and FIGS. 13 and 14. Nonetheless, the overall reduction of L. monocytogenes was significantly lower than with JI 9. The treatments with the lower concentration of LI 5 also provided better results with a more significant reduction of Listeria. While the effect of LI 5 treatments on drains mostly provided a decrease at 6 hours and an increase by 24 hours, the stainless steel samples provided a more gradual decrease of Listeria counts from hour 0 to hour 24. While the reduction of Listeria counts varied more with the LI 5 strain compared to the J19 strain, it was able to reduce more logs in some cases. For example, the LI 5 strain was able to reduce up to 2.67 logs by hour 6 in a drain surface. In one aspect, thereduction in pathogenic bacteria is at least a Logw reduction of 0.5, 0.81, 0.87, 1.01, 1.06, 1.14, 1.21, 1.30, 1.33, 1.36, 1.54, 1.73, 1.81, 1.95, or 2.31.

[0136] This example shows that the treatments significantly affected bacterial counts (independently from the strain of LAB). Both strains are effective as interventions, but LI 5 appears to exert a more pronounced effect, possibly due to the greater differences between treatments, even if the difference isn’t significant. In conclusion, J19 provides a consistent, broadly effective intervention, while LI 5 offers a more adaptable but variable approach, with treatment effects strongly influenced by time and surface.

[0137] It is contemplated that any aspects of the disclosure discussed in this specification can be implemented with respect to any method, kit, reagent, or composition of the disclosure, and vice versa. Furthermore, compositions of the disclosure can be used to achieve methods of the disclosure.

[0138] It will be understood that particular aspects described herein are shown by way of illustration and not as limitations of the disclosure. The principal features of this disclosure can be employed in various aspects without departing from the scope of the disclosure. Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, numerous equivalents to the specific procedures described herein. Such equivalents are considered to be within the scope of this disclosure and are covered by the claims.

[0139] All publications and patent applications mentioned in the specification are indicative of the level of skill of those skilled in the art to which this disclosure pertains. All publications and patent applications are herein incorporated by reference to the same extent as if each individual publication or patent application was specifically and individually indicated to be incorporated by reference.

[0140] The use of the word “a” or “an” when used in conjunction with the term “comprising” in the claims and / or the specification may mean “one,” but it is also consistent with the meaning of “one or more,” “at least one,” and “one or more than one.” The use of the term “or” in the claims is used to mean “and / or” unless explicitly indicated to refer to alternatives only or the alternatives are mutually exclusive, although the disclosure supports a definition that refers to only alternatives and “and / or.” Throughout this application, the term “about” is used to indicate that a value includes the inherent variation of error for the device, the method being employed to determine the value, or the variation that exists among the study subjects.

[0141] As used in this specification and claim(s), the words “comprising” (and any form of comprising, such as “comprise” and “comprises”), “having” (and any form of having, such as “have” and “has”), “including” (and any form of including, such as “includes” and “include”) or“containing” (and any form of containing, such as “contains” and “contain”) are inclusive or open- ended and do not exclude additional, unrecited elements or method steps. In aspects of any of the compositions and methods provided herein, “comprising” may be replaced with “consisting essentially of’ or “consisting of’. As used herein, the phrase “consisting essentially of’ requires the specified integer(s) or steps as well as those that do not materially affect the character or function of the claimed invention. As used herein, the term “consisting” is used to indicate the presence of the recited integer (e.g., a feature, an element, a characteristic, a property, a method / process step or a limitation) or group of integers (e.g., feature(s), element(s), characteristic(s), propertie(s), method / process steps or limitation(s)) only.

[0142] The term “or combinations thereof’ as used herein refers to all permutations and combinations of the listed items preceding the term. For example, “A, B, C, or combinations thereof’ is intended to include at least one of: A, B, C, AB, AC, BC, or ABC, and if order is important in a particular context, also BA, CA, CB, CBA, BCA, ACB, BAC, or CAB. Continuing with this example, expressly included are combinations that contain repeats of one or more item or term, such as BB, AAA, AB, BBC, AAABCCCC, CBBAAA, CABABB, and so forth. The skilled artisan will understand that typically there is no limit on the number of items or terms in any combination, unless otherwise apparent from the context.

[0143] As used herein, words of approximation such as, without limitation, “about”, "substantial" or "substantially" refers to a condition that when so modified is understood to not necessarily be absolute or perfect but would be considered close enough to those of ordinary skill in the art to warrant designating the condition as being present. The extent to which the description may vary will depend on how great a change can be instituted and still have one of ordinary skilled in the art recognize the modified feature as still having the required characteristics and capabilities of the unmodified feature. In general, but subject to the preceding discussion, a numerical value herein that is modified by a word of approximation such as “about” may vary from the stated value by at least ±1, 2, 3, 4, 5, 6, 7, 10, 12 or 15%.

[0144] Additionally, the section headings herein are provided for consistency with the suggestions under 37 CFR 1.77 or otherwise to provide organizational cues. These headings shall not limit or characterize the disclosure(s) set out in any claims that may issue from this disclosure. Specifically, and by way of example, although the headings refer to a “Field of Invention,” such claims should not be limited by the language under this heading to describe the so-called technical field. Further, a description of technology in the “Background of the Invention” section is not to be construed as an admission that technology is prior art to any disclosure(s) in this disclosure. Neither is the “Summary” to be considered a characterization of the disclosure(s) set forth in issued claims. Furthermore, any reference in this disclosure to “invention” in the singular should not beused to argue that there is only a single point of novelty in this disclosure. Multiple inventions may be set forth according to the limitations of the multiple claims issuing from this disclosure, and such claims accordingly define the invention(s), and their equivalents, that are protected thereby. In all instances, the scope of such claims shall be considered on their own merits in light of this disclosure but should not be constrained by the headings set forth herein.

[0145] All of the compositions and / or methods disclosed and claimed herein can be made and executed without undue experimentation in light of the present disclosure. While the compositions and methods of this disclosure have been described in terms of preferred aspects, it will be apparent to those of skill in the art that variations may be applied to the compositions and / or methods and in the steps or in the sequence of steps of the method described herein without departing from the concept, spirit and scope of the disclosure. All such similar substitutes and modifications apparent to those skilled in the art are deemed to be within the spirit, scope and concept of the disclosure as defined by the appended claims.

[0146] To aid the Patent Office, and any readers of any patent issued on this application in interpreting the claims appended hereto, applicants wish to note that they do not intend any of the appended claims to invoke paragraph 6 of 35 U.S.C. § 112, U.S.C. § 112 paragraph (f), or equivalent, as it exists on the date of filing hereof unless the words “means for” or “step for” are explicitly used in the particular claim.

[0147] For each of the claims, each dependent claim can depend both from the independent claim and from each of the prior dependent claims for each and every claim so long as the prior claim provides a proper antecedent basis for a claim term or element.

Claims

What is claimed is:

1. A method of treating animals or humans comprising, consisting essentially of, or consisting of: feeding or providing a feed, supplement, additive, or food to animals or humans with a composition comprising one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 provided in an amount sufficient to be at least one of: bactericidal against spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation.

2. The method of claim 1, wherein the probiotic bacteria are selected from 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L28, L3A, L20-B, J7, JI 4, J27, J43, J16, J34, J25, L5-A and JI 9.

3. The method of claim 1, wherein the spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, are selected from at least one of: Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enterobacter ssp.; Enterococcus ssp.; Escherichia coli enter oinvasive strains; Escherichia coli enteropathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.; Plesiomonas shigelloides; Salmonella ssp. ; Shigella ssp. ; Staphylococcus aureus; Streptococcus ssp. ; Vibrio cholerae; Yersinia enlerocolilica: and Listeria monocytogenes.

4. The method of claim 1, wherein the probiotic bacteria are provided in an amount selected from 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product.

5. The method of claim 1, wherein a combination of 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L28, L3A, L20-B, J7, J 14, J27, J43, JI 6, J34, J25, L5-A and J19, comprise at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product.

6. The method of claim 1, wherein the composition is formulated into a liquid, a tablet, a paste, a powder, a fermentate, a supernatant, an extract, a pill, or capsule.

7. The method of claim 1, wherein the composition consist essentially of a feed for livestock, poultry, fish, birds, reptiles, or domestic animals that consists of at least 0.001, 0.01,0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30% w / w one or more probiotic bacteria selected from L3A, L20-B, J7, J14, J27, J43, J 16, J34, J25, L5-A and J19 as a replacement for in-feed antibiotics and treatment for gut infection and inflammation, wherein the composition is formulated for oral delivery, or wherein JI 9 formulated for poultry feed.

8. A method of treating an animal or human comprising: providing the animal or human a composition that comprises one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3 A, L.20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 in an amount sufficient to be at least one of: bactericidal against bacteria selected from at least one of: Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp.; Clostridium botulinum; Clostridium perfringens; Enterobacter ssp. ; Enterococcus ssp. ; Escherichia coli enter oinvasive strains; Escherichia coli enteropathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp.; Plesiomonas shigelloides; Salmonella ssp. ; Shigella ssp. ; Staphylococcus aureus; Streptococcus ssp. ; Vibrio cholerae; Yersinia enlerocolilica: and Listeria monocytogenes in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation, wherein the composition is formulated for oral delivery.

9. A composition comprising, consisting essentially of, or consisting of: one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, JI 4, J27, J43, J16, J34, J25, L5-A and J19 in an amount sufficient to be at least one of: bactericidal against bacteria selected from spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, at least one of: in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation.

10. The composition of claim 9, wherein the probiotic bacteria are selected from 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, JI 6, J34, J25, L5-A and JI 9.

11. The composition of claim 9, wherein the probiotic bacteria are provided in an amount selected from 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product.

12. The composition of claim 9, wherein a combination of 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L3A, L20-B, J7, J 14, J27, J43, JI 6, J34, J25, L5-A and J 19, comprise at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product.

13. The composition of claim 9, wherein the composition is formulated into a liquid, a tablet, a paste, a powder, a fermentate, a supernatant, an extract, a pill, or capsule.

14. The composition of claim 9, wherein the one or more probiotic bacteria are effective against Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp. ; Clostridium botulinum; Clostridium perfringens;Enter obacter ssp.; Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enteropathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp. ; Plesiomonas shigelloides; Salmonella ssp. ; Shigella ssp. ; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enter ocolitica; and Listeria monocytogenes .

15. The composition of claim 9, wherein the composition consist essentially of a feed for livestock, poultry, fish, birds, reptiles, or domestic animals that consists of at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30% w / w one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J 14, J27, J43, J16, J34, J25, L5-A and J19 as a replacement for in-feed antibiotics and treatment for gut infection and inflammation, wherein the composition is formulated for oral delivery, or wherein JI 9 formulated for poultry feed.

16. An antimicrobial composition comprising a human probiotic or a food comprising the human probiotic comprising, consisting essentially of, or consisting of: one or more probiotic lactic acid bacteria, supernatants, extracts, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 in an amount sufficient to be at least one of: bactericidal against bacteria selected from at least one of: spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof, in animal or human intestines, improve gut structure, reduce gut inflammation, improved nutrient digestibility, improve mineral absorption and growth performance, reduce carriage, reduce fecal shedding, or reduce gut infection and inflammation.

17. The composition of claim 16, wherein the probiotic bacteria are selected 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the bacterial strains selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19.

18. The composition of claim 16, wherein the probiotic bacteria are provided in an amount selected from 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product.

19. The composition of claim 16, wherein a combination of 2, 3, 4, 5, 6, 7, 8, 9, 10, or 11 of the probiotic bacteria selected from L28, L3A, L20-B, J7, J 14, J27, J43, JI 6, J34, .125, L5-A and J19, comprise at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 40, 50, 60, 70, 76, 80, or 90% weight to weight (w / w) of a final product.

20. The composition of claim 16, wherein the composition is formulated into a liquid, a tablet, a paste, a powder, a fermentate, a supernatant, an extract, a pill, or capsule.

21. The composition of claim 16, wherein the one or more probiotic bacteria are effective against Aeromonas caviae; Aeromonas hydrophila; Aeromonas sobria; Bacillus cereus; Campylobacter jejuni; Citrobacter ssp. ; Clostridium botulinum; Clostridium perfringens;Enter obacter ssp.; Enterococcus ssp.; Escherichia coli enteroinvasive strains; Escherichia coli enteropathogenic strains; Escherichia coli enterotoxigenic strains; Escherichia coli O157:H7; Klebsiella ssp. ; Plesiomonas shigelloides; Salmonella ssp. ; Shigella ssp. ; Staphylococcus aureus; Streptococcus ssp.; Vibrio cholerae; Yersinia enter ocolitica; and Listeria monocytogenes .

22. The composition of claim 16, wherein the composition consist essentially of a feed for livestock, poultry, fish, birds, reptiles, or domestic animals that consists of at least 0.001, 0.01, 0.1, 0.5, 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 1, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29 or 30% w / w one or more probiotic bacteria selected from L28, L3A, L20-B, J7, J14, J27, J43, J16, J34, J25, L5-A and J19 as a replacement for in-feed antibiotics and treatment for gut infection and inflammation, wherein the composition is formulated for oral delivery, or wherein JI 9 formulated for poultry feed.

23. The composition of claim 16, wherein the composition is formulated as a food additive.

24. The composition of claim 16, wherein a food product is fermented with at least one of the L28, L3A, L20-B, J7, 14, J27, J43, J16, J34, J25, L5-A and J19 lactic acid bacteria.

25. The composition of claim 16, wherein the probiotic lactic acid bacteria, extract, or supernatant thereof is bactericidal.

26. A method of treating a surface to eliminate a pathogenic bacteria comprising: contacting a surface with a composition comprising one or more probiotic bacteria, supernatant, extract, broth, or fermentate thereof, selected from L28, L3A, L20-B, J7, J14, J27, J43, JI 6, J34, J25, L5-A and JI 9 provided in an amount sufficient to be at least one of: reduce oreliminate at least one of: spore-forming bacteria, gram positive bacteria, gram negative bacteria, or combinations thereof on the surface.

27. The method of claim 26, wherein the surface is steel, stainless steel, galvanized steel, carbon steel, alloy steel, chrome, steel chrome-pl ated, steel with nickel / silicon carbide composite coating, brass, brass-chrome plated, brass with nickel / silicon carbide composite, stainless chrome-plated, stainless with nickel / silicon carbide composite coating, carbonitrided steel, nickel carbide plated steel, tempered steel, copper, brass, bronze, aluminum, tinplate, manganese, nickel, zinc, titanium, tin, plastic, polymer, fiberglass, polypropylene, polytetrafluorethylene (PTFE), polyvinyl chloride (PVC), and high-density polyethylene, polycarbonate, carbon fiber, a composite material, ceramic, porcelain, coated with a non-stick surface, stone, rock, granite, glass, a coating, a plastic-coated metal, wood, a thermoplastic material, and polyether ether ketone (PEEK), or combinations thereof.

28. The method of claim 26, wherein the surface is a drain, a pipe, a pot, a dish, a bowl, a pan, tableware, a spoon, a fork, a knife, crockery, glasses, cutlery, colanders, mixers, food presses, food extruders, spreaders, countertops, cooking utensils, baking utensils, serving trays, a floor, or combinations thereof.

29. The method of claim 26, wherein the surface is on a kitchen utensil selected from group consisting of biscuit cutter, cake and pie server, bottle opener, tin opener, cork screw, nut cracker, crab cracker, pizza cutter, potato masher, rolling pin, corer, curler, pitter, peeler, sieve, scissors, scraper, shear, slicer, scaler, sifter, scoop, spatula, spider, chopper, squeezer, grater, grinder, blender, whisk, tong, ladle, bowl, dish, plate, platter, saucer, tray, sauce boat, gravy boat, creamer, salt shakers, pepper shaker, salt cellar, sugar bowl, jug, pitcher, cup, mug, glass, teapot, or coffee pot.

30. The method of claim 26, wherein the reduction in pathogenic bacteria is at least a Logw reduction of 0.5, 0.81, 0.87, 1.01, 1.06, 1.14, 1.21, 1.30, 1.33, 1.36, 1.54, 1.73, 1.81, 1.95, or 2.31.

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